EP3349784A1 - Glycotargeting-therapeutika - Google Patents

Glycotargeting-therapeutika

Info

Publication number
EP3349784A1
EP3349784A1 EP16778870.2A EP16778870A EP3349784A1 EP 3349784 A1 EP3349784 A1 EP 3349784A1 EP 16778870 A EP16778870 A EP 16778870A EP 3349784 A1 EP3349784 A1 EP 3349784A1
Authority
EP
European Patent Office
Prior art keywords
formula
compound
antigen
immune response
integer
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Withdrawn
Application number
EP16778870.2A
Other languages
English (en)
French (fr)
Inventor
Jeffrey A. Hubbell
David Scott WILSON
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Ecole Polytechnique Federale de Lausanne EPFL
Original Assignee
Ecole Polytechnique Federale de Lausanne EPFL
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Priority claimed from US14/859,292 external-priority patent/US10946079B2/en
Priority claimed from US15/185,564 external-priority patent/US10046056B2/en
Application filed by Ecole Polytechnique Federale de Lausanne EPFL filed Critical Ecole Polytechnique Federale de Lausanne EPFL
Publication of EP3349784A1 publication Critical patent/EP3349784A1/de
Withdrawn legal-status Critical Current

Links

Classifications

    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/50—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates
    • A61K47/51—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent
    • A61K47/54—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being an organic compound
    • A61K47/549—Sugars, nucleosides, nucleotides or nucleic acids
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides
    • A61K38/16—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • A61K38/17—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
    • A61K38/22—Hormones
    • A61K38/28—Insulins
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • A61K39/0005—Vertebrate antigens
    • A61K39/001—Preparations to induce tolerance to non-self, e.g. prior to transplantation
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • A61K39/35—Allergens
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/50—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates
    • A61K47/51—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent
    • A61K47/56—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being an organic macromolecular compound, e.g. an oligomeric, polymeric or dendrimeric molecule
    • A61K47/58—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being an organic macromolecular compound, e.g. an oligomeric, polymeric or dendrimeric molecule obtained by reactions only involving carbon-to-carbon unsaturated bonds, e.g. poly[meth]acrylate, polyacrylamide, polystyrene, polyvinylpyrrolidone, polyvinylalcohol or polystyrene sulfonic acid resin
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P17/00—Drugs for dermatological disorders
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P19/00—Drugs for skeletal disorders
    • A61P19/02—Drugs for skeletal disorders for joint disorders, e.g. arthritis, arthrosis
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P25/00—Drugs for disorders of the nervous system
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P27/00—Drugs for disorders of the senses
    • A61P27/02—Ophthalmic agents
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P29/00—Non-central analgesic, antipyretic or antiinflammatory agents, e.g. antirheumatic agents; Non-steroidal antiinflammatory drugs [NSAID]
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P3/00—Drugs for disorders of the metabolism
    • A61P3/08—Drugs for disorders of the metabolism for glucose homeostasis
    • A61P3/10—Drugs for disorders of the metabolism for glucose homeostasis for hyperglycaemia, e.g. antidiabetics
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P37/00—Drugs for immunological or allergic disorders
    • A61P37/02—Immunomodulators
    • A61P37/06—Immunosuppressants, e.g. drugs for graft rejection
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P37/00—Drugs for immunological or allergic disorders
    • A61P37/08—Antiallergic agents
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • A61K2039/57—Medicinal preparations containing antigens or antibodies characterised by the type of response, e.g. Th1, Th2
    • A61K2039/577—Medicinal preparations containing antigens or antibodies characterised by the type of response, e.g. Th1, Th2 tolerising response
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K2319/00—Fusion polypeptide
    • C07K2319/01—Fusion polypeptide containing a localisation/targetting motif

