Neutral glycosylated amides and dianionic glucuron-oxidized acids as biomolecule stabilizers

Abstract

To provide compounds useful for stabilizing biological molecules, in the presence of pH and thermal stress.SOLUTION: The present invention provides a compound having the following structure (where R1 is H, an optionally substituted alkyl, or CONH2, m is 0, 1 or 2, and the optionally substituted alkyl is C1-C4 alkyl or C1-C4 hydroxyalkyl).SELECTED DRAWING: None

Description

[Technical Field]

[0001] This application claims priority to U.S. Provisional Application No. 62 / 585,341, filed on 13 November 2017, the contents of which are incorporated herein by reference.

[0002] Throughout this application, certain publications are referenced in parentheses. Complete citations of these publications are found immediately before the claims. To more fully illustrate the latest art to which this invention relates, the entire disclosures of these publications are incorporated into this application by reference. [Background technology]

[0003] Proteins and other biomolecules degrade when exposed to pH and heat stress through denaturation, aggregation, and other harmful chemical and physical modifications (Chang et al. 2010; Ueda et al. 2001). Such pH and heat stress can occur during the processing, preparation, or storage of biomolecules. In the case of biomolecules with therapeutic applications, degradation results in reduced yield and loss of activity.

[0004] Physical stresses such as heating and freeze-drying can cause loss of the original structure and, consequently, loss of biomolecular activity. Chemical stresses such as low or high pH can also degrade the structure of biomolecules, leading to aggregation, misfolding, or precipitation (Chang et al. 2010). For therapeutic biomolecules, some formulations require low or high pH for specific drug release needs, but such pH conditions can also adversely affect the stability of the therapeutically active molecule and, consequently, its shelf life. When a biomolecule is therapeutically active, its stabilization is particularly important so that its activity does not decrease over time.

[0005] Certain carbohydrates have been shown to stabilize biomolecules exposed to unfavorable temperatures and other stresses (Ueda et al. 2001; Kaushik et al. 2003; Singer et al. 1998; Lin et al. 1996; Jain et al. 2009; Andya et al. 2003 and Khan et al. 2010). One example is trehalose, a glucose disaccharide linked by an α,α-1,1-glycosidic bond, which accumulates in many organisms, including bacteria, yeasts, fungi, plants, and insects, that can withstand prolonged drought and starvation. In some bacteria, trehalose is synthesized in response to osmotic stress (Reed et al. 1986).

[0006] Trehalose is used as an excipient in several biopharmaceuticals, including Avastin, Herceptin, Lucentis, and Rituxan (Ohtak et al. 2011). The addition of trehalose to some proteins and recombinant proteins increases their stability, as evidenced by the increase in their melting temperature (Tm) (Kaushik et al. 2003; Lin et al. 1996; Singer et al. 1998; Ueda et al. 2001). One theory for the thermal stabilization effect of trehalose is that when trehalose is added to a protein solution, it increases the surface tension of the medium, leading to increased protein hydration and, consequently, increased protein stability against degradation. Such compounds can be understood as shifting the equilibrium to the natively folded conformation by increasing the free energy of the unfolded state (Khan et al. 2010; Rajan et al. 2011).

[0007] Trehalose and sucrose have been studied as stabilizers in lyophilized formulations of recombinant monoclonal antibodies (Andya et al. 2003). One theory for the stabilizing effect of trehalose in lyophilized formulations is that it interacts with the protein surface and acts as a water substitute, maintaining the structure of biomolecules while resisting chemical and physical modifications (Kaushik et al. 2003). Sucrose has also been shown to stabilize proteins from thermal decomposition (Lee et al. 1981). Polyamide sugars (PAS) have been shown to stabilize lysozyme against dehydration and freeze stress (Stidham et al. 2014).

[0008] The stabilization of biomolecules under pH stress remains a challenge. A specific challenge is the stabilization of biomolecules under combined thermal and pH stress. Novel stabilizers are needed to prevent the degradation of biomolecules at low or high pH levels, including in the presence of thermal stress. [Overview of the Initiative]

[0009] The present invention provides a compound or salt thereof having the following structure, [ka] During the ceremony, X is a hexosyl group selected from the group consisting of glucosyl, mannosyl, galactosyl, allosyl, altorosyl, grosyl, idosyl and talosyl, or a uronic acid group selected from the group consisting of glucuronic acid, mannuronic acid, galacturonic acid, aluronic acid, altoruonic acid, guluronic acid, iduronic acid and tarronic acid, or a uronic acid amide group selected from the group consisting of glucuronamide, mannuronamide, galacturonamide, aluronamide, altoruronamide, guluronamide, iduronamide and tarronamide. R 1 and R 2Each of these is independently H, halogen, OH, O-alkyl, CONH2, CO2H, CO2-alkyl, or optionally substituted alkyl. Each of Y and Z is independently H, OH, O-alkyl, or optionally substituted alkyl. m is 0, 1, or 2. A = NR3R4 or OR5. Here R 3 and R 4 Each of these is independently H, OH, O-alkyl or optionally substituted alkyl. R 5 These are independently H or optionally substituted alkyl groups. Here If X is glucosyl, R 1 This is an alkyl group which may be substituted in some cases. R 1 and R 2 Each of these is H, and if m is 0, then X is something other than glucuronic acid.

[0010] The present invention also provides a composition comprising a biomolecule and at least one compound of the present invention, and a method for stabilizing a biomolecule, comprising treating the biomolecule with an effective amount of the compound of the present invention, thereby enabling the stabilization of the biomolecule. [Brief explanation of the drawing]

[0011] [Figure 1] The graph shows the increase in the melting temperature (Tm) of lysozyme in the presence of 0.25 mM of various compounds (from left to right: MGlyA, MLA, GBA, GaGlyA, GaLA, b-GGlyA, b-GLA, b-GaLA, b-GaGlyA, b-GaBA) in 25 mM sodium acetate buffer pH 3.6 (black bar) and pH 12 phosphate buffer (gray bar). The melting temperature (Tm) of lysozyme in the absence of the compounds was 71°C in 25 mM sodium acetate buffer pH 3.6 and 55°C in phosphate buffer pH 12. [Figure 2]The HP-SEC chromatograms of 0.4 mg / ml Humira® samples under low pH (stress) conditions are shown. The line with the highest peak corresponds to the control sample, fresh Humira® (T=0h), in pH 5.5 citrate buffer. The line with the second highest peak corresponds to Humira® at pH 3.2 over 12 hours in the presence of GaGlyA, and the line with the third highest peak corresponds to Humira® at pH 3.2 over 12 hours without GaGlyA. [Figure 3] This shows the increase in the melting temperature (Tm) of Humira® in the presence of 0.25 mM of various compounds (from left to right: MGlyA, MLA, GBA, GaGlyA, GaLA, b-GGlyA, b-GLA, b-GaLA, b-GaGlyA, b-GaBA) in pH 12 phosphate buffer. The melting temperature (Tm) of Humira® in the absence of the compounds was 41°C in phosphate buffer. [Figure 4] This shows the increase in the melting temperature (Tm) of ubiquitin in the presence of 0.25 M of various compounds (from left to right: b-GGly, b-GL, b-GB, b-GaGly, b-GaL, a-MGlyA, GBA, b-GBA, b-GGlyA, b-GaGlyA, GaLA, b-GaLA, b-GaBA) in pH 12 phosphate buffer. The melting temperature (Tm) of ubiquitin in the absence of the compounds was 72°C in phosphate buffer. [Figure 5] This shows the increase in the melting temperature (Tm) of factor IX in the presence of various 0.25 M compounds (from left to right: MGlyA, GGlyA, GBA, GaLA, GGlyA) in water. The melting temperature (Tm) of factor IX in the absence of the compounds was 50°C in water. The pH of the assay in the presence of the compounds was between pH 7 and 8. [Modes for carrying out the invention]

[0012] The present invention provides a compound or salt thereof having the following structure: [ka] During the ceremony, X is a hexosyl group selected from the group consisting of glucosyl, mannosyl, galactosyl, allosyl, altrosyl, gulosyl, idosyl and talosyl, or an uronic acid group selected from the group consisting of glucuronic acid, mannuronic acid, galacturonic acid, alluronic acid, alturonic acid, guluronic acid, iduronic acid and taluronic acid, or an uronic acid amide group selected from the group consisting of glucuronamide, mannuronamide, galacturonamide, alluronamide, alturonamide, guluronamide, iduronamide and taluronamide, R 1 and each of R 2 is independently H, halogen, OH, O-alkyl, CONH2, CO2H, CO2-alkyl, or optionally substituted alkyl, Each of Y and Z is independently H, OH, O-alkyl or optionally substituted alkyl, m is 0, 1, or 2, A = NR3R4 or OR5, where R 3 and each of R 4 is independently H, OH, O-alkyl or optionally substituted alkyl, R 5 is independently H or optionally substituted alkyl, where when X is glucosyl, R 1 is optionally substituted alkyl, R 1 and each of R 2 is H and m is 0, X is other than glucuronic acid.

[0013] The present invention provides a compound having the following structure or a salt thereof,

Chemical formula

[0014] In some embodiments, the compound has the following structure. [ka]

[0015] In some embodiments, optionally substituted alkyl groups are either unsubstituted or substituted.

[0016] In some embodiments, the optionally substituted alkyl is a hydroxyalkyl.

[0017] In some embodiments, the optionally substituted alkyl group is an alkyl-OH group.

[0018] In some embodiments, the optionally substituted alkyl is a C1-C4 hydroxyalkyl group.

[0019] In some embodiments, the optionally substituted alkyl group is a C1-C4 alkyl-OH group.

[0020] In some embodiments, X is glucosyl or mannosyl, m=0, and R 1 or R 2 When one of the atoms is an alkyl-OH group and the other is an H group, and A is an NH2 group, the compound is a β-anomeric compound.

[0021] In some embodiments, X is a hexosyl group selected from the group consisting of glucosyl, mannosyl, galactosyl, allosyl, altorosyl, grosyl, idosyl and talosyl, or a uronic acid group selected from the group consisting of glucuronic acid, mannuronic acid, galacturonic acid, aluronic acid, altoruonic acid, guluronic acid, iduronic acid and tarronic acid, or a uronic acid amide group selected from the group consisting of glucuronamide, mannuronamide, galacturonamide, aluronamide, altoruronamide, guluronamide, iduronamide and tarronamide. R 1 and R 2 Each of these is independently H, CONH2, CO2H, CO2-alkyl, or unsubstituted alkyl. Each of Y and Z is independently H. m is 0, 1, or 2. A = NR3R4 or OR5. Here R 3 and R 4Each of these is independently H, OH, O-alkyl or optionally substituted alkyl. R 5 These are independently H or optionally substituted alkyl groups. Here If X is glucosyl, R 1 It is an unsubstituted alkyl group, R 1 and R 2 If each of them is H and m is 0, then X is something other than glucuronic acid or a salt thereof.

[0022] In some embodiments, X is a hexosyl group selected from the group consisting of glucosyl, mannosyl, galactosyl, allosyl, altorosyl, grosyl, idosyl and talosyl, or a uronic acid group selected from the group consisting of glucuronic acid, mannuronic acid, galacturonic acid, aluronic acid, altoruonic acid, guluronic acid, iduronic acid and tarronic acid, or a uronic acid amide group selected from the group consisting of glucuronamide, mannuronamide, galacturonamide, aluronamide, altoruronamide, guluronamide, iduronamide and tarronamide. R 1 and R 2 Each of these is independently H, CONH2, CO2H, CO2-alkyl, alkyl-OH, or unsubstituted alkyl. Each of Y and Z is independently H. m is 0, 1, or 2. A = NR3R4 or OR5. Here R 3 and R 4 Each of these is independently H, OH, O-alkyl or optionally substituted alkyl. R 5 These are independently H or optionally substituted alkyl groups. Here If X is glucosyl, R 1 It is an unsubstituted alkyl group, R 1 and R 2Each of them is H, and when m is 0, X is something other than glucuronic acid. Or its salt.

[0023] The present invention provides a compound having the following structure, [ka] During the ceremony, X is a hexosyl group selected from the group consisting of glucosyl, mannosyl, galactosyl, allosyl, altrosyl, grosyl, idosyl, and thalosyl. R 1 and R 2 Each of these is independently H, halogen, OH, O-alkyl, CONH2, or optionally substituted alkyl. R 3 and R 4 Each of these is independently H, OH, O-alkyl, or optionally substituted alkyl. Each of Y and Z is independently H, OH, O-alkyl, or optionally substituted alkyl. m is 0, 1, or 2. If X is glucosyl, R 1 It is an alkyl group that may be substituted in some cases.

[0024] The present invention provides a compound having the following structure, [ka] During the ceremony, X is a hexosyl group selected from the group consisting of glucosyl, mannosyl, galactosyl, allosyl, altrosyl, grosyl, idosyl, and thalosyl. R 1 and R 2 Each of these is independently H, OH, O-alkyl, or optionally substituted alkyl. R 3 and R 4 Each of these is independently H, OH, O-alkyl, or optionally substituted alkyl. Each of Y and Z is independently H, OH, O-alkyl, or optionally substituted alkyl. m is 0, 1, or 2. If X is glucosyl, R 1 It is an alkyl group that may be substituted in some cases.

[0025] In some embodiments, X is glucosyl, mannosyl, or galactosyl.

[0026] In some embodiments, each of Y and Z is H.

[0027] In some embodiments, R 1 and R 2 Each of them is either H or R 1 is CH3 and R 2 Is H or R 1 is CONH2 and R 2 H is, or R 1 is CO2CH3 and R 2 H is H.

[0028] In some embodiments, R 3 and R 4 Each of them is H.

[0029] In some embodiments, the compound has the following structure: [ka] In the formula, R 1 is H, optionally a substituted alkyl group, or CONH2, and m is 0, 1, or 2.

[0030] In some embodiments, the compound has the following structure: [ka] In the formula, R 1 is H, optionally a substituted alkyl group, or CONH2, and m is 0, 1, or 2.

[0031] In some embodiments, the compound has the following structure: [ka] In the formula, R 1 is H, optionally a substituted alkyl group, or CONH2, and m is 0, 1, or 2.

[0032] In some embodiments of the above compound, R 1 m is H, hydroxyalkyl, or CONH2, and m is 0, 1, or 2.

[0033] In some embodiments, m is 0. In some embodiments, m is 1.

[0034] In some embodiments, the hexosyl group is an α-anomeric. In some embodiments, the hexosyl group is a β-anomeric. In some embodiments, the hexosyl group is a mixture of an α-anomeric and a β-anomeric. In some embodiments, the hexosyl group is a D-hexose. In some embodiments, the hexosyl group is an L-hexose. In some embodiments, the hexosyl group is a mixture of a D-hexose and an L-hexose.

[0035] In some embodiments, this compound has the following structure [ka] It has, in the formula, R 1 is H, CONH2, or optionally a substituted alkyl group.

[0036] In some embodiments of the above compound, R 1 It is a hydroxyalkyl group.

[0037] In some embodiments of the above compound, R 1 It is a C1-C4 hydroxyalkyl group.

[0038] In some embodiments, this compound has the following structure [ka] [ka] It holds.

[0039] In some embodiments, this compound has the following structure [ka] [ka] It holds.

[0040] In some embodiments, this compound has the following structure [ka] [ka] It holds.

