Heparan sulfate glycomimetic compounds, their uses and intermediates for their preparation

Abstract

The invention relates to heparan sulfate glycomimetic compounds that are mimetics of heparan sulfate and the use of these compounds as pharmaceuticals, cosmeceuticals, and as molecular probes. The invention also relates to intermediate compounds useful for the preparation of a wide range of glycomimetic compounds. The invention has particular relevance for the treatment or prevention of diseases, such as cancer, inflammation, diabetic nephropathy, neurodegenerative disorders, for bone healing or wound healing, for cosmeceutical uses, and as molecular probes including imaging agents.

Description

[0001] HEPARAN SULFATE GLYCOMIMETIC COMPOUNDS, THEIR USES AND INTERMEDIATES FOR THEIR PREPARATION

[0002] TECHNICAL FIELD

[0003] This invention relates generally to heparan sulfate glycomimetic compounds that are mimetics of heparan sulfate and the use of these compounds as pharmaceuticals, cosmeceuticals, and as molecular probes. The invention also relates to intermediate compounds useful for the preparation of a wide range of glycomimetic compounds. The invention has particular relevance for the treatment or prevention of diseases, such as cancer, inflammation, diabetic nephropathy, neurodegenerative disorders, for bone healing or wound healing, for cosmeceutical uses, and as molecular probes including imaging agents.

[0004] BACKGROUND

[0005] Heparanase is an endo-p-D-glucuronidase that degrades the heparan sulfate glycosaminoglycan side chains of proteoglycans in extracellular matrix and basement membrane. Heparanase appears to regulate syndecan clustering, shedding and mitogen binding. Heparanase enzymatic activity is known to be important in the promotion of tumor angiogenesis, primary tumor growth, invasion and metastasis. Heparanase cleaves heparan sulfate side chains at sites of low sulfation, thus facilitating structural alterations of the extracellular matrix and basement membrane underlying epithelial and endothelial cells. Importantly, heparanase activity correlates with the metastatic potential of cancer cells. The interaction between heparanase and its substrate, heparan sulfate has been well characterised. Heparanase has been identified as the single predominant heparan sulfatedegrading enzyme in human cancer, sparking considerable interest in the development of heparanase inhibitors for potential therapeutic applications including cancer, inflammation and diabetic nephropathy. The function of heparanase and its therapeutic potential is reviewed in Fux, L., et al., Trends Biochem. Sci., 2009 Oct, 34(10):511-519.

[0006] As populations age, neurodegenerative disorders, such as Alzheimer's disease, multiple sclerosis, Parkinson's disease, meningitis, schizophrenia and traumatic brain injury, become more prevalent. Alzheimer's disease is a common form of dementia, and is progressive and irreversible. The pathogenesis of the disease is thought to involve cerebral deposits of aggregated amyloid p-peptide. The first, and rate-limiting, step in the generation of amyloid P-peptide is cleavage of amyloid precursor protein by p-secretase (P-site amyloid precursor protein cleaving enzyme-1, p-secretase- 1, hereinafter "BACE-1"). This makes BACE-1 an attractive target for new Alzheimer's therapies.

[0007] Heparan sulfate and its highly sulfated analogue heparin have been shown to inhibit BACE-1 activity. Heparan sulfate and heparin are both glycosaminoglycans comprising 1,4- linked disaccharide units of p-D-glucuronic acid or a-L-iduronic acid with / V-acetyl-a-D- glucosamine (dominant in the case of heparan sulfate) or / V-sulfo-a-D-glucosamine (dominant in the case of heparin) and additional O-sulfate ester substituents. Heparin is a well-known pharmaceutical with anti-coagulant activity. However, the anti-coagulant properties of heparin need to be attenuated if it is to be used for other pharmaceutical applications. Otherwise, possible side effects, such as internal bleeding and impaired blood clotting, can be problematic.

[0008] Multiple sclerosis (MS) is the most common neurodegenerative disorder. Neuroinflammation is a key contributor to the disease and the enzyme heparanase is a key regulator of neuroinflammation. Overexpression of heparanase correlates closely with uncontrolled inflammation that destroys the insulating layer of myelin that surrounds nerve fibres, causing MS. Onset is usually seen in young adults and is characterised by increasing impairment of mobility, vision and co-ordination leading, ultimately, to paralysis. MS affects 2.5 million people globally, and there is no cure. New drugs that can effectively halt or delay the progression of MS are urgently needed. Heparan sulfate-based neuroprotective pharmaceuticals can reduce inflammation and enhance repair of the central nervous system (CNS).

[0009] Heparan sulfate is used in a number of cosmeceutical and dermatological formulations. Heparan sulfate oligosaccharides are highly sulfated glycosaminoglycans that play a crucial role in a range of essential physiological processes. Heparan sulfate binds collagen (the main protein of connective tissue), regulates its synthesis and has the ability to organise water molecules and fill the spaces between water molecules. Collagen is responsible for strength, texture and elasticity of tissue, but the amount of collagen in the skin decreases over time by ~1% per year. Various glycosaminoglycan compounds, such as hyaluronic acid and low molecular weight heparin, have already found use in multiple cosmeceutical formulations delivering drug-like benefits and a rejuvenating effect to the skin. However, the exploitation of heparan sulfate oligosaccharides has been hindered by the complexity of their syntheses. The applicants recently developed a robust and scalable method for preparing novel polyvalent displays of small specific heparan sulfate fragments on chemical structures known as dendritic cores. This methodology greatly simplified the synthesis requirements while retaining the desired bioactivity of the heparan sulfate structures. Biologically active heparan sulfate glycomimetic compounds have the potential to dramatically slow, and possibly reverse, the signs and symptoms of skin aging.

[0010] In the search for improved therapies and treatments for the above-mentioned diseases and disorders, the applicants have investigated glycomimetics of heparan sulfate. Certain compounds based on heparan sulfate have been prepared by the applicant and are described in WO 2014 / 084744 as being potentially effective for treating disorders in which BACE-1 is implicated. These compounds are mimetics of heparan sulfate with di- and tetra-saccharide fragments of heparan sulfate attached to dendritic cores. These fragments were prepared from glucose and glucosamine monosaccharides by multistep syntheses involving glycosylations and selective protection-deprotection reactions employing an orthogonal protecting group strategy. Suitably protected heparan sulfate fragments were attached to the core and selective deprotection allowed selective sulfation leading to the target compounds. Although this approach reduced the number of reaction steps compared to other approaches for the synthesis of heparan sulfate oligosaccharides, the synthesis process remained lengthy.

[0011] The applicant has now developed a new class of heparan sulfate mimetic compounds. Some compounds of the class have potential as therapeutic agents, such as anti-cancer agents, cosmeceutical agents, or therapeutics for neurodegenerative disorders.

[0012] The applicant has also found a convenient and effective synthetic route to a variety of similar compounds which have been "tagged" with a chemical group enabling their use for different applications. For example, some of these compounds are useful as molecular probes for biodistribution studies and as imaging agents.

[0013] The invention provides novel heparan sulfate mimetic compounds having potential as therapeutic agents or cosmeceutical agents or compounds useful as molecular probes or imaging agents.

[0014] SUMMARY OF INVENTION

[0015] In a first aspect, the invention provides a compound of formula (I): wherein:

[0016] X is (CH2)Pand p is an integer from 1 to 20;

[0017] Ri is H or CH3; and wherein • is the point of attachment of R2 to the O atom;

[0018] Y is (CH2)nand n is an integer from 1 to 3;

[0019] R3 is H or C1-C3 alkyl;

[0020] Z is (CH2)m and m is an integer from 1 to 6; and

[0021] M is: wherein R4 is H or SO3H, and • is the point of attachment to Z; or a salt thereof.

[0022] In certain embodiments of the invention, p is an integer from 5 to 12. In some embodiments, p is 5. In other embodiments, p is 6. In other embodiments, p is 12.

[0023] In certain embodiments of the invention, n is 2.

[0024] In certain embodiments of the invention, Ri is H. In other embodiments, Ri is methyl.

[0025] In certain embodiments of the invention, R3 is H. In other embodiments, R3 is methyl or ethyl. In certain embodiments of the invention, m is 4, 5 or 6. Preferably m is 6.

[0026] In certain embodiments of the invention, one or more of the R4 groups are SO3H or SChNa. In some embodiments, all R4 groups are SO3H or SChNa.

[0027] In certain embodiments of the invention, the compound of formula (I) is selected from the group comprising:

[0028] and

[0029]

[0030] In a further aspect, the invention provides a compound of formula (II): wherein: Ri is H or CH3; and wherein • is the point of attachment of R2 to the O atom;

[0031] Y is (CH2)n and n is an integer from 1 to 3;

[0032] R3 is H or C1-C3 alkyl;

[0033] Z is (CH2)m and m is an integer from 1 to 6; and wherein R4 is H or SO3H, and • is the point of attachment to Z; and

[0034] A is a Ci-Ce alkyl group substituted with -NH2, -N3, or -NH(C=O)Rs wherein Rs is: a) a C1-C12 alkyl group; b) a biotinyl substituent; c) a group comprising a fluorescent label; d) a group comprising a fluorine-18 label; e) a group comprising a fluorine-19 label; f) a group comprising a crown ether-based caged ligand for rhodium, iridium, actinium-225 or thorium-227; or g) a group comprising an N-acetate or a C-14 radiolabelled N-acetate; or a salt thereof.

[0035] In certain embodiments of the invention, n is 2.

[0036] In certain embodiments of the invention, Ri is H. In other embodiments, Ri is methyl.

[0037] In certain embodiments of the invention, R3 is H. In other embodiments, R3 is methyl or ethyl.

[0038] In certain embodiments of the invention, m is 4, 5 or 6. Preferably, m is 6.

[0039] In certain embodiments of the invention, A is a Ci-Ce alkylamine, for example pentylamine. In other embodiments, A is a Ci-Ce alkylazide, for example pentylazide.

[0040] In certain embodiments of the invention, A is a Ci-Ce alkyl group substituted with NH(C=0)R4 wherein R4 is a C6-C12 alkyl group.

[0041] In certain embodiments of the invention, A is a Ci-Ce alkyl group substituted with NH(C=0)R4 wherein R4 is a biotinyl substituent. For example, A may be biotinyl.

[0042] In certain embodiments of the invention, A is a Ci-Ce alkyl group substituted with NH(C=0)R4 wherein R4 is a C6-C12 alkyl group comprising a fluorescent label. Preferably the fluorescent label is a BODIPY group or a Cyanine?.5 NIR dye.

[0043] In certain embodiments of the invention, A is a Ci-Ce alkyl group substituted with NH(C=0)R4 wherein R4 is a C6-C12 alkyl group comprising a fluorine-18 label, for example (1- [3-(2-[18F]Fluoropyridin-3-yloxy)propyl]pyrrole-2, 5-dione).

[0044] In certain embodiments of the invention, A is a Ci-Ce alkyl group substituted with NH(C=O)Rs wherein Rs is a C6-C12 alkyl group comprising a fluorine-19 label, for example a F-19 3,5-bis(trifluoromethyl)benzyl label.

