阅读说明：本技术 具有抗炎活性的硫酸化寡糖 (Sulfated oligosaccharides with anti-inflammatory activity ) 是由 凯特琳·阿诺尔德 徐定 徐咏梅 拉法尔·波林斯基 刘健 于 2018-11-05 设计创作，主要内容包括：本文提供了小分子化合物,包括非抗凝硫酸乙酰肝素寡糖分子,具有抗炎性质并能够以足以影响HMGB1蛋白与晚期糖基化终末产物(RAGE)的受体之间的相互作用的方式与高迁移率族蛋白1(HMGB1)蛋白相互作用。本文还提供了治疗受试者的扑热息痛(APAP)过量的方法。(Provided herein are small molecule compounds, including non-anticoagulant heparan sulfate oligosaccharide molecules, having anti-inflammatory properties and capable of interacting with high mobility group box 1(HMGB1) protein in a manner sufficient to affect the interaction between the HMGB1 protein and the receptor for advanced glycation end product (RAGE). Also provided herein are methods of treating an excess of paracetamol (APAP) in a subject.)

1. A small molecule compound having anti-inflammatory properties comprising a non-anticoagulant heparan sulfate oligosaccharide molecule, optionally wherein the non-anticoagulant heparan sulfate oligosaccharide molecule is configured to interact with a high mobility group box 1(HMGB1) protein in a manner sufficient to affect the interaction between the HMGB1 protein and the receptor for advanced glycation end product (RAGE).

2. A small molecule compound having anti-inflammatory properties, said small molecule compound comprising a non-anticoagulant heparan sulfate oligosaccharide molecule comprising one of the following structural formulae:

wherein R is³Is H or a detectable label.

3. The small molecule compound according to any of the preceding claims, wherein the oligosaccharide molecule comprises from about 10 to about 20 saccharide units, or from about 12 to about 18 saccharide units.

4. The small molecule compound according to any of the preceding claims, wherein the oligosaccharide molecule comprises about 18 saccharide units.

5. The small molecule compound according to any of the preceding claims, wherein the small molecule compound is provided in the form of a composition further comprising a small molecule compound comprising a disaccharide building block as shown below:wherein R is-H or-SO₃H。

6. The small molecule compound according to claim 6, wherein the small molecule compound comprising a disaccharide building block is selected from the group consisting of non-anticoagulant heparin, non-anticoagulant low molecular weight heparin and O-desulphated heparin (ODSH).

7. The small molecule compound according to any of the preceding claims, wherein the oligosaccharide molecule protects against liver damage in vivo.

8. The small molecule compound according to any one of the preceding claims, wherein the oligosaccharide molecule reduces neutrophil infiltration in vivo.

9. The small molecule compound of any of the preceding claims, wherein the oligosaccharide molecule reduces inflammation in vivo.

10. A method of treating a subject, the method comprising:

providing a subject to be treated, wherein the subject has inflammation; and

administering to the subject a non-anticoagulant heparan sulfate oligosaccharide molecule having anti-inflammatory properties, optionally wherein the non-anticoagulant heparan sulfate oligosaccharide molecule is configured to interact with a high mobility group box 1(HMGB1) protein in a manner sufficient to affect the interaction between the HMGB1 protein and a receptor for advanced glycation end products (RAGE).

11. A method of treating a subject, the method comprising:

providing a subject to be treated, wherein the subject has inflammation; and

administering to the subject a small molecule compound having anti-inflammatory properties, the small molecule compound comprising a non-anticoagulant heparan sulfate oligosaccharide molecule comprising one of the following structural formulae:

wherein R is³Is H or a detectable label.

12. The method of any one of the preceding claims, wherein the subject has any injury that results in increased inflammation.

13. The method of any one of the preceding claims, wherein the subject has liver injury.

14. The method of any one of the preceding claims, wherein the subject in need of treatment is a subject with an excess of paracetamol (APAP).

15. The method of any one of the preceding claims, wherein the subject in need of treatment is a human subject.

16. The method according to any one of the preceding claims, wherein the small molecule compound comprises a disaccharide building block as shown below:wherein R is-H or-SO₃H。

17. The method according to claim 17, wherein said small molecule compound comprising a disaccharide building block is selected from the group consisting of non-anticoagulant heparin, non-anticoagulant low molecular weight heparin and O-desulphated heparin (ODSH).