[0041] In some embodiments, this compound or a salt thereof has the following structure [ka] It has, in the formula, X is a uronic acid group selected from the group consisting of glucuronic acid, mannuronic acid, galacturonic acid, arulonic acid, argulonic acid, guluronic acid, iduronic acid, and tarronic acid. R 1 and R 2 Each of these is independently H, halogen, OH, O-alkyl, CONH2, CO2H, CO2-alkyl, or optionally substituted alkyl. Each of Y and Z is independently H, OH, O-alkyl, or optionally substituted alkyl. R 5 These are independently H or optionally substituted alkyl groups. m is 0, 1, or 2. R 1 and R 2 If each of them is H and m is 0, then X is something other than glucuronic acid.

[0042] In some embodiments, this compound or a salt thereof has the following structure: [ka] During the ceremony, X is a uronic acid group selected from the group consisting of glucuronic acid, mannuronic acid, galacturonic acid, arulonic acid, argulonic acid, guluronic acid, iduronic acid, and tarronic acid. R 1 and R 2 Each of these is independently H, halogen, OH, O-alkyl, CO2H, CO2(alkyl), or optionally substituted alkyl. Each of Y and Z is independently H, OH, O-alkyl, or optionally substituted alkyl. R 5 These are independently H or optionally substituted alkyl groups. m is 0, 1, or 2. R 1 and R 2 If each of them is H and m is 0, then X is something other than glucuronic acid.

[0043] In some embodiments, this compound or a salt thereof has the following structure: [ka] During the ceremony, X is a uronic acid group selected from the group consisting of glucuronic acid, mannuronic acid, galacturonic acid, arulonic acid, argulonic acid, guluronic acid, iduronic acid, and tarronic acid. R 1 and R 2 Each of these is independently H, OH, O-alkyl, or optionally substituted alkyl. Each of Y and Z is independently H, OH, O-alkyl, or optionally substituted alkyl, R 5 is independently H or optionally substituted alkyl, m is 0, 1, or 2, R 1 and R 2 are each H and m is 0, X is other than glucuronic acid.

[0044] In some embodiments, it is the following compound or its salt, wherein X is an uronic acid group selected from the group consisting of glucuronic acid, mannuronic acid, galacturonic acid, alluronic acid, alturonic acid, guluronic acid, iduronic acid, and taluronic acid, R 1 and R 2 are each independently H, CONH2, CO2H, CO2-alkyl, or unsubstituted alkyl, Each of Y and Z is H, R 5 is independently H or optionally substituted alkyl, m is 0, 1, or 2, R 1 and R 2 are each H and m is 0, X is other than glucuronic acid.

[0045] In some embodiments, X is glucuronic acid, mannuronic acid, or galacturonic acid.

[0046] In some embodiments, each of Y and Z is H. <H

[0047] In some embodiments, R 1 and R 2 are each H, or R 1 is CH3 and R 2 is H, or R 1 is CO2CH3 and R 2 is H.

[0048] In some embodiments, each of R 5 is H or CH3.

[0049] In some embodiments, the compound or its salt has the following structure,

Chemical formula

[0050] In some embodiments, the compound or its salt has the following structure,

Chemical formula

[0051] In some embodiments of the above compound, R 1 is H, hydroxyalkyl, or CO2H, R 5 is H, and m is 0, 1, or 2.

[0052] In some embodiments, the compound or its salt has the following structure,

Chemical formula

[0053] In some embodiments, m is 0. In some embodiments, m is 1.

[0054] In some embodiments, the uronic acid group is an α-anomer. In some embodiments, the uronic acid group is a β-anomer. In some embodiments, the uronic acid group is D-uronic acid. In some embodiments, the uronic acid group is L-uronic acid. In some embodiments, the uronic acid group is a mixture of D and L.

[0055] In some embodiments, this compound or a salt thereof has the following structure: [ka] In the formula, R 1 is H, possibly a substituted alkyl group, or CO2H.

[0056] In some embodiments, this compound or a salt thereof has the following structure: [ka] , where R 1 This may be a substituted alkyl group or CO2H group.

[0057] In some embodiments, the compound or a salt thereof has the following structure. [ka] [ka]

[0058] In some embodiments, the compound or a salt thereof has the following structure. [ka] [ka]

[0059] In some embodiments, the compound or a salt thereof has the following structure. [ka] [ka]

[0060] In some embodiments, this compound has the following structure. [ka] [ka]

[0061] In some embodiments, this compound has the following structure. [ka] [ka]

[0062] In some embodiments, this compound has the following structure. [ka] [ka]

[0063] In some embodiments, this compound is a potassium salt, a calcium salt, or a magnesium salt.

[0064] In some embodiments, this compound is a sodium salt.

[0065] In some embodiments, this compound or a salt thereof has the following structure: [ka] During the ceremony, X is a uronic acid amide group selected from the group consisting of glucuronamide, mannuronamide, galacturonamide, aluronamide, altoronamide, glucuronamide, iduronamide, and tallonamide. R 1 and R 2 Each of these is independently H, halogen, OH, O-alkyl, CONH2, CO2H, CO2-alkyl, or optionally substituted alkyl. Each of Y and Z is independently H, OH, O-alkyl, or optionally substituted alkyl. m is 0, 1, or 2. A=NR3R4, Here R 3 and R 4 Each of these is independently H, OH, O-alkyl, or optionally substituted alkyl.

[0066] In some embodiments, this compound or a salt thereof has the following structure: [ka] During the ceremony, X is a uronic acid amide group selected from the group consisting of glucuronamide, mannuronamide, galacturonamide, aluronamide, altoronamide, glucuronamide, iduronamide, and tallonamide. R 1 and R 2 Each of these is independently H, CONH2, CO2H, CO2-alkyl, alkyl-OH, or unsubstituted alkyl. Each of Y and Z is H, m is 0, 1, or 2. A=NR3R4, Here R 3 and R 4 Each of these is independently H, OH, O-alkyl, or optionally substituted alkyl.

[0067] In some embodiments, X is a uronic acid amide group selected from the group consisting of glucuronamide, mannuronamide, and galacturonamide.

[0068] In some embodiments, each of Y and Z is H.

[0069] In some embodiments, R 1 and R 2 Each of them is either H or R 1 is CH3 and R 2 Is H or R 1 is CONH2 and R 2 H is H.

[0070] In some embodiments, R 3 and R 4 Each of them is H.

[0071] In some embodiments, this compound has the following structure [ka] It has, in the formula, R 1 m is H, optionally a substituted alkyl group, or CONH2, and m is 0, 1, or 2.

[0072] In some embodiments, this compound has the following structure [ka] It has, in the formula, R 1 m is H, optionally a substituted alkyl group, or CONH2, and m is 0, 1, or 2.

[0073] In some embodiments, this compound has the following structure [ka] It has, in the formula, R 1m is H, optionally a substituted alkyl group, or CONH2, and m is 0, 1, or 2.

[0074] In some embodiments of the above compound, R 1 is H, hydroxyalkyl, or CONH2, and m is 0, 1, or 2.

[0075] In some embodiments, m is 0.

[0076] In some embodiments, m is 1.

[0077] In some embodiments, the uronic acid amide group is an α-anomer. In some embodiments, the uronic acid amide group is a β-anomer. In some embodiments, the uronic acid amide group is a D-uronic acid amide. In some embodiments, the uronic acid amide group is an L-uronic acid amide. In some embodiments, the uronic acid amide group is a mixture of D and L.

[0078] In some embodiments, this compound or a salt thereof has the following structure [ka] It has, in the formula, R 1 is H, optionally a substituted alkyl group, or CONH2.

[0079] In some embodiments, this compound or a salt thereof has the following structure [ka] [ka] It holds.

[0080] In some embodiments, this compound or a salt thereof has the following structure [ka] [ka] It holds.

[0081] In some embodiments, this compound or a salt thereof has the following structure [ka] [ka] It holds.

[0082] In some embodiments, the composition comprises a biomolecule and at least one compound of the present invention.

[0083] In some embodiments, the composition comprises a biomolecule and a compound having the following structure: [ka] During the ceremony, X is a hexosyl group selected from the group consisting of glucosyl, mannosyl, and galactosyl. R 1 and R 2 Each of these is independently H, halogen, OH, O-alkyl, CONH2, or optionally substituted alkyl. R 3 and R 4 Each of these is independently H, OH, O-alkyl, or optionally substituted alkyl. Each of Y and Z is independently H, OH, O-alkyl, or optionally substituted alkyl. m is 0, 1, or 2.

[0084] In some embodiments, the composition comprises a biomolecule and a compound having the following structure or a salt thereof. [ka] During the ceremony, X is a uronic acid group selected from the group consisting of glucuronic acid, mannuronic acid, and galacturonic acid, R 1 and each of R 2 is independently H, halogen, OH, O-alkyl, CONH2, CO2H, CO2-alkyl, or optionally substituted alkyl, each of Y and Z is independently H, OH, O-alkyl, or optionally substituted alkyl, R 3 is independently H or optionally substituted alkyl, m is 0, 1, or 2.

[0085] In some embodiments, the composition comprises a biomolecule and a compound having the following structure,

Chemical formula

[0086] In some embodiments of this composition, the biomolecule is a biopharmaceutical, protein, nucleotide, polypeptide or antibody.

[0087] In some embodiments of this composition, the biomolecule has therapeutic activity.

[0088] In some embodiments of this composition, the biomolecules include insulin, humulin, novolin, human inhaled insulin, exvera, insulin aspart, novologue (aspart), insulin glulisine, apidra (glucisine), insulin lispro, humalog (lispro), isophane insulin, NPH, insulin detemir, levemir (detemir), insulin glargine, lantus (glargine), sustained-release insulin zinc, lente, ultralente, acetate Ramrintide, Simurin, Growth Hormone (GH), Somatotropin, Genotropin, Humatrope, Norditropin, NorIVitropin, Neutropin, Omnitrope, Protropin, Siazen, Cerostim, Bartropin, Mecasermin, Increlex, Mecasermin Lymphabate, IPlex, Factor VIII, Biocrate, Helixate, Cogenete, Re Recomate, ReFacto, Factor IX, Benefix, Antithromin III (AT-III), Trombate III, Protein C concentrate, Seprotin, β-glucocerebrosidase, Cerezyme, β-glucocerebrosidase, Cereidase (purified from pooled human placenta), Alglucosidase-α, Myozyme, Laronidase (α-l-iduronidase), Aldurazyme, Idurusulphase (iduronic acid-2) -Sulfatase), Elapase, Galsulphas, Naglazyme, Agalsidase-β (Human α-Galactosidase A), Fabrazyme, α-1-proteinase inhibitor, Aralast, Prolastin, Lactase, Lactide, Pancreatic enzymes (Lipase, Amylase, Protease), Arco-Lase, Cotazym, Creon, Donazyme, Pancrease, Biocase, Zymase, Adenosine deaminase (Pegademase Bovine, PEG-ADA), Addagen, Pooled Immunoglobulin, Octagum, Human Albumin, Albumarc, Albumin, Albuminer, AlbuRx, Albutein, Flexbumin, Buminate, Plasbumin, Erythropoietin, Epoetin-α, Epogen, Procrit, Darbepoetin-α,Aranesp, Filgrastim (granulocyte colony-stimulating factor, G-CSF), Neupogen, Pegfilgrastim (Peg-G-CSF), Neulasta, Salgramostim (granulocyte-macrophage colony-stimulating factor, GM-CSF), Leukin, Oprelbekin (interleukin-11, IL-11), Neumega, human follicle-stimulating hormone (FSH), Gonal-F, Follistim, human chorionic gonadotropin (HCG), Ovidrel, Luveli S, type I α interferon, interferon alphacon 1, consensus interferon, infergen, interferon-α2a (IFNα2a), loferon-A, pegylated interferon-α2a, pegasys, interferon-α2b (IFNα2b), intron A, pegylated interferon-α2b, pegylated intron, interferon-αn3 (IFNαn3), alferon N, interferon-β1a (rIFN -β), Avonex, Rebif, Interferon-β1b (rIFN-β), Beta-Celon, Interferon-γ1b (IFNγ), Actimune, Aldesleukin (Interleukin-2 (IL2), Epidermal Thymocyte Activator, ETAF), Proleukin, Alteplase (Tissue Plasminogen Activator, tPA), Activase, Leteplase (tPA Deletion Mutant Protein), Letabase, Tenecteplase, TN K-ase (TNKase), urokinase, avokinase, factor VIIa, NovoSeven, drotoreggin-α (activated protein C), Xigris, salmon calcitonin, Fortical, miacalcin, teriparatide (human parathyroid hormone residues 1-34), Forteo, exenatide, Vietta, octreotide, Sandostatin, dibotermin-α (recombinant human bone morphogenic protein 2, rhBMP2), Infuse, recombinant human bone morphogenic protein 7 (rhBMP7), osteogenic protein 1, histrelin acetate (gonadotropin-releasing hormone, GnRH), suprelin LA, Vantas, parifermin (keratinocyte growth factor KGF), Kebivans,Becaplermin (platelet-derived growth factor, PDGF), Regranex, Trypsin, Granulex, Nesiritide, Natrecor, Botulinum toxin type A, Botox, Botulinum toxin type B, Myoblock, Collagenase, Santil, Stardeoxyribonuclease I, Dornase α, Pulmozyme, Hyaluronidase (bovine, sheep), Anfadase (bovine), Hydase (bovine), Vitrase (sheep), Hyaluronidase (recombinant human), Hylenex Papain, Accuzyme, Panafil, L-asparaginase, ELSPAR, PEG-asparaginase, Oncaspar, Rasburicase, Elitek, Repildin, Lefuldan, Vivalirudin, Angiomax, Streptokinase, Streptase, Anistreplase (Anisoylated Plasminogen Streptokinase Activated Complex, APSAC), Eminase, Bevacizumab, Avastin, Cetuximab, Erbitux, Panitumumab, Vectibix, Alemtuzumab, Campas Rituximab, Rituxan, Trastuzumab, Herceptin, Abatacept, Orencia, Anakinra, Antril, Kineret, Avalimumab, Humira, Etanercept, Enbrel, Infliximab, Remicade, Alefacept, Amevive, Natalizumab, Tysabri, Eculizumab, Soliris, Antithymocyte globulin (rabbit), Thymoglobulin, Basiliximab, Simulect, Daclizumab, Zenapax, Muromonab-CD3, Orthoclone, OKT3, Omalizuma Xolair, palivizumab, Synagis, Enfvirtide, Fuzeon, absiximab, ReoPro, pegvisomant, Somabart, Crotalidae multivalent immune Fab (sheep), Crofab, digoxin immune serum Fab (sheep), Digifab, ranibizumab, Lucentis, denileukin, Diftitox, Ontac, ibritumomab, tiucetan, zevalin, gemtuzumab,Ozogamicin, Mylotarg, Tositumomab and I-Tositumomab, Bexar, Bexar I-131, hepatitis B surface antigen (HBsAg), Engerix, Recombivax HB, HPV vaccine, Gardasil, OspA, LYMErix, anti-rhesus (Rh) immunoglobulin G, Roferon, recombinant purified protein derivative (DPPD), glucagon, GlucaGen, growth hormone-releasing hormone (GHRH), Geref, secretin, ChiRhoStim (human peptide), SecreFlo (porcine peptide), thyroid-stimulating hormone (TSH), thyrotropin, capromab pendetide, ProstaScint, indium-111-octreotide, OctreoScan, satumomab pendetide, OncoScint, arcitumomab, CEA-Scan, nofetumomab, Verluma, aptide, Acutect, pentetreotide imsirmab, Myoscint, technetium fanolesomab, NeutroSpec, HIV antigen, enzyme immunoassay, OraQuick, UniGold, hepatitis C antigen, or recombinant immunoblot assay (RIBA).