[0045] In certain embodiments of the invention, A is a Ci-Ce alkyl group substituted with NH(C=0)R4 wherein R4 is a C6-C12 alkyl group comprising 1,4,7, 10-tetraazacyclododecane- 1,4,7, 10-tetraacetic acid (DOTA), / V, / V'-bis[(6-carboxy-2-pyridil)methyl]-4,13-diaza-18- crown-6 (H2macropa), or H2macropa-NCS.

[0046] In certain embodiments of the invention, one or more of the R4 groups are SO3H or SChNa. In some embodiments, all R4 groups are SO3H or SOsNa.

[0047] In certain embodiments of the invention, the compound of formula (II) is selected from the group comprising:

[0048]

[0049]

[0050]

[0051]

[0052]

[0053]

[0054]

[0055]

[0056] 5 and

[0057] In certain embodiments of the invention, A is a C3-C6 alkylamine, for example pentylamine.

[0058] In another aspect the invention provides a composition comprising an effective amount of a compound of formula (I) or formula (II) and a suitable carrier, diluent or excipient. The composition may be a pharmaceutical or cosmeceutical composition.

[0059] In another aspect the invention provides a method of treating or preventing any one or more of cancer, inflammation, diabetic nephropathy, a neurodegenerative disorder, a dermatological condition, or for promoting bone healing or wound healing, comprising administering a pharmaceutically effective amount of a compound of formula (I) or formula (II) to a patient requiring treatment.

[0060] In another aspect the invention provides a method of rejuvenating skin or preventing skin-aging comprising administering an effective amount of a compound of formula (I) or formula (II) to human skin.

[0061] In another aspect the invention provides a method of using a compound of formula (II) as an imaging agent in an in vitro or in vivo imaging procedure. In certain embodiments of the invention, the imaging procedure is fluorescence imaging. In other embodiments, the imaging procedure is magnetic resonance imaging. In other embodiments, the imaging procedure is positron emission tomography (PET) imaging.

[0062] In another aspect the invention provides a method of using a compound of formula (II) as a molecular probe. In certain embodiments of the invention, the molecular probe is used for biodistribution investigations.

[0063] In another aspect the invention provides the use of a compound of formula (I) or formula (II) for treating or preventing any one or more of cancer, inflammation, diabetic nephropathy, a neurodegenerative disorder, and a dermatological condition, or for promoting bone healing or wound healing.

[0064] In another aspect the invention provides the use of a compound of formula (I) or formula (II) for rejuvenating skin or preventing skin-aging.

[0065] In another aspect the invention provides the use of a compound of formula (I) or formula (II) in the manufacture of a medicament for treating or preventing any one or more of cancer, inflammation, diabetic nephropathy, a neurodegenerative disorder, and a dermatological condition, or for promoting bone healing or wound healing.

[0066] In another aspect the invention provides the use of a compound of formula (I) or formula (II) in the manufacture of a medicament for treating or preventing skin-aging.

[0067] In another aspect the invention provides a pharmaceutical composition comprising a compound of formula (I) or formula (II) for treating or preventing any one or more of cancer, inflammation, diabetic nephropathy, a neurodegenerative disorder, and a dermatological condition, or for promoting bone healing or wound healing.

[0068] Inflammation can include neuroinflammatory diseases such as multiple sclerosis.

[0069] Neurodegenerative disorders include, but are not limited to, senile dementia, pre-senile dementia, multi-infarct dementia, and Alzheimer's disease.

[0070] In another aspect the invention provides a compound of formula (I) or formula (II) in combination with at least one other compound, e.g. a second drug compound. The other compound may be, for example, an oligosaccharide compound, a cyclitol such as scyllo- inositol or D-chiro-inositol, an acetylcholinesterase inhibitor, a nicotinic agonist, an antibody targeting p-amyloid, an inhibitor of 0-amyloid, an inhibitor of tau aggregation, or memantine.

[0071] In another aspect the invention provides the use of a compound of formula (I) or formula (II) in combination with at least one other compound, e.g. a second drug compound, e.g. an oligosaccharide compound, a cyclitol such as scyllo-inositol or D-chiro-inositol, an acetylcholinesterase inhibitor, a nicotinic agonist, an antibody targeting p-amyloid, an inhibitor of tau aggregation, or memantine, for treating or preventing any one or more of cancer, inflammation, diabetic nephropathy, a neurodegenerative disorder, and a dermatological condition, or for promoting bone healing or wound healing.

[0072] In another aspect the invention provides a method of treating or preventing any one or more of cancer, inflammation, diabetic nephropathy, a neurodegenerative disorder, and a dermatological condition, or for promoting bone healing or wound healing, comprising administering a pharmaceutically effective amount of a compound of formula (I) or formula (II) in combination with at least one other compound, e.g. a second drug compound, e.g. an oligosaccharide compound, a cyclitol such as scyllo-inositol or D-chiro-inositol, an acetylcholinesterase inhibitor, a nicotinic agonist, an antibody targeting p-amyloid, an inhibitor of 0-amyloid, an inhibitor of tau aggregation or memantine. The compound of formula (I) or formula (II) and the other compound may be administered separately, simultaneously or sequentially.

[0073] In another aspect the invention provides the use of a compound of formula (II) as a molecular probe.

[0074] In another aspect the invention provides the use of a compound of formula (II) as an imaging agent in an in vitro or in vivo imaging procedure. In certain embodiments of the invention, the imaging procedure is fluorescence imaging. In other embodiments, the imaging procedure is magnetic resonance imaging. In other embodiments, the imaging procedure is positron emission tomography (PET) imaging.

[0075] BRIEF DESCRIPTION OF THE FIGURES

[0076] Figure 1 is a graph showing the binding of sulfated maltose tetramer BODIPY amide 37 to CD4 T helper immune cells over 12h and 24h in a cell culture.

[0077] Figure 2 comprises graphs a. to e. showing a comparison between sulfated maltose tetramer BODIPY amide 37 and unsulfated maltose tetramer BODIPY amide 36 in binding to immune cells in the blood of healthy animals and animals with EAE. The immune cells include CD45+ cells (a), CD4 T helper cells (b), CD8 cytotoxic T cells (c), monocytes (d) and neutrophils (e).

[0078] Figure 3 comprises graphs a. to e. showing a comparison between sulfated maltose tetramer BODIPY amide 37 and unsulfated maltose tetramer BODIPY amide 36 in binding to immune cells isolated from the brains of healthy animals and animals with EAE. The immune cells include CD45+ cells (a), CD4 T helper cells (b), CD8 cytotoxic T cells (c), monocytes (d) and neutrophils (e).

[0079] Figure 4 comprises confocal microscopy images of sections of mouse brain cortex from animals that received no treatment, compound 36 (unsulfated maltose tetramer BODIPY amide), or compound 37 (sulfated maltose tetramer BODIPY amide). The brain slices were stained to indicate nuclei (blue), endothelial cells (red), and compound 36 or 37 (green).

[0080] Figure 5 comprises images (A) of bEnd.3 cells incubated with sulfated maltose tetramer BODIPY amide 37 showing binding to the cells and graphs (B and C) showing a binding comparison between sulfated and unsulfated maltose tetramer BODIPY amides 36 and 37.

[0081] Figure 6 comprises confocal microscopy images of brain slices from healthy mice incubated with no treatment, compound 36 (unsulfated maltose tetramer BODIPY amide), or compound 37 (sulfated maltose tetramer BODIPY amide).

[0082] Figure 7 comprises confocal microscopy images of the cerebral cortex (A, B, E, F) and cerebellum (C, D, G, H) of mouse brain from healthy and EAE animals.

[0083] Figure 8 comprises graphs A to C showing a comparison in the binding of compound 37 (sulfated maltose tetramer BODIPY amide) to immune cells (A and B) or free compound 37 (C) in the blood of animals administered 60 or 10 pig of compound 37 by intraperitoneal injection (i.p.) or oral gavage (p.o.) 45 minutes previously. The immune cells include CD45+ cells (A) and monocytes (B). Graph C shows fluorescence levels for compound 37 in plasma 45 minutes after i.p. or oral administration of 10 pg and 60 pg doses.

[0084] Figure 9 comprises graphs A and B showing fluorescence levels for compound 37 (sulfated maltose tetramer BODIPY amide) in plasma over 12 hours after i.p. or oral administration of 60 pg and graphs C and D showing the binding of compound 37 to effector memory CD4 T cells in the blood and brains over 12 hours after i.p. administration of 60 pg of compound 37.

[0085] Figure 10 shows fluorescence microscopy images of fluorescently labelled mimetics. Red fluorescence staining (phalloidin) highlights the cytoskeleton, blue fluorescence staining (Hoechst) visualises cell nuclei, and green fluorescence shows the sulfated maltose tetramer BODIPY amide 37.

[0086] Figure 11 comprises19F-MRI images showing the detection of19F-labelled compounds 20 and 24 with 480 averages and acquisition times between 6 and 213 minutes.

[0087] Figure 12 is a graph showing signal-to-noise ratios (SNR) of19F-labelled compounds 20 and 24 at different concentrations.

[0088] Figure 13 comprises MRI images showing a tube containing compound 24 placed beside a mouse cadaver with the:H anatomical scan (left) and19F signal overlaid (right).

[0089] Figure 14 is a graph showing heparanase inhibition activity for compound 7a.

[0090] Figure 15 is a graph showing heparanase inhibition activity for compound 7c.

[0091] DETAILED DESCRIPTION

[0092] Definitions

[0093] The term "Ci-Ce alkyl" means any saturated hydrocarbon radical having up to 6 carbon atoms and is intended to include both straight- and branched-chain alkyl groups. Examples of alkyl groups include: methyl group, ethyl group, n-propyl group, / so-propyl group, n-butyl group, / so-butyl group, sec-butyl group, t-butyl group, n-pentyl group, 1,1-dimethylpropyl group, 1,2-dimethylpropyl group, 2,2-dimethylpropyl group, 1-ethylpropyl group, 2- ethylpropyl group, n-hexyl group and l-methyl-2-ethylpropyl group.

[0094] The term "prodrug" means a pharmacologically acceptable derivative of the compounds of formula (I) such that an in vivo biotransformation of the derivative gives the compound as defined in formula (I). Prodrugs of compounds of formulae (I) may be prepared by modifying functional groups present in the compounds in such a way that the modifications are cleaved in vivo to give the parent compound. Typically, prodrugs of the compounds of formula (I) will be ester prodrug forms. The term "cosmeceutical" means the combination of a pharmaceutical and a cosmetic, and refers generally to a cosmetic product containing one or more biologically active ingredient that has, or is purported to have, a pharmaceutical effect.

[0095] The term "pharmaceutically acceptable salts" is intended to apply to non-toxic salts such as ammonium salts, metal salts, e.g. sodium salts, or salts of organic cations, or a mixture thereof.

[0096] The term "protecting group" means a group that selectively protects an organic functional group, temporarily masking the chemistry of that functional group and allowing other sites in the molecule to be manipulated without affecting the functional group. Suitable protecting groups are known to those skilled in the art and are described, for example, in Protective Groups in Organic Synthesis (3rdEd.), T. W. Greene and P. G. M. Wuts, John Wiley & Sons Inc (1999). Examples of protecting groups include, but are not limited to: O-benzyl, O-benzhydryl, O-trityl, O-tert-butyldimethylsilyl, O-tert-butyldiphenylsilyl, 0-4- methylbenzyl, O-acetyl, O-chloroacetyl, O-methoxyacetyl, O-benzoyl, 0-4-bromobenzoyl, O- 4-methylbenzoyl, O-fluorenylmethoxycarbonyl and O-levulinoyl.

