Compounds for inhibiting fucosylation and methods for using the same

Abstract

To provide compounds for inhibiting fucosylation and methods for using the same.SOLUTION: Provided is a method of inhibiting fucosyltransferase, comprising contacting the fucosyltransferase with a compound of, or a composition comprising, epicatechin monogallate (ECG), epicatechin-3-monogallate (ECGC), purpurogallin, hexachlorophene, acriflavinum, baicalein, epicatechin-3,5-digallate, theaflavin monogallate, tannic acid, methacycline, anthralin, mitoxanthrone hydrochloride, hycanthone, ethcridine lactate, aurin tricarboxylic acid, carboplatin, cisplatin, primuletin, chrysin, diometin, suramin, hematin, gossypol, or any combination thereof.SELECTED DRAWING: Figure 1

Description

[Technical field]

[0001] The disclosure generally provides compounds, and their uses, for inhibiting fucosylation of proteins, such as antibodies. [Background technology]

[0002] L-fucose, also called 6-deoxy-L-galactose or simply fucose, is a monosaccharide that is a component of some N- and O-linked glycans and glycolipids in animals. Fucose is typically added as a terminal modification of glycans, including those attached to blood group antigens, selectins, and antibodies. Protein fucosylation is thought to play a role in mammalian development, and aberrant protein fucosylation has been proposed to be associated with human disease, including upregulation in cancer and rheumatoid arthritis. Removal of fucosylation has been shown to improve antibody binding and activity. Thus, there is a need for compounds that can be used to inhibit fucosylation. The present embodiments meet this need, as well as others. Summary of the Invention

[0003] In some embodiments, a method for inhibiting fucosyltransferase is provided.In some embodiments, the method comprises contacting fucosyltransferase with a composition comprising purpurogallin, hexachlorophene, acriflavinum, baicalein, epicatechin monogallate, epicatechin-3-monogallate, epicatechin-3,5-digallate, theaflavin monogallate, tannic acid, methacycline, anthralin, mitoxantrone hydrochloride, hycanthone, esculidine lactate, aurintricarboxylic acid, carboplatin, cisplatin, primuletin, chrysin, diometin, suramin, hematin, gossypol, or any combination thereof.In some embodiments, the fucosyltransferase is present in the cell that produces antibody.

[0004] In some embodiments, a method for inhibiting protein fucosylation in a cell is provided. In some embodiments, the method includes contacting the cell with a composition comprising purpurogallin, hexachlorophene, acriflavinum, baicalein, epicatechin monogallate, epicatechin-3-monogallate, epicatechin-3,5-digallate, theaflavin monogallate, tannic acid, methacycline, anthralin, mitoxantrone hydrochloride, hycanthone, esculidine lactate, aurintricarboxylic acid, carboplatin, cisplatin, primuletin, chrysin, diometin, suramin, hematin, gossypol, or any combination thereof to inhibit the fucosylation of the protein.

[0005] In some embodiments, a method for producing an antibody with reduced fucosylation is provided. In some embodiments, the method includes contacting a cell producing the antibody with a composition comprising purpurogallin, hexachlorophene, acriflavinum, baicalein, epicatechin monogallate, epicatechin-3-monogallate, epicatechin-3,5-digallate, theaflavin monogallate, tannic acid, methacycline, anthralin, mitoxantrone hydrochloride, hycanthone, esculidine lactate, aurintricarboxylic acid, carboplatin, cisplatin, primuletin, chrysin, diometin, suramin, hematin, gossypol, or any combination thereof. In some embodiments, the cell is contacted with the composition in vitro or in vivo. In some embodiments, the antibody is secreted from the cell. In some embodiments, a compound is contacted with a fucosyltransferase prior to the secretion of the antibody.

[0006] In some embodiments, a method for producing an antibody with an altered glycosylation pattern is provided. In some embodiments, the method comprises contacting a cell producing the antibody with a composition comprising purpurogallin, hexachlorophene, acriflavinum, baicalein, epicatechin monogallate, epicatechin-3-monogallate, epicatechin-3,5-digallate, theaflavin monogallate, tannic acid, methacycline, anthralin, mitoxantrone hydrochloride, hycanthone, esculidine lactate, aurintricarboxylic acid, carboplatin, cisplatin, primuletin, chrysin, diometin, suramin, hematin, gossypol, or any combination thereof. In some embodiments, the cell is contacted with the composition in vitro or in vivo. In some embodiments, the antibody is secreted from the cell. In some embodiments, a compound is contacted with a fucosyltransferase prior to the secretion of the antibody. "Altered glycosylation pattern" with respect to a protein, such as an antibody produced according to any of the methods described herein, refers to a protein that has a different glycosylation pattern compared to the protein produced from the same cell that has not been contacted with a composition or compound described herein.

[0007] In some embodiments, a composition is provided comprising purpurogallin, hexachlorophene, acriflavinum, baicalein, epicatechin monogallate, epicatechin-3-monogallate, epicatechin-3,5-digallate, theaflavin monogallate, tannic acid, methacycline, anthralin, mitoxantrone hydrochloride, hycanthone, esculidine lactate, aurintricarboxylic acid, carboplatin, cisplatin, primuletin, chrysin, diometin, suramin, hematin, gossypol, or any combination thereof, and a cell.

[0008] In some embodiments, a composition is provided comprising purpurogallin, hexachlorophene, acriflavinum, baicalein, epicatechin monogallate, epicatechin-3-monogallate, epicatechin-3,5-digallate, theaflavin monogallate, tannic acid, methacycline, anthralin, mitoxantrone hydrochloride, hycanthone, esculidine lactate, aurintricarboxylic acid, carboplatin, cisplatin, primuletin, chrysin, diometin, suramin, hematin, gossypol, or any combination thereof, and an antibody.

[0009] In some embodiments, a cell-free medium is provided that includes purpurogallin, hexachlorophene, acriflavinum, baicalein, epicatechin monogallate, epicatechin-3-monogallate, epicatechin-3,5-digallate, theaflavin monogallate, tannic acid, methacycline, anthralin, mitoxantrone hydrochloride, hycanthone, esculidine lactate, aurintricarboxylic acid, carboplatin, cisplatin, primuletin, chrysin, diometin, suramin, hematin, gossypol, or any combination thereof. In some embodiments, the medium further includes an antibody. [Brief description of the drawings]

[0010] [Figure 1] FIG. 1 shows a graph of inhibition of fucosylation using Purpurogallin. [Diagram 2] FIG. 1 shows a graph of inhibition of fucosylation using hexachlorophene. [Diagram 3] FIG. 1 shows a graph of inhibition of fucosylation using Acriflavinum. [Figure 4] FIG. 1 shows a graph of inhibition of fucosylation using epicatechin monogallate. [Diagram 5] FIG. 1 shows a graph of inhibition of fucosylation using epicatechin-3-monogallate. [Figure 6] FIG. 1 shows a graph of inhibition of fucosylation using epicatechin-3,5-digallate. [Figure 7] FIG. 1 shows a graph of inhibition of fucosylation using theaflavin monogallate. [Figure 8] FIG. 1 shows a graph of inhibition of fucosylation using tannic acid. [Figure 9] FIG. 1 shows a graph of inhibition of fucosylation using methacycline. [Figure 10] FIG. 1 shows a graph of inhibition of fucosylation using mitoxantrone hydrochloride. [Figure 11] FIG. 1 shows a graph of inhibition of fucosylation using hycanthone. [Figure 12] FIG. 1 shows a graph of inhibition of fucosylation using esculidine lactate. [Figure 13] FIG. 1 shows a graph of inhibition of fucosylation using aurintricarboxylic acid. [Figure 14] FIG. 1 shows a graph of inhibition of fucosylation using carboplatin. [Figure 15] FIG. 1 shows a graph of inhibition of fucosylation using cisplatin. [Figure 16] FIG. 1 shows a graph of inhibition of fucosylation using diomethin. [Figure 17] FIG. 1 shows a graph of inhibition of fucosylation using suramin. [Figure 18] FIG. 1 shows a graph of inhibition of fucosylation using hematin. [Figure 19] FIG. 1 shows a graph of inhibition of fucosylation using gossypol. [Figure 20] FIG. 1 shows reduction of antibody fucosylation by epicatechin monogallate (ECG). [Figure 21] FIG. 1 shows reduction of antibody fucosylation by epicatechin-3-monogallate (ECGC). DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0011] As used herein, unless otherwise indicated, the term "about" is intended to mean ±5% of the value it modifies. Thus, about 100 means 95 to 105.