[0089] In some embodiments of this composition, the biomolecule is lysozyme, adalimumab (Humira®), ubiquitin or factor IX.

[0090] In some embodiments of this composition, a buffer is further included.

[0091] In some embodiments of this composition, the composition is acidic.

[0092] In some embodiments of this composition, the pH is less than 6.8.

[0093] In some embodiments of this composition, the pH is less than 4.

[0094] In some embodiments of this composition, the pH is about 3.

[0095] In some embodiments of this composition, the pH is between 5 and 7.

[0096] In some embodiments of this composition, the composition is basic.

[0097] In some embodiments of this composition, the pH is above 7.2.

[0098] In some embodiments of this composition, the pH is above 10.

[0099] In some embodiments of this composition, the pH is about 12.

[0100] In some embodiments of this composition, the pH is between 7 and 8.

[0101] In some embodiments of this composition, the pH is less than 12.

[0102] In some embodiments of this composition, the pH is above 3.

[0103] In some embodiments of this composition, the pH is greater than 3 and less than 12.

[0104] In some embodiments of the composition, the composition is freeze-dried, lyophilized, a solution, a liquid, a solid, or a suspension.

[0105] In some embodiments of the composition, the compound is present at a concentration between 0.1 mM and about 5 M.

[0106] In some embodiments of the composition, the compound is present at a concentration between about 0.01 M and about 1 M.

[0107] The present invention also provides a composition comprising any compound of the present invention.

[0108] The present invention also provides a pharmaceutical composition comprising a compound of the present invention.

[0109] The present invention also provides a pharmaceutical composition comprising a compound of the present invention and at least one pharmaceutically acceptable carrier.

[0110] A method for stabilizing a biomolecule, the method comprising treating the biomolecule with an effective amount of a compound of the present invention, thereby stabilizing the biomolecule.

[0111] In some embodiments, in this method, the biomolecule is a protein, nucleotide, polypeptide or antibody.

[0112] In some embodiments, in this method, the biomolecule has therapeutic activity.

[0113] In some embodiments, in this method, the biomolecule is a biopharmaceutical.

[0114] In some embodiments, in this method, the biomolecules are insulin, humulin, Novolin, human inhaled insulin, Exvera, insulin aspart, Novolog (aspart), insulin glulisine, Apidra (glucisine), insulin lispro, Humalog (lispro), isophane insulin, NPH, insulin detemir, Levemir (detemir), insulin glargine, Lantus (glargine), sustained-release insulin zinc, Lente, Ultralente, and vinegar. Plumrintide acid, Simurin, Growth hormone (GH), Somatotropin, Genotropin, Humatrope, Norditropin, NorIVitropin, Neutropin, Omnitrope, Protropin, Siazen, Celostim, Bartropin, Mecasermin, Increlex, Mecasermin Lymphabate, IPlex, Factor VIII, Biocrate, Helixate, Cogenete, Recomminate, ReFacto, Factor IX, Benefix, Antithromin III (AT-III), Trombate III, Protein C concentrate, Seprotin, β-glucocerebrosidase, Cerezyme, β-glucocerebrosidase, Cereidase (purified from pooled human placenta), Alglucosidase-α, Myozyme, Laronidase (α-l-iduronidase), Aldurazyme, Idurusulphase (iduronic acid- 2-sulfatase), elapase, galsulfase, naglazyme, agalsidase-β (human α-galactosidase A), fabrazyme, α-1-proteinase inhibitor, aralast, prolastin, lactase, lactide, pancreatic enzymes (lipase, amylase, protease), Arco-Lase, Cotazym, Creon, Donazyme, pancrease, biocase, zymase, adenosine deaminase (bovine pegademase, PEG-ADA), adagen, pooled immunoglobulin, octagum, human albumin, Albumarc, albumin, albuminer, AlbuRx, albutein, flexbumin, buminate, prasbumin, erythropoietin, epoetin-α, epogen, procrit,Darbepoetin-α, Alanesp, Filgrastim (granulocyte colony-stimulating factor, G-CSF), Neupogen, Pegfilgrastim (Peg-G-CSF), Neurasta, Salgramostim (granulocyte-macrophage colony-stimulating factor, GM-CSF), Leukin, Oprelbekin (interleukin-11, IL-11), Neumega, Human follicle-stimulating hormone (FSH), Gonal-F, Follistim, Human chorionic gonadotropin (HCG), Obi Dorel, Luveris, Type I α-interferon, Interferon Alphacon 1, Consensus Interferon, Infergen, Interferon-α2a (IFNα2a), Loferon-A, PegInterferon-α2a, Pegasys, Interferon-α2b (IFNα2b), Intron A, PegInterferon-α2b, PegIntron, Interferon-αn3 (IFNαn3), Alferon N, Interferon-β1a (rIFN-β), Avonex, Rebif, Interferon-β1b (rIFN-β), Beta-Celon, Interferon-γ1b (IFNγ), Actimune, Aldesleukin (Interleukin-2 (IL2), Epidermal Thymocyte Activating Factor, ETAF), Proleukin, Alteplase (Tissue Plasminogen Activating Factor, tPA), Activase, Leteplase (tPA Deletion Mutant Protein), Letabase, Tenecteplase TNKase, urokinase, avokinase, factor VIIa, NovoSeven, drotoreggin-α (activated protein C), Xigris, salmon calcitonin, Fortical, miacalcin, teriparatide (human parathyroid hormone residues 1-34), Forteo, exenatide, Vietta, octreotide, Sandostatin, dibotermin-α (recombinant human bone morphogenic protein 2, rhBMP2), Infuse, recombinant human bone morphogenic protein 7 (rhBMP7), osteogenic protein 1, histrelin acetate (gonadotropin-releasing hormone, GnRH), suprelin LA, Vantas,Palifermin (keratinocyte growth factor KGF), Kebivans, Becaplermin (platelet-derived growth factor, PDGF), Regranex, Trypsin, Granulex, Nesiritide, Natrecor, Botulinum toxin type A, Botox, Botulinum toxin type B, Myoblock, Collagenase, Santil, Stardeoxyribonuclease I, Dornase α, Pulmozyme, Hyaluronidase (bovine, sheep), Anfadase (bovine), Hydase (bovine), Vitrase (Sheep), Hyaluronidase (recombinant human), Hylenex, Papain, Accuzyme, Panafil, L-asparaginase, ELSPAR, Peg-asparaginase, Oncaspar, Rasburicase, Elitek, Repildin, Lefludan, Vivalirudin, Angiomax, Streptokinase, Streptase, Anistreplase (Anisoylated Plasminogen Streptokinase Activated Complex, APSAC), Eminase, Bevacizumab, Avastin, Cetuximab, Erbitux Panitumumab, Vectibix, Alemtuzumab, Campas, Rituximab, Rituxan, Trastuzumab, Herceptin, Abatacept, Orencia, Anakinra, Antril, Kineret, Avalimumab, Humira, Etanercept, Enbrel, Infliximab, Remicade, Alefacept, Amevive, Natalizumab, Tysabri, Eculizumab, Soliris, Antithymocyte globulin (rabbit), Thymoglobulin, Basiliximab, Simulect, Daclizumab, Zenapax, Muromonab-CD3, Orthoclone, OKT3, Omalizumab, Xolair, Palivizumab, Synagis, Enfvirtide, Fuzeon, Absiximab, ReoPro, Pegvisomant, Somavert, Crotalidae Multivalent Immunotherapy Fab (Sheep), Crofab, Digoxin Immunotherapy Fab (Sheep), Digifab, Ranibizumab, Lucentis, Denileukin, Diftitox, Ontac,Ibritumomab, Tiucetan, Zevalin, Gemtuzumab, Ozogamicin, Mylotarg, Tositumomab and I-Tositumomab, Vexar, Vexar I-131, Hepatitis B surface antigen (HBsAg), Engerix, Recombivax HB, HPV vaccine, Gardasil, OspA, Lymerix, Anti-rhesus (Rh) immunoglobulin G, Rophyllac, Recombinant purified protein derivative (DPPD), Glucagon, GlucaGen, Growth hormone-releasing hormone (GHRH), Geref, Secretin, ChiRhoStim (human peptide), Secretflow Flo) (porcine peptide), thyroid-stimulating hormone (TSH), thyrotropin, capromab penetide, prostaScint, indium-111-octreotide, OctreoScan, satumomab penetide, oncoScint, arcitumomab, CEA-Scan, nofetumomab, Verluma, aptide, Acutect, pentiminate imusilomab, myosinto, technetium phanoresomab, NeutroSpec, HIV antigen, enzyme immunoassay, OraQuick, Unigold, hepatitis C antigen, or recombinant immunoblot assay (RI BA).

[0115] In some embodiments, the biomolecule in this method is lysozyme, adalimumab (Humira®), ubiquitin, or factor IX.

[0116] In some embodiments, the biomolecules are stabilized in the presence of pH stress in this method.

[0117] In some embodiments, the pH stress in this method is an acidic environment.

[0118] In some embodiments, the acidic environment in this method has a pH of less than 6.8. In some embodiments, the acidic environment in this method has a pH of less than 4. In some embodiments, the acidic environment in this method has a pH of about 3. In some embodiments, the acidic environment in this method has a pH between 5 and 7.

[0119] In some embodiments, the pH stress in this method is a basic environment.

[0120] In some embodiments, the basic environment in this method is higher than 7.2. In some embodiments, the basic environment in this method has a pH greater than 10. In some embodiments, the basic environment in this method has a pH of about 12.

[0121] In some embodiments, in this method, the biomolecules are treated with the compound before being subjected to pH stress.

[0122] In some embodiments, the biomolecules are stabilized in the presence of thermal stress in this method.

[0123] In some embodiments, the thermal stress in this method is freeze-drying, lyophilization, or heating of biomolecules.

[0124] In some embodiments, the thermal stress in this method is heating the biomolecules to a glass transition state or melting point.

[0125] In some embodiments, in this method, the biomolecules are treated with the compound before being subjected to thermal stress.

[0126] In some embodiments, in this method, the biomolecules are treated with compounds at concentrations ranging from 0.1 mM to approximately 5 M.

[0127] In some embodiments, in this method, the biomolecules are treated with a compound at a concentration between 0.1 M and about 1 M.

[0128] In some embodiments, any compound or mixture thereof of the present invention is for use in any method of the present invention.

[0129] In some embodiments, the composition or pharmaceutical composition comprises any compound or mixture thereof of the present invention.

[0130] The compounds of the present invention include neutral glycosylated amides (neutral amide type) and dianionic glucuronic acid-treated acids (diionic uronic acid type). Preferably, the neutral amide type stabilizer contains only non-ionizable functional groups, such as amide and hydroxyl functional groups, while the dianionic uronic acid type stabilizer contains two ionizable carboxylic acid functional groups, one at C-6 of the hexose moiety and the other linked to the hexose via a glycosidic bond. The dianionic uronic acid type stabilizers of the present invention include stabilizers containing two ionizable acid groups that are neutral, as well as salts derived from their monoanionic and dianionic forms, such as monosodium salts, disodium salts, monopotassium salts, dipotassium salts, calcium salts, magnesium salts, and the like. In certain embodiments, the compound is a dipotassium salt.

[0131] The compounds of the present invention include all hydrates, solvates, and complexes of the compounds used in the present invention. Where a chiral center or another form of isomeric center is present in the compounds of the present invention, all forms of such isomers (plural), including enantiomers and diastereomers, are intended to be covered herein. Compounds containing a chiral center can be used as a racemic mixture, an enantiomer-rich mixture, or the racemic mixture can be separated using known techniques, and the individual enantiomers can be used individually. The enantiomers can be separated using known techniques such as those described in Pure and Applied Chemistry 69, 1469-1474, (1997) IUPAC.

[0132] Unless otherwise specified, if the structure of a compound of the present invention contains a chiral carbon atom, it is understood that the compound exists as a racemic compound, a racemic mixture, and an isolated single enantiomer. All such isomers of these compounds are clearly included in the present invention. Unless otherwise specified, each stereogenic carbon can be in an R or S configuration. Therefore, isomers resulting from such asymmetry (e.g., all enantiomers and diastereomers) should be understood to be included within the scope of the present invention unless otherwise indicated. Such isomers can be obtained in substantially pure form by classical separation techniques and stereochemically controlled synthesis, as described in “Enantiomers, Racemates and Resolutions” by J. Jacques, A. Collet and S. Wilen, Pub. John Wiley & Sons, NY, 1981. For example, resolution can be performed by preparative chromatography on a chiral column.

[0133] The compounds of the present invention may have spontaneous tautomers. Where a compound may exist in tautomers such as keto-enol tautomers, each tautomer is intended to be included in the present invention, whether it exists in equilibrium or primarily in one form.

[0134] In the compound structures shown herein, hydrogen atoms are not shown for carbon atoms with fewer than four bonds to non-hydrogen atoms. However, it is understood that there are enough hydrogen atoms on the above carbon atoms to satisfy the octet rule.

[0135] Where numerical ranges are described herein, unless otherwise specified, the present invention is understood to include integers between the upper and lower limits, and to include the upper and lower limits.

[0136] The present invention also provides isotopic variants of the compounds disclosed herein. Thus, in the compounds provided herein, hydrogen can be enriched with deuterium isotopes. Any notation of carbon in the overall structure of this application, when used without further notation, 12 C, 13 C, or 14 It should be noted that this is intended to represent all isotopes of carbon, such as C. 13 C or 14 Any compound containing C may specifically have the structure of any compound disclosed herein. Furthermore, any representation of hydrogen in the structures of this application, when used without further designation, 1 H, 2 H, or 3 It should also be noted that this is intended to represent all isotopes of hydrogen, such as H. 2 H or 3 Any compound containing H may specifically have any of the structures of the compounds disclosed herein. Isotope-labeled compounds can generally be prepared by conventional techniques known to those skilled in the art, using appropriate isotope-labeled reagents instead of the unlabeled reagents used. It is understood that the present invention encompasses all such isotopic forms.

[0137] It is understood that the substituents and substitution patterns of the compounds used in the methods of the present invention can be selected by those skilled in the art to provide compounds that are chemically stable and can be readily synthesized from starting materials readily available by the art. It is understood that if the substituent itself is substituted with more than one group, these multiple groups may be on the same carbon or different carbons, as long as a stable structure is obtained.

[0138] As used herein, "alkyl" is intended to include both branched, linear, and cycloalkyl saturated aliphatic hydrocarbon groups having a specified number of carbon atoms. Thus, alkyl specifically includes methyl, ethyl, propyl, cyclopropyl, isopropyl, butyl, pentyl, hexyl, heptyl, isopropyl, isobutyl, sec-butyl, and the like. One embodiment is C1-C 12 Alkyl, C1-C3 alkyl, C2-C 12 Alkyl, C3-C 12 Alkyl, C4-C 12 This may include alkyl groups. Adjacent alkyl substituents may be linked to form a saturated carbon ring. In this specification, "cycloalkyl" means a cyclic ring of an alkane having a total of 3 to 8 carbon atoms, or any number within this range (i.e., cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, or cyclooctyl). The alkyl group may be optionally substituted. For example, the alkyl group may be optionally substituted with an oxygen, nitrogen, or sulfur atom. As another example, the alkyl group may be substituted with phenyl, alcohol, halogen (i.e., F, Cl, Br, and I), alkoxy groups such as methoxy, ethoxy, n-propoxy, and isopropoxy, alkylthio groups such as methylthio and ethylthio, carboxylate, or acetate groups.