[0097] The term "experimental autoimmune encephalomyelitis" or "EAE" means the animal model of MS that entails the induction by immunisation of myelin-specific immune cells, which enter into the CNS causing demyelination and MS-like symptoms (i.e. paralysis).

[0098] The term "patient" includes human and non-human animals.

[0099] The terms "treatment", "treating" and the like include the alleviation of one or more symptoms, or improvement of a state associated with the disease or disorder, for example, improvement in cognition, improvement in memory function.

[0100] The terms "preventing", "prevention" and the like include the prevention of one or more symptoms associated with the disease or disorder.

[0101] The compounds of the invention are useful in both free base form and in the form of salts and / or solvates.

[0102] The Compounds of the Invention

[0103] Compounds of formula (I) and formula (II) are herein described as "compounds of the invention". A compound of the invention includes a compound in any form, e.g. in free form or in the form of a salt or a solvate.

[0104] The compounds of the invention include compounds of formula (I): wherein:

[0105] X is (CH2)Pand p is an integer from 1 to 20; Ri is H or CH3; and wherein • is the point of attachment of R2 to the O atom;

[0106] Y is (CH2)n and n is an integer from 1 to 3;

[0107] R3 is H or C1-C3 alkyl;

[0108] Z is (CH2)m and m is an integer from 1 to 6; and

[0109] M is: wherein R4 is H or SO3H, and • is the point of attachment to Z; or a salt thereof.

[0110] The compounds of the invention also include compounds of formula (II) : wherein: Ri is H or CH3; and wherein • is the point of attachment of R2 to the O atom;

[0111] Y is (CH2)n and n is an integer from 1 to 3;

[0112] R3 is H or C1-C3 alkyl;

[0113] Z is (CH2)m and m is an integer from 1 to 6; and

[0114] M is: wherein R4is H or SO3H, and • is the point of attachment to Z; and

[0115] A is a Ci-Ce alkyl group substituted with -NH2, -N3, or -NH(C=O)Rs wherein Rs is: a) a C1-C12 alkyl group; b) a biotinyl substituent; c) a group comprising a fluorescent label; d) a group comprising a fluorine-18 label; e) a group comprising a fluorine-19 label; f) a group comprising a crown ether-based caged ligand for rhodium, iridium, actinium-225 or thorium-227; or g) a group comprising an N-acetate or a C-14 radiolabelled N-acetate; or a salt thereof.

[0116] Synthesis of the compounds of the invention

[0117] The compounds of the invention may be prepared by a variety of different methods. The following are representative non-limiting general methods for synthesising compounds of the invention.

[0118] Compounds of formula (I) may be prepared according to the procedures outlined in Scheme 1 and Scheme 2.

[0119] Scheme 1 shows the preparation of the useful intermediate tetraethyl ester compounds (3a,b,c) from a known diester (1) and dicarboxylic acids (2a,b,c) via peptide coupling using the peptide coupling agent HATU. Compounds (3a,b,c) can then be de-esterified to the respective tetracarboxylic acids (4a,b,c).

[0120] Scheme 1

[0121] Scheme 2 shows the coupling of tetracarboxylic acids (4a,b,c) with the maltose alkylamine (5) using the peptide coupling agent PyBOP to give the tetrameric compounds (6a,b,c). Over-sulfation using sulfur trioxide-trimethylamine complex in DMF or DMF / toluene gives the sulfated tetramers (7a,b,c).

[0122] Scheme 2

[0123] The compounds of formula (II) maybe prepared using the intermediate compound (14). The preparation of this compound is shown in Scheme 3. 0-Glutamic acid diethyl ester (10) is prepared from diethyl 3-oxoglutarate (8). Coupling of the diester (10) with the azidoacid (9) gives the diester azido-amide (11). De-esterification leads to the dicarboxylic acid (12). Coupling with the diester (1) using HATU gives the tetraethyl ester azide (13). Deesterification gives the tetra -carboxylic acid azide (14). Scheme 3

[0124] Treatment of the tetra-carboxylic acid (14) with the maltose alkylamine (5), as shown in Scheme 4, provides the maltose tetramer azide (15). The azide (15) can be reduced to the maltose tetramer amine (16) or sulfated to form the sulfated maltose tetramer azide (17), as shown in Scheme 5. The sulfated azide (17) can be reduced to the sulfated maltose tetramer amine (18), as shown in Scheme 6. The azides (15) and (17), and the amines (16) and (18), have proven to be versatile intermediates for the synthesis of a variety of compounds having interesting potential uses. For example, the primary amine of compound (16) presents a useful functional group for the attachment of alkyl chains to produce glycolipids and isotopically labelled moieties useful as imaging agents for biodistribution studies. Further, the primary amine of compound (18) presents a useful functional group for the attachment of fluorophores or a biotin moiety to produce fluorescent-labelled or biotinylated compounds useful as imaging agents for biodistribution studies or as vehicles for drug delivery.

[0125] Scheme 4

[0126] Scheme 5

[0127] The maltose tetramer amine (16) can be treated with alkyl or aromatic carboxylic acids of varying alkyl chain length or aromatic ring arrangement to produce glycolipids. For example, amine (16) can be treated with undecanoic acid to give the maltose tetramer undecanamide (25), which can then be persulfated to give the sulfated maltose tetramer undecanamide (26). Glycolipids of this type have potential as therapeutic agents or as cosmeceuticals.

[0128] The maltose tetramer amine (16) can also be modified to incorporate a F-19 label or a BODIPY label for use in imaging procedures. For example, treatment of the amine (16) with F-19 labelled 3,5-bis(trifluoromethyl)benzoic acid gives the F6 labelled compound (19) which can then be persulfated to compound (20). In another example, F-19 labelled (23) provides the F27 labelled compound (24) after persulfation. BODIPY-labelling is achieved by

[0129] SUBSTITUTE SHEET (RULE 26) reacting the amine (16) with BDP-COOH followed by persulfation to give the BODIPY-labelled compound (37).

[0130] Biotinylated compounds can be useful for drug delivery and for biodistribution studies. The amine (16) can be biotinylated by treatment with biotin- / V-hydroxysuccinimide ester to give compound (34) following persulfation. In another example, the biotinylated compound (35) can be prepared from sulfated maltose tetramer amine (18). Biotin is the compound named 5-[3aS,4S,6aR)-2-oxohexahydro-lH-thieno[3.4-d]imidazole-4-yl]pentanoic acid and having the chemical structure:

[0131] Therapeutic uses

[0132] The compounds of formula (I), and certain compounds of formula (II), are potential inhibitors of heparanase and therefore have potential for the treatment or prevention of a range of diseases including cancer, inflammation, diabetic nephropathy, heart diseases, and are potentially useful for bone healing or wound healing.

[0133] The compounds are also anticipated to be inhibitors of BACE-1 and may therefore be useful for the treatment or prevention of neurodegenerative disorders, senile dementia, pre- senile dementia, multi-infarct dementia or Alzheimer's disease.

[0134] Glycolipid compounds of the invention, according to formula (II), were prepared with the objective of improving their potency and bioavailability while retaining ease of manufacture and safety. The new class of glycolipid compounds was prepared by 'decorating' sugar fragments with lipophilic "tails" and various other moieties. Glycolipid molecules have a ’polar’ sugar end that is soluble in water and a ’nonpolar’ tail that is soluble in fat. This helps to orient and cluster glycolipid compounds into biological systems in a specific way to maximise their interactions. The lipid moiety of a glycolipid is buried in the cell membrane bilayer, leaving the sugar moiety prominently exposed on the surface of living cells for recognition by proteins.

[0135] Examples 36 to 41, discussed under "Use as imaging agents and molecular probes" below, indicate the potential for compounds structurally related to sulfated maltose tetramer BODIPY amide (37) to be used as therapeutic agents for diseases associated with inflammatory processes and neurodegeneration due to the ability of the BODIPY amide (37) to have sustained interactions with immune cells in the blood and brain and its ability to enter the CNS within 45 minutes of administration. Cosmeceutical uses

[0136] In addition to the therapeutic uses referred to above, the compounds of the invention have also been identified as having potential cosmeceutical uses.

[0137] The reduction of fibrillar type I collagen is a characteristic feature of chronologically aged or "unhealthy" skin, although the destruction of existing collagen is, undoubtedly, central to the deleterious changes observed in aged skin. The failure to replace damaged collagen with newly synthesised material is also critical to the overall pathophysiology. These observations have provided the skin care industry with a strong rationale to develop topical formulations that stimulate skin to produce more collagen which in turn can slow or reverse the pathophysiology of skin aging.

[0138] The diminution of extracellular matrix is a common event during the aging of connective tissues. Human skin fibroblasts from older donors have increased levels of collagenase mRNA and protein, relative to younger donors, whereas expression of the collagen type I and III genes decrease in an age dependent way. Replicative senescence of human skin fibroblasts appears to correlate with a loss of regulation and overexpression of collagenase activity. As a consequence of the age-related increase in dermal collagenase, skin care companies are constantly trying to develop compounds that can inhibit collagenase with the objective of boosting the content of dermal collagen which in turn provides the aesthetic appearance of "younger skin".

[0139] Skin aging is also associated with the loss of skin moisture. The key molecule involved in skin moisture is hyaluronan or hyaluronic acid (HA), a glycosaminoglycan (GAG) with a unique capacity to bind and retain water molecules. The synthesis of epidermal HA is influenced by the underlying dermis and is under separate controls from the synthesis of dermal HA. Progressive reduction in the size of the HA polymers in skin is a result of aging. Thus, the epidermis loses the principal molecule responsible for binding and retaining water molecules, resulting in the loss of skin moisture. In the dermis, the major age-related change is the increasing avidity of HA with tissue structures with the concomitant loss of HA extractability. This parallels the progressive cross-linking of collagen and the steady loss of collagen extractability with age. All of these age-related phenomena contribute to the apparent dehydration, atrophy and loss of elasticity that characterizes aged skin.

[0140] Abnormal hyperpigmentations such as melasma, freckles and senile lentigines and other forms of melanin hyperpigmentation show satisfactory subjective improvement when treated with depigmenting agents, such as hydroquinone, ascorbic acid derivatives, kojic acid, azelaic acid, electron rich phenols, corticosteroids, retinoids and others. Among these agents, hydroquinone drugs are effective in up to 80% of users. They are often formulated with steroids because they may cause irritation and dermatitis. There is therefore a constant search for safe compounds that can reduce skin pigmentation. The rate-limiting steps of mammalian melanin synthesis are regulated by tyrosinase, which catalyses the conversions of L-tyrosine to L-dopa and L-dopa to L-dopa-quinone. Therefore, determining the effect of a compound on tyrosinase activity could provide a strong indicator of its ability to act as a "skin whitening" component.