Definitions

  • a Sequence Listing submitted as an ASCII text file via EFS-Web is hereby incorporated by reference in accordance with 35 U.S.C. ⁇ 1.52(e).
  • the name of the ASCII text file for the Sequence Listing is ANOK001 P1WO_ST25.TXT, the date of creation of the ASCII text file is September 14, 2016, and the size of the ASCII text file is 47.4 KB.
  • compositions that are useful in the treatment of transplant rejection, autoimmune disease, allergy (e.g., food allergy), and immune response against a therapeutic agent.
  • compositions configured to target one or more cell types in the liver and, as a result, deliver an antigen to which tolerance is desired to the one or more cell types targeted, and thereby induce a processing of the antigen and induce immune tolerance to the antigen.
  • the antigen as disclosed in more detail below, can comprise a therapeutic agent, a protein, a protein fragment, an antigenic mimic of a protein or protein fragment (e.g., a mimotope). Additional types of antigens are discussed in more detail below. Methods and uses of such compositions are also provided for, in several embodiments.
  • a compound comprising Formula 1 :
  • X comprises molecule comprising an antigenic region
  • Y is of a linker moiety having a formula selected from the group consisting of:
  • n is an integer from about 1 to 100
  • present p is an integer from about 2 to 150
  • present q is an integer from about 1 to 44
  • present R 8 is -CH 2 - or -CH 2 -CH 2 -C(CH 3 )(CN)-
  • present R 9 is a direct bond or -CH 2 -CH 2 — NH-C(O)-
  • Z comprises a liver-targeting moiety.
  • X is a protein or protein fragment comprising an antigenic region.
  • Z is galactose, while in some embodiments Z is glucose.
  • Z is galactosamine, while in some embodiments Z is glucosamine.
  • the alpha anomer of glucose or galactose is used in the composition.
  • the beta anomer of glucose or galactose is used in the composition.
  • mixtures of the alpha and beta anomers are used, including optionally mixtures of glucose and galactose.
  • Z is N-acetylgalactosamine, while in some embodiments Z is N-acetylglucosamine.
  • the alpha anomer of glucosamine or galactosamine is used in the composition.
  • the alpha anomer of glucosamine or galactosamine is used in the composition. In several embodiments, mixtures of the alpha and beta anomers are used, including optionally mixtures of glucosamine and galactosamine. Likewise, in several embodiments the alpha anomer, the beta anomer, or combinations of the alpha and beta anomers of N-acetylgalactosamine or N- acetylglucosamine. Combinations of any of the alpha or beta anomeric forms of any of the liver targeting sugars, and any combinations of the sugars can be employed in various embodiments.
  • Y comprises Y m or Y n , m is between 1 and 5, n is between 75 and 85, p is between 85 and 95, and q is between 2 and 6. In several embodiments m is between 1 and 3, n is 79, p is 90, and q is 4.
  • X is selected from the group consisting of insulin, proinsulin, preproinsulin, gluten, gliadin, myelin basic protein, myelin oligodendrocyte glycoprotein and proteolipid protein, Factor VIII, Factor IX, asparaginase, uricase and fragments of any of the preceding. In several embodiments, the antigen X is not a full length protein.
  • the antigen is not full length gliadin, insulin, or proinsulin.
  • the antigen X is not a fragment of a protein.
  • m is not greater than 3
  • n is not greater than 80
  • p is not greater than 100
  • q is not more than 5.
  • Y is a linker moiety having a formula of:
  • Y is a linker moiety having a formula of:
  • Y is a linker moiety having a formula of:
  • combinations of the linkers disclosed herein may be used, just as combinations of the liver targeting moieties can be employed.
  • antigens to which tolerance may be desired. These may include, but are not limited to, exogenous antigens that result in an adverse immune response when a subject is exposed to the antigen.
  • the adverse immune response could be a result of ingestion of the antigen, e.g., orally or nasally, or via some other mucosal route. These routes could be the case, for example, with food antigens.
  • the antigen may be purposefully administered to a subject, for example, with the administration of a therapeutic composition to treat a disease or condition that the subject is affected by.
  • the antigen may be produced by the subject, e.g., an autoimmune antigen.
  • X comprises a foreign transplant antigen against which transplant recipients develop an unwanted immune response or a tolerogenic portion thereof.
  • X comprises a foreign food, animal, plant or environmental antigen against which patients develop an unwanted immune response or a tolerogenic portion thereof.
  • X comprises a foreign therapeutic agent against which patients develop an unwanted immune response or a tolerogenic portion thereof.
  • X comprises a synthetic self-antigen against the endogenous version of which patients develop an unwanted immune response or a tolerogenic portion thereof.
  • X is a food antigen.
  • X is one or more of conarachin (Ara h 1), allergen II (Ara h 2), arachis agglutinin, conglutin (Ara h 6), a-lactalbumin (ALA), lactotransferrin, Pen a 1 allergen (Pen a 1 ), allergen Pen m 2 (Pen m 2), tropomyosin fast isoform, high molecular weight glutenin, low molecular weight glutenin, alpha- gliadin, gamma-gliadin, omega-gliadin, hordein, seclain, and avenin.
  • X is selected from the group consisting of gluten, high molecular weight glutenin, low molecular weight glutenin, alpha- gliadin, gamma-gliadin, omega-gliadin, hordein, seclain, and avenin and fragments thereof.
  • X is selected from the group consisting of gluten, high molecular weight glutenin, low molecular weight glutenin, alpha- gliadin, gamma-gliadin, and omega-gliadin and fragments thereof.
  • X is gluten or fragment thereof.
  • X is gliadin or fragment thereof.
  • X is a therapeutic agent.
  • X is selected from the group consisting of Factor VII, Factor IX, asparaginase, and uricase and fragments thereof.
  • X is a therapeutic agent selected from the group consisting of Factor VII and Factor IX and fragments thereof.
  • X is a therapeutic agent selected from the group consisting of Factor VIII or fragment thereof.
  • the compound when X is a therapeutic agent, can be used in the treatment, prevention, reduction, or otherwise amelioration of an immune response developed against a therapeutic agent for hemophilia.
  • mimotopes of any antigenic portion of the antigens above can be used in several embodiments.
  • X comprises asparaginase or a fragment thereof. In several embodiments, X comprises uricase or a fragment thereof. In several such embodiments, the compound can be used in the treatment, prevention, reduction, or otherwise amelioration of an immune response developed against an anti-neoplastic agent. As discussed herein, mimotopes of any antigenic portion of the antigens above can be used in several embodiments.
  • X is associated with an autoimmune disease.
  • the associated autoimmune disease is one or more of Type I diabetes, multiple sclerosis, rheumatoid arthritis, vitiligo, uveitis, pemphis vulgaris and neuromyelitis optica.
  • the autoimmune disease is Type I diabetes and X comprises insulin or a fragment thereof. In several embodiments, the autoimmune disease is Type I diabetes and X comprises proinsulin or a fragment thereof. In several embodiments, the autoimmune disease is Type I diabetes and X comprises preproinsulin or a fragment thereof.
  • mimotopes of any antigenic portion of the antigens above can be used in several embodiments. In several embodiments, combinations of these antigens can be incorporated into the tolerogenic compound which may aid in reducing immune responses to self-antigens at multiple points along the insulin pathway.
  • the autoimmune disease is multiple sclerosis and X comprises myelin basic protein or a fragment thereof. In several embodiments, the autoimmune disease is multiple sclerosis and X comprises myelin oligodendrocyte glycoprotein or a fragment thereof. In several embodiments, the autoimmune disease is multiple sclerosis and X comprises myelin proteolipid protein or a fragment thereof.
  • mimotopes of any antigenic portion of the antigens above can be used in several embodiments. In several embodiments, combinations of these antigens can be incorporated into the tolerogenic compound which may aid in reducing immune responses to self- antigens at multiple points along the enzymatic pathways that control myelination or myelin repair.
  • the autoimmune disease is rheumatoid arthritis and X is selected from the group consisting of fibrinogen, vimentin, collagen type II, alpha enolase and fragments thereof.
  • the autoimmune disease is vitiligo and X is selected from the group consisting of Pmel17, tyrosinase and fragments thereof.
  • the autoimmune disease is uveitis and X is selected from the group consisting of retinal arrestin and interphotoreceptor retinoid-binding protein (IRBP) and fragments thereof.
  • IRBP interphotoreceptor retinoid-binding protein
  • the autoimmune disease is pemphigus vulgaris and X is selected from the group consisting of desmoglein 3, 1 and 4, pemphaxin, desmocollins, plakoglobin, perplakin, desmoplakins, acetylcholine receptor and fragments thereof.
  • the autoimmune disease is neuromyelitis optica and X is aquaporin-4 or a fragment thereof.
  • mimotopes of any antigenic portion of the self-antigens above can be used in several embodiments.
  • compositions comprising a compound disclosed above (or otherwise disclosed herein).
  • the pharmaceutically acceptable composition consists of, or consists essentially of a compound wherein X is a food antigen, therapeutic agent, a self antigen, or fragment thereof, a linker Y, and a liver targeting moiety Z selected from glucose, galactose, glucosamine, galactosamine, N-acetylglucosamine, and N- acetylgalactosamine.
  • the compound is administered prior to the subject being exposed to the antigen.
  • the compound is administered after the subject has been exposed to the antigen.
  • the administration comprises at least one intravenous administration of the compound (e.g., a bolus dose followed by a series of optional maintenance doses).
  • compositions for inducing immune tolerance in a subject and methods and uses of the compositions for achieving the same.
  • immune tolerance is desired because a subject develops an unwanted immune response to an antigen.
  • the antigen may be one or more of a variety of antigens, for example a foreign antigen such as a food antigen that is ingested, or an antigenic portion of a therapeutic drug given to a subject.
  • the antigen may be a self-antigen that the subject's immune system fails to recognize (or only recognizes as self to a limited degree) and therefore mounts an immune response against, leading to autoimmune disorders.
  • composition comprising Formula 1 :
  • X comprises a food antigen, a therapeutic agent, a self- antigen, a fragment of any of such antigens, or a mimotope of any of such antigens
  • Y is of a linker moiety having the following formula:
  • n is an integer from about 70 to 85
  • present p is an integer from about 85 to 95
  • present q is an integer from about 1 to 10
  • present R 8 is -CH 2 - or -CH 2 -CH 2 -C(CH 3 )(CN)-
  • present R 9 is a direct bond or -CH 2 -CH 2 — NH-C(O)-
  • Z comprises a liver-targeting moiety comprising glucose or galactose.
  • m is between 1 and 3
  • n is 79
  • p is 90
  • q is 4.
  • X is selected from the group consisting of insulin, proinsulin, preproinsulin, gluten, gliadin, myelin basic protein, myelin oligodendrocyte glycoprotein and proteolipid protein, Factor VIII, Factor IX, asparaginase, uricase and fragments of any of the preceding.
  • the composition comprises, consists of, or consists essentially of the antigen X, the linker Y and the liver targeting moiety Z.
  • X is selected from the group consisting of insulin, proinsulin, preproinsulin, gluten, gliadin, myelin basic protein, myelin oligodendrocyte glycoprotein and proteolipid protein, Factor VIII, Factor IX, asparaginase, uricase and fragments of any of the preceding
  • Y is of a linker moiety having the following formula:
  • n is an integer from about 70 to 85, where present p is an integer from about 85 to 95, where present q is an integer from about 1 to 10, where present R 8 is -CH 2 - or -CH 2 -CH 2 -C(CH 3 )(CN)-; and where present R 9 is a direct bond or -CH 2 -CH 2 — NH-C(O)-, and Z comprises a liver-targeting moiety comprising a sugar moiety.
  • m is between 1 and 3
  • n is 79
  • p is 90
  • q is 4.
  • Z is selected from the group consisting of glucose, glucosamine, galactose, galactosamine, N-acetylgalactosamine and N-acetylglucosamine.
  • 2,5-dioxopyrrolidin-1 -yl propyl carbonate-linkers and/or 2- (ethyldisulfanyl)ethyl ethylcarbamate-linkers can be used.
  • composition comprising a compound of
  • n is an integer from about 1 to 10;
  • X comprises an antigen to which patients develop an unwanted immune response, wherein the antigen is a food antigen, a therapeutic agent, a self-antigen, or a fragment of any of such antigens;
  • Y is of a linker moiety having a formula selected from the group consisting of:
  • Formula YL wherein the left bracket "(" indicates a bond to X, where present the right ")” indicates a bond to Z, where present the bottom ")" indicates a bond to Z, where present n is an integer from about 1 to about 80, where present q is an integer from about 1 to about 4, where present p is an integer from about 1 to about 90, where present R 8 is -CH 2 - or -CH2-CH2-C(CH 3 )(CN)-, and Z comprises one or more liver- targeting moieties that specifically target liver cells expressing asialoglycoprotein receptors.
  • m is 1 to 4
  • Y is of a linker moiety having a formula of:
  • ci- torm i na Y anc l ⁇ comprises a liver-targeting moiety comprising one or more of galactose, galactosamine, or N-acetyl galactosamine.
  • n is resolved to an integer from 1 to 4
  • Y is of a linker moiety having a
  • Formula Ya and Z comprises a liver-targeting moiety comprising one or more of glucose, glucosamine, or N-acetyl glucosamine.
  • compositions of Formula 1 (X-f— Y— Z] m ), where m is an integer from about 1 to 100, X comprises an antigen against which a patient develops an unwanted immune response, or a tolerogenic portion thereof or X comprises an antibody, antibody fragment or ligand that specifically binds a circulating protein or peptide or antibody, which circulating protein or peptide or antibody is causatively involved in transplant rejection, immune response against a therapeutic agent, autoimmune disease, hypersensitivity and/or allergy, Y comprises a linker moiety, and Z comprises a liver-targeting moiety.
  • Z comprises galactose, galactosamine, N-acetylgalactosamine, glucose, glucosamine or N-acetylglucosamine.
  • Y is selected from N-hydroxysuccinamidyl linkers, malaemide linkers, vinylsulfone linkers, pyridyl di-thiol-poly(ethylene glycol) linkers, pyridyl di-thiol linkers, n- nitrophenyl carbonate linkers, NHS-ester linkers, and nitrophenoxy poly(ethylene glycol)ester linkers.
  • Y comprises an antibody, antibody fragment, peptide or other ligand that specifically binds X, a disulfanyl ethyl ester, a structure represented by one of Formulae Ya to Yp:
  • Y has a portion represented by Formula Y'-CMP:
  • the left bracket "(" indicates the bond between X and Y
  • the right or bottom bracket and ")" indicates the bond between Y and Z
  • n is an integer from about 1 to 100
  • q is an integer from about 1 to 44
  • R 8 is -CH 2 - or -CH 2 -CH 2 -C(CH 3 )(CN)-
  • Y' represents the remaining portion of Y
  • W represents a polymer of the same W 1 group, or W is a copolymer or a random copolymer of the same or different W 1 and W 2 groups, where:
  • R is a direct bond, -CH 2 -CH 2 — NH-C(O)- or -CH 2 -CH2-(0-CH2-CH 2 )t-NH-C(0)-, t is an integer from 1 to 5; and R 1 ° is an aliphatic group, an alcohol or an aliphatic alcohol.
  • m is 1 to 3
  • Y is represented by Formula Ym, wherein R 8 is -CH 2 -CH 2 -C(CH 3 )(CN)-, and W is represented by a block copolymer of W 1 and W 2 where R 9 is -CH 2 -CH 2 -(0-CH 2 -CH 2 ) r NH-C(0)-, t is 1 , and R 1 ° is 2-hydroxypropyl; and Z comprises a liver- targeting moiety comprising one or more of galactose, galactosamine, N-acetylgalactosamine, glucose, glucosamine, N-acetylglucosamine. In several embodiments, Z is the ⁇ -anomer of the corresponding sugar.
  • compositions for inducing tolerance to an antigen to which a subject develops an unwanted immune response, the compositions comprising a compound of Formula 1 (Formula 1 (X-f-Y— Z] m ), where m is an integer from about 1 to 10, X comprises an antigen to which patients develop an unwanted immune response, wherein the antigen is a food antigen, a therapeutic agent, a self-antigen, or a fragment of any of such antigens, Y is of a linker moiety having a formula selected from the group consisting of:
  • n is an integer from about 1 to 100
  • present p is an integer from about 2 to 150
  • present q is an integer from about 1 to 44
  • present R 8 is -CH 2 - or -CH2-CH2-C(CH 3 )(CN)-
  • present R 9 is a direct bond or -CH 2 -CH 2 — NH-C(O)-
  • Z comprises galactose, galactosamine, or N-acetylgalactosamine.
  • m is 1 to 3
  • Y is of a linker moiety having a formula of:
  • Z comprises a liver-targeting moiety comprising one or more of galactose, galactosamine, or N-acetylgalactosamine.
  • Z is the ⁇ -anomer of the selected moiety.
  • X is a self-antigen and the unwanted immune response is an autoimmune response.
  • X is myelin oligodendrocyte glycoprotein or myelin proteolipid protein.
  • the unwanted immune response experienced by the subject is associated with multiple sclerosis.
  • X is insulin, proinsulin, or preproinsulin and wherein the unwanted immune response is associated with diabetes mellitus. It shall be appreciated that being associated with multiple sclerosis, diabetes mellitus or other auto-immune disease need not necessarily require a formal diagnosis of such auto-immune condition, but rather can be associated with common symptoms or characteristics of a particular auto-immune disorder.
  • an unwanted immune response can be raised against a therapeutic agent, such as a protein drug or drug derived from non-human and/or non- mammalian species.
  • a therapeutic agent such as a protein drug or drug derived from non-human and/or non- mammalian species.
  • X is a therapeutic agent, such as Factor VIII, Factor IX, or other hemostasis-inducing agent.
  • the unwanted immune response is against the agent and the associated disease is hemophilia, which fails to improve (in the absence of the composition) because of the autoimmune response.
  • the hemophilia can improve because the composition aids in inducing tolerance to the agent, reducing the response to agent, and allowing reduced symptoms of hemophilia.
  • X is a therapeutic agent such as asparaginase and uricase.
  • an unwanted immune response can result from administration of such agents, as they are derived from non-human sources.
  • the ability of the compositions disclosed herein to induce tolerance to these agents allows these agents to continue to be used by a subject in need of therapy, while the side effects from an immune reaction are reduced, lessened, eliminated or otherwise ameliorated.
  • X is a food antigen.
  • Many food antigens are known to cause allergies upon ingestion, however, in several embodiments, X is selected from the group consisting of conarachin (Ara h 1 ), allergen II (Ara h 2), arachis agglutinin, conglutin (Ara h 6), a-lactalbumin (ALA), lactotransferrin, Pen a 1 allergen (Pen a 1), allergen Pen m 2 (Pen m 2), tropomyosin fast isoform, high molecular weight glutenin, low molecular weight glutenin, alpha- gliadin, gamma-gliadin, omega-gliadin, hordein, seclain, and avenin.
  • treatment with the compositions disclosed herein where X is a food antigen allows the subject to have a significantly reduced immune response to the antigen, e.g., many peanut allergies are so severe that exposure to peanut dust or oil can cause anaphylaxis.
  • treatment reduces and/or eliminates responses to such incidental exposure to the antigen.
  • treatment allows the subject to ingest the food from which the antigen is derived with limited or no adverse immune response.
  • administration of the composition to the subject results in a greater degree of proliferation of antigen-specific T cells as compared to proliferation of antigen-specific T cells resulting from administration of the antigen alone.
  • the proliferation of antigen- specific T cells indicates that delivery of the antigen (via the composition) to the molecular processing machinery that processes antigens as self/non-self is enhanced versus administration of the antigen alone. In other words, in such embodiments the targeted delivery is effective.
  • compositions disclosed herein results in a greater expression of exhaustion markers or markers of apoptosis on antigen-specific T cells as compared to expression of exhaustion markers or markers of apoptosis on antigen-specific T cells resulting from administration of the antigen alone. This result in indicative of specific reduction in activity of T cells directed against the antigen of interest and/or deletion of T cells directed against the antigen of interest.
  • these molecular hallmarks of induction of tolerance are the precursor of the reduction or amelioration of immune response symptoms that the subject would have previously experienced when exposed to the antigen.
  • Z comprises a liver-targeting moiety that is a carbohydrate.
  • the carbohydrate is a short-chain carbohydrate.
  • Z is a sugar.
  • Z is galactose, galactosamine, N-acetylgalactosamine, glucose, glucosamine, or N-acetylglucosamine.
  • the induction of immune tolerance is greater when a glucose, glucosamine, or N-acetylglucosamine is used for Z.
  • enhancements in induction of immune tolerance can be achieved when the liver targeting moiety is a sugar and the sugar is in the ⁇ -anomer configuration.
  • Z is galactose, galactosamine, N-acetylgalactosamine, glucose, glucosamine, or N-acetylglucosamine and conjugated at its C1 , C2 or C6 to Y.
  • compositions can be via a variety of methods, including, but not limited to intravenous, intramuscular, oral, transdermal, or other infusion route. Administration can be daily, weekly, multiple times per day, or on an as needed basis (e.g., prior to an anticipated exposure).
  • compositions disclosed herein for the treatment of unwanted immune responses after exposure to an antigen.
  • such use can be for prophylactic effects and/or for reducing symptoms from prior exposure to antigens (or prior adverse immune effects, such as those in the auto-immune setting).
  • compositions according to Formula 1 for the treatment of unwanted side effects due to exposure to a therapeutic antigen, exposure to a food antigen, or an adverse effect from an immune response against a self-antigen.
  • the compositions disclosed herein are suitable for administration to a subject in conjunction with such use, for example by oral, IV, IM, or other suitable route. Uses of the compositions disclosed herein, in several embodiments, unexpectedly result in the reduction, elimination or amelioration of adverse immune responses to antigens of interest.
  • compositions and methods of using them are provided herein.
  • a pharmaceutically acceptable composition for inducing tolerance to a therapeutic protein in a subject having an deficiency in production of a functional analogous native protein comprising a compound of Formula 1 (X-f— Y— Z] m ), where m is an integer from about 1 to 10, X comprises an antigenic protein or protein fragment, Y is of a linker moiety having a formula selected from the group consisting of Formula Ya, Formula Yc, Formula Ym, Formula Yn, wherein, the left bracket "(" indicates a bond to X, the right or bottom bracket and ")" indicates the bond between Y and Z, n is an integer from about 1 to 100, where present p is an integer from about 2 to 150, where present q is an integer from about 1 to 44, where present R 8 is -CH 2 - or -CH 2 -CH 2 -C(CH 3 )(CN)-, where present