[0012] As used in this specification and the appended claims, the singular forms "a," "an," and "the" include plural referents unless the context clearly dictates otherwise.

[0013] As used herein, the terms "comprising" (and any form of "comprising" such as "comprise," "comprises," and "comprised"), "having" (and any form of "having" such as "have" and "has"), "including" (and any form of "including" such as "includes" and "including"), or "containing" (and any form of "containing" such as "contains" and "contain") are inclusive or open-ended and do not exclude additional, unrecited elements or method steps. Any composition or method using the term "comprising" should also be understood to describe such composition or method as a composition "consisting of," "consisting of," or "consisting essentially of" the recited components or elements.

[0014] The term "antibody" encompasses immunoglobulins or antibody molecules, including polyclonal antibodies, monoclonal antibodies, including murine, human, humanized, and chimeric monoclonal antibodies, as well as antibody fragments. In some embodiments, the antibody is a recombinant antibody. "Recombinant antibody" refers to an antibody expressed from cells that have been genetically engineered to produce a particular antibody. Methods for producing recombinant antibodies are known, and any such method can be used.

[0015] In some embodiments, an antibody refers to a polypeptide that exhibits binding specificity to a specific antigen or target molecule. An intact antibody is a heterotetrameric glycoprotein composed of two light chains and two heavy chains. Usually, each light chain is linked to a heavy chain by one covalent disulfide bond, but the number of disulfide bonds varies among the heavy chains of different immunoglobulin isotypes. Each heavy and light chain also has regularly spaced intrachain disulfide bridges. Each heavy chain contains a variable domain (V H ) followed by several constant domains. Each light chain has a variable domain (V L ) at its other end and a constant domain, with the light chain constant domain aligned with the first constant domain of the heavy chain and the light chain variable domain aligned with the variable domain of the heavy chain. The light chains of antibodies of any vertebrate species can be assigned to one of two clearly distinct types, namely kappa and lambda, based on the amino acid sequence of their constant domains. In some embodiments, the antibodies are IgA, IgD, IgE, IgG, and IgM type antibodies. In some embodiments, the antibodies are IgA1, IgA2, IgG1, IgG2, IgG3, and IgG4 type antibodies.

[0016] In some embodiments, the antibody is an antibody fragment. The term "antibody fragment" refers to a portion of the intact antibody, typically the antigen-binding or variable region of the intact antibody. Examples of antibody fragments include, but are not limited to, Fab, Fab', F(ab')2, and Fv fragments, diabodies, single-chain antibody molecules, and multispecific antibodies that bind to multiple targets or different epitopes of the same target.

[0017] In some embodiments, the antibody is an antibody-drug conjugate. The term "antibody-drug conjugate" refers to an antibody or antibody fragment bound to the drug. Examples of antibody-drug conjugates include, but are not limited to, brentuximab vedotin, cantuzumab ravtansine, gemtuzumab ozogamicin, ibritumomab tiuxetan, polatuzumab vedotin, sacituzumab govitecan, trastuzumab duocarmazine, trastuzumab emtansine, and inotuzumab ozogamicin.

[0018] Polyclonal antibodies are heterogeneous populations of antibody molecules derived from the sera of animals immunized with an antigen. See, e.g., Kohler and Milstein, Nature 256:495 497 (1975); U.S. Patent No. 4,376,110; Ausubel et al. al., eds., Current Protocols in Molecular Biology, Greene Publishing Assoc. and Wiley Interscience, NY, (1987, 1992);Harlow and Lane ANTIBODIES: A Laboratory Manual Cold Spring Harbor Laboratory (1988); and Colligan et al., eds., Current Protocols in Immunology, Greene Publishing Assoc. and Wiley Interscience, NY, (1992, 1993), the contents of which are incorporated herein in their entireties. Such antibodies may be of any immunoglobulin class, including IgG, IgM, IgE, IgA, IgD, and any subclass thereof. Hybridomas producing the mAbs of the invention can be cultivated in vitro, in situ, or in vivo. Production of high titers of mAbs in vivo or in situ is currently the preferred method of production.

[0019] As used herein, the term "monoclonal antibody" (mAb) refers to an antibody (or antibody fragment) obtained from a population of substantially homogeneous antibodies. Monoclonal antibodies are usually , directed against a single antigenic determinant. The modifier "monoclonal" indicates that the antibody is substantially homogeneous in character and does not require production of the antibody by any particular method. For example, murine mAbs can be made by the hybridoma method of Kohler et al., Nature 256:495-497 (1975). Chimeric mAbs, comprising light and heavy chain variable regions derived from a donor antibody (such as, but not limited to, a mouse) associated with light and heavy chain constant regions derived from an acceptor antibody (such as, but not limited to, another mammal, including, but not limited to, a human), can be prepared by the method disclosed in U.S. Pat. No. 4,816,567 or any other method. Humanized mAbs, having CDRs derived from a non-human donor immunoglobulin (such as, but not limited to, mouse) and the remaining immunoglobulin-derived portions of the molecule derived from one or more human immunoglobulins, optionally with altered framework support residues to maintain binding affinity, can be obtained by the techniques disclosed in Queen et al., Proc. Natl. Acad. Sci. (USA), 86:10029-10032 (1989) and Hodgson et al., Bio / Technology, 9:421 (1991). Methods for humanizing antibodies are conventional and are not critical to the methods described herein.

[0020] As used herein, the term "in combination with" means that the described agents can be administered to or contacted with a cell, animal, or target together in a mixture, simultaneously as single agents, or sequentially as single agents in any order.

[0021] As used herein, the term "contact" refers to bringing two components into close proximity to each other. In some embodiments, the components may be mixed together or simply placed in the same container. A composition may also be considered to be in contact with another component (e.g., a subject, a cell, etc.) when it is administered to that other component. Administration may be by injection, addition by pipette, and other conventional administration methods.

[0022] As used herein, the term "inhibit" with respect to enzyme activity means to decrease the activity of the enzyme by any measurable amount.

[0023] As used herein, the term "reduce" in reference to a protein modification means to reduce the amount of said modification by a measurable amount.

[0024] In some embodiments, the compounds described herein are salts of the compounds. In some embodiments, the salts are pharma- ceutically acceptable salts of the compounds. As used herein, the phrase "pharma- ceutically acceptable salt(s)" includes, but is not limited to, salts of acidic or basic groups. Compounds that are basic can form a wide variety of salts with a variety of inorganic and organic acids. Acids that can be used to prepare pharma- ceutically acceptable acid addition salts of such basic compounds include sulfate, thiosulfate, citrate, maleate, acetate, oxalate, hydrochloride, hydrobromide, hydroiodide, nitrate, sulfate, bisulfate, bisulfite, phosphate, acid phosphate, isonicotinate, borate, acetate, lactate, salicylate, citrate, acid citrate, tartrate, oleate, tannate, pantothenate, bitartrate, ascorbate, succinate, maleate, gentisate. non-limiting examples of salts of the carboxylic acid salts include, but are not limited to, fumarate, gluconate, glucuronate, saccharate, formate, benzoate, glutamate, methanesulfonate, ethanesulfonate, benzenesulfonate, p-toluenesulfonate, bicarbonate, malonate, mesylate, esylate, napsydisylate, tosylate, besylate, orthophosphate, trifluoroacetate, and pamoate (i.e., 1,1'-methylene-bis-(2-hydroxy-3-naphthalene)) salts. Toxic acid addition salts are acids that form salts with pharmacologically acceptable anions. Compounds that contain an amino moiety can form pharmaceutically acceptable salts with various amino acids in addition to the acids mentioned above. Compounds that are acidic can form base salts with various pharmacologically acceptable cations. Examples of such salts include, but are not limited to, alkali metal salts or alkaline earth metal salts, particularly calcium, magnesium, ammonium, sodium, lithium, zinc, potassium, and iron salts. The present invention also encompasses quaternary ammonium salts of compounds described herein when the compounds have one or more tertiary amine moieties.