[0141] Use as drug delivery agents

[0142] Compounds of the invention can be useful as vitamin-guided anticancer drug delivery vehicles. They steer the drug to the right location. Vitamins are necessary for certain metabolic processes in all mammalian cells. Water-soluble vitamins biotin (B7) and folic acid (B9) are essential for normal cellular function, growth and development. Biotin functions as a coenzyme in metabolic reactions. Exogenous biotin is taken up via a Sodium-dependent Multi-Vitamin Transporter (SMVT). Various cancer cells overexpress SMVT, thus making biotin a tumour-targeting vector.

[0143] Tetrameric egg white avidin and its bacterial counterpart streptavidin have the highest affinity towards biotin (Kd~1013-1015M'1) known in nature. They can be utilised in drug delivery applications as the avidin-biotin complex induces only a low immune response. Biotin demand in rapidly growing tumours is higher than in normal tissues. Biotin on various constructs can enhance cellular uptake of therapeutic molecules by active recognition by the transporter. Therefore, biotinylation of anticancer agents can improve selective delivery to cancer cells.

[0144] Examples 31-33 describe the synthesis of biotinylated compounds of the invention.

[0145] Use as imaging agents and molecular probes

[0146] Fluorescently labelled derivatives of heparan sulfate glycomimetic compounds can be used to study the biodistribution of heparan sulfate glycomimetic compounds as heparanase inhibitors. This may assist with a greater understanding of the mechanism of action through which the compounds obtain their anti-cancer activity. Developing knowledge of how the compounds are distributed on a cellular level and, importantly, whether they are internalised by cells, may lead to the narrowing of their potential interactome and identification of lectin targets. This, in turn, may enable the development of biological assays, the screening of the potential protein targets, and the development of rational drug-design approaches to the synthesis of therapeutically effective compounds. An improved understanding of the mechanism of action could lead to improvements in efficacy, bioavailability and specificity of glycomimetic compounds.

[0147] Fluorescent labelling may also help identify whether a compound is tissue or cell-type specific in vitro and in vivo. Such selectivity for certain tissues, or perhaps regions of inflammation, may provide the basis for the use of glycomimetic compounds as drug-delivery systems. BODIPY-based compounds are chemically stable and have excellent fluorescent properties. Examples of the synthesis of BODIPY compounds of the invention are described in Examples 34 and 35. Examples 36-41 describe investigations into the cell interactions of sulfated maltose tetramer BODIPY amide (37) and its biodistribution when administered in vivo.

[0148] Example 36 shows that cell-bound BODIPY amide (37) is detected on immune cells in the blood within 45 minutes of IP injection and the binding is similar in healthy and EAE mice. Mice were immunised for experimental autoimmune encephalomyelitis (EAE) or were healthy. 24 days after EAE immunisation, mice received a single i.p. dose of sulfated maltose tetramer BODIPY amide (37) or non-sulfated maltose tetramer BODIPY amide (36) (60 pg in 100 pl PBS). The mimetic was allowed to circulate for 45 minutes before mice were sacrificed and tissues (blood and brain, Figures 2 and 3, respectively) collected for analysis by flow cytometry. BODIPY expression is displayed as BODIPY mean fluorescent intensity (MFI) as a percentage of BODIPY MFI detected in a sham injection control animal (Figures 2a-2c) or raw MFI (Figures 2d and 2e and Figures 3a-e). Data were pooled from two independent experiments (n=5-8 per group) and displayed as mean ± SEM. *p<0.05, **p<0.01 by two- way ANOVA with Holm-Sidak's multiple comparisons test. This Example not only shows that sulfated BODIPY amide (37) binds immune cells in the blood and neutrophils in the brain but also that non-sulfated BODIPY amide (36) does not bind immune cells in the blood or brain. Binding is therefore dependent on highly sulfated saccharide dendrites that interact with a range of extracellular proteins and components found on immune cells. The Example also shows that sulfated BODIPY amide (37) binds and interacts with immune cells in the blood and neutrophils in the brain within 45 after IP injection, and is able to bind and interact with a wide range of immune cells in the blood including T helper cells, cytotoxic T cells, monocytes, and neutrophils. The Example also shows that sulfated BODIPY amide (37) is able to bind to immune cells in the blood and brain of healthy as well as EAE animals.

[0149] Example 37 shows that sulfated BODIPY amide (37) readily and stably binds immune cells in vitro. Binding remained stable for at least 24 hours and more cells became positive for BODIPY over 24 hours compared to 12 hours. Splenocytes were isolated from healthy mice and cultured in vitro for 12 or 24 hours in the presence of sulfated BODIPY amide (37) (0, 1.2, 4, 12 pM). Cells were then assessed by flow cytometry to determine the % of immune cells positive for sulfated BODIPY amide (37). Means and SEM for BDP+CD4+T cells are shown in Figure 1. This finding is consistent with the binding of sulfated BODIPY amide (37) to many different types of immune cells in the blood of mice administered sulfated BODIPY amide (37) i.p. (Figure 2).

[0150] Example 38 shows that sulfated BODIPY amide (37) can be detected in the in the blood and brain bound to immune cells and free in the plasma (see Figures 8 and 9). Mice were injected i.p. with 60 pg sulfated BODIPY amide (37) (Figures 8 and 9) or 10 pg sulfated BODIPY amide (37) (Figure 8). Additionally, mice were treated p.o. with 60 pg sulfated BODIPY amide (37) (Figures 8 and 9) or 10 p.g sulfated BODIPY amide (37) (Figure 8). Control mice were untreated. Sulfated BODIPY amide (37) treatments were 45 minutes (Figures 8 and 9), 4 hours (Figure 9), or 12 hours (Figure 9) before euthanasia. Plasma was collected and fluorescence units measured using a fluorescent plate reader (Figures 8C and 9A; n = 3- 11 mice per group). Binding of sulfated BODIPY amide (37) to immune cells in the blood (Figures 8A, 8B, and 9B) and brain (Figure 9B) was assessed by flow cytometry (n=3-ll mice per group).

[0151] Example 38 shows that sulfated BODIPY amide (37) can be detected on peripheral blood immune cells (CD45+ cells (Figure 8A) and monocytes (Figure 8B)) providing further support for Example 36. Example 38 further reveals that sulfated BODIPY amide (37) can be detected on immune cells in the blood when administered i.p. or orally (Figure 8) and at a 60 and 10 jig i.p. dose after 45 minutes (*p<0.05 by one-tailed Student's t test). Example 38 also demonstrates that sulfated BODIPY amide (37) can be detected free in the plasma 45 minutes after i.p. administration of 60 ng (Figure 8C).

[0152] The graph of Figure 8A shows that sulfated BODIPY amide (37) is detected on immune cells in the blood (BDP+CD45+ cells) and is presented as a % of immune cells (CD45+ cells) in peripheral blood. IP administration of 60 pg leads to the highest number of immune cells bound by compound 36 while 10 pg i.p and 60 pg p.o. of sulfated BODIPY amide (37) led to similar levels of binding concluding that a 10 pg i.p dose delivers a similar level of sulfated BODIPY amide (37) to the blood as 60 pg oral 45 minutes after administration.

[0153] The graph of Figure 8B shows that sulfated BODIPY amide (37) is detected on monocytes in the blood (BDP+ monocytes) and is presented as a % of monocytes. IP administration of 60 pg leads to the highest number monocytes bound by sulfated BODIPY amide (37) while 10 pg i.p and 60 pg p.o. of sulfated BODIPY amide (37) led to similar levels of binding concluding that a 10 pg i.p dose delivers a similar level of sulfated BODIPY amide (37) to the blood as 60 pg 45 minutes after oral administration.

[0154] The graph of Figure 8C shows that sulfated BODIPY amide (37) can be detected in the plasma at the highest i.p. dose (60 pg). Sulfated BODIPY amide (37) is not detected in the plasma at the lower i.p. dose (10 pg) or at either oral dose although it is detect bound to immune cells in the blood and brain (Figures 3, 8A, and 8B). The detection of sulfated BODIPY amide (37) in plasma, which does not contain any cells, indicates free or "non-cell associated" compound 36.

[0155] Example 38 further reveals that sulfated BODIPY amide (37) can be detected on immune cells (CD4 effector memory helper T cells) in the blood when administered at 60 pg i.p. or orally (Figure 9B) for at least 12 hours (***p<0.001, **p<0.01, *p<0.05 by 2-way ANOVA) n = 4 mice per group. Example 38 also demonstrates that sulfated BODIPY amide (37) can be detected free in the plasma for up to 4 hours after i.p. administration of 60 ng (Figure 9A).

[0156] The graph of Figure 9A shows that sulfated BODIPY amide (37) is detected in the plasma up to but not over 4 hours after i.p. administration of 60 pg, which indicates that the "non-cell associated" compound 36 binds to cellular or extracellular constituents or is removed from the plasma by 4 hours post administration.

[0157] The graph of Figure 9B shows that sulfated BODIPY amide (37) is detected on immune cells (CD4 effector memory T cells) in the blood for at least 12 hours after i.p. administration of 60 pg, which indicates that although the "non-cell associated" sulfated BODIPY amide (37) is no longer detected, the cell-associated sulfated BODIPY amide (37) maintains a stable interaction. This finding is supported by Example 37 (Figure 1) which shows stable binding of sulfated BODIPY amide (37) to CD4 T cells for at least 24 hours in in vitro cell cultures. Example 38 (Figure 9B) shows that at later timepoints (4 and 12 hours) there is an increase in the number of sulfated BODIPY amide (37)+ effector memory CD4 T cells in the brain indicating recent trafficking into the CNS. These Examples (36 and 38) provide clear evidence that sulfated BODIPY amide (37) can bind immune cells and can cross the blood-brain barrier. Passive diffusion and attaching to migrating cells are possible mechanisms.

[0158] Overall, Example 38 shows that sulfated BODIPY amide (37) interacts with multiple cell types in vivo. Furthermore, this binding is specifically through the sulfated regions of sulfated BODIPY amide (37) as no non-specific binding was exhibited by unsulfated sulfated BODIPY amide (36). It can be delivered through i.p. and oral administration at several doses and while free sulfated BODIPY amide (37) is no longer detected within 4 hours of administration, sulfated BODIPY amide (37) forms stable interactions with immune cells that last over 12 hours in vivo. The use of sulfated BODIPY amide (37) provides insights into the mechanism in mouse models of MS disease.

[0159] Example 39 (Figure 4) shows that sulfated BODIPY amide (37) can be detected in the parenchyma of EAE mice 45 minutes after IP treatment. Brains from EAE mice treated with BODIPY labelled Tet-29, non-sulfated BODIPY amide (36) (60 pg / mouse; both green), or untreated were fixed in 4% PFA and sectioned in 20 pm sagittal sections. Sections were stained with collagen IV (red, blood vessels) and DAPI (blue, cell nuclei), and were imaged at 20X magnification by confocal microscopy. Figure 4 shows that non-sulfated BODIPY amide (36) is not detected in the brain parenchyma concluding that binding is dependent on highly sulfated saccharide dendrites. In these animals, sulfated BODIPY amide (37) was not associated with CNS blood vessels, and it preferentially bound neural and immune cell bodies. The detection of sulfated BODIPY amide (37) in the brain parenchyma within 45 minutes of IP administration also indicates that sulfated BODIPY amide (37) is able to enter the CNS and the entrance is sulfate-dependent. Example 40 (Figure 6) shows that sulfated BODIPY amide (37) binds to cells in a brain slice via its sulfated regions. Brain slices from healthy mice were stained with DAPI (blue, cell nuclei), collagen IV (red, blood vessels), and sulfated or non-sulfated sulfated BODIPY amide (green). After washing, brain sections were imaged at 20X magnification by confocal microscopy. Scalebar = 50 pm. Figure 6A is a control that shows that without sulfated BODIPY amide (37), no green fluorescence is detected in the brain. Figure 6B shows that when non-sulfated BODIPY amide (36) is added, no green fluorescence is detected in the brain slice. Thus, non-sulfated BODIPY amide (36) does not bind brain tissue. Figure 6C shows that when sulfated BODIPY amide (37) is incubated with brain tissue, green fluorescence can be detected in the brain slice. Therefore, compound 36 binds to brain tissue.