18. The method of any one of the preceding claims, wherein the oligosaccharide molecule comprises from about 10 to about 20 saccharide units, or from about 12 to about 18 saccharide units.

19. The method of any one of the preceding claims, wherein the oligosaccharide molecule comprises about 18 saccharide units.

20. The method of any one of the preceding claims, wherein administration of the oligosaccharide molecule reduces neutrophil infiltration in the subject.

21. The method of any one of the preceding claims, wherein administration of the oligosaccharide molecule reduces inflammation in the subject.

22. The method of any one of the preceding claims, wherein administration of the oligosaccharide molecule protects the subject from liver damage and multiple organ system failure.

23. The method of any one of the preceding claims, wherein the subject to be treated has drug-induced inflammation.

24. A method of treating an excess of paracetamol (APAP) in a subject, the method comprising:

providing a subject in need of treatment for an APAP excess;

administering to the subject a non-anticoagulant heparan sulfate oligosaccharide molecule having anti-inflammatory properties, optionally wherein the non-anticoagulant heparan sulfate oligosaccharide molecule is configured to interact with a high mobility group box 1(HMGB1) protein in a manner sufficient to affect the interaction between the HMGB1 protein and a receptor for advanced glycation end products (RAGE),

wherein the subject's impairment from APAP overdose is alleviated.

25. A method of treating an excess of paracetamol (APAP) in a subject, the method comprising:

providing a subject in need of treatment for an APAP excess;

administering to the subject a small molecule compound having anti-inflammatory properties, the small molecule compound comprising a non-anticoagulant heparan sulfate oligosaccharide molecule comprising one of the following structural formulae:

wherein R is³Is H or a detectable label, and wherein the subject is alleviated from an excessive amount of APAP damage.

26. The method according to any one of the preceding claims, wherein the treatment of the subject for an APAP excess is effective between 0 and 24 hours after the excess, or at least within 12 hours after the excess.

27. The method of any one of the preceding claims, wherein the treatment of the subject for an APAP excess comprises protection against liver injury and/or multiple organ system failure.

28. The method of any one of the preceding claims, wherein the treatment of the subject for an APAP excess comprises reducing neutrophil infiltration in the subject.

29. The method of any one of the preceding claims, wherein treatment of the subject for an APAP excess comprises blocking the interaction between HMGB1 protein and RAGE.

30. The method according to any one of the preceding claims, wherein the small molecule compound comprises a disaccharide building block as shown below:

31. The method according to claim 32, wherein said small molecule compound comprising a disaccharide building block is selected from the group consisting of non-anticoagulant heparin, non-anticoagulant low molecular weight heparin and O-desulphated heparin (ODSH).

32. The method of any one of the preceding claims, wherein the oligosaccharide molecule comprises from about 10 to about 20 saccharide units, or from about 12 to about 18 saccharide units.

33. The method of any one of the preceding claims, wherein the oligosaccharide molecule comprises about 18 saccharide units.

Technical Field

In some embodiments, disclosed herein are novel sulfated heparan sulfate oligosaccharide compounds having anti-inflammatory activity. For example, in some embodiments, the compounds are observed to exhibit protective effects against drug-induced liver damage. While not wishing to be bound by any particular theory of operation, these compounds may inhibit the HMGB1/RAGE interaction, which is a key signaling interaction associated with sterile inflammatory injury. This finding is useful in a variety of diseases caused by an excessive inflammatory response.

Background

Sterile inflammation is a natural process that initiates tissue repair in response to cellular injury. However, excessive inflammation following initial damage often damages surrounding healthy tissue and is a key factor in many disease processes. High mobility group box 1 protein (HMGB1) is a DNA binding protein that regulates transcription and is released from the nucleus upon necrotic cell death (Zitvogel, 2010; Chen, 2010). Extracellularly, HMGB1 is a damage-associated molecular pattern (DAMP) protein that acts as a pro-inflammatory molecule, coordinating the migration and activation of inflammatory cells to the site of injury (Bianchi, 2017). HMGB 1-dependent inflammation is associated with ischemia-reperfusion and drug-induced liver injury (Huebener, 2015).