[0139] The "O-alkyl" group means an (oxygen)-R radical, where R is an alkyl group as defined above. For example, an O-alkyl group can be an oxygen atom bonded to a C1-C6 linear or branched alkyl group.

[0140] The "hexosyl" group is a hexose radical. Hexosyl groups may be, but are not limited to, glucosyl, mannosyl, and galactosyl. Other hexosyl groups include allosyl, altrosyl, grosyl, idosyl, and thalosyl. Hexosyl includes non-oxidized hexosyl groups, but may also include oxidized hexosyl groups such as uronic acid groups. Uronic acid groups are uronic acid radicals that may be, but are not limited to, glucuronsyl, mannuronsyl, and galacturonsyl. Hexose or oxidized hexose groups may be D or L stereoisomers. Hexosyl groups may be α- or β-anomers. Hexosyl groups are bonded to the parent substrate via oxygen to C-1, C-2, C-3, C-4, or C-6. Hexosyl groups may be α- or β-anomers.

[0141] The "glucosyl" group is a radical of the glucose molecule. The glucose molecule can be D or L mannose. The glucosyl group is bonded to the parent substrate via oxygen at C-1, C-2, C-3, C-4, or C-6. The glucosyl group can be an α- or β-anomer. Unless otherwise specified, the glucosyl group is bonded to the oxygen away from the anomeric C-1. For example, glucosyl can be defined as follows: [ka]

[0142] The "galactosyl" group is a radical of the galactose molecule. The galactose molecule can be D or L mannose. The galactosyl group is bonded to the parent substrate via oxygen at C-1, C-2, C-3, C-4, or C-6. The galactosyl group can be an α- or β-anomer. Unless otherwise specified, the galactosyl group is bonded to the oxygen away from the anomeric C-1. For example, galactosyl can be defined as follows: [ka]

[0143] The "mannosyl" group is a radical of the mannose molecule. The mannose molecule can be D or L mannose. The mannosyl group is bonded to the parent substrate via oxygen at C-1, C-2, C-3, C-4, or C-6. The mannosyl group can be an α- or β-anomer. Unless otherwise specified, the mannosyl group is bonded to oxygen away from the anomeric C-1. For example, mannosyl is defined as follows: [ka] A "glucuronosyl" or "glucuronide group" is a radical of the glucuronic acid molecule. The glucuronide group can be D or L glucuronic acid. The glucuronide group is bonded to the parent substrate via oxygen at C-1, C-2, C-3, C-4, or C-6. The glucuronide group can be α- or β-anomer. Unless otherwise specified, the glucuronide group is bonded to oxygen away from the anomeric C-1. For example, the glucuronide group can be defined as follows: [ka]

[0144] A "galacturonosyl" or "galacturonic acid group" is a radical of the galacturonic acid molecule. The galacturonic acid group can be D or L-galacturonic acid. The galacturonic acid group is bonded to the parent substrate via oxygen at C-1, C-2, C-3, C-4, or C-6. The galacturonic acid group can be an α- or β-anomer. Unless otherwise specified, the galacturonic acid group is bonded to oxygen away from the anomeric C-1. For example, the galacturonic acid group can be defined as follows: [ka]

[0145] A "mannuronosyl" or "mannuronic acid group" is a radical of the mannuronic acid molecule. The mannuronic acid group can be D or L mannuronic acid. The mannuronic acid group is bonded to the parent substrate via oxygen at C-1, C-2, C-3, C-4, or C-6. The mannuronic acid group can be α- or β-anomer. Unless otherwise specified, the mannuronic acid group is bonded to oxygen away from the anomeric C-1. For example, the mannuronic acid group can be defined as follows: [ka]

[0146] As used herein, the term "halogen" refers to F, Cl, Br, and I.

[0147] A "possibly substituted" group refers to a functional group in which one or more bonds to hydrogen atoms are replaced by bonds to non-hydrogen or non-carbon atoms, provided that the normal valence is maintained and the substitution results in a stable compound. Substituted groups also include groups in which one or more bonds to carbon or hydrogen atoms are replaced by one or more bonds, including double or triple bonds, to heteroatoms. Examples of substituents include halogens (i.e., F, Cl, Br, and I), alkyl groups such as methyl, ethyl, n-propyl, isopropyl, n-butyl, tert-butyl, and trifluoromethyl, alkoxy groups such as hydroxyl, methoxy, ethoxy, n-propoxy, and isopropoxy, aryloxy groups such as phenoxy, arylalkyloxy groups such as benzyloxy(phenylmethoxy) and p-trifluoromethylbenzyloxy(4-trifluoromethylphenylmethoxy), heteroaryloxy groups, sulfonyl groups such as trifluoromethanesulfonyl, methanesulfonyl, and p-toluenesulfonyl, sulfanyl groups such as nitro, nitrosyl, mercapto, methylsulfanyl, ethylsulfanyl, and propylsulfanyl, amino groups such as cyano, amino, methylamino, dimethylamino, ethylamino, and diethylamino, and carboxyls. If multiple substituents are disclosed or requested, the substituted compound may be substituted individually or in combination with one or more of the disclosed or requested substituents. To be independently substituted means that the (two or more) substituents may be the same or different.

[0148] The compounds of the present invention also include any of the compounds disclosed herein that are modified with common protecting groups. For example, the compounds of the present invention include glycosylated amides as described herein, but are modified if the amide is protected by an amide protecting group, e.g., BOC, and the hydroxyl group is protected by a hydroxyl protecting group, e.g., benzyl. Another example is that the compounds of the present invention include glucuronized acids in which the carboxylic acid moiety is protected as an ester. Common protecting groups are known to those skilled in the art, as described in Greene's Protective Groups in Organic Synthesis (Wuts (2006)).

[0149] When selecting compounds of the present invention, those skilled in the art will recognize that various substituents should be selected according to well-known principles of chemical structure bonding.

[0150] The compounds used in the methods of the present invention may be in the form of salts. As used herein, “salt” refers to a salt of the compound modified by producing an acidic or basic salt of the compound. In the case of compounds used to stabilize therapeutic biomolecules, the salt may be a pharmaceutically acceptable salt. Examples of pharmaceutically acceptable salts include, but are not limited to, mineral or organic acid salts of basic residues such as amines, and alkali or organic salts of acidic residues such as carboxylic acids. Salts can be produced using organic or inorganic acids. Such salts include, but are not limited to, alkali metal and alkaline earth metal salts such as lithium, sodium, potassium, beryllium, magnesium, and calcium salts. Salts also include alkylammonium salts, ammonium salts, and salts derived from amino acids. In this regard, the term “pharmaceutically acceptable salt” refers to relatively non-toxic inorganic and organic acid or base addition salts of the compounds of the present invention. These salts can be prepared in situ during the final isolation and purification of the compounds of the present invention, or by separately reacting the purified compounds of the present invention in the form of free bases or free acids with appropriate organic or inorganic acids or bases and isolating the salts thus formed. (See, for example, Berge et al. (1977) “Pharmaceutical Salts”, J. Pharm. Sci. 66:1-19.)

[0151] The compounds of the present invention may be used in pharmaceutical compositions containing therapeutic biomolecules mixed with suitable pharmaceutical diluents, expanders, excipients, or carriers.

[0152] "Biomolecules" are proteins, nucleotides, polypeptides, antibodies including monoclonal antibodies, enzymes, or fragments or mixtures of any of the above. Biomolecules may also be fragments of cells, viruses, liposomes, or tissues. In another embodiment, biomolecules may or may not have therapeutic activity.

[0153] In embodiments of the present invention, the therapeutic biomolecule may be one of the following: insulin, humuline, novolin, human inhaled insulin, exvera, insulin aspart, novologue (aspart), insulin glulisine, apidra (glucisine), insulin lispro, humalog (lispro), isophane insulin, NPH, insulin detemir, levemir (detemir), insulin glargine, lantus (glargine), sustained-release insulin zinc, lente, ultralente, pramulintide acetate, simulin, growth hormone (GH), somatotropin, genotropin, humatrope, norditropin, norivitropin, neutropin, omnitrope, protropin, siazen, celostim, bartropin, mecasermin, increlex, mecasermin lymphabate, iplex, factor VIII, biocrate, helixate, cogenete, recominate nate), ReFacto, Factor IX, Benefix, Antithromin III (AT-III), Trombate III, Protein C concentrate, Seprotin, β-glucocerebrosidase, Cerezyme, β-glucocerebrosidase, Ceredase (purified from pooled human placenta), Alglucosidase-α, Myozyme, Laronidase (α-l-iduronidase), Aldurazyme, Idurusulfate (iduronic acid-2-sulfate) -ase), elapase, galusulphase, naglazyme, agalsidase-β (human α-galactosidase A), fabrazyme, α-1-proteinase inhibitor, aralast, prolastin, lactase, lactide, pancreatic enzymes (lipase, amylase, protease), alkolase, Cotazym, Creon, Donazyme, pancrease, biocase, zymase, adenosine deaminase (pegademase bovine, PEG-ADA), adagen, pooled immunoglobulin, octagum, human albumin, Albumarc, albumin, albuminer, AlbuRx, albutin, flexbumin, buminate, prasbumin, erythropoietin, epoetin-α, epogen, procrit,Darbepoetin-α, Alanesp, Filgrastim (granulocyte colony-stimulating factor, G-CSF), Neupogen, Pegfilgrastim (Peg-G-CSF), Neurasta, Salgramostim (granulocyte-macrophage colony-stimulating factor, GM-CSF), Leukin, Oprelbekin (interleukin-11, IL-11), Neumega, Human follicle-stimulating hormone (FSH), Gonal-F, Follistim, Human chorionic gonadotropin (HCG), Obi Dorel, Luveris, Type I α-interferon, Interferon Alphacon 1, Consensus Interferon, Infergen, Interferon-α2a (IFNα2a), Loferon-A, PegInterferon-α2a, Pegasys, Interferon-α2b (IFNα2b), Intron A, PegInterferon-α2b, PegIntron, Interferon-αn3 (IFNαn3), Alferon N, Interferon-β1a (rIFN-β), Avonex, Rebif, Interferon-β1b (rIFN-β), Beta-Celon, Interferon-γ1b (IFNγ), Actimune, Aldesleukin (Interleukin-2 (IL2), Epidermal Thymocyte Activating Factor, ETAF), Proleukin, Alteplase (Tissue Plasminogen Activating Factor, tPA), Activase, Leteplase (tPA Deletion Mutant Protein), Letabase, Tenecteplase TNKase, urokinase, avokinase, factor VIIa, NovoSeven, drotoreggin-α (activated protein C), Xigris, salmon calcitonin, Fortical, miacalcin, teriparatide (human parathyroid hormone residues 1-34), Forteo, exenatide, Vietta, octreotide, Sandostatin, dibotermin-α (recombinant human bone morphogenic protein 2, rhBMP2), Infuse, recombinant human bone morphogenic protein 7 (rhBMP7), osteogenic protein 1, histrelin acetate (gonadotropin-releasing hormone, GnRH), suprelin LA, Vantas,Palifermin (keratinocyte growth factor KGF), Kebivans, Becaplermin (platelet-derived growth factor, PDGF), Regranex, Trypsin, Granulex, Nesiritide, Natrecor, Botulinum toxin type A, Botox, Botulinum toxin type B, Myoblock, Collagenase, Santil, Stardeoxyribonuclease I, Dornase α, Pulmozyme, Hyaluronidase (bovine, sheep), Anfadase (bovine), Hydase (bovine), Vitrase (Sheep), Hyaluronidase (recombinant human), Hylenex, Papain, Accuzyme, Panafil, L-asparaginase, ELSPAR, Peg-asparaginase, Oncaspar, Rasburicase, Elitek, Repildin, Lefludan, Vivalirudin, Angiomax, Streptokinase, Streptase, Anistreplase (Anisoylated Plasminogen Streptokinase Activated Complex, APSAC), Eminase, Bevacizumab, Avastin, Cetuximab, Erbitux Panitumumab, Vectibix, Alemtuzumab, Campas, Rituximab, Rituxan, Trastuzumab, Herceptin, Abatacept, Orencia, Anakinra, Antril, Kineret, Avalimumab, Humira, Etanercept, Enbrel, Infliximab, Remicade, Alefacept, Amevive, Natalizumab, Tysabri, Eculizumab, Soliris, Antithymocyte globulin (rabbit), Thymoglobulin, Basiliximab, Simulect, Daclizumab, Zenapax, Muromonab-CD3, Orthoclone, OKT3, Omalizumab, Xolair, Palivizumab, Synagis, Enfvirtide, Fuzeon, Absiximab, ReoPro, Pegvisomant, Somavert, Crotalidae Multivalent Immunotherapy Fab (Sheep), Crofab, Digoxin Immunotherapy Fab (Sheep), Digifab, Ranibizumab, Lucentis, Denileukin, Diftitox, Ontac,Ibritumomab, Tiucetan, Zevalin, Gemtuzumab, Ozogamicin, Mylotarg, Tositumomab and I-Tositumomab, Vexar, Vexar I-131, Hepatitis B surface antigen (HBsAg), Engerix, Recombivax HB, HPV vaccine, Gardasil, OspA, Lymerix, Anti-rhesus (Rh) immunoglobulin G, Rophyllac, Recombinant purified protein derivative (DPPD), Glucagon, GlucaGen, Growth hormone-releasing hormone (GHRH), Geref, Secretin, ChiRhoStim (human peptide), Secretflow Flo) (porcine peptide), thyroid-stimulating hormone (TSH), thyrotropin, capromab pendetide, prostaScint, indium-111-octreotide, OctreoScan, satumomab pendetide, oncoScint, arcitumomab, CEA-Scan, nofetumomab, Verluma, aptide, Acutect, pentiminate imusilomab, myosinto, technetium phanoresomab, NeutroSpec, HIV antigen, enzyme immunoassay, OraQuick, Unigold, hepatitis C antigen, or recombinant immunoblot assay (RI BA).

[0154] As used herein, “degradation” of biomolecules includes, but is not limited to, aggregation, denaturation, misfolding, and precipitation of biomolecules. Degradation may be caused by physical stress or chemical stress. Physical stress includes high or low temperatures, heating above the thermal unfolding temperature, freezing, stirring, shaking, surface, and pressure. Chemical stress includes low or high pH, ​​pH deviation from the ideal pH environment for natively folded proteins (e.g., deviation by pH 1, 2, 3, 4, or 5), dehydration, organic solvents, and the presence of impurities such as detergents and chaotropic agents.

[0155] Stabilized biomolecules retain their original structure and activity for longer periods or under a wider range of conditions than unstabilized biomolecules. Furthermore, or alternatively, stabilized biomolecules do not degrade under conditions that degrade the unstable form of the same biomolecule. Stabilized biomolecules have a higher melting temperature than unstabilized biomolecules.