[0160] Example 40 (Figure 7) also shows that sulfated BODIPY amide (37) specifically binds cell bodies in healthy and EAE brains. Sagittal sections were taken from the brains of healthy and EAE mice and stained with DAPI (blue, cell nuclei), collagen IV (red, blood vessels), and sulfated BODIPY amide (37) (green). Images of the cerebral cortex (A, B, E, F) and cerebellum (C, D, G, H) were taken at 20x magnification.

[0161] Example 41 (Figure 5) shows that sulfated BODIPY amide (37) binds to bEnd.3 cells in a concentration-dependent manner. bEND.3 brain endothelial cells were seeded on a coverslip and incubated with different concentrations of sulfated (37) and unsulfated BODIPY amide (36). Binding of sulfated BODIPY amide (37) is dependent on its highly sulfated saccharide dendrites that interact with a range of proteins and ECM components. Both MFI (b) and area (c) show a concentration-dependant attachment of sulfated BODIPY amide (37), whereas the attachment of non-sulfated BODIPY amide (36) remains steady between concentrations. Results were analysed using a 2-way ANOVA with Sidak's multiple comparisons test ****p < 0.0001 and *p < 0.05 compared to unsulfated Tet-29. n = 3-4 technical replicates per group. This finding indicates that sulfated BODIPY amide (37) binds brain endothelial cells.

[0162] Example 43 describes the use of Magnetic Resonance Imaging (MRI) for detecting compounds labelled with fluorine-19. Fluorine-19 (F-19 or19F) is a natural non-radioactive isotope of fluorine that is essentially absent in biological tissues, making it an excellent contrast agent for biological probes. Examples of19F-tagged HS glycomimetics include compounds (20) and (24), which are suitable for biodistribution studies in vivo. As these compounds retain their pharmaceutical activity, their preferential localisation to tumour tissue may expand their utility as "theranostics", acting as both therapeutic drugs for the treatment of cancer and as probes for the detection of metastatic disease. As shown in Example 43, both compounds were detected down to 5 mM concentrations with linear signal-to-noise ratios (SNR). As expected, the densely19F-labelled compound (24) was more sensitive than compound (20). See Figures 12 and 13. These data confirm 19F-labelled mimetics can be successfully detected using MRI and demonstrate their potential for application as molecular probes, e.g. for cancer detection.

[0163] Figure 13 shows a tube containing 10 mM compound (24) placed beside a mouse cadaver with the ’■H anatomical scan (left) and19F signal overlaid (right). A strongi9F signal from compound (24) was detected with no19F signal from tissue. Reduction of background noise in19F scans to detect compounds internally when injected into tissues is anticipated. Imaging in live anaesthetised mice is expected to demonstrate the detection of novel19F- labelled molecular probes in vivo. Studies in normal healthy and tumour-bearing mice to assess their pharmacokinetics and whether they selectively accumulate in tumour tissue are expected to provide useful information on the biodistribution of compounds of the invention, which will be essential for further clinical development.

[0164] Glycomimetic compounds radio-labelled with fluorine-18 (F-18 or18F) are potentially useful imaging agents for biodistribution studies. Typically, one disaccharide arm of a glycomimetic compound is replaced with a hydrophobic silicon-fluoride acceptor (SiFA) group. This hydrophobic SiFA arm may act similarly to the cholestanol aglycone in PG545, which improves the inhibitor's efficacy and pharmacokinetics through its added lipophilicity. Fluorine-18 (F-18) can be readily introduced to the SiFA group using well-established isotopic exchange protocols to form the PET tracer. F-18 labelled heparanase inhibitors may have potential in diagnostic oncological PET imaging.

[0165] Formulations and administration

[0166] The compounds of the invention may be administered to a patient by a variety of routes, including orally, parenterally, by inhalation spray, topically, rectally, nasally, buccally or via an implanted reservoir. The compounds may also be administered by intracerebral, intracerebroventricular or intrathecal delivery. For parenteral administration, injections may be given intravenously, intra-arterially, intramuscularly or subcutaneously.

[0167] The amount of a compound of the invention to be administered to a patient will vary widely according to the nature of the patient and the nature and extent of the disorder to be treated. Typically, the dosage for an adult human will be in the range of about 0.01 pg / kg to about 1 g / kg, preferably about 0.01 mg / kg to about 100 mg / kg. The specific dosage required for any particular patient will depend upon a variety of factors, such as the patient's age, body weight, general health, gender and diet. Optimal doses will depend on other factors such as mode of administration and level of progression of the disease or disorder. Doses may be given once daily, or two or more doses may be required per day. For example, a dosage regime for an Alzheimer's patient might require one dose in the morning and one in the evening. Alternatively, a dosage regime for such a patient might require four hourly doses.

[0168] For oral administration the compounds can be formulated into solid or liquid preparations, for example tablets, capsules, granules, powders, solutions, suspensions, syrups, elixirs and dispersions. Such preparations are well known in the art as are other oral dosage regimes not listed here.

[0169] For parenteral administration, compounds of the invention can be formulated into sterile solutions, emulsions and suspension.

[0170] Compounds of the invention may be mixed with suitable vehicle and then compressed into the desired shape and size. The compounds may be tableted with conventional tablet bases such as lactose, sucrose and corn starch, together with a binder, a disintegration agent and a lubricant. The binder may be, for example, corn starch or gelatin, the disintegrating agent may be potato starch or alginic acid, and the lubricant may be magnesium stearate. For oral administration in the form of capsules, diluents such as lactose and dried corn starch may be employed. Other components such as colourings, sweeteners or flavourings may be added. Tablets, capsules or powders for oral administration may contain up to about 99% of a compound of the invention.

[0171] When liquid preparations are required for oral use, a compound of the invention may be combined with a pharmaceutically acceptable carriers such as water, an organic solvent such as ethanol, or a mixture of both, and optionally other additives such as emulsifying agents, suspending agents, buffers, preservatives, and / or surfactants may be used. Colourings, sweeteners or flavourings may also be added.

[0172] The compounds may also be administered by injection in a pharmaceutically acceptable diluent such as water or saline. The diluent may comprise one or more other ingredients such as ethanol, propylene glycol, an oil or a pharmaceutically acceptable surfactant.

[0173] The compounds of the invention may also be administered topically. Carriers for topical administration of the compounds include mineral oil, liquid petrolatum, white petrolatum, propylene glycol, polyoxyethylene, polyoxypropylene compound, emulsifying wax and water. The compounds may be present as ingredients in lotions or creams, for topical administration to skin or mucous membranes. Such creams may contain the active compounds suspended or dissolved in one or more pharmaceutically acceptable carriers. Suitable carriers include mineral oil, sorbitan monostearate, polysorbate 60, cetyl ester wax, cetearyl alcohol, 2-octyldodecanol, benzyl alcohol and water.

[0174] The compounds of the invention may further be administered by means of sustained release systems. For example, they may be incorporated into a slowly dissolving tablet or capsule.

[0175] EXAMPLES

[0176] The following examples further illustrate the invention. It is to be appreciated that the invention is not limited to the examples. Abbreviations

[0177] NMR nuclear magnetic resonance

[0178] HRMS high resolution mass spectrometry

[0179] ESI electrospray ionisation

[0180] TLC thin layer chromatography

[0181] RT room temperature

[0182] DCM dichloromethane

[0183] THF tetra hydrofuran

[0184] DMF dimethylformamide

[0185] BODIPY 4,4-difluoro-l,3-dimethyl-4-bora-3a,4a-diaza-s-indacene

[0186] PyBOP benzotriazol- 1-yloxytripyrrolidinophosphonium hexafluorophosphate

[0187] HATU (l-[bis(dimethylamino)methylene]-lH-l,2,3-triazolo[4,5- b]pyridinium 3-oxide hexafluorophosphate

[0188] General

[0189] Air sensitive reactions were performed under argon. Organic solutions were dried over anhydrous MgS04 and the solvents were evaporated under reduced pressure. Anhydrous and chromatography solvents were obtained commercially and used without any further purification. Thin layer chromatography was performed on glass or aluminium sheets coated with 60 F254 silica gel. Organic compounds were visualized under UV light or use of a dip of ammonium molybdate (5 wt%) and cerium(IV) sulfate 4 H2O (0.2 wt%) in aq. H2SO4 (2M). Chromatography (flash column, or an automated system with continuous gradient facility) was performed on silica gel (40-63 pm).XH NMR spectra were measured in CDCI3, CD3OD or D2O (HOD, 8 4.79), and13C NMR spectra in CDCI3 (centre line, 8 77.0), CD3OD (centre line, 8 49.0) or D2O (no internal reference, 8 1.47 where stated). Assignments ofXH and13C resonances were based on 2D (1H-1H DQF-COSY, 1H-13C HSQC, HMBC) and DEPT experiments. NMR abbreviations used: b, broad; s, singlet; d, doublet; t, triplet; m, multiplet. High resolution electrospray mass spectra (ESI-HRMS) were recorded on a Q-TOF Tandem Mass Spectrometer.

[0190] Glutaric acid 2a (55 mg, 0.42 mmol), ethyl 3-[2-amino-3-(3-ethoxy-3-oxo-propoxy)- 2-methylpropoxy]propanoate 1 (O'Donovan, L.; De Bank, P.A.; Org. Biomol. Chem. 2014, 12, 7290-7296 ) (50 mg, 1.6 mmol, 3.9 eq.) and HATU (440 mg, 1.16 mmol, 2.8 eq.) were brought under argon atmosphere. Then anhydrous DMF (5 mL) and DIPEA (400 pL, 2.3 mmol, 5.6 eq.) were added and the reaction was stirred at r.t. for 4 hours. Then the reaction mixture was diluted with EtOAc and washed with saturated NFUCI, water and brine. Then the organic phase was dried over MgS04 and concentrated in vacuo, before being purified via flash chromatography (0%-100% EtOAc in hex) to afford tetraethyl ester 3a (270 mg, 0.380 mmol, 92% yield) as a yellow oil. 1H NMR (500 MHz, CDCI3) 6 = 6.07 (s, 2H, NH), 4.13 (q, J = 7.1 Hz, 8H, C(10)H2), 3.69 (t, J = 6.3 Hz, 8H, C(7)H2), 3.54 (dd, J = 81.7, 9.1 Hz, 8H, C(6)H2), 2.53 (t, J = 6.3 Hz, 8H, C(8)H2), 2.16 (t, J = 7.2 Hz, 4H, C(2)H2), 1.86 (p, J = 7.2 Hz, 2H, C(l)H), 1.31 (s, 6H, C(5)H2), 1.25 (t, J = 7.2 Hz, 12H, C(11)H2). 13C{1H} NMR (125 MHz, CDCI3) 6 = 172.8 (C3), 171.7 (C9), 73.0 (C6), 66.9 (C7), 60.6 (CIO), 56.7 (C4), 36.2 (C8), 35.1 (C2), 22.0 (Cl), 19.2 (C5), 14.4 (Cll). HRMS (ESI-TOF) m / z calcd. for C33H58N2Oi4Na [M + Na]+: 729.3780, observed 729.3780.