  • Y is of a linker moiety having a formula of: wherein CH 2 -CH 2 — NH-C(O)-, and Z comprises a liver-targeting moiety comprising one or more of glucose, glucosamine, N- acetylglucosamine, galactose, galactosamine, or N-acetylgalactosamine.
  • the galactose, galactosamine, or N-acetylgalactosamine are the ⁇ -anomers.
  • combinations of galactose, galactosamine, N-acetylgalactosamine, glucose, glucosamine, or N- acetylglucosamine are used.
  • a pharmaceutically acceptable composition for inducing tolerance to a therapeutic protein in a subject having an deficiency in production of a functional analogous native protein comprising a compound of Formula 1 (X- ⁇ -Y— Z] m ), where m is an integer from about 1 to 10, X comprises a antigenic protein or protein fragment, Y is of a linker moiety having a formula selected from the group consisting of Formula Ya, Formula Yc, Formula Ym, or Formula Ym, wherein the left bracket "(" indicates a bond to X, where present the right ")" indicates a bond to Z, where present the bottom ")" indicates a bond to Z, where present n is an integer from about 1 to about 80, where present q is an integer from about 1 to about 4, where present p is an integer from about 1 to about 90, where present R 8 is -CH 2 - or -CH 2 -CH 2 -C(CH 3 )(CN)-, and where present W represents
  • R 10 is an aliphatic group, an alcohol or an aliphatic alcohol
  • Z comprises glucose, glucosamine, N-acetylglucosamine, galactose, galactosamine, or N-acetylgalactosamine.
  • the galactose, galactosamine, or N-acetylgalactosamine are the ⁇ -anomers.
  • combinations of galactose, galactosamine, N-acetylgalactosamine, glucose, glucosamine, or N-acetylglucosamine are used.
  • m is 1 to 3
  • Y is represented by Formula Ym, wherein R 8 is -CH 2 -CH 2 -C(CH 3 )(CN)-, and W is represented by a block copolymer of W 1 and W 2 where R 9 is -CH2-CH 2 -(0-CH 2 -CH 2 )t-NH-C(0)-, t is 1 , and R 10 is 2-hydroxypropyl; and Z comprises a liver- targeting moiety comprising one or more of glucose, glucosamine, N-acetylglucosamine, galactose, galactosamine, or N-acetylgalactosamine.
  • the galactose, galactosamine, or N- acetylgalactosamine are the ⁇ -anomers. In several embodiments, combinations of galactose, galactosamine, N-acetylgalactosamine, glucose, glucosamine, or N-acetylglucosamine are used.
  • X comprises an antigenic region of myelin basic protein, myelin oligodendrocyte glycoprotein, or myelin proteolipid protein. In additional embodiments, X comprises an antigenic region of Factor VIII, Factor IX, insulin, uricase, PAL, or asparaginase.
  • X comprises a foreign antigen such as conarachin (Ara h 1 ), allergen II (Ara h 2), arachis agglutinin, conglutin (Ara h 6), a-lactalbumin (ALA), lactotransferrin, Pen a 1 allergen (Pen a 1 ), allergen Pen m 2 (Pen m 2), tropomyosin fast isoform, high molecular weight glutenin, low molecular weight glutenin, alpha- gliadin, gamma-gliadin, omega-gliadin, hordein, seclain, and avenin.
  • a foreign antigen such as conarachin (Ara h 1 ), allergen II (Ara h 2), arachis agglutinin, conglutin (Ara h 6), a-lactalbumin (ALA), lactotransferrin, Pen a 1 allergen (Pen a 1 ), allergen Pen
  • compositions comprising a compound of Formula 1 (X-[- Y— Z]m), where m is an integer from about 1 to 100, X comprises an antigen against which a patient develops an unwanted immune response, or a tolerogenic portion thereof, or X comprises an antibody, antibody fragment or ligand that specifically binds a circulating protein or peptide or antibody, which circulating protein or peptide or antibody is causatively involved in transplant rejection, immune response against a therapeutic agent, autoimmune disease, hypersensitivity and/or allergy, Y comprises a linker moiety, and Z comprises a liver-targeting moiety.
  • X comprises an antigen against which a patient develops an unwanted immune response, or a tolerogenic portion thereof
  • X comprises an antibody, antibody fragment or ligand that specifically binds a circulating protein or peptide or antibody, which circulating protein or peptide or antibody is causatively involved in transplant rejection, immune response against a therapeutic agent, autoimmune disease, hypersensitivity and/or allergy
  • Y comprises
  • Z galactose, galactosamine, N-acetylgalactosamine, glucose, glucosamine or N-acetylglucosamine are examples of galactose, galactosamine,
  • N-acetylgalactosamine, glucose, glucosamine or N-acetylglucosamine may also be used, in several embodiments. Further, in several embodiments, the galactose, galactosamine, N-acetylgalactosamine, glucose, glucosamine or N-acetylglucosamine are optionally the ⁇ anomer. In several embodiments, Z is conjugated at its C1 , C2 or C6 to Y.
  • Y is selected from N-hydroxysuccinamidyl linkers, malaemide linkers, vinylsulfone linkers, pyridyl di-thiol-poly(ethylene glycol) linkers, pyridyl di-thiol linkers, n- nitrophenyl carbonate linkers, NHS-ester linkers, and nitrophenoxy poly(ethylene glycol)ester linkers.
  • Y comprises an antibody, antibody fragment, peptide or other ligand that specifically binds X, a disulfanyl ethyl ester, a structure represented by one of Formulae Ya to Yp, or Y has a portion represented by Formula Y'-CMP:
  • Formula Y'-CMP where the left bracket “(" indicates the bond between X and Y, the right or bottom bracket and ")" indicates the bond between Y and Z, n is an integer from about 1 to 100, q is an integer from about 1 to 44, R 8 is -CH 2 - or -CH 2 -CH 2 -C(CH 3 )(CN)-, Y' represents the remaining portion of Y, and W represents a polymer of the same W 1 group, or W is a copolymer or a random copolymer of the same or different W 1 and W 2 groups, where:
  • R 9 is a direct bond, -CH 2 -CH 2 — NH-C(O)- or -CH 2 -CH 2 -(0-CH2-CH 2 )t-NH-C(0)-, t is an integer from 1 to 5; and R 10 is an aliphatic group, an alcohol or an aliphatic alcohol.
  • n is about 40 to 80, p is about 10 to 100, q is about 3 to 20, R 8 is -CH 2 -CH 2 -C(CH3)(CN)-, when R9 is -CH2-CH2— NH-C(O)-, Z is glucose, galactose, N- acetylgalactosamine or N-acetylglucosamine conjugated at its C1 , and when W is a copolymer, R10 is 2-hydroxypropyl.
  • Y comprises Formula Ya, Formula Yb, Formula Yc, Formula Yf, Formula Yg, Formula Yh, Formula Yi, Formula Yk, Formula Ym or Formula Yn.
  • Y comprises Formula Ya, Formula Yb, Formula Yc, Formula Ym or Formula Yn.
  • Y comprises Formula Ya, Formula Yb, Formula Yc, Formula Ym or Formula Yn.
  • X comprises a foreign transplant antigen against which transplant recipients develop an unwanted immune response, a foreign food, animal, plant or environmental antigen against which patients develop an unwanted immune response, a foreign therapeutic agent against which patients develop an unwanted immune response, or a synthetic self-antigen against the endogenous version of which patients develop an unwanted immune response, or a tolerogenic portion thereof.
  • a foreign transplant antigen against which transplant recipients develop an unwanted immune response a foreign food, animal, plant or environmental antigen against which patients develop an unwanted immune response, a foreign therapeutic agent against which patients develop an unwanted immune response, or a synthetic self-antigen against the endogenous version of which patients develop an unwanted immune response, or a tolerogenic portion thereof.
  • Also provided for herein is methods of treatment for an unwanted immune response against an antigen by administering to a mammal in need of such treatment an effective amount of a composition comprising a compound of Formula 1 (X-[- Y— Z]m), where m is an integer from about 1 to 100, X comprises an antigen against which a patient develops an unwanted immune response, or a tolerogenic portion thereof or X comprises an antibody, antibody fragment or ligand that specifically binds a circulating protein or peptide or antibody, which circulating protein or peptide or antibody is causatively involved in transplant rejection, immune response against a therapeutic agent, autoimmune disease, hypersensitivity and/or allergy, Y comprises a linker moiety, and Z comprises a glucosylated liver-targeting moiety.
  • X comprises an antigen against which a patient develops an unwanted immune response, or a tolerogenic portion thereof
  • Y comprises, an antibody, antibody fragment, peptide or other ligand that specifically binds X, a disulfanyl ethyl ester, a structure represented by one of Formulae Ya to Yp or Y has a portion represented by Formula Y'-CMP where, the left bracket "(" indicates the bond between X and Y, the right or bottom bracket and ")" indicates the bond between Y and Z, n is an integer from about 1 to 100, q is an integer from about 1 to 44, R 8 is - CH 2 - or -CH 2 -CH2-C(CH 3 )(CN)-, Y' represents the remaining portion of Y, and W represents a polymer of the same W 1 group, or W is a copolymer or a random copolymer of the same or different W 1 and W 2 groups, where:
  • X comprises the antibody, antibody fragment or ligand, and the composition is administered for clearance of a circulating protein or peptide or antibody that specifically binds to X, which circulating protein or peptide or antibody is causatively involved in transplant rejection, immune response against a therapeutic agent, autoimmune disease, hypersensitivity and/or allergy.
  • X comprises the antibody, antibody fragment or ligand, and the composition is administered in an amount effective to reduce a concentration of the antibodies that are causatively involved in transplant rejection, immune response against a therapeutic agent, autoimmune disease, hypersensitivity and/or allergy in blood of the patient by at least 50% w/w, as measured at a time between about 12 to about 48 hours after the administration.
  • compositions are administered for tolerization of the patient with respect to antigen moiety X.
  • X comprises a foreign transplant antigen against which transplant recipients develop an unwanted immune response, a foreign food, animal, plant or environmental antigen against which patients develop an unwanted immune response, a foreign therapeutic agent against which patients develop an unwanted immune response, or a synthetic self-antigen against the endogenous version of which patients develop an unwanted immune response, or a tolerogenic portion thereof.
  • compositions comprising a compound of Formula 1 :
  • n is an integer from about 1 to 100;
  • X comprises an antigen against which a patient develops an unwanted immune response, or a tolerogenic portion thereof; or X comprises an antibody, antibody fragment or ligand that specifically binds a circulating protein or peptide or antibody, which circulating protein or peptide or antibody is causatively involved in transplant rejection, immune response against a therapeutic agent, autoimmune disease, hypersensitivity and/or allergy;
  • Y comprises a linker moiety
  • Z comprises a liver-targeting moiety.
  • Z can also comprise galactose, galactosamine, N-acetylgalactosamine, glucose, glucosamine or N-acetylglucosamine, for example, conjugated at its C1 , C2 or C6 to Y.
  • N- acetylglucosamine and glucose bind to different lectin receptors as do N-acetylgalactosamine and galactose.
  • the experimental data indicate that the selection of Z as N-acetylglucosamine leads to elevated levels of regulatory T cell responses compared to those achieved with N-acetylgalactosamine. In several embodiments, this results in unexpectedly enhanced induction of immune tolerance and/or clearance of antigens from the blood of a subject.
  • Y can be selected from N-hydroxysuccinamidyl linkers, malaemide linkers, vinylsulfone linkers, pyridyl di-thiol-poly(ethylene glycol) linkers, pyridyl di-thiol linkers, n-nitrophenyl carbonate linkers, NHS-ester linkers, and nitrophenoxy poly(ethylene glycol)ester linkers.
  • Y can also comprise: an antibody, antibody fragment, peptide or other ligand that specifically binds X; a disulfanyl ethyl ester; a structure represented by one of Formulae Ya to Yp:
  • Y has a portion represented by Formula Y'-CMP:
  • n is an integer from about 1 to 100;
  • q is an integer from about 1 to 44;
  • R 8 is -CH 2 - or -CH 2 -CH 2 -C(CH 3 )(CN)-;
  • Y' represents the remaining portion of Y (e.g., HS-PEG).
  • W represents a polymer of the same W 1 group, or W is a copolymer (preferably a random copolymer) of the same or different W 1 and W 2 groups, where:
  • p is an integer from 2 to about 150;
  • R 9 is a direct bond, -CH 2 -CH 2 — NH-C(O)- (i.e., an ethylacetamido group or "EtAcN”) or -CH 2 -CH 2 -(0-CH 2 -CH 2 ) t -NH-C(0)- (i.e., a pegylated ethylacetamido group or "Et- PEGt-AcN”)
  • t is an integer from 1 to 5, (particularly 1 to 3, and more particularly 1 or 2);
  • R 10 is an aliphatic group, an alcohol or an aliphatic alcohol.
  • R 10 is a C f alkyl or C f alkylOH g where f is independently an integer between 0 and 10 and g is independently an integer between 0 and 10.
  • R 10 is 2- hydroxypropyl.
  • linkers according to Ym yield unexpectedly effective tolerance endpoints.
  • linkers according to formula Yn yield unexpectedly effective tolerance endpoints.
  • formulations of Fl m'-OVA-m ⁇ -nyg-pgo-q ⁇ CMP-poly-iEtPEGiAcN-I NAcGLUao- ran-HPMA 60 achieve particularly effective tolerance-associated endpoints.
  • combinations of these linkers lead to synergistic results and still further unexpected increases in immune tolerance induction.
  • n is about 40 to 80
  • p is about 10 to 100
  • q is about 3 to 20
  • R 8 is -CH 2 -CH 2 -C(CH 3 )(CN)-
  • R 9 is -CH 2 -CH 2 — NH-C(O)-
  • Z is galactose or N- acetylgalactosamine conjugated at its C1.
  • Y comprises Formula Ya, Formula Yb, Formula Yh, Formula Yi, Formula Yk, Formula Ym or Formula Yn, particularly Formula Ya, Formula Yb, Formula Ym or Formula Yn.
  • X can further comprise: a foreign transplant antigen against which transplant recipients develop an unwanted immune response; a foreign food, animal, plant or environmental antigen against which patients develop an unwanted immune response; a foreign therapeutic agent against which patients develop an unwanted immune response; or a synthetic self-antigen against the endogenous version of which patients develop an unwanted immune response, or a tolerogenic portion thereof.
  • the disclosure also pertains to a method of treatment for an unwanted immune response against an antigen by administering to a mammal in need of such treatment an effective amount of a composition comprising a compound of Formula 1 as disclosed herein.
  • the composition can be administered for clearance of a circulating protein or peptide or antibody that specifically binds to antigen moiety X, which circulating protein or peptide or antibody is causatively involved in transplant rejection, immune response against a therapeutic agent, autoimmune disease, hypersensitivity and/or allergy.
  • the composition can be administered in an amount effective to reduce a concentration of the antibodies that are causatively involved in transplant rejection, immune response against a therapeutic agent, autoimmune disease, hypersensitivity and/or allergy in blood of the patient by at least 50% w/w, as measured at a time between about 12 to about 48 hours after the administration.
  • the composition can administered for tolerization of a patient with respect to antigen moiety X.
  • Figs. 1A-1 D are a series of graphs showing differential cellular uptake of galactose conjugates.
  • Figure 1A depicts that F1 aA-PE-m 4 -n 80 (Gal-PE) preferentially targets PE to sinusoidal endothelial cells (LSECs) of the liver.
  • Figure 1 B depicts that F1 aA-PE-m 4 -n 80 (Gal-PE) preferentially targets PE to Kupffer cells (KC) of the liver.
  • Figure 1 C depicts that F1 aA-PE-m 4 -n 80 (Gal-PE) preferentially targets PE to hepatocytes.
  • Figure 1 D depicts that F1 aA-PE-m 4 -n 80 (Gal-PE) preferentially targets PE to other antigen presenting cells (APCs) of the liver.
  • * P ⁇ 0.05.
  • Fig. 2 is a graph showing proliferation of OT-I CD8+ T cells in mice treated with F1 aA-OVA- m 4 -n 80 (Gal-OVA), OVA or saline (i.e. naive), with greatest proliferation seen in the Gal-OVA treated group.
  • FIGs. 3A-3B are a series of graphs depicting data related to marker expression on T cells.
  • Figure 3A shows the percentage of OT-I CD8 + T cells expressing PD-1 ("PD1 +") in generations of proliferating T cells treated with saline, OVA or F1 aA-OVA-m 4 -n 80 (GAL-OVA), with greatest level of PD- 1 in the gal-OVA-treated group.
  • Figure 3B shows the percentage of OT-I CD8 + T cells expressing phosphatidylserine (stained as "Annexin V+”) in generations of proliferating T cells treated with saline, OVA or F1 aA-OVA-m 4 -n 80 (GAL-OVA), with greatest level of Annexin-V+ cells in the gal-OVA-treated group.
  • Fig. 4 is a graph showing that galactose conjugation [F1 aA-OVA-m 4 -n 80 (Gal-OVA)] decreases the immunogenicity of OVA as determined by OVA-specific antibody titers (shown in Ab titers log "1 ).
  • FIG. 5 shows that administration of F1 aA-OVA-m 4 -n 80 (Gal-OVA) in repeated doses over time is able to deplete OVA-specific antibodies from the serum of mice.
  • Figure 6A shows the immune response in mice challenged with OVA and LPS.
  • Figure 6B shows the immune response in mice treated with OVA, while Figure 6C shows the immune response in naive mice.
  • Figures 6D and 6E show that F1 aA-OVA-m 4 -n 80 (mGal-OVA; 6D) and F1 b-OVA-m 1 -n 4 4-p 3 4 (pGal-OVA; 6E) are able to mitigate the OVA-specific immune response in draining lymph nodes after intradermal challenge with OVA and the adjuvant LPS.
  • Fig 6F is from a parent application and does not form a part of the present disclosure.
  • Figs. 7A-7B shows the characterization of F1 aA-OVA-m 4 -n 80 and F1 b-OVA-m 1 -n 44 -p 34 .
  • Fig. 7A shows size-exclusion HPLC traces of F1 aA-OVA-m 4 -n 80 (open triangles), F1 b-OVA-m 1 -n 44 -p 34 (filled circles) and unconjugated OVA (solid line). Shift to the left represents an increase in molecular weight.
  • Fig. 7A-7B shows the characterization of F1 aA-OVA-m 4 -n 80 and F1 b-OVA-m 1 -n 44 -p 34 .
  • Fig. 7A shows size-exclusion HPLC traces of F1 aA-OVA-m 4 -n 80 (open triangles), F1 b-OVA-m 1 -n 44 -p 34 (filled circles) and unconjugated OVA (solid
  • Figs. 8A-8B depict data related to the reduction in antigen-specific immune response after administration of Fl m'-OVA-m ⁇ a-nyg-pgo-q ⁇ CMP-poly-iEtPEGiAcN-I NAcGLUao-ran-HPMAeo [labeled OVA-p(Glu-HPMA) and shown as filled circles] or Fl m'-OVA-m ⁇ -nyg-pgo-q ⁇ CMP-poly-iEtPEGiAcN- 1 NAcGAL 3 o-ran-HPMA6o [labeled OVA-p(Gal-HPMA) and shown as filled diamonds].
  • FIG. 8A depicts flow cytometric detection of OTI CD8+ T-cell populations (CD3e + /CD8a + /CD45.2 + ) quantified from the draining lymph nodes (inguinal and popliteal) 4 days following antigen challenge in CD45.1 + mice.
  • Significant reductions in OT-I CD8+ T-cells were detected following administration of OVA-p(Gal-HPMA) and OVA-p(Glu-HPMA).
  • Fig. 8B depicts flow cytometric detection of OTII CD4+ T-cell populations (CD3e + /CD4 + /CD45.2 + ) quantified from the draining lymph nodes (inguinal and popliteal) 4 d following antigen challenge in CD45.1 + mice.
  • Figs. 9A-9B depict data related to the increase in antigen-specific regulatory T-cells in the lymph nodes and spleen of mice after antigen challenge.
  • Fig. 9A depicts flow cytometric detection of an Fl m'-OVA-mLa-nyg-pgo-q ⁇ CMP-poly-iEtPEdAcN-I NAcGLUso-ran-HPMAeo [labeled OVA-p(Glu-HPMA) and shown as filled circles] and Fl m'-OVA-m ⁇ -nyg-pgo-q ⁇ CMP-poly-iEtPEGiAcN-I NAcGALso-ran- HPMA 60 [labeled OVA-p(Gal-HPMA) and shown as filled diamonds]-induced increase in OTII T-regulator cells (CD3e + CD4+ CD45.2+ CD25+ FoxP3+) collected from the lymph nodes 4 d following antigen challenge in CD45.1 + mice.
  • Fig. 10 depicts flow cytometry data related to a decrease in the percentage of antigen-specific effector cells (IFNy+ OTI CD8+ T-cells (CD3e + CD8a+ CD45.2+ IFNy+) 4 d following antigen challenge in CD45.1 + mice.
  • IFNy+ OTI CD8+ T-cells CD3e + CD8a+ CD45.2+ IFNy+
  • Figs. 1 1A-1 1 B depict data related to T cell deletion and regulation in an OTII adoptive transfer model, in which OTII cells (CD4 + T cells from a CD45.2 + mouse) are adoptively transferred into a CD45.1 + recipient, which is treated with Fl m'-OVA-m ⁇ -nyg-pgo-q ⁇ CMP-poly-iEtPEGiAcN- I NAcGALso-ran-HPMAgo ["OVA-p(Gal-HPMA)”] or F1 m'-OVA-m ⁇ -nyg-pgo-q ⁇ CMP-poly- EtPEGiAcN- I NAcGLUso-ran-HPMAgo ["OVA-p(Glu-HPMA)”], or OVA not linked to a polymer ["OVA”] to induce T regulatory responses and prevent subsequent responses to vaccine-mediated antigen challenge.
  • OTII cells CD4 + T cells from a CD45.2 + mouse
  • pGal-OVA and pGlu-OVA were each administered in other groups at the same dosings of 2.5 pg at day 1 , 2.5 pg at day 4, and 16 pg at day 7 or 7 pg at day 1 , 7 pg at day 4, and 7 pg at day 7, all doses being on an OVA equivalent dose basis.
  • saline was administered on the same days.
  • the recipient mice were then challenged with OVA (10 pg) adjuvanted with lipopolysaccharide (50 ng) by intradermal injection. Characterization of the draining lymph nodes was done on day 19, to allow determination as to whether or not deletion actually took place and whether regulatory T cells were induced from the adoptively transferred cells.
  • Fig. 1 1 A shows the number of OTII cells present after challenge
  • Fig. 1 1 B shows the frequency of FoxP3 + CD25 + cells (markers of T regulatory cells). * and # indicate p ⁇ 0.05, ** and ## indicate p ⁇ 0.01 , and ### indicates P ⁇ 0.001.
  • Figs. 12A-12B depicts data related to T cell deletion and regulation in an OTI adoptive transfer model, in which OTI cells (CD8 + T cells from a CD45.2 + mouse) are adoptively transferred into a CD45.1 + recipient, which is treated with Fl m'-OVA-m ⁇ -nyg-pgo-q ⁇ CMP-poly- EtPEG ⁇ cN-I NAcGALso- ran-HPMA 60 ["OVA-p(Gal-HPMA)"] or F1 m'-OVA-m ⁇ -nyg-pgo-q ⁇ CMP-poly- EtPEG ⁇ cN-l NAcGLU 30 - ran-HPMA 60 [OVA-p(Glu-HPMA)"], or OVA not linked to a polymer ["OVA”] to induce T regulatory responses and prevent subsequent responses to vaccine-mediated antigen challenge.
  • pGal-OVA and pGlu-OVA were each administered in other groups at the same dosings of 2.5 pg at day 1 , 2.5 pg at day 4, and 16 pg at day 7 or 7 pg at day 1 , 7 pg at day 4, and 7 pg at day 7, all doses being on an OVA equivalent dose basis.
  • saline was administered on the same days.
  • the recipient mice were then challenged with OVA (10 pg) adjuvanted with lipopolysaccharide (50 ng) by intradermal injection.
  • Fig. 12A shows the number of OTI cells present after challenge
  • Fig. 12B shows the frequency of IFNy- expressing cells (lack thereof indicating anergy). * and # indicate p ⁇ 0.05, ** and ## indicate p ⁇ 0.01 ).
  • FIG. 13 depicts data related to blood glucose levels. Mice were treated with F1 m , -P31 -m 1 ⁇ -n 7 9-P9o-q4-CMP-poly-(EtPEGiAcN-1 NAcGLU3o-ran-HPMA 60 [labeled P31 -p(Glu-HPMA)], Fl m'-PSI -m ⁇ a-nyg-pgo-q -CMP-poly-iEtPEG ⁇ cN-I NAcGALao-ran-HPMAeo [labeled P31-p(Gal-HPMA) conjugates (or saline).
  • Fig. 14 depicts data related to the generation of spontaneous diabetes in non-obese diabetic (NOD) mice.
  • Mice treated with Fl c'-lnsulin-B-m n ⁇ pgo-CMP-poly- EtPEGiAcN-I NAcGLUso-ran- HPMA 60 are shown as filled squares.
  • Mice treated with Fl c'-lnsulin-B-nvn ⁇ pgo-CMP-poly-iEtPEG ⁇ cN- 1 NAcGALso-ran-HPMAeo are shown as filled triangles.
  • Mice treated with saline are shown as filled diamonds.
  • Treating animals with the compounds of Formula 1 reduced the incidences of diabetes onset in NOD mice as compared to animals treated with saline.