[0025] The term "antigen" refers to a target to which an antibody binds.

[0026] The term "purified" with respect to a protein or antibody refers to a protein or antibody that is substantially free of other materials associated with the molecule in its natural or cellular environment. For example, a purified protein is substantially free of cellular material or other proteins from the cell or tissue from which the protein is derived. The term refers to a sample in which the isolated protein is sufficiently pure to analyze it, or is at least 70%-80% (w / w) pure, at least 80%-90% (w / w) pure, 90-95% pure; and at least 95%, 96%, 97%, 98%, 99%, or 100% (w / w) pure. In some embodiments, the antibodies produced according to any of the embodiments described herein are further purified after production.

[0027] The embodiments provided herein provide compositions and methods that can be used, for example, to reduce fucosylation of proteins. "Reduced fucosylation" in the context of a protein generally refers to the reduction of the addition of fucose to glycans via α(1,2), α(1,3), α(1,4), and α(1,6) linkages. Fucosylation in the context of an antibody refers to the addition of fucose to N-acetylglucosamine ("GlcNAc") at the reducing end of the antibody's N-linked glycans ("fucosylation"). "Reduced fucosylation" in the context of an antibody refers to the reduction of fucose molecules attached to N-acetylglucosamine ("GlcNAc") at the reducing end of the antibody's N-linked glycans compared to an untreated antibody.

[0028] Antibody-dependent cellular cytotoxicity (ADCC) is a mechanism of cellular immunity whereby effector cells of the immune system actively lyse target cells bound to a specific antibody. ADCC is one of the mechanisms by which antibodies act as part of the immune response to limit and contain infections and diseases. It has been shown that monoclonal antibodies with reduced amounts of fucose in their glycosylation pattern exhibit significantly higher ADCC activity compared to normal fucosylated antibodies. Without wishing to be bound by any particular theory, the mechanism behind the improved ADCC of low-fucose or fucose-free antibodies appears to be mediated by the increased affinity of the modified Fc region of the antibody for FcγR on target cells. Thus, the compounds, compositions, and methods provided herein can provide antibodies with higher ADCC activity.

[0029] Certain compounds or their salts are referred to herein. Table 1 shows the chemical structures of these compounds. Also included are salts of these compounds, such as the pharma- ceutically acceptable salts of the compounds shown. In some embodiments, these salts are HCl salts of these compounds.

[0030] [Table 1] [Table 2] [Table 3] [Table 4] [Table 5] [Table 6]

[0031] In some embodiments, provided herein is a composition comprising one or more of the above compounds. In some embodiments, the composition comprises one or more of the above compounds and a cell. In some embodiments, the cell is a eukaryotic or prokaryotic cell. In some embodiments, the cell is capable of producing a protein. In some embodiments, the cell is a recombinant genetically modified cell. A "recombinant genetically modified cell" is a cell that has been modified with nucleic acid material that is exogenous to the cell, such as by transfection, transduction, or infection. In some embodiments, the cell is stably transduced or transfected with a nucleic acid molecule that encodes an antibody. In some embodiments, the cell produces an antibody.

[0032] A variety of cells or cell types can be used. Non-limiting examples of cells that can be present in the compositions or used in any of the methods provided herein include CHO cells, NS0 cells, Sp2 / 0 cells, HEK293 cells, PER.C6 cells, etc.

[0033] In some embodiments, the composition further comprises a cell culture medium. In some embodiments, the composition comprises a selection agent. In some embodiments, the composition comprises one or more antibiotics. A selection agent is an agent, such as an antibiotic, used to select cells that have been transfected or transduced with a particular nucleic acid molecule. A variety of selection agents can be used. Examples include Geneticin™ (G418; (2R,3S,4R,5R,6S)-5-amino-6-[(1R,2S,3S,4R,6S)-4,6-diamino-3-[(2R,3R,4R,5R)-3,5-dihydroxy-5-methyl-4-methylaminooxan-2-yl]oxy-2-hydroxycyclohexyl]oxy-2-(1-hydroxyethyl)oxane-3,4-diol), hygromycin b (O-6-amino-6-deoxy-L-glycero-D-galacto-heptopyranosylidene), and the like. -(1-2-3)-O-β-D-talopyranosyl(1-5)-2-deoxy-N3-methyl-D-streptamine), neomycin, blasticidin (4-amino-1-[4-({(3S)-3-amino-5-[[amino(imino)methyl](methyl)amino]pentanoyl}amino)-2,3,4-trideoxy-β-D-erythro-hex-2-enopyranuronosyl]pyrimidin-2(1H)-one) , mycophenolic acid ((E)-6-(4-hydroxy-6-methoxy-7-methyl-3-oxo-1H-2-benzofuran-5-yl)-4-methylhex-4-enoic acid), Zeocin™ (2-({2-[2-{[(6-amino-2-{3-amino-1-[(2,3-diamino-3-oxopropyl)amino]-3-oxopropyl}-5-methyl-4-pyrimidinyl)carbonyl]amino}-3 -[(5-{[1-({2-[4-({4-[(diaminomethylene)amino]butyl}carbamoyl)-4',5'-dihydro-2,4'-bi-1,3-thiazol-2'-yl]ethyl}amino)-3-hydroxy-1-oxo-2-butanyl]amino}-3-hydroxy-4-methyl-5-oxo-2-pentanyl)amino]-1-(1H-imidazol-5-yl)-3-oxopropoxy]-4,5

[0033] Examples of suitable hydroxylase inhibitors include, but are not limited to, N,N-dihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-3-yl}oxy)-3,5-dihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-4-ylcarbamate, and puromycin (3'-deoxy-N,N-dimethyl-3'-[(O-methyl-L-tyrosyl)amino]adenosine), or any combination thereof.

[0034] In some embodiments, a composition comprising one or more compounds provided herein and an antibody. In some embodiments, the antibody is an antibody secreted and / or produced by a cell. In some embodiments, the antibody is an antibody selected from the group consisting of adalimumab, alemtuzumab, alirocumab, atezolizumab, avelumab, belimumab, benralizumab, bevacizumab, bezlotoxumab, blinatumomab, brentuximab vedotin, canakinumab, cantuzumab mertansine, cantuzumab ravtansine, carotuximab, certolizumab pegol, cetuximab, claudiximab, claudiximab, daclizumab, daratumumab, denosumab, depatuxizumab mafodotin, dinutuximab, durvalumab, elotuzumab, enfortumab vedotin, enoblitzumab, gemtuzumab ozogamicin, golimumab, guselkumab, ibalizumab, ibritumomab tiuxetan, ifavotuzumab, inebilizumab, infliximab, inotuzumab Buozogamicin, ipilimumab, isatuximab, margetuximab, mepolizumab, necitumumab, nimotuzumab, nivolumab, obiltoxaximab, obinutuzumab, ocrelizumab, ofatumumab, olaratumab, omalizumab, palivizumab, pembrolizumab, pertuzumab, polatuzumab vedotin, racotumomab, ramucirumab, ranibizumab, raxibacumab, rituximab, ximab, rovalpituzumab tesirine, sacituzumab, sacituzumab govitecan, sarilumab, secukinumab, siltuximab, sirukumab, teprotumumab, tildrakizumab, tocilizumab, tositumomab, trastuzumab, trastuzumab duocarmazine, trastuzumab emtansine, ublituximab, ustekinumab, vedolizumab, or their biosimilars.

[0035] In some embodiments, the compositions provided herein are cell-free or substantially cell-free. A composition is substantially cell-free when the number of cells present in the composition is less than 50,000, less than 40,000, less than 30,000, less than 20,000, less than 10,000, or less than 5,000. In some embodiments, the composition comprises a cell-free medium comprising one or more compounds or salts thereof as described herein. The cell-free medium may also comprise any other components as described herein.