[0156] Techniques and compositions for producing such compositions are described in the following references. 7 Modern Pharmaceutics, Chapters 9 and 10 (Banker & Rhodes, Editors, 1979); Pharmaceutical Dosage Forms: Tablets (Lieberman et al., 1981); Ansel, Introduction to Pharmaceutical Dosage Forms 2nd Edition (1976); Remington's Pharmaceutical Sciences, 17th ed. (Mack Publishing Company, Easton, Pa., 1985); Advances in Pharmaceutical Sciences (David Ganderton, Trevor Jones, Eds., 1992); Advances in Pharmaceutical Sciences Vol. 7. (David Ganderton, Trevor Jones, James McGinity, Eds., 1995); Aqueous Polymeric Coatings for Pharmaceutical Dosage Forms (Drugs and the Pharmaceutical Sciences, Series 36 (James McGinity, Ed., 1989); Pharmaceutical Particulate Carriers: Therapeutic Applications: Drugs and the Pharmaceutical Sciences, Vol 61 (Alain Rolland, Ed., 1993); Drug Delivery to the Gastrointestinal Tract (Ellis Horwood Books in the Biological Sciences. Series in Pharmaceutical Technology; JG Hardy, SS Davis, Clive G. Wilson, Eds.); Modem Pharmaceutics Drugs and the Pharmaceutical Sciences, Vol 40 (Gilbert S. Banker, Christopher T. Rhodes, Eds.) All of the aforementioned publications are incorporated herein by reference.

[0157] The present invention also includes embodiments in which the glucosyl, mannosyl, or galactosyl group is replaced with allosyl, altrosyl, grosyl, idosyl, or talosyl, or any corresponding uronic acid.

[0158] Each embodiment disclosed herein is intended to be applicable to each of the other embodiments disclosed herein. Therefore, all combinations of the various elements described herein fall within the scope of the present invention.

[0159] The present invention will be better understood by referring to the details of the following experiments, but those skilled in the art will readily understand that the specific detailed experiments are merely illustrative of the present invention, which will be better described in the subsequent claims.

[0160] Abbreviations of compound names The abbreviations for the compounds of the present invention are as follows: MglyA - Mannosyl glycolamide MLA - Mannosyl-lactamide GBA-3-glucosylbutanamide GaBA-3-galactosyl-butanamide GGlyA - Glucosyl glycolamide GaGlyA - Galactosyl glycolamide GLA - Glucosilulactamide GaLA - Galactosyl-lactamide b-GGlyA - β-glucosyl glycolamide b-GLA-β-glucosyl-lactamide b-GaGlyA - β-galactosyl glycolamide b-GBA-β-3-glucosylbutanamide a-MglyA-α-mannosyl glycolamide a-MLA-α-mannosyl-lactamide b-GGly - β-glucosyl-glycolate b-GL - β-glucosyl-lactic acid b-GB-β-3-glucosylbutyrate b-GaGly-β-galactosyl-glycolate b-GaL-β-galactosyl-lactic acid

[0161] Compound synthesis In general, the compounds of the present invention can be prepared using several methods known in the chemical art, in combination with the knowledge of those skilled in the art, particularly in light of the description contained herein. Various starting materials, intermediates, and reagents can be purchased from commercial sources or prepared according to methods or applications thereof in the literature. Other reagents, compounds, or methods can be used in practice or in testing, but generalized methods for the preparation of the compounds of the present invention are illustrated by the following description and reaction schemes. The methods disclosed herein, including those outlined in the schemes, descriptions, and examples, are intended for illustrative purposes and should not be construed in any way as limitations thereto. Given the merits of this disclosure, various changes and modifications will be apparent to those skilled in the art and are considered to fall within the technical spirit and scope of this disclosure, as further defined in the appended claims.

[0162] Specific embodiments of various aspects of the present invention are described with reference to schemes, preparations, and / or examples, but it should be understood that such embodiments are for illustrative purposes only and are merely a few examples of many possible specific embodiments that can represent the application of the principles of this disclosure. The starting materials used in the synthesis of the compounds described herein can be obtained from commercial sources such as Aldrich Chemical Co. (Milwaukee, Wis.) and Sigma Chemical Co. (St. Louis, Mo.), or the starting materials can be synthesized. The compounds described herein, and other related compounds having various substituents, can be synthesized using techniques and materials known to those skilled in the art, such as those described in March's Advanced Organic Chemistry: Reactions, Mechanisms, and Structure (Smith (2013)), Design and Strategy in Organic Synthesis (Hanessian (2013)), Greene's Protective Groups in Organic Synthesis (Wuts (2006)), and Fiesers' Reagents for Organic Synthesis (Volumes 1-27) (Ho (2013)), each of which is incorporated by reference as a whole.

[0163] The general methods for preparing the compounds disclosed herein can be derived from known reactions in the art, and these reactions can be modified by the use of appropriate reagents and conditions to introduce the various parts found in the formulas provided herein, as will be recognized by those skilled in the art. (Trombotto et al. 2000; Matsumura et al. 1997; Krajewski et al. 1997; Faria et al. 2008; Xue et al. 2009; Moynihan et al. 2013; WO2008 / 007153 A2; WO2012 / 109263 A1 and WO2015 / 137838)

[0164] The intermediate products described may be recovered by extraction, evaporation, or other techniques known in the art. The crude material may then be optionally purified by chromatography, HPLC, recrystallization, grinding, distillation, or other techniques known in the art.

[0165] As will be understood by those skilled in the art, some methods useful for preparing such compounds, as described above, may require protection of certain functional groups, for example, to prevent interference by the functional group in reactions at other sites, or to maintain the integrity of the molecule or functional group. The need and type of such protection is readily determined by those skilled in the art and varies, for example, depending on the nature of the functional group and the chosen preparation method. Methods for introducing and removing protecting groups are well known to those skilled in the art and are described in Greene's Protective Groups in Organic Synthesis (Wuts (2006)). As with methods for optimizing or adapting the procedures described herein, alternative reagents and starting materials are also readily determined by those skilled in the art.

[0166] Preparation of neutral glycosylated amides Amides from gluco-, manno-, and galactosides were prepared in quantitative yield by reaction with ammonia in methanol. Due to the presence of amide and hydroxyl groups, the resulting stabilizers lack the same charge as trehalose and saccharose.

[0167] Synthesis of 2-O-(α-D-mannopyranosyl)methyl acetate (5) The synthesis of compound 5 was carried out according to the procedure described in the following literature: Carbohydrate Research 343 (2008), 3025-3033.

[0168] Synthesis of (2S)-2-O-(α-D-mannopyranosyl)-3-methyl propanoate (11) The synthesis of compound 11 was carried out according to the procedure described in the literature Carbohydrate Research 343 (2008), 3025-3033. [ka] a) NH3, MeOH, -78℃ / rt, ≥99%

[0169] Each starting material was treated with ammonium in methanol at -78°C, then warmed to room temperature, and the desired product was obtained by ester amide decomposition in quantitative or near-quantitative yield (≥99%).

[0170] The starting materials (1, 3, 5, 7, 9, and 11) and other related compounds can be synthesized using techniques and materials known to those skilled in the art. Furthermore, the starting materials for Scheme 1 are reported in the literature and are therefore accessible as described above. Compound 1 is reported in Carbohydrate Res. 2009, 344, 1646 (β-anomeric); J. Org Chem. 2003, 68, 6672; Tetrahedron Lett. 2000, 41, 8273 (α-anomeric). The β-anomeric of Compound 3 is reported in WO2008 / 007153 A2 and WO2015 / 137838 A1. The α-anomer of compound 5 is reported in Carbohydrate Res. 2008, 343, 3025, WO2015 / 137838 A1 and WO2012 / 109283 A1. The starting materials for Scheme 2 (7, 9, and 11) are prepared in the literature and are therefore available by at least the same method described above. Compound 7 is reported in WO2015 / 137838 A1 (both configurations). The β-anomer of compound 9 is reported in WO2008 / 007153 A2. The α-anomer of compound 11 is reported in Carbohydrate Res. 2008, 343, 3025. The β-anomer of amide 2 (b-GGlyA) was disclosed in Carbohydrate Res. 2013, 374, 29, and its structure has been studied, but importantly, the specific function or effect of the compound was not disclosed. [ka] a) NH3, MeOH, -78℃ / rt, ≥99%

[0171] Each starting material, either ethyl or methyl ester, was treated with ammonium in methanol at -78°C, warmed to room temperature, and yielded the desired amide product via amide decomposition of the ester in quantitative or near-quantitative yield. Compound 14 was produced from starting material 13 in 99% yield.

[0172] The starting materials (13, 15, and 17) and other related compounds can be synthesized using techniques and materials known to those skilled in the art. Furthermore, the starting materials for Scheme 2 are reported in the literature and are therefore accessible as described above. Compound 13 (as ethyl ester) is reported in Phytochemistry 1997, 45 and Biotechnol. Lett., 1995, 17, 1169. Compound 15 (as ethyl and methyl esters) is reported in Biotechnol. Lett. 1997, 19, 583. The α-anomeric compound 17 (as ethyl ester) is reported in WO2015 / 137838 A1.

[0173] Preparation of dianionic glucuron-oxidized acids Gluco-, galact-, and mannuronic acid (compounds 37-42) are prepared as described in Scheme 3. In contrast to trehalose and saccharose, which lack charge due to the presence of two carboxylic acid moieties, the resulting stabilizers are ionizable at two positions. [ka] a)BAIB / TEMPO,CH2Cl2 / H2O b)H2,Pd / C,50 Psi,AcOEt, c)NaOH,H2O

[0174] The primary hydroxyl group at C-6 is efficiently oxidized to the corresponding carboxylic acid using a BAIB / Tempo reagent combination. The benzyl ether protecting group is then removed with Pd / C and 50 psi of H2. Hydrolysis of the methyl ester with NaOH in water yielded the sodium salt of the final product in quantitative yield. Under basic conditions, the final compound exhibited two charges derived from the two carboxylic acid functional groups. Disodium salt 37 was prepared from starting material 19 in a total yield of 70%. Disodium salt 38 was prepared from starting material 20 in a total yield of 66%.

[0175] The starting materials (19-24) and other related compounds can be synthesized using techniques and materials known to those skilled in the art. Furthermore, the starting materials for Scheme 2 are reported in the literature and are therefore accessible as described above. Compounds 19-22 are reported in WO2015 / 137838 A1. Disodium salt 37 was previously disclosed in Carbohydrate Res. 1967, 5, 453, but the specific function or effect of the compound was not disclosed.

[0176] Gluco-, galact-, and mannuronic acid (compounds 37-42) are prepared as described in Scheme 3. Due to the presence of the two carboxylic acid moieties, the resulting stabilizers are ionizable at two positions, in contrast to the uncharged trehalose and saccharose.

[0177] Preparation of diamidogluco-, manno-, and galactoside Diamides from gluco-, manno-, and galactosides 46-48 were prepared by reacting 43-45 with ammonia in methanol. [ka]

[0178] Preparation of additional dianionic glucuron-oxidized acids Gluco-, galact-, and mannuronic acid (compounds 55-60) are prepared as described in Scheme 5. [ka]

[0179] Preparation of uronic acid amides The gluco-, galact-, and mannuronates (compounds 61-66 and 67-72) were prepared as described in schemes 6 and 7. Uronamides 61-66 were prepared by reacting 31-36 with ammonia in methanol. Uronamides 67-72 were prepared by reacting 49-54 with ammonia in methanol. [ka] [ka]

[0180] Synthesis and Characterization of Compounds General procedure 1 The 1H NMR spectrum was obtained using CDCl3 at 400 MHz, and the chemical shift value (δ) from tetramethylsilane was acquired in ppm downfield. 13 ¹³C NMR spectra were acquired at 100.61 MHz using CDCl3. Medium-pressure preparative column chromatography: Silica Gel Merck 60H. Analytical TLC: Aluminum-backed silica gel Merck 60 F254. Reagents and solvents were purified and dried according to WLF Armarego, CLL Chai, Purification of Laboratory Chemicals, 5th ed.; 2003 Elsevier. Specific rotation ([α]D2O) was measured using a Perkin-Elmer D241 automatic polarimeter. All reactions, except those in water, were carried out under an inert atmosphere (argon).

[0181] Experiment 1. Synthesis of 2-O-(α-D-mannopyranosyl)acetamide (6) A solution of 5 (3.01 g, 11.9 mmol) in MeOH (15 mL) was placed in a sealed tube and saturated with NH3 at -78°C. The reaction mixture was stirred at room temperature for 3 days. After evaporating the excess NH3 and concentrating the mixture, product 6 was obtained as a white foam (quantitative yield). 1 H NMR (D2O, 400 MHz): δ4.85 (d, J = 1.8 Hz, 1H), 4.17 (d, J = 15.7 Hz, 1H), 4.05 (d, J = 15.7 Hz, 1H), 4.00 (dd, J = 3.5, 1.7 Hz, 1H), 3.84-3.81 (m, 2H), 3.69 (dd, J = 12.2, 5.7 Hz, 1H), 3.64-3.57 (m, 2H) ppm. 13 C NMR (D2O, 100.61 MHz): δ178.5, 99.9, 73.2, 70.3, 69.7, 66.6, 65.4, 60.8 ppm.

[0182] Experiment 2. Synthesis of (2S)-2-O-(α-D-mannopyranosyl)-3-propanamide (12) The procedure of Experiment 1 was applied to compound 11 (2.10 g, 7.9 mmol) to obtain compound 12 as a white foamy substance (quantitative yield). 1 H NMR (D2O, 400 MHz): δ4.94 (d, J = 1.6 Hz, 1H), 4.20 (q, J = 6.8 Hz, 1H), 3.93 (dd, J = 3.4, 1.7 Hz, 1H), 3.83 (dd, J = 9.4, 3.4 Hz, 1H), 3.76 (dd, J = 12.3, 2.4 Hz, 1H), 3.70-3.60 (m, 2H), 3.55 (m, J = 10.0, 5.3, 2.4 Hz, 1H), 1.34 (d, J = 6.8 Hz, 3H) ppm. 13 C NMR (D2O, 100.61 MHz): δ178.2, 98.7, 73.5, 72.7, 70.4, 70.1, 66.5, 60.7, 17.0 ppm.

[0183] Experiment 3.2 - Synthesis of (α / β-D-glucopyranosyl)acetamide (2) The procedure of Experiment 1 was applied to Compound 1 (0.676 g, 2.68 mmol) to obtain Compound 2 as a white foamy substance (quantitative yield). 1 H NMR (D2O, 400 MHz): δ4.91 (d, J = 3.8 Hz), 4.45 (d, J = 7.9 Hz), 4.31 (d, J = 16.0 Hz), 4.19 (d, J = 15.9 Hz), 4.03 (d, J = 15.9 Hz), 3.86-3.59 (m), 3.54 (dd, J = 9.9, 3.8 Hz), 3.47-3.28 (m) ppm. 13 C NMR (D2O, 100.61 MHz): δ174.9, 102.4, 98.6, 76.0, 75.5, 72.93, 72.79, 72.2, 71.1, 69.45, 69.38, 67.7, 65.9, 60.56, 60.40 ppm.

[0184] Experiment 4. Synthesis of (2S)-2-(α / β-D-glucopyranosyl)propanamide (8) The procedure of Experiment 1 was applied to compound 7 to obtain product 8 as a white foamy substance (quantitative yield, α / β = 11:1). 1 H NMR (D2O, 400 MHz): δ5.01 (d, J = 3.9 Hz), 4.45 (d, J = 8.0 Hz), 4.38 (q, J = 7.0 Hz), 4.18 (q, J = 6.8 Hz), 3.87-3.77 (m), 3.75-3.65 (m), 3.62-3.55 (m), 3.54-3.49 (m), 3.48-3.32 (m), 1.38 (d, J = 7.0 Hz), 1.35 (d, J = 6.8 Hz) ppm. 13 C NMR (D2O, 100.61 MHz): δ178.3, 101.6, 96.9, 76.1, 75.6, 75.2, 73.0, 72.8, 72.5, 71.6, 71.20, 71.03, 69.5, 69.3, 60.6, 60.2, 18.7, 16.9 ppm.