  • Figs. 15A-15B depicts data related to biodistribution of the model antigen OVA tethered to the synthetic glycopolymers, showing uptake in the liver while limiting uptake in the spleen.
  • A Fluorescent signal of perfused livers taken from animals treated with OVA (1 ) or OVA conjugated to various glycopolymers (2-5).
  • B Fluorescent images of spleens taken from animals treated with OVA (1 ) or OVA conjugated to various glycopolymers (2-5).
  • Formulations are as follows: 1. OVA, 2. OVA- p(Ga ⁇ -HPMA), 3. OVA-p(Gal-HPMA), 4. OVA-p(Glu -HPMA), 5. OVA-p(Glu-HPMA).
  • FIGS. 16A-16F depict data related to experiments comparing linker moieties.
  • OVA-p(Gal- HPMA), OVA-p(Glu-HPMA), OVA-p(Gai -HPMA), and OVA-p(Glu -HPMA) conjugates were synthesized and tested for their ability to induce antigen-specific T cell anergy and eliminate the T cell population responsible for long term memory.
  • Fig. 16A shows a schematic of the treatment regimen for 7-day experiment.
  • Fig. 16B depicts the percentage of proliferating OTI splenic T cells as assayed by CSFE dilution.
  • FIG. 16C depicts the percentage of Annexin V+ OTI T cells in the spleens of animals treated with OVA-glycopolymer conjugates or free OVA.
  • Fig. 16D depicts the percentage of PD-1 + splenic OTI cells.
  • Fig. 16E depicts the percentage of T memory cells in the OTI population, where T memory cells was defined as CD62L+ and CD44+.
  • Fig. 16 F depicts the percentage of OTI cells expressing CD127.
  • Fig. 17 depicts data related to development of symptoms of diabetes in naive, control, and experimental groups.
  • Fig. 18 depicts the various treatment groups and the experimental timeline used in an experiment related to evaluation of induction of long-lasting tolerance in transferred T cells.
  • Figures 19A-19B depict data related to T cell composition in the lymph node.
  • Figure 19A depicts the remaining OTI cells after antigen challenge (as a percentage of total CD8 + cells) in the various treatment groups.
  • Figure 19B depicts the remaining OTI I cells after antigen challenge (as a percentage of total CD4 + cells) in the various treatment groups.
  • Fig. 20 depicts the various treatment groups and the experimental timeline used in an experiment related to evaluation of induction of long-lasting tolerance in endogenous T cells.
  • Figures 21 A-21 B depict data related to T cell composition in the lymph node.
  • Figure 21 A depicts the remaining OTI cells after antigen challenge (as a percentage of total CD8 + cells) in the various treatment groups.
  • Figure 21 B depicts the remaining OTII cells after antigen challenge (as a percentage of total CD4 + cells) in the various treatment groups.
  • Fig. 22 depicts the experimental design used to evaluate the ability of compositions as disclosed herein to prophylactically reduce antibody response.
  • Fig. 23 depicts experimental data related to the amount of anti-asparaginase antibodies from mice in the various treatment groups.
  • Figures 24A-24B depict the experimental design and composition used in evaluating tolerance to myelin oligodendrocyte glycoprotein (MOG).
  • Fig. 24A shows the experimental protocol used in immunizing donor mice and treating recipient mice.
  • Fig. 24B shows one example of a tolerogenic composition in accordance with several embodiments disclosed herein.
  • Figures 25A-25B depict experimental data related to induction of tolerance against MOG using a first concentration of the tolerogenic composition.
  • Fig. 25A depicts data related to delay of disease onset in the various treatment groups.
  • Fig. 25B depicts data related to reduction of weight loss in the various treatment groups.
  • Figures 26A-26B depict experimental data related to induction of tolerance against MOG using an additional concentration of the tolerogenic composition.
  • Fig. 26A depicts data related to delay of disease onset in the various treatment groups.
  • Fig. 26B depicts data related to reduction of weight loss in the various treatment groups.
  • FIG.27A depicts targeting of OVA via various conjugates to LSECs in the liver.
  • Fig.27B depicts targeting of OVA via various conjugates to Kupffer cells in the liver.
  • Fig.27C depicts targeting of OVA via various conjugates to CD11 c+ cells in the liver.
  • Fig.27D depicts targeting of OVA via various conjugates to hepatocytes in the liver.
  • Fig.27E depicts targeting of OVA via various conjugates to stellate cells in the liver.
  • ASGPRs asialoglycoprotein receptors
  • LSECs liver sinusoidal endothelial cells
  • Other galactose/galactosamine/N-acetylgalactosamine receptors can be found in various forms on multiple cell types [e.g., dendritic cells, hepatocytes, LSECs, and Kupffer cells].
  • Dendritic cells are considered "professional antigen presenting cells," because their primary function is to present antigens to the immune system for generating immune responses. Some cells within the liver are known to be able to present antigens, but the liver is more known to be involved in tolerogenesis. The liver is understood to be a tolerogenic organ.
  • glycosylation i.e., the presence of significant numbers of glycosylated proteins.
  • glycophorins e.g., glycophorin A
  • Glycophorins are proteins with many covalently attached sugar chains, the terminus of which is sialic acid. As an erythrocyte ages and becomes ripe for clearance, the terminal sialic acid of its glycophorins tends to be lost, leaving N-acetylgalactosamine at the free end.
  • N-acetylgalactosamine is a ligand selectively received by the ASGPR associated with hepatic cells, leading to binding of N-acetylgalactosamine-containing substances by hepatic cells and their subsequent uptake and processing in the liver.
  • the present disclosure provides, in several embodiments, certain therapeutic compositions that are targeted for delivery to (and for uptake by) the liver, particularly hepatocytes, LSECs, Kupffer cells and/or stellate cells, more particularly hepatocytes and/or LSECs, and even more particularly to specifically bind ASGPR.
  • Liver-targeting facilitates two mechanisms of treatment: tolerization and clearance.
  • Tolerization takes advantage of the liver's role in clearing apoptotic cells and processing their proteins to be recognized by the immune system as "self," as well as the liver's role in sampling peripheral proteins for immune tolerance. Clearance takes advantage of the liver's role in blood purification by rapidly removing and breaking down toxins, polypeptides and the like.
  • compositions to the liver is accomplished by a galactosylating moiety (e.g., galactose, galactosamine and N-acetylgalactosamine, particularly conjugated at C1 , C2 or C6, though some embodiments involved conjugation at other or any carbon in the molecule), by a glucosylating moiety (e.g., glucose, glucosamine and N-acetylglucosamine, particularly conjugated at C1 , C2 or C6, though some embodiments involved conjugation at other or any carbon in the molecule), or by de-sialylating a polypeptide for which such liver-targeting is desired.
  • a galactosylating moiety e.g., galactose, galactosamine and N-acetylgalactosamine, particularly conjugated at C1 , C2 or C6, though some embodiments involved conjugation at other or any carbon in the molecule
  • a glucosylating moiety e
  • the galactosylating or glucosylating moiety can be chemically conjugated or recombinantly fused to an antigen, whereas desialylation exposes a galactose- like moiety on an antigen polypeptide.
  • the antigen can be endogenous (a self-antigen) or exogenous (a foreign antigen), including but not limited to: a foreign transplant antigen against which transplant recipients develop an unwanted immune response (e.g., transplant rejection), a foreign food, animal, plant or environmental antigen to which patients develop an unwanted immune (e.g., allergic or hypersensitivity) response, a therapeutic agent to which patients develop an unwanted immune response (e.g., hypersensitivity and/or reduced therapeutic activity), a self-antigen to which patients develop an unwanted immune response (e.g., autoimmune disease), or a tolerogenic portion (e.g., a fragment or an epitope) thereof; these compositions are useful for inducing tolerization to the antigen.
  • a foreign transplant antigen against which transplant recipients develop an unwanted immune response e.g., transplant rejection
  • a foreign food, animal, plant or environmental antigen to which patients develop an unwanted immune (e.g., allergic or hypersensitivity) response e.g., a therapeutic agent to which patients develop an unwanted immune
  • the galactosylating or other liver-targeting moiety can be conjugated to an antibody, antibody fragment or ligand that specifically binds a circulating protein or peptide or antibody, which circulating protein or peptide or antibody is causatively involved in transplant rejection, immune response against a therapeutic agent, autoimmune disease, and/or allergy (as discussed above); these compositions are useful for clearing the circulating protein, peptide or antibody. Accordingly, the compositions of the present disclosure can be used for treating an unwanted immune response, e.g., transplant rejection, an immune response against a therapeutic agent, an autoimmune disease, and/or an allergy, depending on the embodiment.
  • an unwanted immune response e.g., transplant rejection, an immune response against a therapeutic agent, an autoimmune disease, and/or an allergy, depending on the embodiment.
  • compositions containing a therapeutically effective amount of a composition of the disclosure admixed with at least one pharmaceutically acceptable excipient.
  • the disclosure provides methods for the treatment of an unwanted immune response, such as transplant rejection, response against a therapeutic agent, autoimmune disease or allergy.
  • an "antigen” is any substance that serves as a target for the receptors of an adaptive immune response, such as the T cell receptor, major histocompatibility complex class I and II, B cell receptor or an antibody.
  • an antigen may originate from within the body (e.g., "self,” “auto” or "endogenous").
  • an antigen may originate from outside the body ("non-self,” “foreign” or “exogenous”), having entered, for example, by inhalation, ingestion, injection, or transplantation, transdermal ⁇ , etc.
  • an exogenous antigen may be biochemically modified in the body.
  • Foreign antigens include, but are not limited to, food antigens, animal antigens, plant antigens, environmental antigens, therapeutic agents, as well as antigens present in an allograft transplant.
  • an "antigen-binding molecule” as used herein relates to molecules, in particular to proteins such as immunoglobulin molecules, which contain antibody variable regions providing a binding (specific binding in some embodiments) to an epitope.
  • the antibody variable region can be present in, for example, a complete antibody, an antibody fragment, and a recombinant derivative of an antibody or antibody fragment.
  • Antigen-binding fragments containing antibody variable regions include (without limitation) “Fv”, “Fab”, and “F(ab') 2 " regions, “single domain antibodies (sdAb)", “nanobodies”, “single chain Fv (scFv)” fragments, “tandem scFvs” (V H A-V L A-V H B-V L B), “diabodies”, “triabodies” or “tribodies”, “single- chain diabodies (scDb)", and “bi-specific T-cell engagers (BiTEs)".
  • a "chemical modification” refers to a change in the naturally occurring chemical structure of one or more amino acids of a polypeptide. Such modifications can be made to a side chain or a terminus, e.g., changing the amino-terminus or carboxyl terminus. In some embodiments, the modifications are useful for creating chemical groups that can conveniently be used to link the polypeptides to other materials, or to attach a therapeutic agent.
  • Constant changes can generally be made to an amino acid sequence without altering activity. These changes are termed “conservative substitutions” or mutations; that is, an amino acid belonging to a grouping of amino acids having a particular 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.
  • the nonpolar (hydrophobic) amino acids include alanine, leucine, isoleucine, valine, proline, phenylalanine, tryptophan, methionine, and tyrosine.
  • the polar neutral amino acids include glycine, serine, threonine, cysteine, tyrosine, asparagine and glutamine.
  • the positively charged (basic) amino acids include arginine, lysine and histidine.
  • the negatively charged (acidic) amino acids include aspartic acid and glutamic acid. Such substitutions are not expected to substantially affect apparent molecular weight as determined by polyacrylamide gel electrophoresis or isoelectric point. Conservative substitutions also include substituting optical isomers of the sequences for other optical isomers, specifically D amino acids for i_ amino acids for one or more residues of a sequence. Moreover, all of the amino acids in a sequence can undergo a D to L isomer substitution.
  • Exemplary conservative substitutions include, but are not limited to, Lys for Arg and vice versa to maintain a positive charge; Glu for Asp and vice versa to maintain a negative charge; Ser for Thr so that a free -OH is maintained; and Gin for Asn to maintain a free -NH 2 .
  • Yet another type of conservative substitution constitutes the case where amino acids with desired chemical reactivities are introduced to impart reactive sites for chemical conjugation reactions, if the need for chemical derivatization arises.
  • Such amino acids include but are not limited to Cys (to insert a sulfhydryl group), Lys (to insert a primary amine), Asp and Glu (to insert a carboxylic acid group), or specialized noncanonical amino acids containing ketone, azide, alkyne, alkene, and tetrazine side-chains.
  • Conservative substitutions or additions of free -NH 2 or -SH bearing amino acids can be particularly advantageous for chemical conjugation with the linkers and galactosylating moieties of Formula 1.
  • point mutations, deletions, and insertions of the polypeptide sequences or corresponding nucleic acid sequences can in some cases be made without a loss of function of the polypeptide or nucleic acid fragment.
  • Substitutions can include, e.g., 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 those listed).
  • a variant usable in the present invention may exhibit a total number of up to 200 (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, 1 10, 120, 130, 140, 150, 160, 170, 180, 190, or 200, including any number in between those listed) changes in the amino acid sequence (e.g., exchanges, insertions, deletions, N-terminal truncations, and/or C-terminal truncations).
  • the number of changes is greater than 200.
  • the variants include polypeptide sequences or corresponding nucleic acid sequences that exhibit a degree of functional equivalence with a reference (e.g., unmodified or native sequence). In several embodiments, the variants exhibit about 80%, about 85%, about 90%, about 95%, about 97%, about 98%, about 99% functional equivalence to an unmodified or native reference sequence (and any degree of functional equivalence between those listed).
  • the amino acid residues described herein employ either the single letter amino acid designator or the three-letter abbreviation in keeping with the standard polypeptide nomenclature, J. Biol. Chem., (1969), 243, 3552-3559. All amino acid residue sequences are represented herein by formulae with left and right orientation in the conventional direction of amino- terminus to carboxy-terminus.
  • an effective amount refers to that amount of a composition of the disclosure that is sufficient to effect treatment, as defined below, when administered to a mammal in need of such treatment. This amount will vary depending upon the subject and disease condition being treated, the weight and age of the subject, the severity of the disease condition, the particular composition of the disclosure chosen, the dosing regimen to be followed, timing of administration, manner of administration and the like, all of which can readily be determined by one of ordinary skill in the art.
  • An “epitope”, also known as antigenic determinant, is the segment of a macromolecule, e.g. a protein, which is recognized by the adaptive immune system, such as by antibodies, B cells, major histocompatibility complex molecules, or T cells.
  • An epitope is that part or segment of a macromolecule capable of binding to an antibody or antigen-binding fragment thereof.
  • binding in particular relates to a specific binding.
  • the term “epitope” refers to the segment of protein or polyprotein that is recognized by the immune system.
  • galactose refers to a monosaccharide sugar that exists both in open-chain form and in cyclic form, having D- and L- isomers.
  • cyclic form there are two anomers, namely alpha and beta.
  • alpha form the C1 alcohol group is in the axial position
  • beta form the C1 alcohol group is in the equatorial position.
  • galactose refers to the cyclic six-membered pyranose, more in particular the D-isomer and even more particularly the alpha-D-form (a-D- galactopyranose) the formal name for which is (2R,3R,4S,5R,6R)-6-(hydroxymethyl)tetrahydro-2H- pyran-2,3,4,5-tetraol.
  • Glucose is an epimer of galactose; the formal name is (2R,3R,4S,5S,6R)-6- (hydroxymethyl)tetrahydro-2/- -pyran-2,3,4,5-tetraol.
  • the structure and numbering of galactose and glucose are shown giving two non-limiting examples of stereochemical illustration.
  • galactosylating moiety refers to a particular type of liver-targeting moiety.
  • Galactosylating moieties include, but are not limited to a galactose, galactosamine and/or N- acetylgalactosamine residue.
  • a "glucosylating moiety” refers to another particular type of liver-targeting moiety and includes, but is not limited to glucose, glucosamine and/or N-acetylglucosamine.
  • liver-targeting moiety refers to moieties having the ability to direct, e.g., a polypeptide, to the liver.
  • the liver comprises different cell types, including but not limited to hepatocytes, sinusoidal epithelial cells, Kupffer cells, stellate cells, and/or dendritic cells.
  • a liver-targeting moiety directs a polypeptide to one or more of these cells.
  • receptors are present which recognize and specifically bind the liver-targeting moiety.
  • Liver-targeting can be achieved by chemical conjugation of an antigen or ligand to a galactosylating or glucosylating moiety, desialylation of an antigen or ligand to expose underlying galactosyl or glucosyl moieties, or specific binding of an endogenous antibody to an antigen or ligand, where the antigen or ligand is: desialylated to expose underlying galactosyl or glucosyl moieties, conjugated to a galactosylating or a glucosylating moiety.
  • Naturally occurring desialylated proteins are not encompassed within the scope of certain embodiments of the present disclosure.
  • n can be an integer from about 1 to 100 (e.g., 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 1 1 , 12, 13, 14, 15, 16, 17, 18, 19, 20, 22, 25, 30, 34, 35, 37, 40, 41 , 45, 50, 54, 55, 59, 60, 65, 70, 75, 80, 82, 83, 85, 88, 90, 95, 99, 100, 105 or 1 10, or any between those listed, including the endpoints of the range) and that the disclosed mixture encompasses ranges such as 1 -4, 2-4, 2-6, 3-8,
  • a peptide that specifically binds a particular target is referred to as a "ligand" for that target.
  • polypeptide is a term that refers to a chain of amino acid residues, regardless of post- translational modification (e.g., phosphorylation or glycosylation) and/or complexation with additional polypeptides, and/or synthesis into multisubunit complexes with nucleic acids and/or carbohydrates, or other molecules. Proteoglycans therefore also are referred to herein as polypeptides.
  • polypeptides can be produced by a number of methods, many of which are well known in the art. For example, polypeptides can be obtained by extraction (e.g., from isolated cells), by expression of a recombinant nucleic acid encoding the polypeptide, or by chemical synthesis. Polypeptides can be produced by, for example, recombinant technology, and expression vectors encoding the polypeptide introduced into host cells (e.g., by transformation or transfection) for expression of the encoded polypeptide
  • pharmaceutically acceptable carrier or “pharmaceutically acceptable excipient” includes any and all solvents, dispersion media, coatings, antibacterial and antifungal agents, isotonic and absorption delaying agents and the like.
  • the use of such media and agents for pharmaceutically active substances is well known in the art. Except insofar as any conventional media or agent is incompatible with the active ingredient, its use in the therapeutic compositions is contemplated. Supplementary active ingredients can also be incorporated into the compositions.
  • purified refers to a polypeptide that has been chemically synthesized and is thus substantially uncontaminated by other polypeptides, or has been separated or isolated from most other cellular components by which it is naturally accompanied (e.g., other cellular proteins, polynucleotides, or cellular components).
  • An example of a purified polypeptide is one that is at least 70%, by dry weight, free from the proteins and naturally occurring organic molecules with which it naturally associates.
  • a preparation of a purified polypeptide therefore can be, for example, at least 80%, at least 90%, or at least 99%, by dry weight, the polypeptide.
  • Polypeptides also can be engineered to contain a tag sequence (e.g., a polyhistidine tag, a myc tag, a FLAG ® tag, or other affinity tag) that facilitates purification or marking (e.g., capture onto an affinity matrix, visualization under a microscope).
  • a purified composition that comprises a polypeptide refers to a purified polypeptide unless otherwise indicated.
  • isolated indicates that the polypeptides or nucleic acids of the disclosure are not in their natural environment. Isolated products of the disclosure can thus be contained in a culture supernatant, partially enriched, produced from heterologous sources, cloned in a vector or formulated with a vehicle, etc.
  • random copolymer refers to the product of simultaneous polymerization of two or more monomers in admixture, where the probability of finding a given monomeric unit at any given site in a polymer chain is independent of the nature of the neighboring units at that position (Bernoullian distribution).
  • the chain can comprise any sequence from 2 up to about 150 W 1 and W 2 groups, such as: -W 1 -W 2 -W 1 -W 2 -; -W 2 -W 1 -W 2 -W 1 -; -W 1 -W 1 -W 1 -W 2 -; -W 1 -W 1 -W 2 -W 2 -; -W 1 -W 2 -W 2 -W 1 -; -W 1 -W 2 -W 2 -W 1 -; -W 1 -W 2 -W 1 -W 2 -W 2 -W 1 -W 2 -W 1 -; -W 1 -W 1 -W 2 -W 2 -W 1 -W 2 -W 1 -; -W 1 -W 1 -W 2 -W 2 -W 1 -W 2 -W 1 -; and W 2 -W 2 -W 1 -W 2 -W 1 -
  • sequence identity is used with regard to polypeptide (or nucleic acid) sequence comparisons. This expression in particular refers to a percentage of sequence identity, for example 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% to the respective reference polypeptide or to the respective reference polynucleotide.
  • the polypeptide in question and the reference polypeptide exhibit the indicated sequence identity over a continuous stretch of 20, 30, 40, 45, 50, 60, 70, 80, 90, 100 or more amino acids or over the entire length of the reference polypeptide.
  • Specific binding refers to a molecule that binds to a target with a relatively high affinity as compared to non-target tissues, and generally involves a plurality of non-covalent interactions, such as electrostatic interactions, van der Waals interactions, hydrogen bonding, and the like. Specific binding interactions characterize antibody-antigen binding, enzyme-substrate binding, and certain protein-receptor interactions; while such molecules might bind tissues besides their specific targets from time to time, to the extent that such non-target binding is inconsequential, the high-affinity binding pair can still fall within the definition of specific binding.