[0036] In some embodiments, the antibody is selected from the group consisting of ErbB1, also known as HER1 or EGFR, ErbB2, also known as HER2 or HER2 / neu, and ErbB3, also known as HER3. The antibody can bind to ErbB proteins, such as ErbB3, also known as ErbB1, and ErbB4, also known as HER4. In some embodiments, the antibody binds to transforming growth factor proteins, such as, but not limited to, TGFB1, TGFB2, TGFB3, and TGFB4. In some embodiments, the antibody binds to vascular endothelial growth factor proteins, such as, but not limited to, VEGFR1, Flt-1, VEGFR2, Flk-1 / KDR, VEGFR3, and Flt4. In some embodiments, the antibody binds to receptor activator of nuclear factor kappa beta (RANK), also known as TRANCE receptor or TNFRSFIIA. In some embodiments, the antibody binds to proteins related to the signaling lymphocyte activation molecule family (SLAMF) proteins, such as, but not limited to, SLAMF1, SLAMF2, SLAMF3, SLAMF4, SLAMF5, SLAMF6, SLAMF7, and SLAMF8. In some embodiments, the antibody binds to platelet-derived growth factor receptor proteins, such as, but not limited to, PDGFRα and PDGFRβ. In some embodiments, the antibody binds to proteins of the killer cell immunoglobulin-like receptor (KIR) family, such as, but not limited to, KIR3DL3, KIR3DP1, KIR3DL4, and KIR3DL2. In some embodiments, the antibody binds to major histocompatibility complex class I chain-related proteins (MIC), such as, but not limited to, MICA and MICB. In some embodiments, the antibody binds to proteins of the tumor necrosis factor (TNF) family, such as, but not limited to, LT alpha, LT beta, FASL, 4-IBBL, OX40L, and TNF-related apoptosis-inducing ligand (TRAIL). In some embodiments, the antibody binds to proteins of the death receptor (DR) family, such as, but not limited to, DR1, DR2, DR3, DR4, DR5, DR6, DR7, and DR8.In some embodiments, the antibody binds to a protein of the cytotoxic T lymphocyte antigen family, such as, but not limited to, CTLA4. In some embodiments, the antibody binds to a programmed death receptor, such as, but not limited to, PD1. In some embodiments, the antibody binds to a protein of the carcinoembryonic antigen family, such as, but not limited to, CD66a, CD66b, CD66c, CD66d, CD66e, and CD66f. In some embodiments, the antibody binds to a receptor of the T cell immunoglobulin and mucin domain family, such as, but not limited to, TIM1, TIM2, TIM3, and TIM4. In some embodiments, the antibody binds to lymphocyte activation genes (LAG), such as, but not limited to, FAG3. In some embodiments, the antibody binds to a group of differentiation antigens, including, but not limited to, CD2, CD3, CD19, CD20, CD22, CD25, CD27, CD30, CD33, CD38, CD39, CD52, CD70, CD73, CD94, CD134, CD137, CD252, and CD340. In some embodiments, the antibody binds to a tumor-associated carbohydrate antigen, including, but not limited to, mucin-associated GalNAc-O-ser / thr, also known as Tn, and Neu5Acα2-6GalNAcα-O-Ser / Thr, also known as sialyl Tn. In some embodiments, the antibody binds to a developmental stage-specific embryonic antigen, including, but not limited to, SSEA-1 (also known as Lewis X), SSEA-2, SSEA-3, and SSEA-4. In some embodiments, the antibody binds to a Thomsen-Freidenreich antigen, also known as Gal-Gal-NAc. In some embodiments, the antibody binds to a Lewis-related antigen, such as, but not limited to, Lewis Y, sialyl Lewis X, or sialyl Lewis A.In some embodiments, the antibody is directed to glycosphingolipid carbohydrates classified into the series, including, but not limited to, -Globo, -Isoglobo, -Ganglio, -Isoganglio, -Lacto, -Neolacto, -LactoGanglio, -Muco, -Neogala, -Mollu, -Arthro, -Schisto, and -Spirometo. In some embodiments, the antibody binds to gangliosides, such as, but not limited to, GD1, GD2, GD3, GM1, GM2, fucosyl Gm1, Neu5GcGm3, and polysialic acid.

[0037] In some embodiments, the composition may also include one or more buffers, stabilizers, emulsifiers, or any combination thereof. In some embodiments, the composition includes water.

[0038] In some embodiments, the composition comprises one or more cell-permeable compounds. In some embodiments, the cell-permeable compounds include hypotonic buffers; organic compounds such as methanol, ethanol, acetone, toluene, DMSO, and alkyltrimethylammonium bromides; detergents such as saponin, TritonX-100, Tween-20, Tween-80, digitonin, and sodium dodecyl sulfate (SDS); and pore-forming cytolysins, e.g., beta-hemolytic cytolysins such as streptolysin-O (SLO) and perfringolysin-O (PFO); or any combination thereof. In some embodiments, the composition comprises at least one cell permeabilizing agent selected from the group consisting of hypotonic buffer, methanol, ethanol, acetone, toluene, DMSO, alkyltrimethylammonium bromide, saponin, TritonX-100, Tween-20, Tween-80, digitonin, sodium dodecyl sulfate (SDS), beta-hemolytic cytolysin, streptolysin-O (SLO), perfringolysin-O (PFO), or any combination thereof.

[0039] Thus, in some embodiments, a method for inhibiting fucosyltransferase is provided.As used herein, "fucosyltransferase" refers to an enzyme that catalyzes the addition of fucose to a target, such as a protein.An example of fucosyltransferase includes, but is not limited to, α1,6 fucosyltransferase, which may also be referred to as FUT8.Thus, in some embodiments, the fucosyltransferase that is inhibited is FUT8. In some embodiments, the method includes contacting a fucosyltransferase with a composition comprising purpurogallin, hexachlorophene, acriflavinum, baicalein, epicatechin monogallate, epicatechin-3-monogallate, epicatechin-3,5-digallate, theaflavin monogallate, tannic acid, methacycline, anthralin, mitoxantrone hydrochloride, hycanthone, esculidine lactate, aurintricarboxylic acid, carboplatin, cisplatin, primuletin, chrysin, diometin, suramin, hematin, gossypol, or any combination thereof, or any salt thereof. The fucosyltransferase may be present in a cell or in a cell-free system.

[0040] A variety of cells can contain fucosyltransferase. Examples of cells include, but are not limited to, CHO cells, NS0 cells, Sp2 / 0 cells, HEK293 cells, or PER.C6 cells. These cells are non-limiting examples and other cell types and cell lines can be used.

[0041] In some embodiments, cells contacted with the compositions and compounds described herein produce an antibody. In some embodiments, the antibody is a recombinant antibody. In some embodiments, the antibody is directed to the following targets: IL8, ErbB, ErbB1, ErbB2, ErbB3, ErbB4, HER1, HER2, HER3, HER4, TGF, TGFB1, TGFB2, TGFB3, TGFB4, VEGF, VEGFR1, Flt-1, VEGFR2, Flk-1 / KDR, VEGFR3, Flt4, RANK, TNFRSFIIA, SLAMF, SLAMF1, SLAMF2, SLAMF3, SLAMF4, SLAMF5, SLAMF6, SLAMF7, SLAMF8, PDGFRα, P DGFRβ, KIR3DL3, KIR3DP1, KIR3DL4, KIR3DL2, MIC, MICA, MICB, TNF, LTα, LTβ, FASL, 4-IBBL, OX40L, TNF-related apoptosis-inducing ligand (TRAIL), DR1, DR2, DR3, DR4, DR5, DR6, DR7, DR8, CTLA4, PD1, CD66a, CD66b, CD66c, CD66d, CD66e, CD66f, TIM1, TIM2, TIM3, LAG, LAG3, CD2, CD3, CD19, CD20, CD22, CD25, CD27, CD30, CD33, CD38, CD39, CD52, CD70, CD73, CD94, CD134, CD137, CD252, Binds to one or more of CD340, GalNAc-O-ser / thr, Neu5Acα2-6GalNAcα-O-Ser / Thr, SSEA-, SSEA-2, SSEA-3, SSEA-4, Gal-Gal-NAc, Lewis Y, Sialyl Lewis X, Sialyl Lewis A, -Globo, -Isoglobo, -Ganglio, -Isoganglio, -Lacto, -Neolacto, LactoGanglio, -Muco, -Neogala, -Mollu, -Arthro, -Schisto, -Spirometo, GD1, GD2, GD3, GM1, GM2, Fucosyl Gm1, Neu5GcGm3, and polysialic acid.