Paracetamol (APAP), also known as paracetamol, is a widely used analgesic agent that isThe active pharmaceutical ingredient of (1). Ingestion of a supratherapeutic dose can lead to Acute Liver Failure (ALF) (Heard, 2008). Misuse

Or

(co-formulations of opioids and APAP) may also lead to ALF. In the united states, nearly 50% of pharmaceutical liver injury is attributed to APAP toxicity (Lee, 2007), with approximately 80,000 emergency visits per year (Blieden, 2014). The mechanism of APAP toxicity begins with its metabolic conversion to the reactive chemical N-acetyl-p-benzoquinone imine (NAPQI), which causes hepatocyte necrosis. (Tacke, 2015). Necrotic hepatocytes release HMGB1, which drives neutrophil chemotaxis through receptors for advanced glycation end products (RAGE), activating sterile inflammation and amplifying liver injury (Huebener 2015).

However, there remains a need in the art for additional compositions and methods for treating inflammation in a subject in need thereof.

Disclosure of Invention

This summary lists several embodiments of the presently disclosed subject matter, and in many cases, lists variations and permutations of these embodiments. This summary is merely an example of the many varying embodiments. Reference to one or more representative features of a given implementation is also exemplary. Such embodiments may or may not generally have the mentioned features. Likewise, these features can be applied to other embodiments of the presently disclosed subject matter, whether listed in this summary or not. To avoid excessive repetition, this summary does not list or suggest all possible combinations of such functions.

In some embodiments, provided herein are small molecule compounds having anti-inflammatory properties comprising a non-anticoagulant heparan sulfate oligosaccharide molecule, optionally wherein the non-anticoagulant heparan sulfate oligosaccharide molecule is configured to interact with a high mobility group box 1(HMGB1) protein in a manner sufficient to affect the interaction between the HMGB1 protein and a receptor for advanced glycation end products (RAGE). The oligosaccharide molecule can comprise from about 10 to about 20 saccharide units, or from about 12 to about 18 saccharide units. The oligosaccharide molecule may comprise about 18 saccharide units.

In some embodiments, provided herein are small molecule compounds comprising a disaccharide structural unit shown below:

wherein R is-H or-SO₃H. The disaccharide building block may be selected from the group consisting of non-anticoagulant heparin, non-anticoagulant low molecular weight heparin and O-desulphated heparin (ODSH). In some embodiments, the oligosaccharide molecule protects against liver damage in vivo. In some embodiments, the oligosaccharide molecule reduces neutrophil infiltration in vivo. In some embodiments, the oligosaccharide molecule reduces inflammation in vivo.

Provided herein are methods of treating a subject comprising providing to a subject to be treated (wherein the subject has inflammation) and administering to the subject a non-anticoagulated heparan sulfate oligosaccharide molecule having anti-inflammatory properties, optionally wherein the non-anticoagulated heparan sulfate oligosaccharide molecule is configured to interact with a high mobility group box 1(HMGB1) protein in a manner sufficient to affect the interaction between the HMGB1 protein and a receptor for advanced glycation end products (RAGE). In some embodiments, the subject is suffering from any injury that results in an increase in inflammation. In some embodiments, the subject has liver injury. In some embodiments, the subject in need of treatment is a subject suffering from an excess of paracetamol (APAP). In some embodiments, the subject in need of treatment is a human subject. In some embodiments, the small molecule compound comprises a disaccharide structural unit selected from the group consisting of non-anticoagulant heparin, non-anticoagulant low molecular weight heparin, and O-desulphated heparin (ODSH). In some embodiments, the oligosaccharide molecule comprises from about 10 to about 20 saccharide units, or from about 12 to about 18 saccharide units. In some embodiments, the oligosaccharide molecule comprises about 18 saccharide units. Administration of the oligosaccharide molecule can reduce neutrophil infiltration in a subject. Administration of the oligosaccharide molecule can reduce inflammation in the subject. Administration of the oligosaccharide molecule can protect a subject from liver damage and multiple organ system failure. In some aspects, the subject to be treated has drug-induced inflammation.