  • treatment means any treatment of a disease or disorder in a mammal, including:
  • the term "unwanted immune response” refers to a reaction by the immune system of a subject, which in the given situation is not desirable.
  • the reaction of the immune system is unwanted if such reaction does not lead to the prevention, reduction, or healing of a disease or disorder but instead causes, enhances or worsens, or is otherwise associated with induction or worsening of a disorder or disease.
  • a reaction of the immune system causes, enhances or worsens a disease if it is directed against an inappropriate target.
  • an unwanted immune response includes but is not limited to transplant rejection, immune response against a therapeutic agent, autoimmune disease, and allergy or hypersensitivity.
  • variant is to be understood as a protein (or nucleic acid) which differs in comparison to the protein from which it is derived by one or more changes in its length, sequence, or structure.
  • polypeptide from which a protein variant is derived is also known as the parent polypeptide or polynucleotide.
  • variant comprises "fragments” or “derivatives” of the parent molecule. Typically, “fragments” are smaller in length or size than the parent molecule, whilst “derivatives” exhibit one or more differences in their sequence or structure in comparison to the parent molecule.
  • modified molecules such as but not limited to post-translationally modified proteins (e.g.
  • variants glycosylated, phosphorylated, ubiquitinated, palmitoylated, or proteolytically cleaved proteins
  • modified nucleic acids such as methylated DNA.
  • variants Naturally occurring and artificially constructed variants are to be understood to be encompassed by the term "variant" as used herein.
  • variants usable in the present invention may also be derived from homologs, orthologs, or paralogs of the parent molecule or from artificially constructed variant, provided that the variant exhibits at least one biological activity of the parent molecule, e.g., is functionally active.
  • a variant can be characterized by a certain degree of sequence identity to the parent polypeptide from which it is derived. More precisely, a protein variant in the context of the present disclosure may exhibit at least 80% sequence identity to its parent polypeptide. Preferably, the sequence identity of protein variants is over 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 to an unmodified or native reference sequence (and any degree of functional equivalence between those listed).
  • compositions, pharmaceutical formulations, and methods of treatment employing such compositions as represented by Formula 1 :
  • n is an integer from about 1 to 100, particularly from about 1 to 20, and most particularly 1 to about 10;
  • X is an antigen moiety, particularly a foreign antigen or self-antigen against which a patient develops an unwanted immune response, or a tolerogenic portion (e.g., a fragment or an epitope) of such an antigen moiety;
  • Y is a linker moiety or a direct bond, or an antibody, antibody fragment, peptide or other ligand that specifically binds X;
  • Z is a liver-targeting moiety, in particular galactosylating or a glucosylating moiety.
  • m in Formula 1 will depend upon the nature of X, in that each antigen, antibody, antibody fragment or ligand will have an individual number and density of sites (predominantly the N-terminal amine, lysine residues and cysteine residues) to which a linker, a galactosylating moiety or a glucosylating moiety can be bound.
  • Antigens having a limited number of such sites can be derivatized, for example, at the N or C terminus, by adding lysine or cysteine residues (optionally via a cleavable linker, particularly a linker having an immunoproteosome cleavage site).
  • compositions of Formula 1 Generally, it is preferred to provide an adequate degree of galactosylation/glucosylation in compositions of Formula 1 so as to facilitate uptake by liver cells.
  • Pharmaceutical formulations and methods of the disclosure can employ a cocktail of compositions of Formula 1 , respectively bearing different X moieties (e.g., several epitopes associated with a particular unwanted immune response).
  • compositions of Formula 1 include the sub-genuses where X is a foreign transplant antigen against which transplant recipients develop an unwanted immune response (e.g., transplant rejection), a foreign food, animal, plant or environmental antigen against which patients develop an unwanted immune (e.g., transplant rejection), a foreign food, animal, plant or environmental antigen against which patients develop an unwanted immune (e.g.
  • Y is a linker of Formulae Ya through Yp; and/or where Z is galactose, galactosamine, N-acetylgalactosamine, glucose, glucosamine or N-acetylglucosamine as illustrated by Formulae 1 a through 1 p as described below with reference to the Reaction Schemes.
  • X in the compositions of Formula 1 , can be an antibody, antibody fragment or ligand that specifically binds a circulating protein or peptide or antibody, which circulating protein or peptide or antibody is causatively involved in transplant rejection, immune response against a therapeutic agent, autoimmune disease, hypersensitivity and/or allergy.
  • the antigen employed as X in the compositions of Formula 1 can be a protein or a peptide, e.g. the antigen may be a complete or partial therapeutic agent, a full-length transplant protein or peptide thereof, a full-length autoantigen or peptide thereof, a full-length allergen or peptide thereof, and/or a nucleic acid, or a mimetic of an aforementioned antigen.
  • the antigen may be a complete or partial therapeutic agent, a full-length transplant protein or peptide thereof, a full-length autoantigen or peptide thereof, a full-length allergen or peptide thereof, and/or a nucleic acid, or a mimetic of an aforementioned antigen.
  • a listing of any particular antigen in a category or association with any particular disease or reaction does not preclude that antigen from being considered part of another category or associated with another disease or reaction.
  • Antigens employed in the practice of the present disclosure can be one or more of the following:
  • Therapeutic agents that are proteins, peptides, antibodies and antibody-like molecules, including antibody fragments and fusion proteins with antibodies and antibody fragments. These include human, non-human (such as mouse) and non-natural (i.e., engineered) proteins, antibodies, chimeric antibodies, humanized antibodies, and non-antibody binding scaffolds, such as fibronectins, DARPins, knottins, and the like.
  • Self-antigens that cause an unwanted, autoimmune response.
  • the polypeptides employed in the disclosed compositions are typically synthesized exogenously (as opposed to being purified and concentrated from a source of origin).
  • the antigen can be a complete protein, a portion of a complete protein, a peptide, or the like, and can be derivatized (as discussed above) for attachment to a linker and/or galactosylating moiety, can be a variant and/or can contain conservative substitutions, particularly maintaining sequence identity, and/or can be desialylated.
  • specific antigens can be selected from: Abatacept, Abciximab, Adalimumab, Adenosine deaminase, Ado-trastuzumab emtansine, Agalsidase alfa, Agalsidase beta, Aldeslukin, Alglucerase, Alglucosidase alfa, a-1 -proteinase inhibitor, Anakinra, Anistreplase (anisoylated plasminogen streptokinase activator complex), Antithrombin III, Antithymocyte globulin, Ateplase, Bevacizumab, Bivalirudin, Botulinum toxin type A, Botulinum toxin type B, C1 -esterase inhibitor, Canakinumab, Carboxypeptidase G2 (Glucarpidase
  • Particular therapeutic protein, peptide, antibody or antibody-like molecules include Abciximab, Adalimumab, Agalsidase alfa, Agalsidase beta, Aldeslukin, Alglucosidase alfa, 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 varying formats.
  • Another particular group includes the hemostatic agents (Factor VIII and IX), Insulin (including rHu Insulin), and the non-human therapeutics uricase, PAL and asparaginase.
  • Unwanted immune response in hematology and transplant includes autoimmune aplastic anemia, transplant rejection (generally), and Graft vs. Host Disease (bone marrow transplant rejection).
  • the antigen is a human allograft transplantation antigen
  • specific sequences can be selected from: subunits of the various MHC class I and MHC class II haplotype proteins (for example, donor/recipient differences identified in tissue cross-matching), and single-amino-acid polymorphisms on minor blood group antigens including RhCE, Kell, Kidd, Duffy and Ss.
  • Such compositions can be prepared individually for a given donor/recipient pair.
  • antigen is a self-antigen
  • specific antigens can be selected from:
  • insulin In type 1 diabetes mellitus, several main antigens have been identified: insulin, proinsulin, preproinsulin, glutamic acid decarboxylase-65 (GAD-65 or glutamate decarboxylase 2), GAD- 67, glucose-6 phosphatase 2 (IGRP or islet-specific glucose 6 phosphatase catalytic subunit related protein), insulinoma-associated protein 2 (IA-2), and insulinoma-associated protein 2 ⁇ ( ⁇ -2 ⁇ ); other antigens include ICA69, ICA12 (SOX-13), carboxypeptidase H, Imogen 38, GLIMA 38, chromogranin-A, HSP-60, carboxypeptidase E, peripherin, glucose transporter 2, hepatocarcinoma-intestine-pancreas/pancreatic associated protein, S100p, glial fibrillary acidic protein, regenerating gene II, pancreatic duodenal homeobox 1 , dystrophia myotonica kinase
  • insulin is an example of an antigen that can be characterized both as a self-antigen and a therapeutic protein antigen.
  • rHu Insulin and bovine insulin are therapeutic protein antigens (that are the subject of unwanted immune attack), whereas endogenous human insulin is a self-antigen (that is the subject of an unwanted immune attack). Because endogenous human insulin is not available to be employed in a pharmaceutical composition, a recombinant form is employed in certain embodiments of the compositions of the disclosure.
  • o Human insulin including an exogenously obtained form useful in the compositions of the disclosure, has the following sequence (UNIPROT P01308):
  • o GAD-65 including an exogenously obtained form useful in the compositions of the disclosure, has the following sequence (UNIPROT Q05329):
  • o IGRP including an exogenously obtained form useful in the compositions of the disclosure, has the following sequence (UNIPROT QN9QR9):
  • main antigens include thyroglobulin (TG), thyroid peroxidase (TPO) and thyrotropin receptor (TSHR); other antigens include sodium iodine symporter (NIS) and megalin.
  • TG thyroglobulin
  • TPO thyroid peroxidase
  • TSHR thyrotropin receptor
  • NIS sodium iodine symporter
  • an antigen is insulin-like growth factor 1 receptor.
  • a main antigen is calcium sensitive receptor.
  • main antigens include 21 -hydroxylase, 17a-hydroxylase, and P450 side chain cleavage enzyme (P450scc); other antigens include ACTH receptor, P450c21 and P450c17.
  • P450scc P450 side chain cleavage enzyme
  • main antigens include FSH receptor and a-enolase.
  • main antigens include pituitary gland-specific protein factor (PGSF) 1a and 2; another antigen is type 2 iodothyronine deiodinase.
  • PGSF pituitary gland-specific protein factor
  • main antigens include myelin basic protein (“MBP”), myelin oligodendrocyte glycoprotein (“MOG”) and myelin proteolipid protein (“PLP”).
  • MBP myelin basic protein
  • MOG myelin oligodendrocyte glycoprotein
  • PGP myelin proteolipid protein
  • o MBP including an exogenously obtained form useful in the compositions of the disclosure, has the following sequence (UNIPROT P02686):
  • o MOG including an exogenously obtained form useful in the compositions of the disclosure, has the following sequence (UNIPROT Q16653):
  • o PLP including an exogenously obtained form useful in the compositions of the disclosure, has the following sequence (UNIPROT P60201 ):
  • o Peptides/epitopes useful in the compositions of the disclosure for treating multiple sclerosis include some or all of the following sequences, individually in a composition of Formula 1 or together in a cocktail of compositions of Formula 1 :
  • ⁇ MBP13-32 KYLATASTMDHARHGFLPRH (SEQ ID NO:7);
  • ⁇ MBP146-170 AQGTLSKIFKLGGRDSRSGSPMARR (SEQ ID NO: 10);
  • MOG1 -20 GQFRVIGPRHPIRALVGDEV (SEQ ID NO: 1 1);
  • MOG35-55 MEVGWYRPPFSRWHLYRNGK (SEQ ID NO: 12);
  • ⁇ PLP139-154 HCLGKWLGHPDKFVGI (SEQ ID NO: 13).
  • main antigens include collagen II, immunoglobulin binding protein, the fragment crystallizable region of immunoglobulin G, double-stranded DNA, and the natural and cirtullinated forms of proteins implicated in rheumatoid arthritis pathology, including fibrin/fibrinogen, vimentin, collagen I and II, and alpha-enolase.
  • a main antigen is H+,K+-ATPase.
  • main antigens are tissue transglutaminase and the natural and deamidated forms of gluten or gluten-like proteins, such as alpha-, gamma-, and omega-gliadin, glutenin, hordein, secalin, and avenin.
  • alpha gliadin turns more immunogenic in the body through deamidation by tissue glutaminase converting alpha gliadin's glutamines to glutamic acid.
  • alpha gliadin is originally a foreign food antigen, once it has been modified in the body to become more immunogenic it can be characterized as a self-antigen.
  • a main antigen is tyrosinase, and tyrosinase related protein 1 and 2.
  • o Tyrosinase including an exogenously obtained form useful in the compositions of the disclosure, has the following sequence (UNIPROT P14679):
  • o Melanocyte protein PMEL, gp100 including an exogenously obtained form useful in the compositions of the disclosure, has the following sequence (UNIPROT P40967):
  • acetylcholine receptor In myasthenia gravis, a main antigen is acetylcholine receptor.
  • main antigens are desmoglein 3, 1 and 4; other antigens include pemphaxin, desmocollins, plakoglobin, perplakin, desmoplakins, and acetylcholine receptor.
  • main antigens include BP180 and BP230; other antigens include plectin and laminin 5.
  • main antigens include endomysium and tissue transglutaminase.
  • a main antigen is collagen VII.
  • main antigens include matrix metalloproteinase 1 and 3, the collagen- specific molecular chaperone heat-shock protein 47, fibrillin-1 , and PDGF receptor; other antigens include Scl-70, U1 RNP, Th/To, Ku, Jo1 , NAG-2, centromere proteins, topoisomerase I, nucleolar proteins, RNA polymerase I, II and III, PM-SIc, fibrillarin, and B23.
  • a main antigen is Ul snRNP.
  • the main antigens are nuclear antigens SS-A and SS-B; other antigens include fodrin, poly(ADP-ribose) polymerase and topoisomerase, muscarinic receptors, and the Fc-gamma receptor 1Mb.
  • main antigens include nuclear proteins including the "Smith antigen," SS-A, high mobility group box 1 (HMGB1 ), nucleosomes, histone proteins and double- stranded DNA (against which auto-antibodies are made in the disease process).
  • SS-A nuclear proteins
  • HMGB1 high mobility group box 1
  • nucleosomes nucleosomes
  • histone proteins double- stranded DNA (against which auto-antibodies are made in the disease process).
  • main antigens include glomerular basement membrane proteins including collagen IV.
  • a main antigen is cardiac myosin.
  • autoimmune polyendocrine syndrome type 1 antigens include aromatic L-amino acid decarboxylase, histidine decarboxylase, cysteine sulfinic acid decarboxylase, tryptophan hydroxylase, tyrosine hydroxylase, phenylalanine hydroxylase, hepatic P450 cytochromes P4501A2 and 2A6, SOX-9, SOX-10, calcium-sensing receptor protein, and the type 1 interferons interferon alpha, beta and omega.
  • AQP4 AQP4.
  • o Aquaporin-4 including an exogenously obtained form useful in the compositions of the disclosure, has the following sequence (UNIPROT P55087):
  • VHVI DVDRGEEKKGKDQSGEVLSSV (SEQ ID NO: 17).
  • main antigens include Retinal S-antigen or "S-arrestin” and interphotoreceptor retinoid binding protein (IRBP) or retinol-binding protein 3.
  • IRBP interphotoreceptor retinoid binding protein
  • o S-arrestin including an exogenously obtained form useful in the compositions of the disclosure, has the following sequence (UNIPROT P10523):
  • o IRBP including an exogenously obtained form useful in the compositions of the disclosure, has the following sequence (UNIPROT P10745):
  • antigen is a foreign antigen against which an unwanted immune response can be developed, such as food antigens
  • specific antigens can be:
  • conarachin (Ara h 1 ), allergen II (Ara h 2), arachis agglutinin, conglutin (Ara h 6); o conarachin, for example has the sequence identified as UNIPROT Q6PSU6
  • high molecular weight glutenin from wheat and/or other cereals: high molecular weight glutenin, low molecular weight glutenin, alpha-, gamma- and omega-gliadin, hordein, secalin and/or avenin;
  • o peptides/epitopes useful in the compositions of the disclosure for treating Celiac Disease include some or all of the following sequences, individually in a composition of Formula 1 or together in a cocktail of compositions of Formula 1 :
  • LQLQPFPQPELPYPQPELPYPQPELPYPQPQPF (SEQ ID NO:21 )
  • antigen is a foreign antigen against which an unwanted immune response is developed, such as to animal, plant and environmental antigens
  • specific antigens can, for example, be: cat, mouse, dog, horse, bee, dust, tree and goldenrod, including the following proteins or peptides derived from:
  • weeds including ragweed allergens amb a 1 , 2, 3, 5, and 6, and Amb t 5; pigweed Che a 2 and 5; and other weed allergens Par j 1 , 2, and 3, and Par o 1 );
  • grass including major allergens Cyn d 1 , 7, and 12; Dac g 1 , 2, and 5; Hoi I 1.01203; Lol p 1 , 2, 3, 5, and 1 1 ; Mer a 1 ; Pha a 1 ; Poa p 1 and 5);
  • pollen from ragweed and other weeds including curly dock, lambs quarters, pigweed, plantain, sheep sorrel, and sagebrush
  • grass including Bermuda, Johnson, Kentucky, Orchard, Sweet vernal, and Timothy grass
  • trees including catalpa, elm, hickory, olive, pecan, sycamore, and walnut
  • pets including cats, dogs, rodents, and farm animals;
  • major cat allergens include Fel d 1 through 8, cat IgA, BLa g 2, and cat albumin;
  • major dog allergens include Can f 1 through 6, and dog albumin);
  • bee stings including major allergens Api m 1 through 12;
  • fungus including allergens derived from, species of Aspergillus and Penicillium, as well as the species Alternaria alternata, Davidiella tassiana, and Trichophyton rubrum.
  • a patient can be tested to identify an antigen against which an unwanted immune response has developed, and a protein, peptide or the like can be developed based on that antigen and incorporated as X in a composition of the present disclosure.
  • antigens antibodies, antibody fragments having sialylation that can be removed to leave glycosylation specifically targeting the ASGPR: follicle stimulating hormone (FSH), human chorionic gonadotropin (HCG), luteinizing hormone (LH), osteopontin, thyroid stimulating hormone (TSH), agalsidase alfa, agalsidase beta (FABRAZYME®; Genzyme), epoetin alfa and epoetin beta, follitropin alfa (GONAL-F®; Merck/Serono) and follitropin beta (FOLLISTIM®; Schering-Plough), insulin growth factor binding protein 6 (IGFBP-6), lutropin alfa (LUVERIS®; Merck/Serono), transforming growth factor ⁇ 1 , antithrombin (ATry n®/TRO M BATE-I II®; Genzyme/Talecris Bio
  • FSH follicle stimulating hormone
  • Pharmaceutically relevant proteins that have previously been hyperglycosylated and can be desialylated for hepatocyte-ASGPR targeting include: interferon alfa and gamma, luteinizing hormone, Fv antibody fragments, asparaginase, cholinesterase, darbepoetin alfa (AraNESP®; Amgen), thrombopoietin, leptin, FSH, IFN-a2, serum albumin, and corifollitropin alfa.
  • Proteins with glycans that do not normally terminate in sialic acids including proteins produced in bacteria or yeast (such as arginase, some insulins, and uricase) would not be amenable to desialylation.
  • such moieties are chosen to specifically bind a targeted circulating protein or peptide or antibody, and result in hepatic uptake of the circulating targeted moiety, possibly as an adduct with the targeting moiety, ultimately resulting in the clearance and inactivation of the circulating targeted moiety.
  • a targeted circulating protein or peptide or antibody for example, liver-targeted Factor VIII will bind and clear circulating anti-Factor VIII antibodies. Procedures for the identification of such moieties will be familiar to those skilled in the art.
  • the linkers employed in the compositions of the present disclosure (“Y" in Formula 1 ) can include N-hydroxysuccinamidyl linkers, malaemide linkers, vinylsulfone linkers, pyridyl di-thiol- poly(ethylene glycol) linkers, pyridyl di-thiol linkers, n-nitrophenyl carbonate linkers, NHS-ester linkers, nitrophenoxy poly(ethylene glycol)ester linkers and the like.
  • linkers comprises linkers of Formula Y'-CMP below (where Y' indicates the remaining portion of the linker and R 9 and Z are as defined). More particularly, in the group of linkers including Formula Y'-CMP, in several embodiments the R 9 substituent is an ethylacetamido group, and even more particularly the ethylacetamido is conjugated with C1 of N-acetylgalactosamine or N-acetylglucosamine.
  • Di-thiol-containing linkers particularly disulfanylethyl carbamate-containing linkers (named including a free amine of X, otherwise named a "disulfanyl ethyl ester" without including the free amine of X) are particularly advantageous in the present compositions as having the ability to cleave and release an antigen in its original form once inside a cell, for example as illustrated below (where Y' indicates the remaining portion of the linker and X' and Z are as defined).
  • n is an integer representing a mixture including from about 1 to 100, particularly about 8 to 90 (e.g., 7, 8, 9, 10, 1 1 , 12, 13, 14, 15, 16, 17, 18, 19, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65 70, 75, 80, 85, 90 or 95), more particularly about 40 to 80 (e.g., 39, 40, 43, 45, 46, 48, 50, 52, 53, 55, 57, 60, 62, 65, 66, 68, 70, 73, 75, 78, 80 or 81 ) ethylene glycol groups, where the mixture typically encompasses the integer specified as n ⁇ 10%;
  • p is an integer representing a mixture including from about 2 to 150, particularly about 20 to 100 (e.g., 18, 19, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65 70, 75, 80, 85, 90, 95, 100 or 105) and more particularly about 30 to 40 (e.g., 28, 29, 30, 31 , 32, 33, 34, 35, 36, 37, 38, 39, 40, 41 , 42, 43 or 44), where the mixture typically encompasses the integer specified as p ⁇ 10%;
  • q is an integer representing a mixture including from about 1 to 44, particularly about 3 to 20 (e.g., 3, 4, 5, 6, 7, 8, 9, 10, 1 1 , 12, 13, 14, 15, 16, 17, 18, 19, 20, 21 or 22) and more particularly about 4 to 12 (e.g., 4, 5, 6, 7, 8, 9, 10, 11 , 12 or 13), where the mixture typically encompasses the integer specified as q ⁇ 10%; and
  • R 8 is -CH 2 - ("methyl”) or -CH 2 -CH 2 -C(CH 3 )(CN)- ("1 -cyano-1 -methyl-propyl" or "CMP").
  • Y' represents the remaining portion of Y (e.g., HS-PEG).
  • W represents a polymer of the same W 1 group, or W is a copolymer (preferably a random copolymer) of the same or different W 1 and W 2 groups, where:
  • p is an integer from 2 to about 150;
  • R 9 is a direct bond, -CH 2 -CH 2 — NH-C(O)- (i.e., an ethylacetamido group or "EtAcN”) or -CH2-CH 2 -(0-CH2-CH 2 )t-NH-C(0)- (i.e., a pegylated ethylacetamido group or "Et- PEGt-AcN”)
  • t is an integer from 1 to 5, (particularly 1 to 3, and more particularly 1 or 2);
  • R 10 is an aliphatic group, an alcohol or an aliphatic alcohol, particularly N-(2- hydroxypropyl)methylacrylamide;