[0042] In some embodiments, the antibody produced by the cell is selected from the group consisting of adalimumab, alemtuzumab, alirocumab, atezolizumab, avelumab, belimumab, benralizumab, bevacizumab, bezlotoxumab, blinatumomab, brentuximab vedotin, canakinumab, cantuzumab mertansine, cantuzumab vedotin, carotuximab, certolizumab pegol, cetuximab, , claudiximab, daclizumab, daratumumab, denosumab, depatuxizumab mafodotin, dinutuximab, durvalumab, elotuzumab, enfortumab vedotin, enoblitzumab, gemtuzumab ozogamicin, golimumab, guselkumab, ibalizumab, ibritumomab tiuxetan, ifavotuzumab, inebilizumab, infliximab , inotuzumab ozogamicin, ipilimumab, isatuximab, margetuximab, mepolizumab, necitumumab, nimotuzumab, nivolumab, obiltoxaximab, obinutuzumab, ocrelizumab, ofatumumab, olaratumab, omalizumab, palivizumab, pembrolizumab, pertuzumab, polatuzumab vedotin, racotumomab, ramucirumab, ranibizumab, raxibacumab , rituximab, rovalpituzumab tesirine, sacituzumab, sacituzumab govitecan, sarilumab, secukinumab, siltuximab, sirukumab, teprotumumab, tildrakizumab, tocilizumab, tositumomab, trastuzumab, trastuzumab duocarmazine, trastuzumab emtansine, ublituximab, ustekinumab, vedolizumab, or their biosimilars.

[0043] A biosimilar is an antibody that has the same or similar sequence as an antibody named herein, but may not be identical. A biosimilar antibody may also be an antibody that is so similar to an existing FDA-approved reference product, including those described herein, that there are no clinically meaningful differences. A biosimilar is clinically meaningful when there are no meaningful (significant) differences in safety, purity, and potency (safety and effectiveness) compared to the reference product.

[0044] Without wishing to be bound by any particular theory, the result is that the composition can be used to reduce the level of fucosylation of proteins in cells by inhibiting fucosyltransferase with the composition. The fucosylation can be reduced for proteins produced by the cells. In some embodiments, the protein to be reduced in fucosylation is an antibody. Non-limiting examples of antibodies are described herein. In some embodiments, the composition reduces the level of fucosylation by about 1% to about 99%. In some embodiments, the composition reduces the level of fucosylation by about 1% to about 99%. In some embodiments, the composition reduces the level of fucosylation by about 1% to about 95%. In some embodiments, the composition reduces the level of fucosylation by about 1% to about 90%. In some embodiments, the composition reduces the level of fucosylation by about 1% to about 85%. In some embodiments, the composition reduces the level of fucosylation by about 1% to about 80%. In some embodiments, the composition reduces the level of fucosylation by about 1% to about 75%. In some embodiments, the composition reduces the level of fucosylation by about 1% to about 70%. In some embodiments, the composition reduces the level of fucosylation by about 1% to about 65%. In some embodiments, the composition reduces the level of fucosylation by about 1% to about 60%. In some embodiments, the composition reduces the level of fucosylation by about 1% to about 55%. In some embodiments, the composition reduces the level of fucosylation by about 1% to about 50%. In some embodiments, the composition reduces the level of fucosylation by about 1% to about 45%. In some embodiments, the composition reduces the level of fucosylation by about 1% to about 40%. In some embodiments, the composition reduces the level of fucosylation by about 1% to about 35%. In some embodiments, the composition reduces the level of fucosylation by about 1% to about 30%. In some embodiments, the composition reduces the level of fucosylation by about 1% to about 25%. In some embodiments, the composition reduces the level of fucosylation by about 1% to about 20%. In some embodiments, the composition reduces the level of fucosylation by about 1% to about 15%. In some embodiments, the composition reduces the level of fucosylation by about 1% to about 10%. In some embodiments, the composition reduces the level of fucosylation by about 1% to about 5%. In some embodiments, the composition reduces the level of fucosylation by about 5% to about 99%. In some embodiments, the composition reduces the level of fucosylation by about 10% to about 99%. In some embodiments, the composition reduces the level of fucosylation by about 15% to about 99%. In some embodiments, the composition reduces the level of fucosylation by about 20% to about 99%.In some embodiments, the composition reduces the level of fucosylation by about 25% to about 99%. In some embodiments, the composition reduces the level of fucosylation by about 30% to about 99%. In some embodiments, the composition reduces the level of fucosylation by about 35% to about 99%. In some embodiments, the composition reduces the level of fucosylation by about 40% to about 99%. In some embodiments, the composition reduces the level of fucosylation by about 45% to about 99%. In some embodiments, the composition reduces the level of fucosylation by about 50% to about 99%. In some embodiments, the composition reduces the level of fucosylation by about 55% to about 99%. In some embodiments, the composition reduces the level of fucosylation by about 60% to about 99%. In some embodiments, the composition reduces the level of fucosylation by about 65% to about 99%. In some embodiments, the composition reduces the level of fucosylation by about 70% to about 99%. In some embodiments, the composition reduces the level of fucosylation by about 75% to about 99%. In some embodiments, the composition reduces the level of fucosylation by about 80% to about 99%. In some embodiments, the composition reduces the level of fucosylation by about 85% to about 99%. In some embodiments, the composition reduces the level of fucosylation by about 90% to about 99%. In some embodiments, the composition reduces the level of fucosylation by about 95% to about 99%. In some embodiments, the level of fucosylation of a protein or pool of proteins is reduced by at least 5, 10, 20, 30, 40, 50, 60, 70, 80, 90, or 95%. In some embodiments, the composition reduces but does not eliminate fucosylation. The reduction in protein fucosylation is relative to the level of fucosylation of a protein produced from a particular cell type, and the cells are contacted with the compositions and compounds described herein. For example, when an antibody is produced from a CHO cell and then the same or similar CHO cells producing said antibody are contacted with the compositions and compounds provided herein, the level of fucosylation on the antibody is reduced.The above reductions occur upon contact with the compositions and compounds described herein. This is a comparison with the antibody produced from the CHO cells described above, which has not been tested.

[0045] In some embodiments, a method for inhibiting or reducing protein fucosylation in a cell is provided. In some embodiments, the method comprises contacting the cell with any of the compositions or compounds provided herein. In some embodiments, the cell is contacted with the composition in vitro or in vivo. In some embodiments, the composition inhibits fucosylation by inhibiting fucosyltransferase.

[0046] In some embodiments, the cell is any of the cell types disclosed herein, although the cell types presented herein are non-limiting examples. In some embodiments, the cell produces an antibody. In some embodiments, the antibody is a recombinant antibody. In some embodiments, the antibody is directed against the following targets: IL8, ErbB, ErbB1, ErbB2, ErbB3, ErbB4, HER1, HER2, HER3, HER4, TGF, TGFB1, TGFB2, TGFB3, TGFB4, VEGF, VEGFR1, Flt-1, VEGFR2, Flk-1 / KDR, VEGFR3, Flt4, RANK, TNFRSFIIA, SLAMF, SLAMF1, SLAMF2, SLAMF3, SLAMF4, SLA MF5, SLAMF6, SLAMF7, SLAMF8, PDGFRα, PDGFRβ, KIR3DL3, KIR3DP1, KIR3DL4, KIR3DL2, MIC, MICA, MICB, TNF, LTα, LTβ, FASL, 4-IBBL, OX40L, TNF-related apoptosis-inducing ligand (TRAIL), DR1, DR2, DR3, DR4, DR5, DR6, DR7, DR8, CTLA4, PD1, CD66a, CD66b, CD66c, CD66d, CD66e , CD66f, TIM1, TIM2, TIM3, LAG, LAG3, CD2, CD3, CD19, CD20, CD22, CD25, CD27, CD30, CD33, CD38, CD39, CD52, CD70, CD73, CD94, C D134, CD137, CD252, CD340, GalNAc-O-ser / thr, Neu5Acα2-6GalNAcα-O-Ser / Thr, SSEA-, SSEA-2, SSEA-3, SSEA-4, Gal-Gal-NAc , Lewis Y, Sialyl Lewis X, Sialyl Lewis A, -Globo, -Isoglobo, -Ganglio, -Isoganglio, -Lacto, -Neolacto, LactoGanglio, -Muco, -Neogala, -Mollu, -Arthro, -Schisto, -Spirometo, GD1, GD2, GD3, GM1, GM2, Fucosyl Gm1, Neu5GcGm3, and polysialic acid.