Provided herein are methods of treating a subject comprising providing to a subject to be treated (wherein the subject has inflammation) and administering to the subject a non-anticoagulated heparan sulfate oligosaccharide molecule having anti-inflammatory properties, optionally wherein the non-anticoagulated heparan sulfate oligosaccharide molecule is configured to interact with a high mobility group box 1(HMGB1) protein in a manner sufficient to affect the interaction between the HMGB1 protein and a receptor for advanced glycation end products (RAGE). In some embodiments, the subject is suffering from any injury that results in an increase in inflammation. In some embodiments, the subject has liver injury. In some embodiments, the subject in need of treatment is a subject suffering from an excess of paracetamol (APAP). In some embodiments, the subject in need of treatment is a human subject. In some embodiments, the small molecule compound comprises a disaccharide structural unit selected from the group consisting of non-anticoagulant heparin, non-anticoagulant low molecular weight heparin, and O-desulphated heparin (ODSH). In some embodiments, the oligosaccharide molecule comprises from about 10 to about 20 saccharide units, or from about 12 to about 18 saccharide units. In some embodiments, the oligosaccharide molecule comprises about 18 saccharide units. Administration of the oligosaccharide molecule can reduce neutrophil infiltration in a subject. Administration of the oligosaccharide molecule can reduce inflammation in the subject. Administration of the oligosaccharide molecule can protect a subject from liver damage and multiple organ system failure. In some aspects, the subject to be treated has drug-induced inflammation.

In some aspects, provided herein are methods of treating an excess of paracetamol (APAP) in a subject, the method comprising providing treatment to a subject in need of treatment for an excess of APAP, administering to the subject a non-anticoagulated heparan sulfate oligosaccharide molecule having anti-inflammatory properties, optionally wherein the non-anticoagulated heparan sulfate oligosaccharide molecule is configured to interact with a high mobility group box 1 protein (HMGB1) protein in a manner sufficient to affect the interaction between the HMGB1 protein and a receptor for advanced glycation end product (RAGE), wherein the impairment of APAP excess in the subject is reduced. In some embodiments, the treatment of the subject for an APAP excess may be effective from 0 hours to 24 hours, or at least within 12 hours, after the excess. Treatment of APAP overdose in a subject may include prevention of liver injury and/or multiple organ system failure. Treatment of an APAP excess in a subject may include a reduction in neutrophil infiltration in the subject. Treatment of APAP excess in a subject may comprise blocking the interaction between HMGB1 protein and RAGE. In some aspects, the small molecule compound comprises a disaccharide structural unit shown below:

wherein R is-H or-SO₃H. The small molecule compound may comprise a disaccharide building block selected from the group consisting of non-anticoagulant heparin, non-anticoagulant low molecular weight heparin and O-desulphated heparin (ODSH). The oligosaccharide molecule comprises from about 10 to about 20 saccharide units, or from about 12 to about 18 saccharide units. In some aspects, the oligosaccharide molecule comprises about 18 saccharide units.

It is therefore an object of the presently disclosed subject matter to provide sulfated heparan sulfate oligosaccharide compounds having anti-inflammatory activity and methods of making and using the same.

This and other objects are achieved in whole or in part by the subject matter of this disclosure. Further, objects and advantages of the presently disclosed subject matter have been set forth above, and will become apparent to those skilled in the art upon a study of the following description, drawings and examples.

Drawings

The subject matter of the present disclosure may be better understood by reference to the following drawings. The components in the figures are not necessarily to scale, emphasis instead being placed upon illustrating the principles of the disclosed subject matter (which are generally schematic). In the drawings, like reference numerals designate corresponding parts throughout the different views. A further understanding of the subject matter of the present disclosure may be obtained by reference to the embodiments illustrated in the figures of the accompanying drawings. While the illustrated embodiments are merely examples of systems for carrying out the disclosed subject matter, the organization and method of operation of the disclosed subject matter, as well as further objects and advantages thereof, will be more readily understood by reference to the drawings and to the following description, taken in general. The drawings are not intended to limit the scope of the disclosed subject matter, which is particularly set forth in the appended or subsequently amended claims, but rather merely to illustrate and exemplify the disclosed subject matter.