[0191] Example 2: Maltose tetramer 6a

[0192] Tetraethyl ester 3a (255 mg, 0.361 mmol, 1.00 eq.) was dissolved in MeOH (2 mL) and NaOH (2.7 mL, 2.0 mol / L, 15 eq.) was added and the reaction was stirred at r.t. for 16 hours. Then the solvent was removed in vacuo and the material was co-evaporated with H2O once. The solid white material was dissolved in H2O (5 mL) and HCI (2.2 mL, 2.5 mol / L, 15 eq.) was added and the mixture was once more concentrated in vacuo and co-evaporated with acetone 3 times. The solid white material was re-suspended in acetone and filtered through a sinter funnel. The filtrate was concentrated in vacuo to afford tetra carboxylic acid 4a (214 mg, 0.361 mmol, 99% yield) as a cream coloured amorphous solid. The material was used without further purification.

[0193] Tetra carboxylic acid 4a (214 mg, 0.360 mmol, 1.00 eq.), PyBOP (930 mg, 1.80 mmol, 5.00 eq.) and maltose alkylamine 5 (950 mg, 2.16 mmol, 6.00 eq.) were brought under argon atmosphere before anhydrous DMF (10 mL) and DIPEA (0.5 mL, 3 mmol, 8 eq.) were added and the reaction was stirred at r.t. for 5 hours. The mixture was diluted in MeOH and concentrated in vacuo, co-evaporating 5 times with toluene. The viscous crude was dissolved in 50% H2O in ACN and purified by flash chromatography (0%-50% H2O in ACN) affording the maltose tetramer 6a (405 mg, 0.177 mmol, 49% yield) as a fluffy white solid after lyophilisation. 1H NMR (500 MHz, D2O) 6 = 5.45 (d, J = 3.9 Hz, 4H, C(1")H), 4.52 (d, J = 7.9 Hz, 4H C(l')H), 4.05 - 3.55 (m, 64H), 3.48 (t, J = 9.5 Hz, 4H), 3.36 (dd, J = 9.5, 8.0 Hz, 4H, C(2')H), 3.26 (t, J = 6.9 Hz, 8H, C(10)H2), 2.55 (t, J = 5.9 Hz, 8H, C(8)H2), 2.28 (t, J = 7.3 Hz, 4H, C(2)H2), 1.87 (p, J = 7.6 Hz, 2H, C(l)H), 1.69 (p, J = 6.8 Hz, 8H, C(14)H2), 1.58 (p, J = 7.1 Hz, 8H, C(11)H2), 1.49 - 1.38 (m, 16H, C(12)H2) and C(13)H2), 1.34 (s, 6H, C(5)H3). 13C{1H} NMR (125 MHz, D2O) 6 = 175.5 (C3), 173.9 (C9), 102.1 (Cl'), 99.7 (Cl"), 77.0, 76.3, 74.6, 73.1 (C2'), 72.9, 72.8, 72.3 (C6), 71.7, 70.5 (C15), 69.4, 67.6 (C7), 60.8, 60.6, 57.0 (C4), 39.4 (CIO), 36.3 (C8), 35.7 (C2), 28.8 (C14), 28.4 (Cll), 25.9 (C12 or 13), 24.8 (C12 or 13), 22.0 (Cl), 18.9 (C5). HRMS (ESI-TOF) m / z calcd. for C97H175N6O54 [M + H]+: 2288.1132, observed 2288.1155.

[0194] Maltose tetramer 6a (267 mg, 0.117 mmol, 1 eq.) and sulfur trioxide trimethylamine (2.39 g, 16.3 mmol, 140 eq.) were brought together under vacuum for 2 hours. Then the materials were placed under argon atmosphere before anhydrous DMF (10 mL) was added. The reaction was stirred for 60 hours at 60 °C then cooled to r.t., diluted with MeOH and concentrated in vacuo. The mixture was co-evaporated 5 times with toluene, dissolved in minimal MeOH and purified via flash chromatography (0%-50% H2O in ACN, both with 1% NH4OH (v / v)). The collected material was concentrated in vacuo and passed through an Amberlyst® Na+ ion exchange column to afford sulfated maltose tetramer 7a (583 mg, 0.113 mmol, 97% yield) as a fluffy white solid after lyophilisation. 1H NMR (500 MHz, D2O) 6 = 5.57 (d, J = 3.4 Hz, 4H, C(1")H), 4.94 (d, J = 4.5 Hz, 4H, C(l')H), 4.86 (t, J = 7.7 Hz, 4H, C(3")H)), 4.77 (t, J = 4.8 Hz, 4H, C(3')H), 4.58 (dd, J = 8.3, 3.4 Hz, 4H, C(2")H), 4.54 - 4.45 (m, 8H), C(2')H and C(4")H), 4.41 (d, J = 11.4 Hz, 4H, 1 / 2C(6")H2), 4.37 - 4.25 (m, 12H, 1 / 2C(6")H2 and C(6')H2), 4.24 - 4.16 (m, 8H, C(4')H) and C(5")H), 4.15 - 4.09 (m, 4H), C(5')H), 3.94 - 3.85 (m, 4H), 1 / 2C(15)H2), 3.77 - 3.71 (m, 8H, C(7)H2), 3.70 - 3.52 (m, 12H, 1 / 2C(15)H2 and C(6)H2), 3.17 (t, J = 7.0 Hz, 8H, C(10)H2), 2.49 (t, J = 6.0 Hz, 8H, C(8)H2), 2.21 (t, J = 7.6 Hz, 4H, C(2)H2), 1.78 (p, J = 7.7 Hz, 2H, C(1)H2), 1.62 (p, J = 7.1 Hz, 8H, C(14)H2), 1.51 (p, J = 7.2 Hz, 8H, C(11)H2),1 1.42 - 1.29 (m, 16H), C(12)H2 and C(13)H2), 1.26 (s, 6H, C(5)H3) 13C{1H} NMR (125 MHz, D2O) 6 = 175.7 (C3), 173.9 (C9), 99.9 (Cl'), 94.3 (Cl"), 77.4 (C3'), 76.3 (C2'), 75.0 (C3"), 73.6 (C2"), 73.4 (C4"), 72.3 (C5'), 72.2 (C6), 72.1 (C4'), 70.3 (C15), 70.1 (C5"), 67.9 (C6'), 67.6 (C7), 66.3 (C6"), 57.1 (C4), 39.6 (CIO), 36.2 (C8), 35.7 (C2), 28.6 (C14), 28.4 (Cll), 26.0 (C12 or 13), 24.9 (C12 or 13), 21.9 (Cl), 19.0 (C5). HRMS (ESI-TOF) m / z calcd. for C97Hi46N6Oi38Na2iS28 [M - 7 Na]7’: 711.4946 (40%), 711.6379 (50%), 711.7804 (90%), 711.9234 (90%), 712.0660 (100%), 712.2089 (80%), 712.3516 (70%), 712.4944 (50%), observed 711.5020 (40%), 711.6447 (50%), 711.7870 (90%), 711.9290 (90%), 712.0721 (100%), 712.2150 (80%), 712.3576 (70%), 712.4998 (50%).

[0195] Example 4: Tetraethyl ester 3b

[0196] 3-[2-Amino-3-(3-ethoxy-3-oxo-propoxy)-2-methylpropoxy]propanoate 1 (334 mg, 1 mmol) was dissolved in anhydrous DMF (3 mL), and the reaction mixture was stirred under argon at r.t. Adipic acid (40 mg, 0.273 mmol) and HATU (438 mg, 1 mmol, 4 eq.) were added followed by DIPEA (191 pL, 1 mmol, 4 eq.). The reaction mixture was stirred at r.t. for 17 hours. The mixture was diluted with EtOAc and washed with aq. ammonium chloride, then with water twice, dried over magnesium sulfate and concentrated. The residue was purified by flash chromatography (0 -100% EtOAc in hexane) to yield tetraethyl ester 3b (180 mg, 0.273 mmol, 91%) as a pale yellow oil. TLC (DCM : MeOH, 9: 1, v / v): Rf = 0.5.XH NMR (500 MHz, Chloroform-d) 6 4.15 (q, J = 7.1 Hz, 8H), 3.71 (t, J = 6.1 Hz, 8H), 3.61 (d, J = 9.1 Hz, 8H), 3.47 (d, J = 9.1 Hz, 8H), 2.56 (t, J = 6.1 Hz, 8H), 2.16 (t, J = 3.6 Hz, 2H), 1.63 - 1.53 (m, 2H), 1.34 - 1.23 (m, 16H).13C NMR (125 MHz, CDCI3) 6 172.4, 172.1, 72.8, 66.8, 60.8, 60.6, 56.6, 56.5, 36.5, 36.5, 34.8, 24.8, 18.6, 14.1. HRMS (ESI) calcd. for C34H6oN2Oi4Na [M + Na]+m / z, 743.3942; found, 743.3934.

[0197] A solution of tetraethyl ester 3b (72 mg, 0.243 mmol) in MeOH (1.5 mL) and 2M NaOH (1.5 mL) was stirred at 60 °C for 72 hours. After cooling to 0 °C, the reaction mixture was diluted with water (3 mL) and acidified with 2M HCI till pH~3. The solvent was removed in vacuo, and the aqueous layer was extracted with EtOAc twice, dried over magnesium sulfate and concentrated to afford tetra carboxylic acid 4b (60 mg, 0.099 mmol, 98%), that was used in the next step without further purification.

[0198] Tetramer carboxylic acid 4b (59 mg, 0.097 mmol, 1.00 eq.), PyBOP (250 mg, 0.480 mmol, 5.00 eq.) and maltose alkylamine 5 (340 mg, 0.780 mmol, 8.00 eq.) were brought under argon atmosphere before anhydrous DMF (10 mL) and DIPEA (0.13 mL, 0.78 mmol, 8.0 eq.) were added and the reaction was stirred at r.t. for 5 hours. The mixture was diluted in MeOH and concentrated in vacuo, co-evaporating 5x with toluene. The viscous crude was dissolved in 50% H2O in ACN and purified by flash chromatography (0%-50% H2O in ACN) affording maltose tetramer 6b (80 mg, 0.034 mmol, 36% yield) as a fluffy white solid after lyophilisation.XH NMR (500 MHz, D2O) 6 5.31 (d, J = 3.8 Hz, 4H), 4.37 (d, J = 7.9 Hz, 4H), 3.89 - 3.73 (m, 14H), 3.74 - 3.44 (m, 49H), 3.34 (t, J = 9.5 Hz, 16H), 3.21 (t, J = 9.5, 8.0 Hz, 8H), 3.11 (t, J = 6.9 Hz, 8H), 2.40 (t, J = 5.9 Hz, 8H), 2.18 - 2.11 (m, 8H), 1.60 - 1.40 (m, 24H), 1.35 - 1.22 (m, J = 5.3, 3.8 Hz, 22H), 1.20 (s, 6H).13C NMR (126 MHz, D2O) 6 176.1, 173.9, 102.1, 99.7, 77.0, 76.3, 74.6, 73.1, 72.9, 72.8, 72.3, 71.7, 70.5, 69.4, 67.6, 60.8, 60.6, 57.0, 39.4, 36.3, 36.2, 28.8, 28.4, 26.0, 24.8, 18.9. HRMS (ESI-TOF) m / z calcd. for C98H177N6O54 [M + H]+: 2302.1289, observed 2302.1274.