[0185] Experiment 5.2 - Synthesis of (α / β-D-galactopyranosyl)acetamide (4) The procedure of Experiment 1 was applied to Compound 3 to obtain Product 4 as a white foamy substance (quantitative yield, α / β = 2:1). 1 H NMR (CDCl3, 400 MHz): δ4.93 (d, J = 3.8 Hz), 4.38 (d, J = 7.6 Hz), 4.30 (d, J = 16.0 Hz), 4.20-4.15 (m), 4.02 (d, J = 15.9 Hz), 3.92-3.91 (m), 3.86 (dt, J = 10.7, 4.3 Hz), 3.79 (dd, J = 10.3, 3.8 Hz), 3.74-3.57 (m), 3.52 (dd, J = 9.9, 7.6 Hz) ppm. 13 13C NMR (CDCl 3, 100,61 MHz): δ175.0, 102.9, 98.7, 75.3, 72.5, 71.4, 70.6, 69.19, 69.12, 68.5, 68.0, 67.7, 65.9, 61.1, 60.9 ppm.

[0186] Experiment 6. Synthesis of (2S)-2-(α / β-D-galactopyranosyl)propanamide (10) The procedure of Experiment 1 was applied to compound 9 to obtain product 10 as a white foamy substance (quantitative yield, α / β = 3:1). 1 H NMR (CDCl3, 400 MHz): 5.02 (d, J = 3.9 Hz), 4.40-4.35 (m), 4.18 (q, J = 6.8 Hz), 3.92 (d, J = 3.1 Hz), 3.84 (dd, J = 10.3, 3.3 Hz), 3.76 (dt, J = 10.0, 4.8 Hz), 3.72-3.56 (m), 3.50 (dd, J = 10.0, 7.8 Hz), 1.37 (d, J = 7.0 Hz), 1.33 (d, J = 6.8 Hz) ppm.

[0187] Experiment 7. Synthesis of 3-O-(α / β-D-glucopyranosyl)-3-hydroxybutylamide (14) The procedure of Experiment 1 was applied to compound 13 (0.396 g, 1.35 mmol) to obtain product 14 as a white foamy substance (0.291 g, 82%). 1 1H NMR (CDCl3, 400 MHz): δ4.95 (d, J = 3.8 Hz), 4.47 (d, J = 7.9 Hz), 4.46 (d, J = 7.9 Hz), 4.21-4.03 (m), 3.81-3.28 (m), 3.18-3.12 (m), 2.65-2.36 (m), 1.23-1.15 (m) ppm.

[0188] Experiment 8. Synthesis of 3-O-(β-D-galactopyranosyl)-3-hydroxybutylamide (16) The procedure of Experiment 1 was applied to compound 15 (0.820 g, 2.8 mmol) to obtain product 16 as a white foamy substance (0.677 g, 90%). 1 1H NMR (CDCl3, 400 MHz): δ4.41 (d, J = 7.9 Hz), 4.40 (d, J = 7.9 Hz), 4.27-4.18 (m), 3.84-3.83 (bs), 3.73-3.55 (m), 3.42-3.38 (m), 2.53-2.38 (m), 1.24 (d, J = 6.4 Hz), 1.18 (d, J = 6.3 Hz) ppm. 13 ¹ 100.61 MHz): δ176.6, 176.5, 102.0, 101.0, 75.2, 75.0, 73.9, 72.7, 72.7, 72.5, 70.8, 70.7, 68.6, 68.5, 60.9, 60.8, 42.7, 41.9, 20.4, 18.8 ppm.

[0189] Experiment 9. Synthesis of 3-O-(α-D-mannopyranosyl)-3-hydroxybutylamide (18) The procedure of Experiment 1 was applied to compound 17 (1.15 g, 3.9 mmol) to obtain product 18 as a white foamy substance (0.711 g, 69%). 1 1H NMR (CDCl3, 400 MHz): δ4.91 (d, J = 1.4 Hz), 4.86 (d, J = 1.7 Hz), 4.19-4.08 (m), 3.83-3.76 (m), 3.73-3.60 (m), 3.58-3.51 (m), 2.45-2.33 (m), 1.22 (d), 1.16 (d) ppm. 13 ¹ 100.61 MHz): δ176.7, 99.9, 96.4, 73.0, 72.8, 72.7, 70.6, 70.5, 70.4, 70.2, 69.3, 66.8, 66.4, 60.9, 60.6, 42.9, 42.4, 20.6, 17.7 ppm.

[0190] Experiment 10.2 - Synthesis of (α / β-D-glucopyranosideurone) disodium acetate (37) To a vigorously stirred solution of 19 (1.44 g, 2.76 mmol) in 9.2 mL of DCM and 9.2 mL of H2O, TEMPO (0.178 g, 0.55 mmol) and BAIB (1.08 g, 6.91 mmol) were added. After complete conversion of the starting materials, the reaction mixture was quenched with a 10% solution of Na2S2O3 (20 mL) and subsequently extracted with RINKAN (3 × 20 mL). The combined organic layers were dried over MgSO4, filtered, and concentrated. Product 25 was obtained as a colorless, viscous foam (1.185 g, 80%) by flash column chromatography using (70:30, RINKAN / Hex). A solution of 25 in RINKAN was hydrogenated at 50 psi in the presence of 10% Pd / C (0.25 equivalents). After 5 hours, the reaction mixture was filtered, and the solvent was evaporated to obtain 31 as a very viscous, colorless foam. A solution of 1 M NaOH (2 equivalents) was added to a stirred solution of compound 31 in H2O (2 mL). After all the starting material was consumed, the pH was adjusted to 7 with 10% HCl, and the solvent was evaporated to obtain compound 37 as a viscous, colorless foam (70%, 4-step total yield).1 1H NMR (CDCl3, 400 MHz): δ4.98 (d, J = 3.9 Hz), 4.40 (d, J = 7.9 Hz), 4.26 (q, J = 7.0 Hz), 4.03 (q, J = 6.8 Hz), 3.96 (d, J = 10.1 Hz), 3.77 (t, J = 9.5 Hz), 3.73-3.69 (m), 3.65-3.63 (m), 3.58 (dd, J = 11.7, 4.3 Hz), 3.51 (dd, J = 9.8, 3.9 Hz), 3.46-3.44 (m), 3.40 (t, J = 9.6 Hz, 1H), 3.32-3.28 (m), 1.34 (d, J = 6.9 Hz), 1.29 ppm (d, J = 6.8 Hz). 13 ¹ 100,61 MHz): δ180.9, 176.8, 101.7, 96.5, 76.7, 76.2, 75.6, 74.6, 73.2, 72.8, 72.4, 72.2, 71.7, 71.2, 18.9, 17.3 ppm.

[0191] Experiment 11. Synthesis of (2S)-2-(α / β-D-glucopyranosidourone) disodium propanoate (38) The procedure of Experiment 10 was applied to compound 20 to obtain product 38 as a viscous, colorless, gum-like substance (66%, total yield in 4 steps). 1 1H NMR (CDCl3, 400 MHz): δ4.90 (d, J = 3.7 Hz), 4.43 (d, J = 7.8 Hz), 4.26 (d, J = 15.5 Hz), 4.09 (d, J = 15.5 Hz), 3.89 (dd, J = 12.9, 2.4 Hz), 3.78-3.62 (m), 3.57-3.33 (m) ppm. 13 ¹ 100,61 MHz): δ177.4, 176.6, 102.1, 98.3, 73.1, 72.9, 72.3, 71.96, 71.76, 71.46, 71.26, 69.5, 66.86, 66.78 ppm.

[0192] Stabilization research The ability of novel compounds to stabilize several proteins, including enzymes and monoclonal antibodies, was evaluated under thermal and / or pH stress using DSF and high-speed size exclusion chromatography (HPSEC) (see Figures 1–5). HPSEC can be used to evaluate the ability of compounds to stabilize biomolecules in the absence of thermal stress. The proteins used in the stabilization assays were lysozyme, adalimumab (Humira®), ubiquitin, and factor IX.

[0193] Differential scanning fluorescence (DSF) assay Protein melting temperature (T M The determination of the unfolding was performed by monitoring protein unfolding using the SYPRO orange fluoroprobe (Molecular Probe), which is completely quenched in an aqueous environment but fluoresces when bound to a hydrophobic patch of the protein. Such an increase in fluorescence can be measured as a function of temperature. The thermal shift assay was performed on an iCycle iQ5 real-time PCR detection system (Bio-Rad) equipped with a charge-coupled device (CCD) camera and a Cy3 filter with excitation and emission wavelengths of 490 nm and 575 nm, respectively. This instrument can be used for parallel thermal stability assays because it can simultaneously detect fluorescence changes in a 96-well plate (low-profile plate, Bio-Rad). The 96-well plate was sealed with optical-quality sealing tape (Bio-Rad) and centrifuged at 2500 g for 1 minute immediately before the assay to remove air bubbles. Subsequently, the plate was heated from 20°C to 90°C, increasing by 1°C with an equilibration time of 60 seconds, followed by fluorescence readings. The protein melting temperature (T) can be determined by using the fluorescence intensity as a function of temperature to determine the first derivative (d(Rfu) / dT).M Calculate the delta T under various conditions and extract the precise transition inflection point. M Value (DT M ) is obtained for each condition T M It was calculated by subtracting the TM value obtained for reference from the value.

[0194] In a typical assay with a total volume of 20 μL, a protein concentration of 0.1–0.5 mg / mL and a 5x dye concentration were used to ensure the best signal-to-noise ratio. Before performing the DSF experiment, protein stock solutions were prepared in the corresponding buffer. Stabilizer solutions and dyes were prepared according to the specific conditions of each assay. Assays were prepared by adding 1–2 μL of protein to 8–9 μL of dye buffer and 10 μL of compound solution. Controls were prepared by replacing the amount of stabilizer with the corresponding buffer.

[0195] High-speed size exclusion chromatography (HPSEC) assay HP-SEC was performed using a Waters 515 pump, Waters 2487 dual absorbance detector (Waters, USA), and Rheodyme 77251 injector (Waters, USA). A TSK Gel G3000 SWXL column (300 mm × 7.8 mm) (Tosoh Biosep, Germany) was used. To inject 50 mg of biomolecular sample, the injection volume was adjusted according to the concentration of each sample, and separation was performed at a flow rate of 1.0 mL / min, or adjusted as appropriate. For Humira® samples, an appropriate running buffer, e.g., 100 mM sodium sulfate, 100 mM dibasic sodium phosphate pH 6.8, was used. UV detection was performed at a wavelength suitable for detecting biomolecules, e.g., 280 nm. No thermal stress was applied, and the assay was performed at room temperature. Absorbance was measured to determine the concentration of biomolecules and therefore the amount of biomolecular degradation.

[0196] Stabilization of lysozyme measured by Experiment 10-DSF The stabilization of lysozyme at pH 3.6 and pH 12 in the presence of several stabilizers at a concentration of 0.25 M was investigated using DSF (Figure 1). At higher pH (12), lysozyme showed lower stability and a decrease in its melting temperature. Neutral amide-containing glycosides, namely mannosyl glycolamide (MglyA), mannosyl lactamide (MLA), 3-glucosyl butanamide (GBA), galactosyl glycolamide (GaGlyA), galactosyl lactamide (GaLA), β-glucosyl glycolamide (b-GGlyA), β-glucosyl lactamide (b-GLA), β-galactosyl glycolamide (b-GaGlyA), β-3-galactosyl butanamide (b-GaBA), and β-galactosyl lactamide (b-GaLA), showed stabilizing effects. At higher pH levels (pH 12), the stabilizing effect was greater for GBA, GaGlyA, b-GGlyA, b-GaLA, b-GaGlyA, and b-GaBA. All new compounds can stabilize lysozyme under both stress conditions.

[0197] Experiment 11 - Stabilization assay of adalimumab (Humira®) measured by HPSEC pH titration of adalimumab (Humira®) to pH 3.2 was performed both in the absence and in the presence of the stabilizer GaGlyA 0.5M, and the results were analyzed by HPSEC (Figure 2). The injection volume was adjusted according to each sample concentration to inject 50 mg of Humira, and separation was performed at a flow rate of 1.0 mL / min. The running buffer consisted of 100 mM sodium sulfate and 100 mM dibasic sodium phosphate pH 6.8. UV detection was performed at 280 nm. Unlike the DSF assay, this assay was performed at room temperature and no thermal stress was applied. Absorbance was measured to determine the concentration of biomolecules and, therefore, the amount of biomolecule degradation. Adalimumab degrades after 12 hours at pH 3.2 (Figure 2, line C), however, this degradation is significantly reduced in the presence of GaGlyA (Figure 2, line B).

[0198] Experiment 12 - Adalimumab Stabilization Assay Measured by DSF The stabilization of adalimumab was also evaluated using DSF in the presence of several stabilizers at pH 12 and 0.25 M (Figure 3). All compounds tested stabilized adalimumab, and higher stability was obtained with β-3-galactosyl-butanamide (b-GaBA), accompanied by a 22°C increase in the melting temperature of the monoclonal antibody.

[0199] Experiment 13 - Ubiquitin Stabilization Assay Measured by DSF The stabilization of ubiquitin using the neutral amide-containing glycosides and charged glycosides of the present invention was measured using DSF. Ubiquitin was equally stable in the presence of several novel compounds at 0.25 M, pH 12 (Figure 4). The charged glycosides, namely β-glucosyl-glycolate (b-Ggly), β-glucosyl-lactic acid (b-GL), β-3-glucosyl-butyric acid (b-GB), β-galactosyl-glycolate (b-GaGly), and the neutral amide-containing glycoside β-galactosyl-lactic acid (b-GaL), resulted in significantly better stability of ubiquitin at pH 12. For charged glycosides, the best result was only a 2°C increase in melting temperature, but neutral amide-containing glycosides, such as 3-glucosylbutanamide (GBA), were able to increase the melting temperature of ubiquitin by as much as 9°C (Figure 4). Neutral amide-containing glycosides were better stabilizers than carboxylic acid-containing glycosides, as shown in Figure 4.

[0200] Experiment 14 - Stabilization assay of factor IX measured by DSF The stabilization of factor IX using the neutral amide-containing glycosides of the present invention was measured using DSF. As shown in Figure 5, the novel stabilizers increased the melting temperature of factor IX at a concentration of 0.25 M in water and a pH of 7-8. Mannosyl glycolamide (MglyA), glucosyl glycolamide (GGlyA), 3-glucosyl butanamide (GBA), and galactosyl lactamide (GaLA) showed stabilization of factor IX.

[0201] Experiment 1: 15-hydroxymethyl derivative Hydroxymethylamide derivatives are prepared by glycosylation of a glycosyl donor (mann-, gluco-, galactopyranose) and its corresponding glycosyl acceptor, using a method described in the literature (Carbohydrate Research 343 (2008), 3025-3033), followed by further reaction of the unprotected sugar with ammonia in methanol.

[0202] Hydroxymethylurone derivatives are prepared by glycosylation of a glycosyl donor (mann-, gluco-, galactopyranose) and its corresponding glycosyl acceptor, using the method described in the literature (Carbohydrate Research 343 (2008), 3025-3033), followed by further oxidation of the C-6 primary hydroxyl group to the corresponding carboxylic acid using a BAIB / Tempo reagent combination.