For a more complete understanding of the subject matter of the present disclosure, reference is now made to the following drawings, in which:

FIGS. 1A-1G are schematic diagrams of the chemoenzymatic synthesis pathway for synthesizing 18-mer oligosaccharides;

FIG. 2 is a schematic representation of the chemical structure of an 18-mer oligosaccharide disclosed herein;

FIGS. 3A to 3D are graphical depictions of data showing the effect of 18-mer oligosaccharides on liver damage following APAP overdose;

FIGS. 4A to 4D are graphical depictions of data showing 18-mer oligosaccharides targeting HMGB1 to reduce inflammation,

FIG. 4E shows the symbol structure of HS oligosaccharides;

FIG. 4F is the illustration of FIG. 4E;

FIGS. 5A to 5D are graphical depictions of data showing the effect of delayed 18-mer oligosaccharide treatment on APAP excess;

FIGS. 6A-6D are graphical depictions of data showing biological parameters of APAP-excess mice;

FIG. 7 is a schematic of the chemical structures of biotinylated and non-biotinylated oligosaccharides: 6-mer, 12-mer, 18-mer and 18-mer AXa;

FIGS. 8A-8D are graphical depictions of data showing the protection of 6-mer, 12-mer, 18-mer, and 18-mer AXa oligosaccharides;

FIGS. 9A and 9B are graphical depictions of data showing that an excess of APAP results in elevated ALT levels, neutrophil migration, and syndecan-1 shedding; and

fig. 10A to 10C are graphical depictions based on analysis of syndecan-1 from ALF patients and data on plasma concentrations of ALT and HMGB1 in ALF patients.

Detailed Description

The presently disclosed subject matter now will be described more fully hereinafter, in which some, but not all embodiments of the presently disclosed subject matter are described. Indeed, the subject matter of the present disclosure may be embodied in many different forms and should not be construed as limited to the embodiments set forth herein; rather, these embodiments are provided so that this disclosure will satisfy applicable legal requirements.

Heparan Sulfate (HS) is a sulfated polysaccharide abundantly present on cell surfaces and in the extracellular matrix. Many DAMP, including HMGB1, are HS binding proteins (Xu, 2011). HS is involved in various aspects of inflammation, including chemokine presentation and neutrophil transendothelial migration (Proudfoot, 2003; Wang, 2005; Axelsson, 2013; Sarris, 2012). HS consists of a disaccharide repeating unit of glucuronic acid (GlcA) or iduronic acid (IdoA) linked to a glucosamine residue bearing a sulfo group. The chain length and sulfation pattern of HS determine its biological function (Gama, 2006). According to an aspect of the presently disclosed subject matter, there is provided the synthesis of specifically designed HS octadecaoses (18-mers) which exert an anti-inflammatory effect. The 18-mer protected APAP-induced acute liver failure by neutralizing the pro-inflammatory activity of HMGB1 in a mouse model. The presented results provide a new chemical space for inhibiting HMGB 1-mediated inflammatory diseases.

In some embodiments, small molecule compounds having anti-inflammatory properties are disclosed. In some embodiments, the small molecule compound comprises a non-anticoagulant heparan sulfate oligosaccharide molecule having anti-inflammatory properties. In some embodiments, the non-anticoagulant heparan sulfate oligosaccharide molecule is configured to interact with a high mobility group family protein 1(HMGB1) protein in a manner sufficient to affect the interaction between the HMGB1 protein and the receptor for advanced glycation end products (RAGE).

In some embodiments, the small molecule compounds according to the presently disclosed subject matter comprise a disaccharide structural unit, or can be part of a composition comprising a disaccharide structural unit, as shown below:

wherein R is-H or-SO₃H。

By way of example and not limitation, the disaccharide composition can be present in non-anticoagulant heparin, non-anticoagulant low molecular weight heparin, or O-desulphated heparin (ODSH). Compositions according to the presently disclosed subject matter may comprise non-anticoagulant heparin, non-anticoagulant low molecular weight heparin, or O-desulphated heparin (ODSH).

In some embodiments, the oligosaccharide molecule comprises from about 10 to about 20 saccharide units, or from about 12 to about 18 saccharide units. In some embodiments, the oligosaccharide molecule comprises about 18 saccharide units. In some embodiments, the general structure of heparan sulfate oligosaccharides that exhibit anti-inflammatory effects comprises:

wherein R is³Is H or a detectable label. In some embodiments, the detectable label comprises p-nitrophenyl.

In some embodiments, the compositions of the presently disclosed subject matter comprise a composition comprising a pharmaceutically acceptable carrier. Any suitable pharmaceutical formulation may be used to prepare the composition for administration to a subject. In some embodiments, the composition and/or carrier may be for use in a human.