[0199] Maltose tetramer 6b (80 mg, 34.7 pmol) was dissolved in anhydrous DMF (4 mL), and the reaction mixture was stirred under argon at r.t. Sulfur trioxide trimethylamine complex (713 mg, 4.86 mmol, 140 eq.) was added, and the reaction mixture was stirred at 60 °C for 72 hours. MeOH (1 mL) was added, the mixture stirred for 30 min and concentrated in vacuo. Chromatography (acetonitrile : water : aq. ammonia, 6:2: 1 -> 5:2: l-> 4:2: 1 -> 3:2: 1) afforded a product as an ammonium salt. This was dissolved in water and passed through a Dowex 50WX8-200 (Na+) resin column. The desired product was eluted with water to furnish sulfated maltose tetramer 7b (151 mg, 34 pmol, 84%) as a sodium salt.XH NMR (500 MHz, D2O) 6 5.59 (d, J = 3.5 Hz, 4H), 4.97 (d, J = 4.2 Hz, 4H), 4.86 - 4.79 (m, 4H), 4.77 (t, J = 4.5 Hz, 4H), 4.56 (dd, J = 7.9, 3.8 Hz, 4H), 4.53 - 4.41 (m, 8H), 4.45 - 4.31 (m, 4H), 4.36 - 4.26 (m, 12H), 4.22 - 4.16 (m, 8H), 4.15 - 4.08 (m, 4H), 3.89 -3.84 (dd, J = 10.0, 6.8 Hz, 4H), 3.74 (t, J = 6.0 Hz, 8H), 3.73 - 3.61 (m, 12H), 3.56 - 3.19 (m, 12H), 2.49 (t, J = 5.9 Hz, 8H), 2.21 (d, J = 6.4 Hz, 4H), 2.09 - 1.98 (m, 4H), 1.63 - 1.55 (m, 8H), 1.59 - 1.48 (m, 16H), 1.44 - 1.33 (m, 8H).13C NMR (125 MHz, D2O) 6 175.20, 172.93, 118.44, 99.00, 93.16, 76.19, 75.19, 74.07, 72.61, 72.30, 71.15, 70.91, 69.12, 66.75, 66.55, 65.20, 55.99, 38.48, 35.28, 35.21, 27.55, 27.32, 24.91, 23.79, 17.91. HRMS (ESI-TOF) m / z calcd. for C98H177N6O54 [M + H]+: 2302.1289, observed 2302.1274. Example 7: Tetraethyl ester 3c

[0200] 3-[2-Amino-3-(3-ethoxy-3-oxo-propoxy)-2-methylpropoxy]propanoate 1 (636 mg, 2.08 mmol, 4 eq.) was dissolved in anhydrous DMF (3 mL), and the reaction mixture was stirred under argon at r.t. Dodecanedioic acid (120 mg, 0.521 mmol) and HATU (834 mg, 2.08 mmol, 4 eq.) were added followed by DIPEA (363 pL, 2.08 mmol, 4 eq.). The reaction mixture was stirred at r.t. for 17 hours. The mixture was diluted with EtOAc and washed with aq. ammonium chloride, then with water twice, dried over magnesium sulfate and concentrated. The residue was purified by flash chromatography (60% EtOAc in hexane) to yield tetraethyl ester 3c (390 mg, 0.521 mmol, 93%) as a yellow oil. TLC (EtOAc): Rf = 0.7.XH NMR (500 MHz, MeOD) 6 4.15 (q, J = 7.1 Hz, 8H), 3.71 (t, J = 6.1 Hz, 8H), 3.61 (d, J = 9.2 Hz, 8H), 3.47 (d, J = 9.2 Hz, 8H), 2.56 (t, J = 6.1 Hz, 8H), 2.13 (t, J = 7.6 Hz, 4H), 1.56 (t, J = 6.9 Hz, 10H), 1.33 - 1.23 (m, 16H).13C NMR (125 MHz, MeOD) 6 174.1, 172.1, 72.8, 66.7, 60.6, 56.5, 48.7, 38.4, 37.3, 34.9, 29.4, 29.2, 29.1, 25.7, 18.7, 14.1. HRMS (ESI) calcd. for C4oH72N2Oi4Na [M+Na]+m / z, 827.4881; found, 827.4883.

[0201] A solution of tetraethyl ester 3c (195 mg, 0.242 mmol) in MeOH (5 mL) and 2M NaOH (1.5 mL) was stirred at 60 °C for 72 hours. After cooling to 0 °C, the reaction mixture was diluted with water (3 mL) and acidified with 2M HCI till pH~3. The solvent was removed in vacuo, and the aqueous layer was extracted with EtOAc twice, dried over magnesium sulfate and concentrated. Crude tetra-acid was used in the next step without further purification. This material (82 mg, 2.08 mmol, 4 eq.) was dissolved in anhydrous DMF (2.5 mL), and the reaction mixture was stirred under argon at r.t. PyBOP (311 mg, 591 mmol, 5 eq.) was added and the reaction mixture was stirred for 20 min at r.t. Maltose alkylamine 5 (313 mg, 0.710 mmol, 6 eq.) was added followed by DIPEA (165 pL, 0.947 mmol, 8 eq.). The reaction mixture was stirred at r.t. for 17 hours, and then DMF was removed in vacuo. The residue was purified by flash chromatography (70% EtOH in water) to yield maltose tetramer 6c (175 mg, 0.118 mmol, 62%) as a solidified syrup.XH NMR (500 MHz, D2O) 6 5.32 (d, J = 4.0 Hz, 4H), 4.39

[0202] (d, J = 7.8 Hz, 4H), 3.90 - 3.83 (m, 12H), 3.82 - 3.70 (m, 40H), 3.69 - 3.52 (m, 8H), 3.54

[0203] - 3.47 (m, 4H), 3.34 (t, J = 9.3 Hz, 4H), 3.27 - 3.18 (m, 8H), 2.48 (t, J = 6.0 Hz, 8H), 2.19

[0204] - 2.11 (m, 6H), 1.60 - 1.54 (m, 22H), 1.33 - 1.20 (m, 28H) 1.12 (s, 6H).13C NMR (125 MHz,

[0205] D2O) 6 175.4, 173.7, 102.1, 99.7, 77.0, 76.3, 74.6, 73.1, 72.9, 72.7, 71.7, 70.4, 69.4, 62.5, 60.8, 60.5, 57.4, 39.8, 28.5, 25.5, 24.7, 16.8. HRMS (ESI) calcd. for CicwHissNeC^Na [M + Na]+m / z, 2408.2047; found, 2408.2053.

[0206] Maltose tetramer 6c (170 mg, 71.2 pmol) was dissolved in anhydrous DMF (5 mL), and the reaction mixture was stirred under argon at r.t. Sulfur trioxide trimethylamine complex (1.46 g, 9.97 mmol, 140 eq.) was added, and the reaction mixture was stirred at 60 °C for 72 hours. MeOH (1 mL) was added, the mixture stirred for 30 min and concentrated in vacuo. Chromatography (acetonitrile:water:aq. ammonia, 6:2: 1 -> 5:2: l-> 4:2: 1 -> 3:2: 1) afforded a product as an ammonium salt. This was dissolved in water and passed through a Dowex 50WX8-200 (Na+) resin column. The desired product was eluted with water to furnish sulfated maltose tetramer 7c (310 mg, 71.2 pmol, 83%) as a sodium salt.XH NMR (500 MHz, D2O) 6 5.60 (d, J = 3.5 Hz, 4H), 4.97 (d, J = 4.2 Hz, 4H), 4.85 - 4.74 (m, 4H), 4.61 - 4.52 (m, 4H), 4.48 (dd, J = 8.8, 6.9 Hz, 4H), 4.53 - 4.41 (m, 8H), 4.42 (dd, J = 11.2, 2.8 Hz, 4H), 4.40 - 4.30 (m, 12H), 4.22 - 4.10 (m, 8H), 4.0 - 3.95 (m, 4H), 3.90 (dd, J = 10.1, 6.8 Hz, 4H), 3.75 (t, J = 5.9 Hz, 8H), 3.72 - 3.61 (m, 12H), 3.56 - 3.19 (m, 12H), 2.50 (t, J = 5.9 Hz, 8H), 2.20 (d, J = 7.3 Hz, 8H), 2.09-1.97 (m, 16H), 1.63 -1.54 (m, 24H), 1.38-1.28 (m, 8H).13C NMR (125 MHz, D2O) 6 175.6, 173.1, 155.97, 100.06, 99.28, 94.23, 77.87, 77.43, 76.41, 76.35, 75.21, 75.09, 73.78, 73.61, 73.46, 73.36, 72.34, 72.07, 71.78, 70.30, 70.08, 69.67, 67.89, 67.82, 66.33, 47.01, 43.69, 42.89, 39.65, 35.63, 29.43, 28.76, 28.60, 28.40, 28.30, 26.59, 26.09, 25.19, 25.00, 24.92, 24.65. HRMS (ESI) calcd. for Cii2Hi6oF6N8Na280i4oS28 [M- 8Na]8“ m / z 635.4516; found: 635.4538.

[0207] Example 10: 0-Giutamic acid diethyl ester 10

[0208] 10

[0209] A solution of diethyl 3-oxoglutarate 8 (0.500 mL, 2.75 mmol) and ammonium acetate (3.00 g, 38.9 mmol) in methanol (10 mL) was stirred at r.t. over molecular sieves (3 A; ~1 g) for 18 h. The mixture was then acidified to ~ pH 3 by the addition of methanolic HCI (3 M, ~ 3.5 mL). Sodium cyanoborohydride (237 mg, 3.58 mmol) was then added in a single portion and the resulting mixture stirred at for a further 1 h. The mixture was then filtered over Celite, the cake washed with excess methanol, and the filtrate concentrated in vacuo. To the residual oil was added saturated aqueous NaHCOs (40 mL) and dichloromethane (40 mL) and the layers separated. The aqueous phase was further extracted with dichloromethane (40 mL), the combined organic extracts washed with brine, dried over MgSCh and concentrated in vacuo yielding p-glutamic acid diethyl ester as a colourless oil (542 mg, 2.67 mmol, 97%) which was used directly without further purification. Lit. J. Chem. Soc., Perkin Trans. 1, 1990, 2363-2369.