[0203] Hydroxymethyldiamide derivatives are prepared from their respective hydroxymethylurone derivatives (man-, gluc-, galacturone) by further reaction of unprotected sugars with ammonia in methanol.

[0204] Consideration There is a great need for novel stabilizers for biomolecules. The assays described above demonstrate that the compounds of the present invention stabilize biomolecules under various conditions. Results from DFS studies (Experiments 10, 12, 13, and 14) show that neutral glycosylated amides stabilize a wide range of biomolecules under both pH and thermal stress. The increase in Tm value corresponds to improved structural stability of the biomolecules. Results from the HPSEC study (Experiment 11) show that neutral glycosylated amides also stabilize biomolecules under pH stress in the absence of thermal stress. The absorbance (mV) values ​​indicate that the compounds of the present invention have the ability to protect biomolecules from degradation caused by pH. Neutral glycosylated amides provide unexpectedly improved protection from pH stress compared to conventional stabilizers. Referring to the comparative data of Experiment 13 and Figure 4, this shows that neutral glycosylated amides stabilize given biomolecules more effectively than charged glycosides, as evidenced by the increased melting temperature of the biomolecules.

[0205] In summary, novel molecules that stabilize biomolecules under both thermal and pH stress have been identified. These molecules were shown to stabilize biomolecules from pH-induced stress even in the absence of thermal stress. Compared to conventional charged glycosides, these compounds were shown to be superior stabilizers of biomolecules under pH stress. The following are the embodiments of the claims originally filed for this application. [1] A compound or salt thereof having the following structure, [ka] (In the formula, X is a hexosyl group selected from the group consisting of glucosyl, mannosyl, galactosyl, allosyl, altrosyl, grosyl, idosyl and talosyl; a uronic acid group selected from the group consisting of glucuronic acid, mannuronic acid, galacturonic acid, aluronic acid, altoluronic acid, guluronic acid, iduronic acid and tarronic acid; or a uronic acid amide group selected from the group consisting of glucuronamide, mannuronamide, galacturonamide, aluronamide, altoluronamide, guluronamide, iduronamide and tarronamide. R 1 and R 2 Each of these is independently H, halogen, OH, O-alkyl, CONH 2 CO 2 H, CO 2 - Alkyl or optionally substituted alkyl, Each of Y and Z is independently H, OH, O-alkyl, or optionally substituted alkyl. m is 0, 1, or 2. A is NR 3 R4 OR 5 And, R 3 and R 4 Each of these is independently H, OH, O-alkyl or optionally substituted alkyl. R 5 These are independently H or optionally substituted alkyl groups. Here, If X is glucosyl, R 1 This is an alkyl group which may be substituted in some cases. R 1 and R 2 If each of them is H and m is 0, then X is something other than glucuronic acid. X is glucosyl or mannosyl, m is 0, R 1 or R 2 One of the atoms is -alkyl-OH and the other is -H, and A is -NH 2 In this case, the compound is a β-anomeric compound. A compound or a salt thereof. [2] The following structure

change

change

change

change

[10] The following structure

change

[11] The compound according to any one of [2] to

[10] , wherein the hexosyl group is an α-anomeric group.

[12] The compound according to any one of [2] to

[10] , wherein the hexosyl group is a β-anomeric group.

[13] The following structure

change

[14] The following structure

change

change

[15] The following structure

change

change

change

[16] The following structure

change

[17] The compound according to

[16] , wherein X is glucuronic acid, mannuronic acid, or galacturonic acid.

[18] The compound described in

[16] , wherein each of Y and Z is H.

[19] The following structure

change

[16] , having the following characteristics.

[20] The following structure

change

[16] , having the following characteristics.

[21] The following structure

change

[16] , having the following characteristics.

[22] The compound according to any one of

[16] to

[21] , wherein the uronic acid group is an α-anomeric group.

[23] The compound according to any one of

[16] to

[21] , wherein the uronic acid group is a β-anomeric group.

[24] The following structure

change

change

[16] , having the following characteristics.

[25] The following structure

change

change

[16] , having the following characteristics.

[26] The following structure

change

change

change

[16] , having the following characteristics.

[27] The following structure

change

change

[16] , having the following characteristics.

[28] The following structure

change

change

change

[16] , having the following characteristics.

[29] The compound according to any one of

[16] to

[28] , wherein the compound is a sodium salt, a potassium salt, a calcium salt, or a magnesium salt.

[30] The following structure

change

[31] The following structure

change

[30] , having the following characteristics.

[32] The following structure

change

[30] , having the following characteristics.

[33] The following structure

change

[30] , having the following characteristics.

[34] The compound according to any one of

[30] to

[33] , wherein the uronic acid amide group is an α-anomeric group.

[35] The compound according to any one of

[30] to

[33] , wherein the uronic acid amide group is a β-anomeric group.

[36] The following structure

change

[30] , having the following characteristics.

[37] The following structure

change

change

[30] , having the following characteristics.

[38] The following structure

change

change

change

[30] , having the following characteristics.

[39] A composition comprising a biomolecule and at least one compound described in any one of [1] to

[38] .

[40] The composition according to

[39] , wherein the biomolecule is a biopharmaceutical, protein, nucleotide, polypeptide, or antibody.

[41] The biomolecules include insulin, humulin, novolin, human inhaled insulin, exvera, insulin aspart, novologue (aspart), insulin glulisine, apidra (glucisine), insulin lispro, humalog (lispro), isophane insulin, NPH, insulin detemir, levemir (detemir), insulin glargine, lantus (glargine), sustained-release insulin zinc, lente, ultralente, pramulintide acetate, simulin, Growth hormone (GH), somatotropin, genotropin, humatrope, norditropin, norivitropin, neutropin, omnitrope, protropin, siazen, celostim, bartropin, mecasermin, increlex, mecasermin lymphabate, iplex, factor VIII, biocrate, helixate, cogenerate, recominate cominate), ReFacto, Factor IX, Benefix, Antithromin III (AT-III), Trombate III, Protein C concentrate, Seprotin, β-glucocerebrosidase, Cerezyme, β-glucocerebrosidase, Ceredase (purified from pooled human placenta), Alglucosidase-α, Myozyme, Laronidase (α-l-iduronidase), Aldurazyme, Idurusulfate (iduronic acid-2-sulfate) (Agalactosidase), elapase, galusulphase, naglazyme, agalsidase-β (human α-galactosidase A), fabrazyme, α-1-proteinase inhibitor, aralast, prolastin, lactase, lactide, pancreatic enzymes (lipase, amylase, protease), arco-lase, cotazym, creon, donazyme, pancrease, biocase, zymase, adenosine deaminase (pegademase) Bovine, PEG-ADA), Adagen, Pooled Immunoglobulin, Octagum, Human Albumin, Albumarc, Albumin, Albuminer, AlbuRx, Albutein, Flexbumin, Buminate, Plasbumin, Erythropoietin, Epoetin-α, Epogen, Procrit, Darbepoetin-α,Aranesp, Filgrastim (granulocyte colony-stimulating factor, G-CSF), Neupogen, Pegfilgrastim (Peg-G-CSF), Neulasta, Salgramostim (granulocyte-macrophage colony-stimulating factor, GM-CSF), Leukin, Oprelbekin (interleukin-11, IL-11), Neumega, human follicle-stimulating hormone (FSH), Gonal-F, Follistim, human chorionic gonadotropin (HCG), Ovidrel, Luveris, type I α-interferon, Interferon Alphacon 1, Consensus Interferon, Infergen, Interferon-α2a (IFNα2a), Loferon-A, PegInterferon-α2a, Pegasys, Interferon-α2b (IFNα2b), Intron A, PegInterferon-α2b, PegIntron, Interferon-αn3 (IFNαn3), Alferon N, Interferon-β1a (rIFN-β), Avonex, Rebif, Interferon Lon-β1b (rIFN-β), Beta-Seron, Interferon-γ1b (IFNγ), Actimune, Aldesleukin (Interleukin-2 (IL2)), Epidermal Thymocyte Activating Factor (ETAF), Proleukin, Alteplase (Tissue Plasminogen Activating Factor (tPA)), Activase, Leteplase (tPA deletion mutant protein), Letabase, Tenecteplase, TNKase, Urokinase, Avokinase, Factor VIIa, Novoseven ( NovoSeven), Drotrecogin-α (Activated Protein C), Xigris, Salmon Calcitonin, Fortical, Miacalcin, Teriparatide (Human Parathyroid Hormone Residues 1-34), Forteo, Exenatide, Vietta, Octreotide, Sandostatin, Dibotermin-α (Recombinant Human Osteogenesis Imperative 2, rhBMP2), Infuse, Recombinant Human Osteogenesis Imperative 7 (rhBMP7), Osteogenic Protein 1, Histrelin Acetate (Gonadotropin-Releasing Hormone, GnRH), Suprelin LA, Vantas, Palifermin (Keratinocyte Growth Factor KGF), Kebivans, Becaprelmin (Platelet-Derived Growth Factor, PDGF), Regranex, Trypsin, Granulex,Nesiritide, Natrecor, Botulinum toxin type A, Botox, Botulinum toxin type B, Myoblock, Collagenase, Santil, Starfish oxyribonuclease I, Dornase α, Pulmozyme, Hyaluronidase (bovine, sheep), Anfadase (bovine), Hydase (bovine), Vitrase (sheep), Hyaluronidase (recombinant human), Hylenex, Papain, Accuzyme, Panafil, L-asparaginase, ELSPAR, PEG-asparaginase, Oncaspar, Rasbrica Elitek, Repildin, Lefuldan, Vivalirudin, Angiomax, Streptokinase, Streptase, Anistreplase (Anisoylated Plasminogen Streptokinase Activated Complex, APSAC), Eminase, Bevacizumab, Avastin, Cetuximab, Erbitux, Panitumumab, Vectibix, Alemtuzumab, Campas, Rituximab, Rituxan, Trastuzumab, Herceptin, Abatacept, Orencia, Anakinra, Antril, Kineret, Avalimumab Humira, etanercept, Enbrel, infliximab, Remicade, Alefacept, Amevive, natalizumab, Tysabri, eculizumab, Soliris, antithymocyte globulin (rabbit), thymoglobulin, basiliximab, Simulect, daclizumab, Zenapax, muromonab-CD3, orthoclone, OKT3, omalizumab, Xolair, palivizumab, Synagis, Enfvirtide, Fuzeon, Absix Mab, ReoPro, Pegvisomant, Somabart, Crotalidae Multivalent Immunotherapy Fab (Sheep), Crofab, Digoxin Immunoserum Fab (Sheep), Digifab, Ranibizumab, Lucentis, Denileukin, Diftitox, Ontac, Ibritumomab, Tiucetan, Zevalin, Gemtuzumab, Ozogamicin, Mylotarg, Tositumomab and I-Tositumomab, Vexar, Vexar I-131, Hepatitis B Surface Antigen (HBsAg),Engerix, Recombivax HB, HPV vaccine, Gardasil, OspA, Lymerix, anti-rhesus (Rh) immunoglobulin G, Rophyllac, recombinant purified protein derivative (DPPD), glucagon, glucagen, growth hormone-releasing hormone (GHRH), Geref, secretin, tyrotropin (human peptide), secreteFlo (porcine peptide), thyroid-stimulating hormone (TSH), thyrotropin, capromab penetide, Pros The composition described in

[39] is ProstaScint, indium-111-octreotide, OctreoScan, satumomab pendetide, OncoScint, arcitumomab, CEA-Scan, nofetumomab, Verluma, aptide, Acutect, pentimate imusilomab, myosinto, technetium phanoresomab, NeutroSpec, HIV antigen, enzyme immunoassay, OraQuick, Unigold, hepatitis C antigen, or recombinant immunoblot assay (RI BA).

[42] The composition according to

[39] , wherein the biomolecule is lysozyme, adalimumab (Humira®), ubiquitin, or factor IX.

[43] The composition according to any one of

[39] to

[42] , wherein the composition is freeze-dried, freeze-dried, a solution, a liquid, a solid, or a suspension.

[44] A method for stabilizing a biomolecule, comprising treating the biomolecule with an effective amount of any one of the compounds described in [1] to

[38] , thereby stabilizing the biomolecule.

[45] The method according to

[44] , wherein the biomolecule is a protein, nucleotide, polypeptide, or antibody.

[0206] References 1. Chang BS, Yeung B. (2010) Physical Stability Of Protein Pharmaceuticals in Formulation And Process Development Strategies For Manufacturing Biopharmaceuticals (Feroz Jameel and Susan Hershenson eds). John Wiley & Sons Inc. pp. 69-104 2. Ueda T, Nagata M, Imoto T. (2001) Aggregation and chemical reaction in hen lysozyme caused by heating at pH 6 are depressed by osmolytes, sucrose and trehalose. 491-496 3. Kaushik JK, Bhat R. (2003) Why is trehalose an exceptional protein stabilizer? An analysis of the thermal stability of proteins in the presence of the compatible osmolyte trehalose. J Biol Chem 278(29):26458-26465 4. Singer MA, Lindquist S. (1998) Multiple effects of trehalose on protein folding in vitro and in vivo. Mol. Cell 1(5):639-648 5. Lin TY, Timasheff SN. (1996) On the role of surface tension in the stabilization of globular proteins. Protein Sci 5(2):372-381 6. Reed RH, Borowitzka LJ, Mackay MA, Chudek JA, Foster R, Warr SCR, Moore DJ, Stewart WDP. (1986) Organic solute accumulation in osmotically stressed cyanobacteria. FEMS Microbiol Rev 39:51-56. 7. Ohtake S, Wang YJ. (2011) Trehalose: Current Use and Future Applications. J. Pharm. Sciences, 100, 2020-2053 8. Lee JC, Timasheff SN. (1981) The stabilization of proteins by sucrose. J Biol Chem. Jul 25;256(14):7193-201. 9. Jain NK, Roy I. (2009) Effect of trehalose on protein structure. Protein Sci. 18, 24 10. Andya JD, Hsu CC, Shire SJ. (2003) Mechanisms of Aggregate Formation and Carbohydrate Excipient Stabilization of Lyophilized Humanized Monoclonal Antibody Formulations. AAPS PharmSci. 5 (2), article 10. 11. Khan SH, Ahmad N, Ahmad F, Kumar R. (2010) Naturally occurring organic osmolytes: From cell physiology to disease prevention. IUBMB Life, 62, 891-895 12. Rajan RS, Tsumoto K, Tokunaga M, Tokunaga H, Kita Y, Arakawa T. (2011) Chemical and pharmacological chaperones: application for recombinant protein production and protein folding diseases, Curr. Med. Chem., 18, 1-15 13. Stidham SE, Chin SL, Dane EL, Grinstaff MW. (2014) Carboxylated glucuronic poly-amido-saccharides as protein stabilizing agents, J. Am. Chem. Soc., 136, 9544-9547 14. Trombotto S, Danel M, Fitremann J, Bouchu A, Queneau Y. (2003) Straightforward Route for Anchoring a Glucosyl Moiety onto Nucleophilic Species: Reaction of Amines and Alcohols with Carboxymethyl 3,4,6-Tri-O-acetyl-r-D-glucopyranoside 2-O-Lactone. J. Org. Chem. 2003, 68, 6672 15. Trombotto S, Bouchu A, Descotes G, Queneau Y. (2000) Hydrogen peroxide oxidation of palatinose and trehalulose: direct preparation of carboxymethyl a-D-glucopyranoside. Tetrahedron Lett. 2000, 41, 8273-8277 16. Fischer L, Bromann R, Wagner F. (1995) Enantioselective Synthesis of Several 1-O-beta-D-glucoconjugates Using Almond beta-Glucosidase (E.C.3.2.1.21). Biotech. Lett. Vol. 17, No. 11, Nov. 1995, pp. 1169-1174 17. Matsumura S, Yamazaki H, Toshima K. (1997) R-Enantioselective Galactosylation Of Secondary Alcohols Using beta-Galactosidase. Biotech. Lett., Vol. 19, No. 6, June 1997, pp. 583-586 18. Krajewski D, Duque C, Schreier P. (1997) Aliphatic Beta-D-Glucosides From Fruits Of Carica Pubsecens. Phytochemistry, Vol. 45, No. 8, pp. 1627-1630 19. Faria TQ, Mingote A, Siopa F, Ventura R, Maycock C, Santos H (2008) Design of new enzyme stabilizers inspired by glycosides of hyperthermophilic microorganisms. Carbohydrate Res. 2008, 343, 3025-3033. 20. Xue JL, Cecioni S, He L, Vidal S, Praly JP. (2009) Variations on the SnCl4 and CF3CO2Ag-promoted glycosidation of sugar acetates: a direct, versatile and apparently simple method with either alpha or beta stereocontrol. Carbohydrate Res. 2009, 344, 1646 21. Moynihan HA, Hayes JA, Eccles KS, Coles SJ, Lawrence SE. (2013) Hydrogen bonding in crystal forms of primary amide functionalized glucose and cellobiose. Carbohydrate Res. 2013, 374, 29 22. PCT International Application Publication No. WO 2008 / 007153 A2 23. PCT International Application Publication No. WO 2012 / 109263 A1 24. PCT International Application Publication No. WO 2015 / 137838 A

Claims

1. The structure below 【Chemistry 1】 (In the formula, R １ is H, optionally substituted alkyl, or CONH ２ The alkyl group is C １ -C ４ Alkyl or C １ -C ４ (It is a hydroxyalkyl group.) A compound having the following properties.