[0047] In some embodiments, the antibody is selected from the group consisting of adalimumab, alemtuzumab, alirocumab, atezolizumab, avelumab, belimumab, benralizumab, bevacizumab, bezlotoxumab, blinatumomab, brentuximab vedotin, canakinumab, cantuzumab mertansine, cantuzumab vedotin, carotuximab, certolizumab pegol, cetuximab, claudiximab, daclizumab, daratumumab, denosumab, depatuxizumab mafodotin, dinutuximab, durvalumab, elotuzumab, enfortumab vedotin, enoblitzumab, gemtuzumab ozogamicin, golimumab, guselkumab, ibali ibritumomab tiuxetan, ifavotuzumab, inebilizumab, infliximab, inotuzumab ozogamicin, ipilimumab, isatuximab, margetuximab, mepolizumab, necitumumab, nimotuzumab, nivolumab, obiltoxaximab, obinutuzumab, ocrelizumab, ofatumumab, olaratumab , omalizumab, palivizumab, pembrolizumab, pertuzumab, polatuzumab vedotin, racotumomab, ramucirumab, ranibizumab, raxibacumab, rituximab, rovalpituzumab tesirin, sacituzumab, sacituzumab govitecan, sarilumab, secukinumab, siltuximab, sirukumab, teprotumumab, tildraki uzumab, tocilizumab, tositumomab, trastuzumab, trastuzumab duocarmazine, trastuzumab emtansine, ublituximab, ustekinumab, vedolizumab, or a biosimilar thereof. The level of fucosylation inhibition or reduction may be as described herein.

[0048] In some embodiments, methods of inhibiting fucosyltransferase are provided. In some embodiments, the methods include contacting the fucosyltransferase with any of the compositions or compounds provided herein. In some embodiments, the fucosyltransferase is present in a cell. In some embodiments, the cell is any of the cell types provided herein. In some embodiments, the cell produces an antibody. In some embodiments, the antibody is a recombinant antibody. In some embodiments, the antibody is any of the antibodies disclosed herein. In some embodiments, the antibody binds to any of the targets or antigens provided herein.

[0049] In some embodiments, methods of producing antibodies are provided. In some embodiments, methods of producing antibodies with reduced fucosylation are provided. In some embodiments, the methods include contacting an antibody-producing cell with any of the compositions and compounds provided herein. The cell may be contacted with the compositions and compounds in vitro or in vivo. The antibody-producing cell is not limited to any particular cell type, and may be, for example, any of the cell types provided herein. In some embodiments, the antibody is a recombinant antibody. In some embodiments, the antibody is any of the antibodies provided herein. In some embodiments, the antibody is directed against any of the antigens disclosed herein.

[0050] In some embodiments, any of the methods provided herein include purifying an antibody with reduced fucosylation. The antibody can be purified, for example, by column chromatography, precipitation, etc. Other purification methods include, but are not limited to, size exclusion chromatography, ammonium sulfate precipitation, ion exchange chromatography, immobilized metal chelate chromatography, thiophilic adsorption, Melon™ Gel chromatography, antibody purification with Protein A, G, and L antibody binding ligands, Protein A, G, and L, etc. These purification methods are non-limiting and other methods can be used.

[0051] In some embodiments, methods of producing antibodies with altered glycosylation patterns are provided. In some embodiments, the methods include contacting a cell producing the antibody with any of the compositions or compounds provided herein. In some embodiments, the composition is contacted with the cell in vitro or in vivo. In some embodiments, the cell is of any of the cell types provided herein. In some embodiments, the antibody is a recombinant antibody. In some embodiments, the antibody is any of the antibodies disclosed herein. In some embodiments, the antibody is directed against any of the antigens disclosed herein.

[0052] The antibodies produced herein can also be used in methods of treating various diseases in a subject, such as cancer, autoimmune diseases, etc., for which the methods of treatment have been developed. Thus, in some embodiments, a method of treating a subject is provided. In some embodiments, the method comprises contacting a cell producing the therapeutic antibody with a compound or composition described herein to produce a therapeutic antibody with reduced fucosylation. Non-limiting examples of therapeutic antibodies are described herein. The reduced fucosylation antibody can be isolated and prepared into a pharmaceutical composition or a therapeutic agent suitable for administration to a subject in need thereof. can be formulated into a formulation. In some embodiments, the subject is being treated for cancer or an autoimmune disease, such as rheumatoid arthritis, Crohn's disease, ulcerative colitis, ankylosing spondylitis, Behcet's disease, fistulizing disease, hidradenitis suppurativa, juvenile idiopathic arthritis, psoriasis, psoriatic arthritis, and relapsing polychondritis.

[0053] The present embodiment will now be described with reference to the following examples. These examples are presented for illustrative purposes only, and the present embodiment should not be construed as being limited to these examples in any way, but should be construed as embracing all variations that become evident as a result of the teachings provided herein. Those skilled in the art will readily recognize a variety of non-critical parameters that can be changed or modified to produce essentially similar results. EXAMPLES

[0054] The following examples are illustrative of the compositions and methods described herein, but are not intended to be limiting thereof. Other suitable modifications and adaptations known to those skilled in the art are within the scope of the following embodiments.

[0055] Example 1: Inhibition of fucosylation by selected compounds The compounds exemplified in this example were found to inhibit FUT8 activity, thereby demonstrating that the compounds are capable of inhibiting fucosyltransferases. The compounds exemplified below were found to be effective in inhibiting FUT8 activity and would be expected to reduce fucosylation of antibodies produced from cells containing fucosyltransferases such as FUT8. Briefly, the compounds were mixed with fucosyltransferase (FUT-8) and an appropriate substrate and incubated for 1 hour. The reaction was stopped and a detection reagent mixture was added to detect the amount of fucosylation of the substrate protein. The inhibition values ​​for each compound are shown in Table 2 and are also shown in Figures 1-19.

[0056] [Table 7]

[0057] Example 2: Fucosyltransferase inhibitors reduce antibody fucosylation The compounds exemplified in this example were found to reduce the level of fucosylation of antibodies produced by cells containing fucosyltransferases such as FUT8. Briefly, epicatechin monogallate (ECG), epicatechin-3-monogallate (ECGC), and aurintricarboxylic acid (ATA) were added to cell culture medium and added to CHO cells (ATCC CRL-12445) obtained from ATCC (also described in U.S. Patent No. 6,025,158, which is incorporated herein in its entirety) producing anti-IL8 IgG monoclonal antibody (mAb). After the indicated period, the cells were harvested and the anti-IL8 mAb was purified by standard methods. The purified anti-IL8 mAb was then reduced by adding dithiothreitol and subjected to FC-MS analysis to measure the level of fucosylation. The reduction in the level of fucosylation of anti-IL8 mAb is shown in Figures 20 and 21. These results indicate that inhibition of fucosyltransferases reduces antibody fucosylation. Furthermore, ECG was shown to cause a dose-dependent decrease in core fucosylated species with a concomitant increase in nonfucosylated species. A 25% reduction in fucosylation was observed at the highest concentration of ECG (100 μM), while a 10-fold lower concentration achieved a 13% reduction. 10 μM ECGC caused a 6% reduction in fucosylation. This was achieved. For this antibody, ATA did not alter the fucosylation level at either concentration tested (data not shown). These results indicate that the use of different types of fucosyltransferase inhibitors may reduce antibody fucosylation.