  • Z (not shown) is galactose, glucose, galactosamine, glucosamine, N- acetylgalactosamine or N-acetylglucosamine.
  • Linkers of Formula Yn can be synthesized via certain precursors that render Yn particularly suitable for conjugation to hydrophobic antigens.
  • linkers shown above as Formulae Yh through Yn are synthesized as isomers that are employed without separation.
  • the linkers of Formulae Yh, Yi, Yj and Yn will be a mixture of the 8,9-dihydro-1H-dibenzo[i),/][1 ,2,3]triazolo[4,5-d]azocin-8yl and 8,9-dihydro-3H- dibenzo[b, ][1 ,2,3]triazolo[4,5-d]azocin-8yl structures illustrated below:
  • linkers of Formulae Yk, YL and Ym will be a mixture of the 8,9-dihydro-1 H- dibenzo[3,4:7,8]cycloocta[1 ,2-d][1 ,2,3]triazol-8-yl and 8,9-dihydro-1 H-dibenzo[3,4:7,8]cycloocta[1 ,2- d][1 ,2,3]triazol-9-yl stru
  • the galactosylating moieties employed in the compositions of the present disclosure serve to target the compositions to liver cells (for example, specifically binding hepatocytes) and can be selected from: galactose, galactosamine or N-acetylgalactosamine.
  • the glucosylating moieties employed in the compositions of the present disclosure serve to target the compositions to liver cells (for example, specifically binding hepatocytes or LSECs) and can be selected from: glucose, glucosamine or N-acetylglucosamine.
  • liver-targeting moieties include galactose or glucose conjugated at C1 or C6, galactosamine or glucosamine conjugated at C2, and N-acetylgalactosamine or N-acetylglucosamine conjugated at C6.
  • Other particular liver-targeting moieties include N-acetylgalactosamine or N- acetylglucosamine conjugated at C2, more particularly conjugated to a linker bearing an R 9 substituent that is CH 2 .
  • liver-targeting moieties include galactose, galactosamine, N-acetylgalactosamine, glucose, glucosamine or N-acetylglucosamine conjugated at C1 , more particularly conjugated to a linker bearing an R 9 substituent that is an ethylacetamido group.
  • compositions of Formula 1 can be named using a combination of lUPAC and trivial names.
  • Y is Formula Ya
  • m is 1
  • n is 4
  • Z is N- acetylgalactosamin-2-yl or N-acetylglucosamin-2-yl:
  • composition of the disclosure where X' is tissue transglutaminase, m is 2, n is 4 and Z' is N- acetylgalactosamin-2-yl or N-acetylglucosamin-2-yl can be named (3Z)-((tissue transgultaminase)-1 ,3- diylbis(1 -oxo-1 -((2,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-3-yl)amino)-4,7, 10, 13-tetraoxa- 16, 17-dithiahenicosan-21 -yl-21 -ylidene))bis(triaz-1 -yn-2-ium) chloride.
  • compositions of Formula 1 can be named using an alternative naming system by reference to X and correspondence to one of Formulae 1 a to 1 p (as illustrated in the reaction schemes) followed by recitation of the integers for variables m, n, p and/or q, R , R and identification of the galactosylating moiety and the position at which it is conjugated.
  • the compounds where W is a copolymer are designated by the letter of the "Y group” followed by a "prime” (e.g., F1 c') and include the number and an identification of the comonomers.
  • composition of Formula 1 a where X is ovalbumin, m is 2, n is 4 and Z is N- acetylgalactosamin-2-yl can be named "F1 a-OVA-m 2 -n 4 -2NAcGAL.”
  • the corresponding composition of Formula 1 a where X is ovalbumin, m is 2, n is 4 and Z is N-acetylglucosamin-2-yl can be named "F1 a- OVA-m 2 -n 4 -2NAcGLU.”
  • both isomers can be named "Fl n-insulin-m ⁇ n ⁇ Pi-q ⁇ CMP-EtAcN-I NAcGAL" (or ""Fl n-insulin-mi-ni-pi-q ⁇ CMP-EtAcN-I NAcGLU" because no stereochemistry is shown for the sugar ring) where CMP indicates that R 8 is 1 -cyano-1 -methyl-propyl, EtAcN indicates that R 9 is ethylacetamido and I NAcGAL indicates Z" is N-acetylgalactosamine conjugated at C1. Absence of the abbreviation EtAcN before the designation for Z would indicate that R 9 is a direct bond.
  • composition of Formula 1 exemplifies compounds where W is a copolymer :
  • the compound can be named "F1 c'-DQ8-relevant Alpha Gliadin-rrvn ⁇ p t-CMP-poly ⁇ EtPEGiAcN-l NAcGLU 2 -HPMA 2 )".
  • compositions of Formula 1 can be prepared, for example, by adjusting the procedures described in Zhu, L, et al., Bioconjugate Chem. 2010, 27, 21 19-2127. Syntheses of certain compositions of Formula 1 are also described below with reference to Reaction Schemes 1 to 14. Other synthetic approaches will be apparent to those skilled in the art.
  • R 1 is a free surface amino (-NH 2 ) or thiol (-SH) moiety positioned on X's three-dimensional structure so as to be accessible for conjugation to a linker, and X' represents the remainder of X excluding the identified free amino group(s) [(X" is used in the reaction schemes to represent the remainder of X excluding free thiol group(s)].
  • X there will be at least one (the N-terminal amine) and can be multiple R 1 groups (predominantly from lysine residues or cysteine residues that are not involved in disulfide bonding), as represented by m, which is an integer from about 1 to 100, more typically 1 or from about 4 to 20, and most typically 1 to about 10.
  • R 1 groups randominantly from lysine residues or cysteine residues that are not involved in disulfide bonding
  • m which is an integer from about 1 to 100, more typically 1 or from about 4 to 20, and most typically 1 to about 10.
  • R 2 is OH or a protecting group
  • R 3 is OH, NH 2 , NHAc, a protecting group or NH-protecting group
  • R 4 is OH or a protecting group
  • R 5 is OH or a protecting group
  • R 6 is OH or a protecting group
  • Z' is galactose or glucose conjugated at C1 or C6, galactosamine or glucosamine conjugated at C2, or N-acetylgalactosamine conjugated or N-acetylglucosamine at C6;
  • R 8 is -CH 2 - or -CH 2 -CH 2 -C(CH 3 )(CN)-;
  • R 9 is a direct bond and Z" is N-acetylgalactosamine conjugated at C2;
  • R 9 is an ethylacetamido or a pegylated ethylacetamido group and Z" is galactose, glucose, galactosamine, glucosamine, N-acetylgalactosamine or N-acetylglucosamine conjugated at C1.
  • solvent inert under the conditions of the reaction being described in conjunction therewith [including, for example, benzene, toluene, acetonitrile, tetrahydrofuran (“THF”), dimethylformamide (“DMF”), chloroform, methylene chloride (or dichloromethane), diethyl ether, methanol, pyridine and the like].
  • solvents used in the reactions of the present disclosure are inert organic solvents.
  • q.s means adding a quantity sufficient to achieve a stated function, e.g., to bring a solution to the desired volume (i.e., 100%).
  • Isolation and purification of the compounds and intermediates described herein can be effected, if desired, by any suitable separation or purification procedure such as, for example, filtration, extraction, crystallization, column chromatography, thin-layer chromatography or thick-layer chromatography, centrifugal size exclusion chromatography, high-performance liquid chromatography, recrystallization, sublimation, fast protein liquid chromatography, gel electrophoresis, dialysis, or a combination of these procedures.
  • suitable separation and isolation procedures can be had by reference to the examples hereinbelow. However, other equivalent separation or isolation procedures can, of course, also be used.
  • Reaction Scheme 1 illustrates the preparation of compositions of Formula 1 where Z can be galactose, glucose, galactosamine, glucosamine, N-acetylgalactosamine or N-acetylglucosamine.
  • Z' as employed in Reaction Scheme 1 encompasses galactose or glucose conjugated at C1 and C6 and corresponding to the following structures according to Formula 1 :
  • a pyridyl di-thiol-poly(ethylene glycol)-NHS ester (Formula 104) is contacted with galactosamine or glucosamine (Formula 105 where R 3 is NH 2 and R 2 , R 4 , R 5 and R 6 are OH) with stirring at about pH 8 for about 1 hour to give the corresponding pyridyl di-thiol- poly(ethylene glycol)-sugar of Formula 106A, which can be used without further purification.
  • Step 4 the acid of Formula 109 is contacted with a protected galactose or N-acetylgalactosamine of Formula 105 where R 2 is OH and R 3 , R 4 , R 5 and R 6 are protecting groups ("PG"), where R 6 is OH and R 2 , R 3 , R 4 and R 5 are PG, or where R 6 is N-acetyl and R 2 , R 3 , R 4 and R 5 are PG to give the corresponding pyridyl di-thiol-poly(ethylene glycol)-sugars of Formulae 106B, 106C and 106D, which can be used following de-protection.
  • R 2 is OH and R 3 , R 4 , R 5 and R 6 are protecting groups
  • PG protecting groups
  • Step 5 to a stirred solution of the product of Step 1 (Formula 103') is added an excess (corresponding to the value of m) of the product of Step 2 or Step 4 (Formula 106, i.e., 106A, 106B, 106C or 106D) for about 1 hour, followed by centrifugal sized exclusion chromatography to remove any free remaining reactants to yield the corresponding product according to Formula 1 a, respectively, Formula 1 aA, Formula 1 aB, Formula 1 aC and Formula 1 aD.
  • compositions corresponding to Formula 1 a can be named, respectively, e.g., as follows:
  • Reaction Schemes 2-14 illustrate preparation of the compounds where W is a polymer of the same W 1 group.
  • Z refers to N-acetylgalactosamine or N-acetylglucosamine conjugated at C2:
  • the C1 conjugated compositions can be protected during synthesis, for example by cyclizing the amine with the C3 hydroxyl and de-protecting following incorporation of the protected galactosamine into the adjacent portion of the linker.
  • the poly(galactose methacrylate) and poly(glucose methacrylate) reactants of Formulae 201 , 401 , 501 , 601 , 701 , 803 and 1401 can be prepared by methacrylating galactose or glucose, e.g., contacting galactosamine or glucosamine and methacrylate anhydride, followed by reversible addition- fragmentation chain transfer (RAFT) polymerization with a corresponding RAFT agent in the presence of azobisisobutyronitrile (AIBN) in a suitable solvent, starting with freeze-thaw cycles followed by heating at about 60-80°C, preferably 70°C for about 5-8, preferably about 6 hours.
  • the polymer can be precipitated in a lower alkanol, preferably methanol.
  • compositions of Formula 1 b can be named as follows:
  • composition of Formula 1 b where X' is uricase, m is 1 , n is 4, p is 4 and Z" is N- acetylgalactosamine conjugated at C2 can be named "Fl b-uricase-nvn ⁇ p ⁇ NAcGAL" or "30-(uricase)- 3,5J,9-tetramethyl-12-oxo-1 -phenyl-1 -thioxo-3,5,7,9-tetrakis((2,4,5-trihydroxy-6- (hydroxymethyl)tetrahydro-2H-pyran-3-yl)carbamoyl)-13, 16, 19,22-tetraoxa-2,25,26-trithiatriacontan-30- iminium".
  • an antigen, antibody, antibody fragment or ligand having native free surface thiol group(s) (cysteines) [Formula 101 " corresponding to Formula 101 and illustrating where X", as the term will be subsequently employed, represents X excluding the identified free surface thiol group(s)] is contacted with an excess (corresponding to the value of m) of a pyridyl di- thiol-poly(ethylene glycol) of Formula 201 to yield the corresponding product according to Formula 1 c.
  • compositions corresponding to Formula 1 c can be named as follows:
  • an antigen, antibody, antibody fragment or ligand having native free surface thiol group(s) of Formula 101 " is contacted with an excess (corresponding to the value of m) of a pyridyl di-thiol of Formula 401 to yield the corresponding product according to Formula 1 d.
  • compositions corresponding to Formula 1 d can be named as follows:
  • compositions corresponding to Formula 1 e can be named as follows:
  • an antigen, antibody, antibody fragment or ligand having native free surface amino group(s) of Formula 101 ' is contacted with an excess (corresponding to the value of m) of a n-nitrophenyl carbonate poly(ethylene glycol)ester of Formula 601 to yield the corresponding product according to Formula 1f.
  • compositions corresponding to Formula 1f can be named as follows:
  • an antigen, antibody, antibody fragment or ligand having native free surface amino group(s) of Formula 101 ' is contacted with an excess (corresponding to the value of m) of a NHS-ester poly(ethylene glycol)ester of Formula 701 to yield the corresponding product according to Formula 1 g.
  • the compositions corresponding to Formula 1 g can be named as follows:
  • Step 1 an antigen, antibody, antibody fragment or ligand having native free surface amino group(s) of Formula 101 ' is contacted with an excess (corresponding to the value of m) of an amine-reactive linker for Click chemistry of Formula 801 to yield the corresponding product according to Formula 802.
  • Step 2 the product of Formula 802 is then contacted with an equivalent amount (again corresponding to the value of m) of a galactos(amine) polymer of Formula 803 to yield the corresponding isomeric product according to Formula 1 h.
  • the two isomers, illustrated above, result from non-specific cyclization of the azide of Formula 803 with the triple bond of Formula 802. Such non-specific cyclization occurs in the synthesis of other compositions where Y is selected from Formulae Yh through Yn, but will not be illustrated in each instance.
  • compositions corresponding to Formula 1 h can be named as follows:
  • Step 1 an antigen, antibody, antibody fragment or ligand having native free surface thiol group(s) of Formula 101 " is contacted with an excess (corresponding to the value of m) of a thiol-reactive linker for Click chemistry of Formula 901 to yield the corresponding product according to Formula 902".
  • Step 2 the product of Formula 902" is then contacted with an equivalent amount (again corresponding to the value of m) of a galactos(amine) polymer of Formula 803 to yield the corresponding isomeric product according to Formula 1 i.
  • compositions corresponding to Formula 1 i can be named as follows:
  • compositions corresponding to Formula 1j can be named as follows:
  • Step 1 an antigen, antibody, antibody fragment or ligand having native free surface thiol group(s) of Formula 101 " is contacted with an excess (corresponding to the value of m) of a thiol-reactive linker for Click chemistry of Formula 1001 to yield the corresponding product according to Formula 1002.
  • Step 2 the product of Formula 1002 is then contacted with an equivalent amount (again corresponding to the value of m) of a galactos(amine) polymer of Formula 803 to yield the corresponding isomeric product according to Formula 1 k.
  • compositions corresponding to Formula 1 k can be named as follows:
  • compositions corresponding to Formula 1 i_ can be named as follows:
  • an azide-modified uRAFT agent (Formula 1 106) is added to a solution of the product of Formula 1 105 with azobisisobutyronitrile, subjected to 4 free-pump-thaw cycles and then stirred at 70°C. After several hours the corresponding polymer product of Formula 1 107 is precipitated by addition of a lower alkanol followed by solvent removal.
  • R 3 is NH-protecting group (e.g., cyclized with R 4 ) the protecting group(s) is(are) removed at this point.
  • an antigen, antibody, antibody fragment or ligand having native free surface amino group(s) of Formula 101 ' is added to a pH 8.0 buffer and contacted with an excess (corresponding to the value of m) of a dioxopyrrolidine of Formula 1 108 with stirring. After 1 hour, unreacted Formula 1 108 is removed and the resulting product of Formula 1 109 is used without further purification.
  • compositions corresponding to Formula 1 m can be named as follows:
  • Reaction Scheme 12 The synthetic approach of Reaction Scheme 12 is particularly suitable for hydrophobic antigens, antibodies, antibody fragments and ligands (e.g., Insulin) due to the use of organic solvents.
  • ligands e.g., Insulin
  • Step 1 an antigen, antibody, antibody fragment or Iigand having native free surface amino group(s) of Formula 101 ' is dissolved in an organic solvent (e.g., DMF) containing triethylamine. To this is added an amount (corresponding to the value of m) of a compound of Formula 1201 followed by stirring and the addition of t-butyl methyl ether. The corresponding product of Formula 1202 is recovered as a precipitate.
  • an organic solvent e.g., DMF
  • the product of Formula 1202 is resuspended in the organic solvent and an amount (corresponding to the value of m) of Formula 1 107 (obtained, e.g., as described with reference to Reaction Scheme 1 1) is added followed by stirring.
  • the reaction product is precipitated via the addition of dichloromethane, followed by filtration and solvent removal. Purification (e.g., resuspension in PBS followed by centrifugal size exclusion chromatography yields the corresponding isomeric product of Formula 1 n.
  • compositions corresponding to Formula 1 n can be named as follows:
  • Step 1 a nitrophenoxycarbonyl-oxyalkyl di-thiol-poly(ethylene glycol)-NHS ester (Formula 1301 ) is contacted with galactose, galactosamine or N-acetylgalactosamine (Formula 105) to give the corresponding product of Formula 1302, along with the other two illustrated products, from which the desired nitrophenoxycarbonyl di-thiol-poly(ethylene glycol)-carboxyethyl galactose, galactosamine or N-acetylgalactosamine of Formula 1302 is isolated before proceeding to the next step.
  • an antigen, antibody, antibody fragment or ligand having native free surface amino group(s) of Formula 101 ' is contacted with an excess (corresponding to the value of m) of the product of Formula 1302 to yield the corresponding product according to Formula 1o.
  • compositions corresponding to Formula 1 o can be named as follows:
  • an antigen, antibody, antibody fragment or ligand having native free surface amino group(s) (Formula 101 ') is contacted with an excess (corresponding to the value of m) of a pyridyl di-thiol-poly(ethylene glycol)-NHS ester of Formula 1401 to yield the corresponding product according to Formula 1 p.
  • compositions corresponding to Formula 1 p can be named as follows:
  • Reaction Schemes 15-18 illustrate preparation of the compounds where W is a copolymer of the same or different W 1 and W 2 groups.
  • Step 1 galactose or glucose (Formula1 101 where R 3 and R 4 are OH), protected galactosamine or protected glucosamine (Formula 1 101 where R 3 is NH- protecting group, e.g., cyclized with R 4 ) or N-acetyl-D-galactosamine or N-acetyl-D-glucosamine (Formula1 101 where R 3 is NHAc and R 4 is OH) is contacted with a 2-(poly-(2- chloroethoxy)ethoxy)ethan-1 -ol of Formula 1501 (where t is 1 to 5) followed by cooling and the dropwise addition of acetylchloride. The solution is warmed to room temperature and then heated to 70°C for several hours. Ethanol is added to the crude product and the resulting solution is stirred in the presence of carbon and then filtered followed by solvent removal to yield the corresponding product of Formula 1502.
  • the product of Formula 1504 is added to a solution of methacrylate anhydride. Triethylamine is added and the reaction stirred for 2 hours followed by solvent removal and isolation to yield the corresponding product of Formula 1505. Alternatively, pentafluorophenyl methacrylate (or another acrylating agent) can be used to prepare the corresponding product of Formula 1505. In some embodiments, the product of formula 1504 is added to DMF. Triethyl amine (e.g., an organic base) is added and the mixture is cooled (e.g., to 4°C using an ice bath).
  • Triethyl amine e.g., an organic base
  • pentafluorophenyl methacrylate (or another acrylating agent) is added (e.g., drop-wise with constant stirring). After a period of time (e.g., 30 minutes), the cooling (e.g., ice-bath) is removed and the reaction is allowed to stir at room temperature for a period of time (e.g., 4 hours). In some embodiments, the solvent is then removed. In some embodiments, the product is purified using flash chromatography.
  • an azide-modified uRAFT agent of Formula 1 106 and a methacrylamide of Formula 1506 are added to a solution of the product of Formula 1505 with azobisisobutyronitrile, subjected to 4 free-pump-thaw cycles and then stirred at 70°C. After several hours the corresponding random copolymer product of Formula 1507 is precipitated by addition of a lower alkanol or acetone followed by solvent removal.
  • R 3 is NH-protecting group (e.g. , cyclized with R 4 ) the protecting group(s) is(are) removed at this point.
  • Step 6 the product of Formula 1507 is added to a pH 8.0 buffer, to which is added the product of Formula 1 109 (prepared, for example, as described with reference to Reaction Scheme 1 1 ). After stirring for 2 hours, the excess Formula 1 109 is removed to yield the corresponding isomeric random copolymer product of Formula 1 m'.
  • compositions corresponding to Formula 1 m' can be named as follows:
  • Step 1 a compound of Formula 1601 is contacted with compounds of Formulae 1505 and 1506 under conditions analogous to those of Reaction Scheme 15, Step 5, to afford the corresponding compound of Formula 1602.
  • t is an integer from about 1 to about 10 or about 1 to about 5.
  • an oligoethylene glycol (1650) is reacted with p-toluenesulfonyl chloride (or some other agent capable of functionalizing 1650 with a leaving group) to form oligoethylene glycol mono p- toluenesulfonate (1651 )(or some other oligoethylene glycol functionalized with a leaving group).
  • compound 1651 can be reacted with potassium thioacetate to form compound
  • compound 1652 is reacted with 2,2-dithiodipyridine to form compound
  • compound 1653 is coupled to compound 1654 to form compound 1601 a.
  • compositions corresponding to Formula 1 c' can be named as follows:
  • Step 1 a compound of Formula 600' is contacted with compounds of Formulae 1505 and 1506 under conditions analogous to those of Reaction Scheme 15, Step 5, to afford the corresponding compound of Formula 601 '.
  • compositions corresponding to Formula 1f can be named as follows:
  • compositions corresponding to Formula 1 g' can be named as follows:
  • a compound of Formula 103' is contacted with an excess (corresponding to the value of m) of a compound of Formula 106 to give the corresponding product of Formula 1 a.
  • a compound of Formula 103' is contacted with an excess (corresponding to the value of m) of a compound of Formula 201 to give the corresponding product of Formula 1 b.
  • a compound of Formula 802, 902 or 1002 is contacted with an excess (corresponding to the value of m) of a compound of Formula 803 to give the corresponding product of Formula 1 h, Formula 1 i or Formula 1 k, respectively.
  • a compound of Formula 1 109 is contacted with an excess (corresponding to the value of m) of a compound of Formula 1 107 to give the corresponding product of Formula 1 m, particularly where n is about 80, p is about 30, q is about 4, and m being a function of the antigen is about 2 to 10.
  • a compound of Formula 1202 is contacted with an excess (corresponding to the value of m) of a compound of Formula 1 107 to give the corresponding product of Formula 1 n, particularly where n is about 1 , p is about 30, q is about 4, and m being a function of the antigen is about 2 to 10.
  • a compound of Formula 1507 is contacted with a compound of Formula 1 109 to give the corresponding product of Formula 1 m', particularly where n is about 4, p is about 90, q is about 4, t is about 1 or 2, R 3 is NHAc, R 4 is OH, R 8 is CMP, R 10 is 2-hydroxypropyl and m being a function of the antigen is about 1 to 10.
  • a compound of Formula 101 " is contacted with a compound of Formula 1602 to give the corresponding product of Formula 1 c', particularly where n is about 4, p is about 90, t is about 1 or 2, R 3 is NHAc, R 4 is OH, R 8 is CMP, R 10 is 2-hydroxypropyl and m being a function of the antigen is about 1 to 10.
  • a compound of Formula 101 ' is contacted with a compound of Formula 601 ' to give the corresponding product of Formula 1f, particularly where n is about 4, p is about 90, t is about 1 or 2, R 3 is NHAc, R 4 is OH, R 8 is CMP, R 10 is 2-hydroxypropyl and m being a function of the antigen is about 1 to 10.