2. The structure is, 【Chemistry 2】 (wherein, R １ is CH ３ , CH ２ OH or CONH ２ and m is 0, 1 or 2) The compound according to claim 1.

3. The structure below 【Transformation 3】 (In the formula, R １ is H, optionally substituted alkyl, or CONH ２ The alkyl group is C １ -C ４ Alkyl or C １ -C ４ (It is a hydroxyalkyl group.) A compound having the following properties.

4. The structure is 【Chemistry 4】 (In the formula, R １ CH ３ ,CH ２ OH or CONH ２ (where m is 0, 1, or 2) The compound according to claim 3.

5. The structure below 【Transformation 5】 (In the formula, R １ is H, optionally substituted alkyl, or CONH ２ The alkyl group is C １ -C ４ Alkyl or C １ -C ４ (It is a hydroxyalkyl group.) A compound having the following properties.

6. The structure is 【Transformation 6】 (In the formula, R １ CH ３ ,CH ２ OH or CONH ２ (where m is 0, 1, or 2) The compound according to claim 5.

7. The compound according to any one of claims 1 to 6, wherein the uronic acid amide group is an α-anomeric group.

8. The compound according to any one of claims 1 to 6, wherein the uronic acid amide group is a β-anomeric group.

9. The structure below 【Transformation 7】 (In the formula, R １ is H, optionally substituted alkyl, or CONH ２ The alkyl substituted in the above case is C １ -C ４ Alkyl or C １ -C ４ (It is a hydroxyalkyl group.) A compound having the following properties.

10. The structure below 【Chemistry 8-1】 【Chemistry 8-2】 A compound having the following properties.

11. The structure below 【Chemistry 9-1】 【Chemistry 9-2】 【Chemistry 9-3】 A compound having the following properties.

12. The structure below 【Chemistry 10】 (In the formula, R １ This may be a substituted alkyl or CO ２ H is H, and the alkyl substituted in the above case is C １ -C ４ Alkyl or C １ -C ４ It is a hydroxyalkyl, R ５ (where is H and m is 0 or 1) A compound or salt thereof having the properties of a compound.

13. The structure below 【Chemistry 11】 (In the formula, R １ H, CH ３ ,CH ２ OH or CO ２ H and R ５ (where R is H and m is 1 or 2, or where R 1 is H, CH 3 or CH 2 OH, R 5 is H and m is 0) A compound or salt thereof having the properties of a compound.

14. The structure below 【Chemistry 12】 (In the formula, R １ is H, optionally a substituted alkyl, or CO ２ H is H, and the alkyl substituted in the above case is C １ -C ４ Alkyl or C １ -C ４ It is a hydroxyalkyl, R ５ (where is H and m is 0, 1, or 2) A compound or salt thereof having the properties of a compound.

15. The structure below 【Chemistry 13】 (In the formula, R １ H, CH ３ ,CH ２ OH or CO ２ H and R ５ (where is H and m is 0, 1, or 2) A compound or salt thereof having the properties of a compound.

16. The structure below 【Chemistry 14】 (In the formula, R １ is H, optionally a substituted alkyl, or CO ２ H is H, and the alkyl substituted in the above case is C １ -C ４ Alkyl or C １ -C ４ It is a hydroxyalkyl, R ５ (where is H and m is 0, 1, or 2) A compound or salt thereof having the properties of a compound.

17. The structure below 【Chemistry 15】 (In the formula, R １ CH ３ ,CH ２ OH or CO ２ H and R ５ (where is H and m is 0 or 1) A compound or salt thereof having the properties of a compound.

18. The compound according to any one of claims 12 to 17, wherein the uronic acid group is an α-anomeric group.

19. The compound according to any one of claims 12 to 17, wherein the uronic acid group is a β-anomeric group.

20. The structure below 【Chemistry 16-1】 (In the formula, R １ This may be a substituted alkyl or CO ２ H is H, and the alkyl substituted in the above case is C １ -C ４ Alkyl or C １ -C ４ (It is a hydroxyalkyl group), or 【Chemistry 16-2】 (In the formula, R1 is optionally a substituted alkyl, and the optionally substituted alkyl is a C1-C4 alkyl or a C1-C4 hydroxyalkyl.) A compound or salt thereof having the properties of a compound.

21. The structure below 【Chemistry 17】 A compound or salt thereof having the properties of a compound.

22. The structure below 【Chemistry 18-1】 【Chemistry 18-2】 【Chemistry 18-3】 A compound or salt thereof having the properties of a compound.

23. The compound according to any one of claims 12 to 22, wherein the compound is a sodium salt, a potassium salt, a calcium salt, or a magnesium salt.

24. The structure below 【Chemistry 19-1】 【Chemistry 19-2】 A compound having the following properties.

25. The structure below 【Chemistry 20-1】 【Chemistry 20-2】 【Chemistry 20-3】 A compound having the following properties.

26. A composition comprising a biomolecule and at least one compound according to any one of claims 1 to 25, wherein the biomolecule is insulin, human inhaled insulin, insulin aspart, insulin glulisine, insulin lispro, isophan insulin, insulin detemir, insulin glargine, sustained-release insulin zinc, plum lynchide acetate, growth hormone (GH), somatotropin, genotropin, humatrope, norditropin, mecasermin, mecasermin lymphabate, factor VIII, factor IX, antithromin III (AT -III), Protein C concentrate, β-glucocerebrosidase, β-glucocerebrosidase, alglucosidase-α, laronidase (α-l-iduronidase), idulsulphaze (iduronic acid-2-sulfatase), galsulphaze, agalsidase-β (human α-galactosidase A), α-1-proteinase inhibitor, lactase, pancreatic enzymes (lipase, amylase, protease), pooled immunoglobulins, human albumin, albumin, buminate, erythropoietin, epoetin-α, darbepoetin-α, filgrastim (Granulocyte colony-stimulating factor, G-CSF), Pegfilgrastim (Peg-G-CSF), Salgramostim (Granulocyte-macrophage colony-stimulating factor, GM-CSF), Oprelbequin (Interleukin 11, IL-11), Human follicle-stimulating hormone (FSH), Human chorionic gonadotropin (HCG), Type I α-interferon, Interferon alphacon 1, Consensus interferon, Infergen, Interferon-α2a (IFNα2a), Peginterferon-α2a, Interferon-α2b (IFNα2b), Pegin Interferon-α2b, interferon-αn3 (IFNαn3), interferon-β1a (rIFN-β), interferon-β1b (rIFN-β), interferon-γ1b (IFNγ), aldesleukin (interleukin-2 (IL2)), epidermal thymocyte activator, ETAF), alteplase (tissue plasminogen activator, tPA), reteplase (tPA deletion mutant protein), tenecteplase, urokinase, avokinase, factor VIIa, drolecogin-α (activated protein C), salmon calcitonin,Teriparatide (human parathyroid hormone residues 1-34), exenatide, octreotide, sandostatin, dibotermin-α (recombinant human bone morphogenetic factor 2, rhBMP2), recombinant human bone morphogenetic factor 7 (rhBMP7), osteogenic protein 1, histrelin acetate (gonadotropin-releasing hormone, GnRH), parifermin (keratinocyte growth factor KGF), Kebivans, becaprelmin (platelet-derived growth factor, PDGF), trypsin, nesilitide, botulinum toxin type A, Botox, botulinum toxin type B, collagenase, star deoxyribonuclease I, dorunase α, hyaluronidase (bovine or sheep), anfadase (bovine), hidase (bovine), papain, Accuzyme, L-as Paraginase, pegu-asparaginase, rasburicase, repirudin, bivalirudin, streptokinase, streptase, anistreplase (anisoylated plasminogen streptokinase activating complex, APSAC), bevacizumab, cetuximab, panitumumab, alemtuzumab, rituximab, trastuzumab, abatacept, anakinra, avalimumab, etanercept, infliximab, remicade, amevive, natalizumab, eculizumab, anti Thymocyte globulin (rabbit), thymoglobulin, basiliximab, daclizumab, muromonab-CD3, OKT3, omalizumab, palivizumab, enfuvirtide, absiximab, pegvisomant, Crotalidae multivalent immunofab (sheep), digoxin immunoserum fab (sheep), ranibizumab, denileukin, Diftitox, ibritumomab, gemtuzumab, ozogamicin, tositumomab and I-tositumomab, hepatitis B surface antigen (HBsA g), HPV vaccine, OspA, anti-rhesus (Rh) immunoglobulin G, recombinant purified protein derivative (DPPD), glucagon, growth hormone-releasing hormone (GHRH), secretin, thyroid-stimulating hormone (TSH), thyrotropin, capromab pendetide, indium-111-octreotide, satumomab pendetide, arcitumomab, nofetumomab, apsitide, pentiminate imusilomab, technetium phanoresomab, HIV antigen, enzyme immunoassay, hepatitis C antigen,Alternatively, a composition that is a recombinant immunoblot assay (RIBA).

27. The composition according to claim 26, wherein the biomolecule is lysozyme, adalimumab, ubiquitin, or factor IX.

28. The composition according to claim 26 or 27, wherein the composition is freeze-dried, freeze-dried, a solution, a liquid, a solid, or a suspension.

29. A method for stabilizing a biomolecule, comprising treating the biomolecule with an effective amount of the compound described in any one of claims 1 to 25 or the composition described in any one of claims 26 to 28, thereby stabilizing the biomolecule. The aforementioned biomolecules include insulin, human inhaled insulin, insulin aspart, insulin glulisin, insulin lispro, isophan insulin, insulin detemir, insulin glargine, sustained-release insulin zinc, plum lynate acetate, growth hormone (GH), somatotropin, genotropin, humatrope, norditropin, mecasermin, mecasermin lynfabe, factor VIII, factor IX, antithromin III (AT-III), protein C concentrate, β-glucocerebrosidase, β-glucocerebrosidase, and algebra. Glucosidase-α, laronidase (α-l-iduronidase), idulosulfase (iduronic acid-2-sulfatase), galsulfase, agalsidase-β (human α-galactosidase A), α-1-proteinase inhibitors, lactase, pancreatic enzymes (lipase, amylase, protease), pooled immunoglobulins, human albumin, albumin, buminate, erythropoietin, epoetin-α, darbepoetin-α, filgrastim (granulocyte colony-stimulating factor, G-CSF), pegfilgrastim (PEG-G-CSF), salgramo Stim (granulocyte-macrophage colony-stimulating factor, GM-CSF), Oprelbekin (interleukin 11, IL-11), human follicle-stimulating hormone (FSH), human chorionic gonadotropin (HCG), type I α-interferon, interferon alphacon 1, consensus interferon, infergen, interferon-α2a (IFNα2a), pegylated interferon-α2a, interferon-α2b (IFNα2b), pegylated interferon-α2b, interferon-αn3 (IFNαn3), interferon-β1a (rIF N-β), interferon-β1b (rIFN-β), interferon-γ1b (IFNγ), aldethleukin (interleukin-2 (IL2)), epidermal thymocyte activator, ETAF), alteplase (tissue plasminogen activator, tPA), reteplase (tPA deletion mutant protein), tenecteplase, urokinase, avokinase, factor VIIa, drolecogin-α (activated protein C), salmon calcitonin, teriparatide (human parathyroid hormone residues 1-34), exenatide, octreotide, sandostatin,Dibotermin-α (recombinant human bone morphogenetic factor 2, rhBMP2), recombinant human bone morphogenetic factor 7 (rhBMP7), osteogenic protein 1, histrelin acetate (gonadotropin-releasing hormone, GnRH), parifermin (keratinocyte growth factor KGF), Kebivans, becaprelmin (platelet-derived growth factor, PDGF), trypsin, nesilitide, botulinum toxin type A, Botox, botulinum toxin type B, collagenase, starfish oxyribonuclease I, dorunase α, hyaluronidase (bovine or sheep), anfadase (bovine), hidase (bovine), papain, Accuzyme, L-asparagina -ase, pegu-asparaginase, rasburicase, repirudin, bivalirudin, streptokinase, streptase, anistreplase (anisoylated plasminogen streptokinase activating complex, APSAC), bevacizumab, cetuximab, panitumumab, alemtuzumab, rituximab, trastuzumab, abatacept, anakinra, avalimumab, etanercept, infliximab, remicade, amevive, natalizumab, eculizumab, antithymocyte globulin (Rabbit), thymoglobulin, basiliximab, daclizumab, muromonab-CD3, OKT3, omalizumab, palivizumab, enfvirtide, absiximab, pegvisomant, Crotalidae multivalent immune Fab (sheep), digoxin immune serum Fab (sheep), ranibizumab, denileukin, Diftitox, ibritumomab, gemtuzumab, ozogamicin, tositumomab and I-tositumomab, hepatitis B surface antigen (HBsAg), HPV vaccine, Osp A method comprising: A. anti-rhesus (Rh) immunoglobulin G, recombinant purified protein derivative (DPPD), glucagon, growth hormone-releasing hormone (GHRH), secretin, thyroid-stimulating hormone (TSH), thyrotropin, capromab pendetide, indium-111-octreotide, satumomab pendetide, arcitumomab, nofetumomab, apsitide, pentimate imusilomab, technetium phanoresomab, HIV antigen, enzyme immunoassay, hepatitis C antigen, or recombinant immunoblot assay (RIBA).

30. The method according to claim 29, wherein the biomolecule is lysozyme, adalimumab, ubiquitin, or factor IX.

31. The method according to claim 29, wherein the composition is freeze-dried, freeze-dried, a solution, a liquid, a solid, or a suspension.

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