[0058] The present description is not limited to the specific processes, compositions, or methodologies described, as these processes, compositions, or methodologies may vary. The terms used in the description are only intended to describe a particular version or embodiment, and are not intended to limit the scope of the embodiments described herein. Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by those skilled in the art. In some cases, terms with commonly understood meanings are defined herein for clarity and / or ease of reference, and the inclusion of such definitions herein should not necessarily be construed as representing a substantial difference to those commonly understood in the art. However, in case of conflict, the patent specification, including definitions, shall prevail.

[0059] From the foregoing, it will be understood that various embodiments of the present disclosure have been described herein for purposes of illustration, and that various modifications can be made without departing from the scope and spirit of the present disclosure. Accordingly, the various embodiments disclosed herein are not intended to be limiting.

Claims

1. 1. A method of inhibiting a fucosyltransferase comprising contacting the fucosyltransferase with a compound selected from the group consisting of epicatechin monogallate (ECG), epicatechin-3-monogallate (ECGC), purpurogallin, hexachlorophene, acriflavinum, baicalein, epicatechin-3,5-digallate, theaflavin monogallate, tannic acid, methacycline, anthralin, mitoxantrone hydrochloride, hycanthone, esculidine lactate, aurintricarboxylic acid, carboplatin, cisplatin, primuletin, chrysin, diometin, suramin, hematin, gossypol, or a composition comprising same, or any combination thereof.

2. The method of claim 1 , wherein the fucosyltransferase is present in a cell.

3. The method of claim 1 , wherein the cells produce an antibody.

4. The method of claim 3 , wherein the antibody is a recombinant antibody.

5. The antibody is directed against the following targets: IL8, ErbB, ErbB1, ErbB2, ErbB3, ErbB4, HER1, HER2, HER3, HER4, TGF, TGFB1, TGFB2, TGFB3, TGFB4, VEGF, VEGFR1, Flt-1, VEGFR2, Flk-1 / KDR, VEGFR3, Flt4, RANK, TNFRSFIIA, SLAMF, SLAMF1, SLAMF2, SLAMF3, SLAMF4, SLAMF5, SLAMF6, SLAMF7, SLAMF8, SLAMF9, SLAMF10, SLAMF11, SLAMF12, SLAMF13, SLAMF14, SLAMF15, SLAMF16, SLAMF17, SLAMF18, SLAMF19, SLAMF20, SLAMF21, SLAMF22, SLAMF23, SLAMF24, SLAMF25, SLAMF26, SLAMF27, SLAMF28, SLAMF29, SLAMF30, SLAMF31, SLAMF32, SLAMF33, SLAMF34, SLAMF35, SLAMF36, SLAMF37, SLAMF38, SLAMF39 ...9, SLAMF39, SLAMF31, SLAMF32, SLAMF33, SLAMF34, SLAMF35, SLAMF36, SLAMF37, SLAMF39, SLAMF39, SLAMF39, SLAMF39, SLAMF39, SLAMF39, SLAMF AMF7, SLAMF8, PDGFRα, PDGFRβ, KIR3DL3, KIR3DP1, KIR3DL4, KIR3DL2, MIC, MICA, MICB, TNF, LTα, LTβ, FASL, 4-IBBL, OX40L, TNF-related apoptosis-inducing ligand (TRAIL), DR1, DR2, DR3, DR4, DR5, DR6, DR7, DR8, CTLA4, PD1, CD66a, CD66b, CD66c, CD66d, CD66e, CD66f, TIM1, T IM2, TIM3, LAG, LAG3, CD2, CD3, CD19, CD20, CD22, CD25, CD27, CD30, CD33, CD38, CD39, CD52, CD70, CD73, CD94, CD134, CD137, CD252, CD340, GalNAc-O-ser / thr, Neu5Acα2-6GalNAcα-O-ser / thr, SSEA-, SSEA-2, SSEA-3, SSEA-4, Gal-Gal-NAc, Lewis Y, Sialyl Lewis 4. The method of claim 3, wherein the sialic acid binds to one or more of: Sialyl Lewis A, -Globo, -Isoglobo, -Ganglio, -Isoganglio, -Lacto, -Neolacto, LactoGanglio, -Muco, -Neogala, -Mollu, -Arthro, -Schisto, -Spirometo, GD1, GD2, GD3, GM1, GM2, Fucosyl Gm1, Neu5GcGm3, and polysialic acid.

6. The antibody produced by the cell is selected from the group consisting of adalimumab, alemtuzumab, alirocumab, atezolizumab, avelumab, belimumab, benralizumab, bevacizumab, bezlotoxumab, blinatumomab, brentuximab vedotin, canakinumab, cantuzumab mertansine, cantuzumab vedotin, carotuximab, certolizumab pegol, cetuximab, claudiximab, daclizumab, daratumumab , denosumab, depatuxizumab mafodotin, dinutuximab, durvalumab, elotuzumab, enfortumab vedotin, enoblitzumab, gemtuzumab ozogamicin, golimumab, guselkumab, ibalizumab, ibritumomab tiuxetan, ifavotuzumab, inebilizumab, infliximab, inotuzumab ozogamicin, ipilimumab, isatuximab, margetuximab, mepolizumab, necitumumab, nimotuzumab, Nivolumab, obiltoxaximab, obinutuzumab, ocrelizumab, ofatumumab, olaratumab, omalizumab, palivizumab, pembrolizumab, pertuzumab, polatuzumab vedotin, racotumomab, ramucirumab, ranibizumab, raxibacumab, rituximab, rovalpituzumab tesirin, sacituzumab, sacituzumab gobi 3. The method of claim 2, wherein the therapeutic agent is selected from the group consisting of tecan, sarilumab, secukinumab, siltuximab, sirukumab, teprotumumab, tildrakizumab, tocilizumab, tositumomab, trastuzumab, trastuzumab duocarmazine, trastuzumab emtansine, ublituximab, ustekinumab, vedolizumab, or biosimilars thereof.

7. 1. A method for inhibiting protein fucosylation in a cell, comprising contacting the cell with a compound selected from the group consisting of epicatechin monogallate, epicatechin-3-monogallate, purpurogallin, hexachlorophene, acriflavinum, baicalein, epicatechin-3,5-digallate, theaflavin monogallate, tannic acid, methacycline, anthralin, mitoxantrone hydrochloride, hycanthone, esculidine lactate, aurintricarboxylic acid, carboplatin, cisplatin, primuletin, chrysin, diometin, suramin, hematin, gossypol, or a composition comprising same, or any combination thereof, to inhibit fucosylation of the protein.

8. The method of claim 7, wherein the compound or composition inhibits fucosylation by inhibiting a fucosyltransferase.

9. 9. The method of claim 7 or 8, wherein the cell is contacted with the compound or composition in vitro or in vivo.

10. The method according to any one of claims 7 to 9, wherein the protein is an antibody.

11. 11. The method of any of claims 7 to 10, wherein the composition comprises at least one cell permeabilizing agent selected from the group consisting of hypotonic buffer, methanol, ethanol, acetone, toluene, DMSO, alkyltrimethylammonium bromide, saponin, Triton X-100, Tween-20, Tween-80, digitonin, sodium dodecyl sulfate (SDS), beta-hemolytic cytolysin, streptolysin-O (SLO), perfringolysin-O (PFO), or any combination thereof.

12. The method of any one of claims 7 to 11, wherein the cell is a CHO cell, an NSO cell, an Sp2 / 0 cell, an HEK293 cell, or a PER.C6 cell.

13. 1. A method for producing an antibody having reduced fucosylation, comprising contacting a cell producing said antibody with a compound selected from the group consisting of epicatechin monogallate, epicatechin-3-monogallate, purpurogallin, hexachlorophene, acriflavinum, baicalein, epicatechin-3,5-digallate, theaflavin monogallate, tannic acid, methacycline, anthralin, mitoxantrone hydrochloride, hycanthone, esculidine lactate, aurintricarboxylic acid, carboplatin, cisplatin, primuletin, chrysin, diometin, suramin, hematin, gossypol, or a composition comprising same, or any combination thereof.