  • a compound of Formula 101 ' is contacted with a compound of Formula 701 ' to give the corresponding product of Formula 1 g', particularly where n is about 4, p is about 90, t is about 1 or 2, R 3 is NHAc, R 4 is OH, R 8 is CMP, R 10 is 2-hydroxypropyl and m being a function of the antigen is about 1 to 10.
  • compositions, pharmaceutical formulations, methods of manufacture and use of the present disclosure are the following combinations and permutations of substituent groups of Formula 1 (sub-grouped, respectively, in increasing order of preference):
  • X is a foreign transplant antigen against which transplant recipients develop an unwanted immune response, a foreign antigen to which patients develop an unwanted immune response, a therapeutic protein to which patients develop an unwanted immune response, a self-antigen to which patients develop an unwanted immune response, or a tolerogenic portion thereof.
  • X is a therapeutic protein to which patients develop an unwanted immune response selected from:
  • X is Abciximab, Adalimumab, Agalsidase alfa, Agalsidase beta, Aldeslukin, Alglucosidase alfa, Factor VIII, Factor IX, Infliximab, L-asparaginase, Laronidase, Natalizumab, Octreotide, Phenylalanine ammonia-lyase (PAL), or Rasburicase (uricase).
  • PAL Phenylalanine ammonia-lyase
  • Rasburicase uricase
  • X is Factor VIII, Factor IX, uricase, PAL or asparaginase.
  • X is a self-antigen polypeptide selected for treating type 1 diabetes mellitus, pediatric multiple sclerosis, juvenile rheumatoid arthritis, celiac disease, or alopecia universalis.
  • X is a self-antigen polypeptide selected for treating new onset type 1 diabetes mellitus, pediatric multiple sclerosis or celiac disease.
  • X is a foreign antigen to which patients develop an unwanted immune response o From peanut, including conarachin (Ara h 1)
  • X is a foreign transplant antigen against which transplant recipients develop an unwanted immune response, e.g. a human leukocyte antigen protein.
  • X is an antibody, antibody fragment or ligand that specifically binds a circulating protein or peptide or antibody, which circulating protein or peptide or antibody gives rise to transplant rejection, immune response against a therapeutic agent, autoimmune disease, and/or allergy.
  • X binds an endogenous circulating protein or peptide or antibody.
  • Y is a linker selected from: Formula Ya, Formula Yb, Formula Yh, Formula Yi, Formula Yk, Formula Ym, Formula Yn, Formula Yo and Formula Yp.
  • n 8 to 90 ⁇ 10%
  • p 20 to 100 ⁇ 10%
  • q is 3 to 20 ⁇ 3.
  • n 40 to 80 ⁇ 10%
  • p is 30 to 40 ⁇ 10%
  • q is 4 to 12 ⁇ 3.
  • Y is Formula Ya, Formula Yb, Formula Ym or Formula Yn.
  • n 8 to 90 ⁇ 10%
  • p 20 to 100 ⁇ 10%
  • q is 3 to 20 ⁇ 3.
  • n 40 to 80 ⁇ 10%
  • p 30 to 40 ⁇ 10%
  • q 4 to 12 ⁇ 3.
  • R 8 is CMP
  • R 8 is CMP
  • Y is a linker selected from: Formula Yc, Formula Yf, Formula Yg and Formula Ym.
  • W p is a random copolymer in which R 9 is Et-PEG t -AcN and R 10 is 2- hydroxypropyl.
  • Z is galactose, galactosamine, N-acetylgalactosamine, glucose, glucosamine or N- acetylglucosamine.
  • Z is galactose or N-acetylgalactosamine conjugated at C1 , C2 or C6.
  • Z is galactose or N-acetylgalactosamine conjugated at C1 or C2.
  • Z is N-acetylgalactosamine conjugated at C1. o Especially where Z is glucose or N-acetylglucosamine conjugated at C1 , C2 or C6. ⁇ Particularly where Z is glucose or N-acetylglucosamine conjugated at C1 or C2. • More particularly where Z is N-acetylglucosamine conjugated at C1.
  • X is a self-antigen polypeptide selected for treating type 1 diabetes mellitus, pediatric multiple sclerosis, juvenile rheumatoid arthritis, celiac disease, or alopecia universalis.
  • X is a self-antigen polypeptide selected for treating new onset type 1 diabetes mellitus, pediatric multiple sclerosis or celiac disease.
  • Y is a linker selected from: Formula Ya, Formula Yb, Formula Yc, Formula Yf, Formula Yg, Formula Yh, Formula Yi, Formula Yk, Formula Ym, Formula Yn, Formula Yo and Formula Yp.
  • W p is a W 1 polymer in which R 9 is Et-PEG t -AcN or a random copolymer in which R 9 is Et-PEG t -AcN and R 10 is 2- hydroxypropyl.
  • W p is a random copolymer and includes about 30 W 1 and 60 W 2 comonomers.
  • n 8 to 90 ⁇ 10%
  • p 20 to 100 ⁇ 10%
  • q 3 to 20 ⁇ 3.
  • n 40 to 80 ⁇ 10%
  • p is 30 to 40 ⁇ 10%
  • q is 4 to 12 ⁇ 3.
  • n 8 to 90 ⁇ 10%
  • p 20 to 100 ⁇ 10%
  • q is 3 to 20 ⁇ 3.
  • n 40 to 80 ⁇ 10%
  • p is 30 to 40 ⁇ 10%
  • q is 4 to 12 ⁇ 3.
  • Z is galactose, galactosamine or N- acetylgalactosamine.
  • Z is galactose or N-acetylgalactosamine conjugated at C1 , C2 or C6.
  • Z is galactose or N- acetylgalactosamine conjugated at C1 or C2.
  • Z is glucose, glucosamine or N-acetylglucosamine.
  • Z is glucose or N-acetylglucosamine conjugated at C1 , C2 or C6.
  • Z is glucose or N- acetylglucosamine conjugated at C1 or C2.
  • Y is a linker selected from: Formula Yc, Formula Yf, Formula Yg and Formula Ym.
  • W p is a random copolymer in which R 9 is Et-PEG r AcN and R 10 is 2-hydroxypropyl.
  • p is about 90 and includes about 30 W 1 and 60 W 2 comonomers.
  • Y is a linker selected from: Formula Yc and Formula Ym.
  • W p is a random copolymer in which R 9 is Et-PEG r AcN and R 10 is 2-hydroxypropyl.
  • p is about 90 and includes about 30 W 1 and 60 W 2 comonomers.
  • Z is galactose, galactosamine or N-acetylgalactosamine.
  • Z is galactose or N-acetylgalactosamine conjugated at C1 , C2 or C6.
  • Z is galactose or N-acetylgalactosamine conjugated at C1 or C2.
  • Z is glucose, glucosamine or N-acetylglucosamine.
  • Z is glucose or N-acetylglucosamine conjugated at C1 , C2 or C6.
  • Z is glucose or N-acetylglucosamine conjugated at C1 or C2.
  • Y is a linker selected from: Formula Ya, Formula Yb, Formula Yh, Formula Yi, Formula Yk, Formula Ym, Formula Yn, Formula Yo and Formula Yp.
  • Y is a linker selected from: Formula Yc, Formula Yf, Formula Yg and Formula Ym.
  • W p is a random copolymer in which R 9 is Et-PEG r AcN and R 10 is 2-hydroxypropyl.
  • p is about 90 and includes about 30 W 1 and 60 W 2 comonomers.
  • Y is a linker selected from: Formula Yc and Formula Ym.
  • W p is a random copolymer in which R 9 is Et-PEG t -AcN and R 10 is 2-hydroxypropyl.
  • p is about 90 and includes about 30 W 1 and 60 W 2 comonomers.
  • n 8 to 90 ⁇ 10%
  • p 20 to 100 ⁇ 10%
  • q is 3 to 20 ⁇ 3.
  • n 40 to 80 ⁇ 10%
  • p 30 to 40 ⁇ 10%
  • q 4 to 12 ⁇ 3.
  • n 8 to 90 ⁇ 10%
  • p 20 to 100 ⁇ 10%
  • q 3 to 20 ⁇ 3.
  • n 40 to 80 ⁇ 10%
  • p is 30 to 40 ⁇ 10%
  • q is 4 to 12 ⁇ 3.
  • Z is galactose, galactosamine or N-acetylgalactosamine.
  • Z is galactose or N-acetylgalactosamine conjugated at C1 , C2 or C6.
  • Z is galactose or N-acetylgalactosamine conjugated at C1 or C2.
  • Y is a linker selected from: Formula Yc, Formula Yf, Formula Yg and Formula Ym.
  • W p is a random copolymer in which R 9 is Et- PEG,-AcN and R 10 is 2-hydroxypropyl.
  • p is about 90 and includes about 30 W 1 and 60 W 2 comonomers.
  • Z is glucose, glucosamine or N-acetylglucosamine.
  • Z is glucose or N-acetylglucosamine conjugated at C1 , C2 or C6.
  • Z is glucose or N-acetylglucosamine conjugated at C1 or C2.
  • Y is a linker selected from: Formula Yc, Formula Yf, Formula Yg and Formula Ym.
  • W p is a random copolymer in which R 9 is Et- PEGt-AcN and R 10 is 2-hydroxypropyl.
  • p is about 90 and includes about 30 W 1 and 60 W 2 comonomers.
  • Y is a linker selected from: Formula Yc and Formula Ym.
  • W p is a random copolymer in which R 9 is Et- PEGt-AcN and R 10 is 2-hydroxypropyl.
  • p is about 90 and includes about 30 W 1 and 60 W 2 comonomers.
  • o m is 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 1 1 , 12, 13, 14, 15, 16, 17, 18, 19, 20, 25, 30, 35, 40, 45,
  • m is from about 1 to 20.
  • ⁇ m is 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 1 1 , 12, 13, 14, 15, 16, 17, 18, 19, 20, 21 or 22.
  • m is about 10. • m is 9, 10 or 1 1.
  • an integer representing a mixture including from about 1 to 100
  • o n is 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 1 1 , 12, 13, 14, 15, 16, 17, 18, 19, 20, 22, 25, 30, 34, 35, 37, 40, 41 , 45, 50, 54, 55, 59, 60, 65, 70, 75, 80, 82, 83, 85, 88, 90, 95, 99, 100, 105 or 1 10.
  • n is about 8 to 90.
  • n is 8, 9, 10, 1 1 , 12, 13, 14, 15, 16, 17, 18, 19, 20, 22, 25, 30, 34, 35, 37, 40, 41 , 45, 50, 54, 55, 59, 60, 65, 70, 75, 80, 82, 83, 85, 88, 90, 95 or 99.
  • n is about 40 to 80.
  • n is 37, 40, 41 , 45, 50, 54, 55, 59, 60, 65, 70, 75, 80, 82, 83 or 88.
  • o n represents a mixture encompassing the ranges 1 -4, 2-4, 2-6, 3-8, 7-13, 6-14, 15-25, 26-30, 42-50, 46-57, 60-82, 85-90, 90-1 10 and 107-1 13.
  • n represents a mixture encompassing the ranges 7-13, 6-14, 15-25, 26-30, 42-50, 46-57, 60-82, 85-90 and 82-99.
  • n represents a mixture encompassing the ranges 36- 44, 42-50, 46-57, 60-82 and 75-85.
  • o p is an integer representing a mixture including from about 2 to 150.
  • ⁇ p is 2, 3, 4, 5, 6, 7, 8, 9, 10, 1 1 , 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, 1 10, 120, 130, 140, 150, 160 or 165.
  • n is an integer representing a mixture including from about 1 to 100.
  • n is 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 1 1 , 12, 13, 14, 15, 16, 17, 18, 19, 20, 22, 25, 30, 34, 35, 37, 40, 41 , 45, 50, 54, 55, 59, 60, 65, 70, 75, 80, 82, 83, 85, 88, 90, 95, 99, 100, 105 or 1 10.
  • n is about 8 to 90.
  • n 8 9, 10, 1 1 , 12, 13, 14, 15, 16, 17, 18, 19,
  • n is about 40 to 80.
  • n is 37, 40, 41 , 45, 50, 54, 55, 59, 60, 65, 70, 75, 80, 82, 83 or 88.
  • p is 18, 19, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65 70, 75, 80, 85, 90, 95, 100 or 110.
  • n is an integer representing a mixture including from about 1 to 100.
  • n is 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 1 1 , 12, 13,
  • n is about 8 to 90.
  • n is 8, 9, 10, 1 1 , 12, 13, 14, 15, 16, 17, 18, 19, 20, 22, 25, 30, 34, 35, 37, 40, 41 , 45, 50, 54, 55, 59, 60, 65, 70, 75, 80, 82, 83, 85, 88, 90, 95 or 99.
  • n is about 40 to 80.
  • n 37, 40, 41 , 45, 50, 54, 55, 59, 60, 65, 70, 75, 80, 82, 83 or 88.
  • p is 27, 28, 29, 30, 31 , 32, 33, 34, 35, 36, 37, 38, 39, 40, 41 , 42, 43, or 44.
  • n is 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 1 1 , 12, 13, 14, 15, 16, 17, 18, 19, 20, 22, 25, 30, 34, 35, 37, 40, 41 , 45, 50, 54, 55, 59, 60, 65, 70, 75, 80, 82, 83, 85, 88, 90, 95, 99, 100, 105 or 1 10.
  • n is about 8 to 90.
  • n is 8, 9, 10, 1 1 , 12, 13, 14, 15, 16, 17, 18, 19, 20, 22, 25, 30, 34, 35, 37, 40, 41 , 45, 50, 54, 55, 59, 60, 65, 70, 75, 80, 82, 83, 85, 88, 90, 95 or 99.
  • n is about 40 to 80.
  • n 37, 40, 41 , 45, 50, 54, 55, 59, 60, 65, 70, 75, 80, 82, 83 or 88.
  • o q is an integer representing a mixture including from about 1 to 44.
  • ⁇ q is 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 1 1 , 12, 13, 14, 15, 16, 17, 18, 19, 20, 25, 30, 35, 40, 44 or 48.
  • compositions of the disclosure find use in a variety of applications including, as will be appreciated by those in the art, treatment of transplant rejection, immune response against a therapeutic agent, autoimmune disease, and food allergy, among other uses.
  • compositions of the disclosure are used to modulate, particularly down-regulate, antigen-specific undesirable immune response.
  • compositions of the disclosure are useful, in additional embodiments, to bind and clear from the circulation specific undesired proteins, including antibodies endogenously generated in a patient (i.e., not exogenous antibodies administered to a patient), peptides and the like, which cause autoimmunity and associated pathologies, allergy, inflammatory immune responses, and anaphylaxis.
  • antigens are targeted to the liver for presentation via antigen-presenting cells to specifically down-regulate the immune system or for clearance of unwanted circulating proteins.
  • This is distinct from previous uses of liver targeting, for example as described in US 2013/0078216, where the purpose of liver-targeting molecules such as DOM26h-196-61 was the delivery of therapeutic agents to treat liver diseases such as fibrosis, hepatitis, Cirrhosis and liver cancer.
  • compositions and methods to treat unwanted immune response to self-antigens and foreign antigens including but not limited to: a foreign transplant antigen against which transplant recipients develop an unwanted immune response (e.g., transplant rejection), a foreign antigen to which patients develop an unwanted immune (e.g., allergic or hypersensitivity) response, a therapeutic agent to which patients develop an unwanted immune response (e.g., hypersensitivity and/or reduced therapeutic activity), a self-antigen to which patients develop an unwanted immune response (e.g., autoimmune disease)
  • Autoimmune disease states that can be treated using the methods and compositions provided herein include, but are not limited to: Acute Disseminated Encephalomyelitis (ADEM); Acute interstitial allergic nephritis (drug allergies); Acute necrotizing hemorrhagic leukoencephalitis; Addison's Disease; Alopecia areata; Alopecia universalis; Ankylosing Spondylitis; Arthritis, juvenile; Arthritis, psoriatic; Arthritis, rheumatoid; Atopic Dermatitis; Autoimmune aplastic anemia; Autoimmune gastritis; Autoimmune hepatitis; Autoimmune hypophysitis; Autoimmune oophoritis; Autoimmune orchitis; Autoimmune polyendocrine syndrome type 1 ; Autoimmune polyendocrine syndrome type 2; Autoimmune thyroiditis; Behcet's disease; Bronchio
  • a particular group of autoimmune disease states that can be treated using the methods and compositions provided herein include, but are not limited to: Acute necrotizing hemorrhagic leukoencephalitis; Addison's Disease; Arthritis, psoriatic; Arthritis, rheumatoid; Autoimmune aplastic anemia; Autoimmune hypophysitis; Autoimmune gastritis; Autoimmune polyendocrine syndrome type 1 ; Bullous pemphigoid; Celiac disease; Coxsackie myocarditis; Dermatitis herpetiformis Duhring; Diabetes mellitus (Type 1); Epidermolysis bullosa acquisita; Giant cell myocarditis; Goodpasture's syndrome; Graves' disease; Hashimoto's thyroiditis; Mixed connective tissue disease; Multiple sclerosis; Myasthenia gravis; Neuromyelitis optica; Pernicious angemis; Pemphigus vulgaris
  • treatment can be provided for reactions against, for example: peanut, apple, milk, egg whites, egg yolks, mustard, celery, shrimp, wheat (and other cereals), strawberry and banana.
  • a patient can be tested to identify a foreign antigen against which an unwanted immune response has developed, and a composition of the disclosure can be developed based on that antigen.
  • compositions and methods of the disclosure specificity in binding to antigen-presenting cells in the liver (particularly binding to hepatocytes and specifically ASGPR) should initially be determined. This can be accomplished, for example, by employing a marker (such as the fluorescent marker phycoerythrin ("PE")) in a composition of the disclosure.
  • PE fluorescent marker
  • the composition is administered to suitable experimental subjects. Controls, e.g., unconjugated PE or vehicle (saline) are administered to other group(s) of subjects.
  • the composition and controls are allowed to circulate for a period of 1 to 5 hours, after which the spleens and livers of the subjects are harvested and measured for fluorescence. The specific cells in which fluorescence is found can be subsequently identified.
  • Compositions of the disclosure when tested in this manner, show higher levels of concentration in the antigen-presenting cells of the liver as compared with unconjugated PE or vehicle.
  • Effectiveness in immune modulation can be tested by measuring the proliferation of OT-I CD8 + cells (transplanted into host mice) in response to the administration of a composition of the disclosure incorporating a known antigen, such as ovalbumin ("OVA"), as compared with administration of the antigen alone or just vehicle.
  • Compositions of the disclosure when tested in this manner, show an increase of OT-I cell proliferation as compared with antigen alone or vehicle, demonstrating increased CD8+ T-cell cross-priming.
  • the proliferating OT-I CD8 + T cells can be phenotypically analyzed for molecular signatures of exhaustion [such as programmed death-1 (PD-1 ), FasL, and others], as well as Annexin-V binding as a hallmark of apoptosis and thus deletion.
  • the OT-I CD8 + T cells can also be assessed for their responsiveness to an antigen challenge with adjuvant in order to demonstrate functional non-responsiveness, and thus immune tolerance, towards the antigen. To do so, the cells are analyzed for inflammatory signatures after administration of compositions of the disclosure into host mice followed by an antigen challenge. Compositions of the disclosure when tested in this manner demonstrate very low (e.g., background) levels of inflammatory OT-I CD8 + T cell responses towards OVA in comparison to control groups, thus demonstrating immune tolerance.
  • Humoral immune response can be tested by administering a composition of the disclosure incorporating a known antigen, such as OVA, as compared with the administration of the antigen alone or just vehicle, and measuring the levels of resulting antibodies.
  • Compositions of the disclosure when tested in this manner show very low (e.g. , background) levels of antibody formation responsive to their administration and the administration of vehicle, with significantly higher levels of antibody formation responsive to administration of the antigen.
  • compositions of the disclosure when tested in this manner show low levels of antibody formation responsive to challenge with the antigen in groups pretreated with such compositions as compared to groups that are not pretreated.
  • NOD non-obese diabetic
  • EAE experimental autoimmune encephalomyelitis
  • the NOD mouse develops spontaneous autoimmune diabetes (similar to type 1 a diabetes in humans).
  • Groups of NOD mice are treated with test compound or a negative control, followed by measurement of blood glucose.
  • Successful treatment corresponds to likelihood of treating diabetes in humans or proof of mechanism for approaches to the treatment of other autoimmune diseases. (See, e.g., Anderson and Bluestone, Annu. Rev. Immunol. 2005;23:447-85.)
  • compositions of the disclosure are administered at a therapeutically effective dosage, e.g., a dosage sufficient to provide treatment for the disease states previously described.
  • Administration of the compounds of the disclosure or the pharmaceutically acceptable salts thereof can be via any of the accepted modes of administration for agents that serve similar utilities.
  • human dosage levels have yet to be optimized for the compounds of the disclosure, these can initially be extrapolated from the about 10 pg to 100 pg doses administered for mice.
  • an individual human dose is from about 0.01 to 2.0 mg/kg of body weight, preferably about 0.1 to 1.5 mg/kg of body weight, and most preferably about 0.3 to 1.0 mg/kg of body weight.
  • Treatment can be administered for a single day or a period of days, and can be repeated at intervals of several days, one or several weeks, or one or several months. Administration can be as a single dose (e.g., as a bolus) or as an initial bolus followed by continuous infusion of the remaining portion of a complete dose over time, e.g., 1 to 7 days.
  • the amount of active compound administered will, of course, be dependent on any or all of the following: the subject and disease state being treated, the severity of the affliction, the manner and schedule of administration and the judgment of the prescribing physician. It will also be appreciated that amounts administered will depend upon the molecular weight of the antigen, antibody, antibody fragment or ligand as well as the size of the linker.
  • compositions of the disclosure can be administered either alone or in combination with other pharmaceutically acceptable excipients. While all typical routes of administration are contemplated (e.g. oral, topical, transdermal, injection (intramuscular, intravenous, or intra-arterial)), it is presently preferred to provide liquid dosage forms suitable for injection.
  • the formulations will typically include a conventional pharmaceutical carrier or excipient and a composition of the disclosure or a pharmaceutically acceptable salt thereof.
  • compositions can include other medicinal agents, pharmaceutical agents, carriers, and the like, including, but not limited to the therapeutic protein, peptide, antibody or antibody-like molecule corresponding to the antigen (X) employed in the composition of the disclosure, and other active agents that can act as immune-modulating agents and more specifically can have inhibitory effects on B-cells, including anti-folates, immune suppressants, cyostatics, mitotic inhibitors, and anti-metabolites, or combinations thereof.
  • active agents that can act as immune-modulating agents and more specifically can have inhibitory effects on B-cells, including anti-folates, immune suppressants, cyostatics, mitotic inhibitors, and anti-metabolites, or combinations thereof.
  • the pharmaceutically acceptable composition will contain about 0.1 % to 95%, preferably about 0.5% to 50%, by weight of a composition of the disclosure, the remainder being suitable pharmaceutical excipients, carriers, etc. Dosage forms or compositions containing active ingredient in the range of 0.005% to 95% with the balance made up from non-toxic carrier can be prepared.
  • Liquid pharmaceutically administrable compositions can, for example, be prepared by dissolving, dispersing, etc. an active composition of the disclosure (e.g. , a lyophilized powder) and optional pharmaceutical adjuvants in a carrier, such as, for example, water (water for injection), saline, aqueous dextrose, glycerol, glycols, ethanol or the like (excluding galactoses), to thereby form a solution or suspension.
  • a carrier such as, for example, water (water for injection), saline, aqueous dextrose, glycerol, glycols, ethanol or the like (excluding galactoses)
  • the pharmaceutical composition to be administered can also contain minor amounts of nontoxic auxiliary substances such as wetting agents, emulsifying agents, stabilizing agents, solubilizing agents, pH buffering agents and the like, for example, sodium acetate, sodium citrate, cyclodextrine derivatives, sorbitan monolaurate, triethanolamine acetate and triethanolamine oleate, etc., osmolytes, amino acids, sugars and carbohydrates, proteins and polymers, salts, surfactants, chelators and antioxidants, preservatives, and specific ligands.
  • auxiliary substances such as wetting agents, emulsifying agents, stabilizing agents, solubilizing agents, pH buffering agents and the like, for example, sodium acetate, sodium citrate, cyclodextrine derivatives, sorbitan monolaurate, triethanolamine acetate and triethanolamine oleate, etc., osmolytes, amino acids, sugars and carbohydrates, proteins and polymers, salts, surfactants,
  • F1aA-OVA-m 4 -n 80 (or F1 a-OVA-m 4 -n 80 -2NGAL)
  • galactosamine (10.0 mg, 0.04638 mmol) was dissolved with stirring in 100 ⁇ of pH 8.0 PBS containing 5 mM EDTA.
  • Pyridyl dithiol-poly(ethylene glycol)-NHS ester (Formula 104 where n is 80) (16.23 mg, 0.00464 mmol) dissolved in 100 ⁇ of pH 7.0 PBS was added to the stirring solution of galactosamine. After 1 hour, the resulting pyridyl dithiol-poly(ethylene glycol)-N- acetylgalactosamine (Formula 106A) was ready to be used without further purification.
  • X Insulin (SEQ ID NO:1) and m is 2,
  • X is MBP13-32 (SEQ ID NO:7) and m is 1,
  • X is MBP111-129 (SEQ ID NO:9) and m is 1,
  • X Tyrosinase (SEQ ID NO: 15) and m is 8,
  • X is Aquaporin-4 (SEQ ID NO:17) and m is 4,
  • X is S-arrestin (SEQ ID NO: 18) and m is 12,
  • X is Alpha-gliadin "33-mer” native (SEQ ID NO:20) and m is 1 ,
  • X Alpha-gliadin (SEQ ID NO:22) and m is 1,
  • X Omega-gliadin (SEQ ID NO:23) and m is 1 ,