For example, suitable formulations may include aqueous and non-aqueous sterile injection solutions which may contain antioxidants, buffers, bacteriostats, bactericidal antibiotics and solutes which render the formulation isotonic with the body fluids of the subject; and aqueous and non-aqueous sterile suspensions which may contain suspending agents and thickening agents. The formulations may be presented in unit-dose or multi-dose containers, for example sealed ampoules and vials, and may be stored in a frozen or freeze-dried (lyophilized) condition requiring only the addition of the sterile liquid carrier, for example water for injections, immediately prior to use. Some exemplary ingredients are Sodium Dodecyl Sulfate (SDS), in one example 0.1 to 10mg/ml, in another example about 2.0 mg/ml; and/or mannitol or another sugar, for example in the range of 10 to 10mg/ml, in another embodiment about 30 mg/ml; and/or Phosphate Buffered Saline (PBS).

It is to be understood that the formulations of the presently disclosed subject matter may include other agents conventional in the art having regard to the type of formulation in question, in addition to the ingredients particularly mentioned above. For example, sterile pyrogen-free aqueous and non-aqueous solutions can be used.

Also provided according to the presently disclosed subject matter are methods of treatment of a subject, including but not limited to methods of treating a disorder comprising inflammation. The treatment methods of the presently disclosed subject matter can comprise administering to a subject a non-anticoagulant heparan sulfate oligosaccharide molecule. In some embodiments, the non-anticoagulant heparan sulfate oligosaccharide molecule is configured to interact with a high mobility group family protein 1(HMGB1) protein in a manner sufficient to affect the interaction between the HMGB1 protein and the receptor for advanced glycation end products (RAGE).

In some embodiments, the oligosaccharide molecule protects against liver damage in vivo. In some embodiments, the oligosaccharide molecule reduces neutrophil infiltration in vivo. In some embodiments, the oligosaccharide molecule reduces inflammation in vivo.

In some embodiments, the subject has inflammation. In some embodiments, the subject suffers from any injury that results in an increase in inflammation. In some embodiments, the subject has liver injury. In some embodiments, the subject in need of treatment is a subject suffering from an excess of paracetamol (acetaminophen or APAP).

In some embodiments, the subject treated in the presently disclosed subject matter is desirably a human subject, although it is understood that the methods described herein are effective for all vertebrate species (e.g., mammals, birds, etc.), which are intended to be encompassed by the term "subject".

More specifically, provided herein are treatments of mammals, such as humans, as well as those mammals of importance to humans due to being endangered (e.g., siberian tigers), of economic importance (animals raised on farms for human consumption) and/or of social importance (animals raised as pets or zoos), such as non-human carnivores (e.g., cats and dogs), swine (pigs, live pigs, and boars), ruminants (e.g., cattle, bulls, sheep, giraffes, deer, goats, bison, and camels), and horses. Thus, the methods described herein include the treatment of livestock, including but not limited to domestic swine (pigs and live pigs), ruminants, horses, and the like.

In some embodiments, the oligosaccharide molecule reduces neutrophil infiltration in the subject. In some embodiments, administration of the oligosaccharide molecule reduces inflammation in the subject. In some embodiments, administration of the oligosaccharide molecule protects the subject from liver damage and multiple organ system failure. In some embodiments, the subject to be treated has drug-induced inflammation.

In some embodiments, the damage caused by an overabundance of APAP in a subject is reduced.

In some embodiments, the treatment of the subject for an APAP excess is effective from 0 to 24 hours post-excess, or in some embodiments at least within 12 hours post-excess, and in some embodiments within 6 hours post-excess. In some embodiments, the treatment of an APAP excess in a subject comprises protection against liver injury and/or multiple organ system failure. In some embodiments, the treatment of an APAP excess in a subject comprises reducing neutrophil infiltration in the subject. In some embodiments, the treatment of an APAP excess in a subject comprises blocking the interaction between HMGB1 protein and RAGE.

In some embodiments, the small molecule compounds according to the presently disclosed subject matter comprise a disaccharide structural unit, as shown below:

wherein R is-H or-SO₃H。

By way of example and not limitation, the disaccharide structural unit can be present in non-anticoagulant heparin, non-anticoagulant low molecular weight heparin or O-desulphated heparin (ODSH). Compositions for use in accordance with the methods of the present disclosure may comprise non-anticoagulant heparin, non-anticoagulant low molecular weight heparin, or O-desulphated heparin (ODSH). In some embodiments, the composition protects against liver damage in vivo, e.g., provides a hepatoprotective effect in vivo. In some embodiments, the oligosaccharide molecule reduces neutrophil infiltration in vivo. In some embodiments, the subject has liver injury. In some embodiments, the subject in need of treatment is a subject suffering from an excess of paracetamol (acetaminophen or APAP).