14. The method of claim 13, wherein the cell is contacted with the compound or composition in vitro or in vivo.

15. The method of claim 13 , wherein the compound or composition inhibits a fucosyltransferase.

16. The method of any one of claims 13 to 15, wherein the antibody is secreted from the cell.

17. The method of claim 16, wherein the compound or composition is contacted with the fucosyltransferase prior to the secretion of the antibody.

18. The method of any of claims 13 to 17, wherein the level of fucosylation of the produced antibody is reduced compared to antibodies produced from cells not contacted with the compound or composition.

19. 18. The method of any of claims 13 to 17, wherein the compound or composition comprises at least one cell permeabilizing agent selected from the group consisting of hypotonic buffer, methanol, ethanol, acetone, toluene, DMSO, alkyltrimethylammonium bromide, saponin, Triton X-100, Tween-20, Tween-80, digitonin, sodium dodecyl sulfate (SDS), beta-hemolytic cytolysin, streptolysin-O (SLO), perfringolysin-O (PFO), or any combination thereof.

20. The method of any one of claims 13 to 19, wherein the cell is a CHO cell, an NSO cell, an Sp2 / 0 cell, an HEK293 cell, or a PER.C6 cell.

21. 1. A method for producing an antibody having an altered glycosylation pattern, comprising contacting a cell producing said antibody with a compound selected from the group consisting of epicatechin monogallate, epicatechin-3-monogallate, purpurogallin, hexachlorophene, acriflavinum, baicalein, epicatechin-3,5-digallate, theaflavin monogallate, tannic acid, methacycline, anthralin, mitoxantrone hydrochloride, hycanthone, esculidine lactate, aurintricarboxylic acid, carboplatin, cisplatin, primuletin, chrysin, diometin, suramin, hematin, gossypol, or a composition comprising same, or any combination thereof.

22. 22. The method of claim 21, wherein the cell is contacted with the composition in vitro or in vivo.

23. The method of claim 21 , wherein the compound or composition inhibits a fucosyltransferase.

24. The method of any one of claims 21 to 23, wherein the antibody is secreted from the cell.

25. 25. The method of claim 24, wherein the compound or composition is contacted with the cell prior to the secretion of the antibody.

26. The method of any one of claims 21 to 25, wherein the glycosylation pattern of the antibody produced is altered compared to an antibody produced from a cell not contacted with the composition.

27. 27. The method of any of claims 21 to 26, wherein the compound or composition comprises at least one cell permeabilizing agent selected from the group consisting of hypotonic buffer, methanol, ethanol, acetone, toluene, DMSO, alkyltrimethylammonium bromide, saponin, Triton X-100, Tween-20, Tween-80, digitonin, sodium dodecyl sulfate (SDS), beta-hemolytic cytolysin, streptolysin-O (SLO), perfringolysin-O (PFO), or any combination thereof.

28. The method of any one of claims 21 to 27, wherein the cell is a CHO cell, an NSO cell, an Sp2 / 0 cell, an HEK293 cell, or a PER.C6 cell.

29. A composition comprising epicatechin monogallate, epicatechin-3-monogallate, purpurogallin, hexachlorophene, acriflavinum, baicalein, epicatechin-3,5-digallate, theaflavin monogallate, tannic acid, methacycline, anthralin, mitoxantrone hydrochloride, hycanthone, esculidine lactate, aurintricarboxylic acid, carboplatin, cisplatin, primuletin, chrysin, diometin, suramin, hematin, gossypol, or any combination thereof, and a cell.

30. The composition of claim 29 , wherein the cells produce a recombinant antibody.

31. 31. The composition of claim 29 or 30, wherein the cell is a CHO cell, an NSO cell, an Sp2 / 0 cell, a HEK293 cell, or a PER.C6 cell.

32. The composition of any of claims 29 to 31, further comprising a cell culture medium.

33. The composition of any one of claims 29 to 32, further comprising one or more antibiotics.

34. The composition of any of claims 29 to 33, further comprising one or more screening agents.

35. The composition of any of claims 29 to 34, further comprising a recombinant antibody produced and secreted from said cells.

36. A composition comprising epicatechin-3-monogallate, epicatechin-3,5-digallate, purpurogallin, hexachlorophene, acriflavinum, baicalein, epicatechin monogallate, theaflavin monogallate, tannic acid, methacycline, anthralin, mitoxantrone hydrochloride, hycanthone, esculidine lactate, aurintricarboxylic acid, carboplatin, cisplatin, primuletin, chrysin, diometin, suramin, hematin, gossypol, or any combination thereof, and an antibody.

37. The antibody is selected from the group consisting of adalimumab, alemtuzumab, alirocumab, atezolizumab, avelumab, belimumab, benralizumab, bevacizumab, bezlotoxumab, blinatumomab, brentuximab vedotin, canakinumab, cantuzumab mertansine, cantuzumab vedotin, carotuximab, certolizumab pegol, cetuximab, and claudiximab. , daclizumab, daratumumab, denosumab, depatuxizumab mafodotin, dinutuximab, durvalumab, elotuzumab, enfortumab vedotin, enoblitzumab, gemtuzumab ozogamicin, golimumab, guselkumab, ibalizumab, ibritumomab tiuxetan, ifavotuzumab, inebilizumab, infliximab, inotuzumab ozogamicin, ipilimumab , isatuximab, margetuximab, mepolizumab, necitumumab, nimotuzumab, nivolumab, obiltoxaximab, obinutuzumab, ocrelizumab, ofatumumab, olaratumab, omalizumab, palivizumab, pembrolizumab, pertuzumab, polatuzumab vedotin, racotumomab, ramucirumab, ranibizumab, raxibacumab, rituximab, rovalpituzumab 37. The composition of claim 36, which is cilizumab, sacituzumab, sacituzumab govitecan, sarilumab, secukinumab, siltuximab, sirukumab, teprotumumab, tildrakizumab, tocilizumab, tositumomab, trastuzumab, trastuzumab duocarmazine, trastuzumab emtansine, ublituximab, ustekinumab, vedolizumab, or a biosimilar thereof.

38. 1. A cell-free medium comprising epicatechin-3-monogallate, epicatechin-3,5-digallate, purpurogallin, hexachlorophene, acriflavinum, baicalein, epicatechin monogallate, theaflavin monogallate, tannic acid, methacycline, anthralin, mitoxantrone hydrochloride, hycanthone, esculidine lactate, aurintricarboxylic acid, carboplatin, cisplatin, primuletin, chrysin, diometin, suramin, hematin, gossypol, or any combination thereof.

39. 39. The medium of claim 38, further comprising an antibody.

40. The antibody is selected from the group consisting of adalimumab, alemtuzumab, alirocumab, atezolizumab, avelumab, belimumab, benralizumab, bevacizumab, bezlotoxumab, blinatumomab, brentuximab vedotin, canakinumab, cantuzumab mertansine, cantuzumab vedotin, carotuximab, certolizumab pegol, cetuximab, and claudiximab. , daclizumab, daratumumab, denosumab, depatuxizumab mafodotin, dinutuximab, durvalumab, elotuzumab, enfortumab vedotin, enoblitzumab, gemtuzumab ozogamicin, golimumab, guselkumab, ibalizumab, ibritumomab tiuxetan, ifavotuzumab, inebilizumab, infliximab, inotuzumab ozogamicin, ipilimumab , isatuximab, margetuximab, mepolizumab, necitumumab, nimotuzumab, nivolumab, obiltoxaximab, obinutuzumab, ocrelizumab, ofatumumab, olaratumab, omalizumab, palivizumab, pembrolizumab, pertuzumab, polatuzumab vedotin, racotumomab, ramucirumab, ranibizumab, raxibacumab, rituximab, rovalpituzumab tesi 39. The cell-free medium of claim 38, wherein the antibody is selected from the group consisting of sucrine, sacituzumab, sacituzumab govitecan, sarilumab, secukinumab, siltuximab, sirukumab, teprotumumab, tildrakizumab, tocilizumab, tositumomab, trastuzumab, trastuzumab duocarmazine, trastuzumab emtansine, ublituximab, ustekinumab, vedolizumab, or a biosimilar thereof.

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