Landscapes

  • Health & Medical Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Chemical & Material Sciences (AREA)
  • Animal Behavior & Ethology (AREA)
  • Veterinary Medicine (AREA)
  • Medicinal Chemistry (AREA)
  • Public Health (AREA)
  • Pharmacology & Pharmacy (AREA)
  • General Health & Medical Sciences (AREA)
  • Engineering & Computer Science (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Epidemiology (AREA)
  • Immunology (AREA)
  • Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
  • Organic Chemistry (AREA)
  • General Chemical & Material Sciences (AREA)
  • Chemical Kinetics & Catalysis (AREA)
  • Mycology (AREA)
  • Microbiology (AREA)
  • Diabetes (AREA)
  • Transplantation (AREA)
  • Biochemistry (AREA)
  • Endocrinology (AREA)
  • Molecular Biology (AREA)
  • Rheumatology (AREA)
  • Hematology (AREA)
  • Emergency Medicine (AREA)
  • Obesity (AREA)
  • Ophthalmology & Optometry (AREA)
  • Pain & Pain Management (AREA)
  • Proteomics, Peptides & Aminoacids (AREA)
  • Zoology (AREA)
  • Neurology (AREA)
  • Neurosurgery (AREA)
  • Physical Education & Sports Medicine (AREA)
  • Orthopedic Medicine & Surgery (AREA)
  • Pulmonology (AREA)
  • Dermatology (AREA)
  • Gastroenterology & Hepatology (AREA)
  • Biomedical Technology (AREA)
EP16778870.2A 2015-09-19 2016-09-16 Glycotargeting-therapeutika Withdrawn EP3349784A1 (de)

Applications Claiming Priority (3)

Application Number Priority Date Filing Date Title
US14/859,292 US10946079B2 (en) 2014-02-21 2015-09-19 Glycotargeting therapeutics
US15/185,564 US10046056B2 (en) 2014-02-21 2016-06-17 Glycotargeting therapeutics
PCT/IB2016/001411 WO2017046652A1 (en) 2015-09-19 2016-09-16 Glycotargeting therapeutics

Publications (1)

Publication Number Publication Date
EP3349784A1 true EP3349784A1 (de) 2018-07-25

Family

ID=57121450

Family Applications (1)

Application Number Title Priority Date Filing Date
EP16778870.2A Withdrawn EP3349784A1 (de) 2015-09-19 2016-09-16 Glycotargeting-therapeutika

Country Status (12)

Country Link
EP (1) EP3349784A1 (de)
JP (3) JP7082045B2 (de)
KR (1) KR20180080195A (de)
CN (1) CN108430497B (de)
AU (2) AU2016325073B2 (de)
BR (1) BR112018005409A2 (de)
CA (1) CA2998066A1 (de)
EA (1) EA036102B9 (de)
HK (1) HK1257845A1 (de)
IL (2) IL306015A (de)
MX (1) MX2018003411A (de)
WO (1) WO2017046652A1 (de)

Families Citing this family (12)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US9517257B2 (en) 2010-08-10 2016-12-13 Ecole Polytechnique Federale De Lausanne (Epfl) Erythrocyte-binding therapeutics
US9518087B2 (en) 2010-08-10 2016-12-13 Ecole Polytechnique Federale De Lausanne (Epfl) Erythrocyte-binding therapeutics
US9850296B2 (en) 2010-08-10 2017-12-26 Ecole Polytechnique Federale De Lausanne (Epfl) Erythrocyte-binding therapeutics
US10946079B2 (en) 2014-02-21 2021-03-16 Ecole Polytechnique Federale De Lausanne Glycotargeting therapeutics
ES2874884T3 (es) 2014-02-21 2021-11-05 Ecole Polytechnique Fed De Lausanne Epfl Epfl Tto Compuestos terapéuticos dirigidos a la glucosa
US10046056B2 (en) 2014-02-21 2018-08-14 École Polytechnique Fédérale De Lausanne (Epfl) Glycotargeting therapeutics
US10953101B2 (en) 2014-02-21 2021-03-23 École Polytechnique Fédérale De Lausanne (Epfl) Glycotargeting therapeutics
CA2998066A1 (en) * 2015-09-19 2017-03-23 Ecole Polytechnique Federale De Lausanne Glycotargeting therapeutics
WO2018232176A1 (en) 2017-06-16 2018-12-20 The University Of Chicago Compositions and methods for inducing immune tolerance
EA202092723A1 (ru) * 2018-05-09 2021-04-09 Зе Юниверсити Оф Чикаго Композиции и способы, касающиеся иммунной толерантности
CN111214461B (zh) * 2020-03-24 2021-04-16 河南大学 糖靶向修饰siRNA纳米粒子的制备及应用
CN116819072B (zh) * 2023-08-28 2023-11-17 迪亚莱博（张家港）生物科技有限公司 一种用于抗aqp4自身抗体检测的抗原蛋白组合物及化学发光检测试剂盒

Family Cites Families (10)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2006002382A2 (en) 2004-06-24 2006-01-05 The Scripps Research Institute Arrays with cleavable linkers
CA2786660A1 (en) 2010-01-14 2011-07-21 Glaxo Group Limited Liver targeting molecules
US9517257B2 (en) 2010-08-10 2016-12-13 Ecole Polytechnique Federale De Lausanne (Epfl) Erythrocyte-binding therapeutics
US9518087B2 (en) 2010-08-10 2016-12-13 Ecole Polytechnique Federale De Lausanne (Epfl) Erythrocyte-binding therapeutics
US8501930B2 (en) * 2010-12-17 2013-08-06 Arrowhead Madison Inc. Peptide-based in vivo siRNA delivery system
WO2012167088A1 (en) 2011-06-03 2012-12-06 3M Innovative Properties Company Heterobifunctional linkers with polyethylene glycol segments and immune response modifier conjugates made therefrom
BR112014020304A8 (pt) * 2012-02-15 2018-01-16 Ecole Polytechnique Fed Lausanne Epfl terapêutica para ligação de eritrócitos
CN109705218B (zh) * 2012-08-09 2022-07-19 罗切格利卡特公司 Asgpr抗体及其用途
ES2874884T3 (es) * 2014-02-21 2021-11-05 Ecole Polytechnique Fed De Lausanne Epfl Epfl Tto Compuestos terapéuticos dirigidos a la glucosa
CA2998066A1 (en) 2015-09-19 2017-03-23 Ecole Polytechnique Federale De Lausanne Glycotargeting therapeutics

Also Published As

Publication number Publication date
EA036102B1 (ru) 2020-09-29
IL306015A (en) 2023-11-01
EA036102B9 (ru) 2020-12-30
JP7082045B2 (ja) 2022-06-07
HK1257845A1 (zh) 2019-11-01
AU2016325073B2 (en) 2023-03-16
CN108430497A (zh) 2018-08-21
IL257958A (en) 2018-06-28
JP7434248B2 (ja) 2024-02-20
IL257958B1 (en) 2023-10-01
WO2017046652A1 (en) 2017-03-23
JP2018532722A (ja) 2018-11-08
MX2018003411A (es) 2018-09-06
AU2023203475A1 (en) 2023-06-29
EA201890778A1 (ru) 2018-08-31
JP2024063005A (ja) 2024-05-10
BR112018005409A2 (pt) 2018-10-09
CN108430497B (zh) 2024-05-07
AU2016325073A1 (en) 2018-04-05
KR20180080195A (ko) 2018-07-11
CA2998066A1 (en) 2017-03-23
IL257958B2 (en) 2024-02-01
JP2022028673A (ja) 2022-02-16

Similar Documents

Publication Publication Date Title
AU2016325073B2 (en) Glycotargeting therapeutics
US11793882B2 (en) Glycotargeting therapeutics
US11801305B2 (en) Glycotargeting therapeutics
US11666638B2 (en) Glycotargeting therapeutics
US11654188B2 (en) Glycotargeting therapeutics

Legal Events

Date Code Title Description
STAA Information on the status of an ep patent application or granted ep patent

Free format text: STATUS: THE INTERNATIONAL PUBLICATION HAS BEEN MADE

PUAI Public reference made under article 153(3) epc to a published international application that has entered the european phase

Free format text: ORIGINAL CODE: 0009012

STAA Information on the status of an ep patent application or granted ep patent

Free format text: STATUS: REQUEST FOR EXAMINATION WAS MADE

17P Request for examination filed

Effective date: 20180411

AK Designated contracting states

Kind code of ref document: A1

Designated state(s): AL AT BE BG CH CY CZ DE DK EE ES FI FR GB GR HR HU IE IS IT LI LT LU LV MC MK MT NL NO PL PT RO RS SE SI SK SM TR

AX Request for extension of the european patent

Extension state: BA ME

DAV Request for validation of the european patent (deleted)
DAX Request for extension of the european patent (deleted)
STAA Information on the status of an ep patent application or granted ep patent

Free format text: STATUS: EXAMINATION IS IN PROGRESS

17Q First examination report despatched

Effective date: 20190506

REG Reference to a national code

Ref country code: HK

Ref legal event code: DE

Ref document number: 1257845

Country of ref document: HK

STAA Information on the status of an ep patent application or granted ep patent

Free format text: STATUS: EXAMINATION IS IN PROGRESS

STAA Information on the status of an ep patent application or granted ep patent

Free format text: STATUS: EXAMINATION IS IN PROGRESS

STAA Information on the status of an ep patent application or granted ep patent

Free format text: STATUS: THE APPLICATION HAS BEEN WITHDRAWN

18W Application withdrawn

Effective date: 20240228