In some embodiments, the oligosaccharide molecule comprises from about 10 to about 20 saccharide units, or from about 12 to about 18 saccharide units. In some embodiments, the oligosaccharide molecule comprises about 18 saccharide units. In some embodiments, the general structure of heparan sulfate oligosaccharides that exhibit anti-inflammatory effects comprises:

wherein R is³Is H or a detectable label. In some embodiments, the detectable label comprises p-nitrophenyl.

Definition of

The terminology used herein is for the purpose of describing particular embodiments only and is not intended to be limiting of the presently disclosed subject matter.

While the following terms are considered well understood by those of ordinary skill in the art, the following definitions are set forth in order to explain the presently disclosed subject matter.

Unless defined otherwise below, all technical and scientific terms used herein are intended to have the same meaning as commonly understood by one of ordinary skill in the art. References to techniques used herein are intended to refer to techniques commonly understood in the art, including variations on those techniques or substitutions of equivalent techniques as would be apparent to one of skill in the art. While the following terms are considered well understood by those of ordinary skill in the art, the following definitions are set forth in order to explain the presently disclosed subject matter.

The term "non-anticoagulated" and the phrase "non-anticoagulative" as used in this application describe the presently disclosed subject matter as having an anticoagulation activity of less than or equal to about 50 units/mg, wherein the anticoagulation activity is attributed to inhibition of factor Xa and/or factor IIa.

In describing the presently disclosed subject matter, it should be understood that a number of techniques and procedures are disclosed. Each of these has its own benefits and each can also be used in combination with one or more, or in some cases all, of the other disclosed techniques.

Thus, for the sake of clarity, the description will avoid repeating every possible combination of steps in an unnecessary fashion. However, it is to be understood that such combinations are entirely within the scope of the invention and the claims when read.

The terms "a", "an" and "the" are used herein, including the claims, to mean "one or more" in accordance with established patent statutory convention. Thus, for example, reference to "a unit cell" includes a plurality of such unit cells and the like.

Unless otherwise indicated, all numbers expressing quantities of ingredients, reaction conditions, and so forth used in the specification and claims are to be understood as being modified in all instances by the term "about". Accordingly, unless indicated to the contrary, the numerical parameters set forth in this specification and attached claims are approximations that may vary depending upon the desired properties sought to be obtained by the presently disclosed subject matter.

As used herein, the term "about," when referring to a value or amount, mass, weight, temperature, time, volume, concentration, percentage, etc., of a composition, is intended to encompass variations from the specified amount by, in some embodiments, ± 20%, in some embodiments ± 10%, in some embodiments ± 5%, in some embodiments ± 1%, in some embodiments ± 0.5%, and in some embodiments ± 0.1%, as such variations are suitable for performing the disclosed methods or employing the disclosed compositions.

The term "comprising" is synonymous with "including," "containing," or "characterized by," and is inclusive or open-ended and does not exclude additional, unrecited elements or method steps. "comprising" is a term of art used in claim language that means that the named element is essential, but that other elements can be added and still form a construct within the scope of the claims.

The phrase "consisting of" as used herein does not include any elements, steps or ingredients not specified in the claims. The phrase "consisting of" when it appears in the clause of the subject matter of the claims, and not immediately after the preamble, it limits only the elements specified in the clause; other elements as a whole are not excluded from the claims.

As used herein, the phrase "consisting essentially of" limits the scope of the claims to the specified materials or steps, as well as those materials or steps that do not materially affect the basic and novel characteristics of the claimed subject matter.

With respect to the terms "comprising," "consisting of," and "consisting essentially of," where one of these terms is used herein, the presently disclosed and claimed subject matter can include the use of either of the other two terms.

As used herein, the term "and/or," when used in the context of a list of entities, refers to entities that exist alone or in combination. Thus, for example, the phrases "A, B, C and/or D" include A, B, C and D, respectively, but also include any and all combinations and subcombinations of A, B, C and D.