[0210] Example 11: Diethyl 3-(6-azidohexanamido)pentanedioate 11

[0211] 11

[0212] To a solution of 0-glutamic acid diethyl ester 10 (400 mg, 1.97 mmol) and PyBOP (1.50 g, 2.88 mmol) in DMF (5 mL) was added 6-azidohexanoic acid 9 (0.45 mL, 3.08 mmol) and DIPEA (1.40 mL, 8.18 mmol) and the resulting mixture stirred at r.t. for 16 h. Thereafter, the reaction mixture was concentrated in vacuo, ethyl acetate (30 mL) and saturated aqueous NH4CI (30 mL) and the layers separated. The organic phase was further washed with saturated aqueous NaHCO3 (30 mL) and brine (30 mL), dried over MgSCh and concentrated in vacuo to afford the crude material. Purification by flash column chromatography on silica gel eluting with ethyl acetate - light petroleum (0%^50% ethyl acetate) gave the diethyl ester 11 as a thick yellow oil (605 mg, 1.77 mmol, 90%). HRMS [ESI, M + Na]+Found: 365.1801 [C15H26N4O5 + Na]+requires 365.1804; 6H (500 MHz, methanol-d4) 4.58-4.53 (1 H, m, CH), 4.12 (4 H, q, J 7.1, 2 x CH2), 3.29 (2 H, t, J 6.9, CH2), 2.64-2.55 (4 H, m, 2 x CH2), 2.16 (2 H, t, J 7.4, CH2), 1.64-1.56 (4 H, m, 2 x CH2), 1.41-1.35 (2 H, m, CH2), 1.25 (6 H, t, J 7.1, 2 x Me); 6c (125 MHz, methanol-d4) 175.3 (C=O), 172.4 (2 x C=O), 61.7 (2 x CH2), 52.3 (CH2), 45.0 (CH), 40.0 (2 x CH2), 36.8 (CH2), 29.6 (CH2), 27.2 (CH2), 26.4 (CH2), 14.5 (2 x Me).

[0213] Example 12: 3-(6-Azidohexanamido)pentanedioic acid 12

[0214] To a solution of diethyl 3-(6-azidohexanamido)pentanedioate 11 (647 mg, 1.89 mmol) in methanol (10 mL) was added 2 M aqueous NaOH (2.2 mL) and the resulting mixture stirred at r.t. for 2 h. Methanol was then removed in vacuo and the resulting solution acidified to ~ pH 2-3 using 2 M aqueous HCI then diluted with water (45 mL). Ethyl acetate (45 mL) was then added, the layers separated, and the aqueous layer further extracted with ethyl acetate (2 x 45 mL). The combined organic extracts were dried over MgSChand concentrated in vacuo supplying 3-(6-azidohexanamido)pentanedioic acid 12 as a white solid (493 mg, 1.72 mmol, 91%).

[0215] Example 13: Tetraethyl ester azide 13

[0216] 3-(6-Azidohexanamido)pentanedioic acid was subsequently solved in DMF (8 mL) and DIPEA (0.65 mL, 3.80 mmol) and HATU (1.469 g, 3.86 mmol) added successively. To the resultant bright orange solution was added diethyl ester 1 (1.118 g, 3.66 mmol) as a solution in DMF (7.5 mL), and stirring continued at r.t. for 2 h. Thereafter, ethyl acetate (100 mL) and saturated aqueous NaHCCh (100 mL) were added, the layers separated, and the organic phase further washed with brine (100 mL), dried over MgSCh, and concentrated in vacuo. Purification of the crude material by flash column chromatography on silica gel eluting with DCM - methanol (0^10% methanol) gave the tetraethyl ester azide 13 as a golden oil (1.288 g, l.50 mmol, 88%). HRMS [ESI, M + Na]+Found: 883.4633 [C39H68N6O15 + Na]+requires 883.4640; 6H (500 MHz, CDCI3) 7.04 (1 H, d, J 8.0, NH), 6.44 (2 H, s, 2 x NH), 4.41-4.29 (1 H, m, CH), 4.13 (8 H, q, J 7.1, 4 x CH2), 3.70 (8 H, t, J 6.3, 4 x CH2), 3.63-3.46 (8 H, m, 4 x CH2), 3.25 (2 H, t, J 6.9, CH2), 2.56-2.50 (10 H, m, 4 x CH2, 2 x1 / 2CH2), 2.29 (2 H, dd, J 14.0, 6.9, 2 x1 / 2CH2), 2.16 (2 H, t, J 7.6, CH2), 1.67-1.57 (4 H, m, 2 x CH2), 1.43-1.36 (2 H, m, CH2), 1.30 (6 H, s, 2 x Me), 1.25 (12 H, t J 7.1, 4 x Me); 6c (125 MHz, CDCI3) 172.1 (C=O), 171.7 (4 x C=O), 171.1 (2 x C=O), 72.83 (2 x CH2), 72.76 (2 x CH2), 67.0 (4 x CH2), 60.7 (4 x CH2), 57.0 (2 x C), 51.4 (CH2), 45.0 (CH), 40.0 (2 x CH2), 36.6 (CH2), 35.1 (4 x CH2), 28.7 (CH2), 26.5 (CH2), 25.2 (CH2), 19.3 (2 x Me), 14.4 (4 x Me).

[0217] To a solution of tetraethyl ester azide 13 (200 mg, 0.232 mmol) in methanol (4 mL) was added 2 M aqueous NaOH (1.4 mL) and the resulting mixture stirred at r.t. for 2 h. Methanol was then removed in vacuo and the resulting solution acidified to ~ pH 2-3 using 2 M aqueous HCI then diluted with water (30 mL). Ethyl acetate (30 mL) was then added, the layers separated, and the aqueous layer further extracted with ethyl acetate (2 x 30 mL). The combined organic extracts were dried over MgSChand concentrated in vacuo supplying tetracarboxylic acid azide 14 as a colourless film (169 mg, 0.226 mmol, 97%). 1H NMR (500 MHz, MeOD) 6 = 4.45 (p, J = 6.7 Hz, 1H, C(7)H2), 3.70 (t, J = 6.1 Hz, 8H, C(13)H2), 3.63 - 3.50 (m, 8H, C(12)H2), 3.29 (d, J = 6.9 Hz, 2H, C(1)H2), 2.54 (t, J = 6.1 Hz, 8H, C(14)H2), 2.41 (d, J = 6.8 Hz, 4H, C(8)H2), 2.20 (t, J = 7.6 Hz, 2H, C(5)H2), 1.68 - 1.56 (m, 4H, C(2)H2 and C(4)H2), 1.46 - 1.38 (m, 2H, C(3)H2), 1.30 (s, 6H, C(11)H3). 13C{1H} NMR (125 MHz, MeOD) 6 = 175.8 (C15), 175.1 (C6), 172.7 (C9), 73.65 (C12), 72.59 (C12), 68.2 (C13), 58.3 (CIO), 52.3 (Cl), 46.4 (C7), 42.0 (C8), 37.1 (C5), 35.9 (C14), 29.6 (C2), 27.4 (C3), 26.4 (C4), 19.5 (Cll). HRMS (ESI) calcd. for CsiHssNeOisNa [M + Na]+m / z, 771.3388; found, 771.3382. Example 15: Maltose tetramer azide 15

[0218] Tetra-carboxylic acid azide 14 was dissolved in DMF (4 mL) and DIPEA (0.31 mL, 1.81 mmol) and HATU (447 mg, 1.14 mmol) added successively. The resultant mixture was stirred at r.t. for 15 mins, during which time a dark brown colour gradually developed. This mixture was then added in a single portion to a solution of maltose alkylamine 5 (O.V. Zubkova et al., ACS Chem. Biol., 2018, 13 (12), 3236-3242) (565 mg, 1.28 mmol) in DMF (4 mL) and stirring continued for 4 h. Following the addition of methanol (~2 mL), the mixture was concentrated in vacuo to give the crude material as an orange oil, which was purified by flash column chromatography on silica gel eluting with acetonitrile - water (0->45% water) delivering the maltose tetramer azide 15 as a glassy white solid (364 mg, 0.149 mmol, 66%). HRMS [ESI, M + 2H]2+Found: 1221.6053 [C103H184N10O55 + 2H]2+requires 1221.6033; 6H (500 MHz, D2O) 5.41 (4 H, d, J 3.9, 4 x CH), 4.51-4.48 (1 H, m, CH), 4.47 (4 H, d, J 8.0, 4 x CH), 3.96-3.54 (64 H, m, 24 x CH + 20 x CH2), 3.44 (4 H, t, J 9.5, 4 x CH), 3.36 (2 H, t, J 6.8, CH2), 3.31 (4 H, dd, J 9.4, 8.1, 4 x CH), 3.21 (8 H, t, J 6.9, 4 x CH2), 2.50 (8 H, t, J 5.7, 4 x CH2), 2.47- 2.40 (4 H, m, 2 x CH2), 2.24 (2 H, d, J 7.5, CH2), 1.68-1.38 (38 H, m, 19 x CH2), 1.28 (6 H, s, 2 x Me); 13C{1H} NMR 6c (125 MHz, D2O) 175.5 (C=O), 173.8 (4 x C=O), 172.0 (2 x C=O), 102.1 (4 x CH), 99.6 (4 x CH), 76.9 (4 x CH), 76.3 (4 x CH), 74.6 (4 x CH), 73.1 (4 x CH), 72.9 (4 x CH), 72.7 (4 x CH), 72.15 (2 x CH2), 72.06 (2 x CH2), 71.7 (4 x CH), 70.5 (4 x CH2), 69.3 (4 x CH), 67.5 (4 x CH2), 60.8 (4 x CH2), 60.5 (4 x CH2), 57.1 (2 x C), 51.0 (CH2), 45.0 (CH), 41.3 (2 x CH2), 39.4 (4 x CH2), 36.3 (4 x CH2), 35.8 (CH2), 28.7 (4 x CH2), 28.4 (4 x CH2), 27.8 (CH2), 25.9 (4 x CH2), 25.7 (CH2), 25.1 (CH2), 24.8 (4 x CH2), 18.7 (2 x Me). Example 16: Maltose tetramer amine 16

[0219] To maltose tetramer azide 15 (100 mg, 0.041 mmol) in methanokwater (1: 1, 5 mL) was added Raney Nickel (50% solution in water, ~0.1 mL) and the resulting mixture stirred under an atmosphere of hydrogen and stirred at r.t. for 16 h. The reaction mixture was then filtered over Celite, the cake washed with methanol: water (1 : 1, 10 mL) and the filtrate concentrated in vacuo to yield maltose tetramer amine 16 as a fluffy white solid (97 mg, 0.040 mmol, >98%). HRMS [ESI, M + 2H]2+Found: 1208.6071 [CiosHiseNsOss + 2H]2+requires 1208.6080; 6H (500 MHz, D2O) 5.41 (4 H, d, J 3.9, 4 x CH), 4.51-4.47 (1 H, m, CH), 4.47 (4 H, d, J 8.0, 4 x CH), 3.96-3.54 (64 H, m, 24 x CH + 20 x CH2), 3.43 (4 H, t, J 9.5, 4 x CH), 3.31 (4 H, dd, J 9.4, 8.1, 4 x CH), 3.21 (8 H, t, J 6.9, 4 x CH2), 2.74 (2 H, t, J 7.3, CH2), 2.50 (8 H, t, J 5.8, 4 x CH2), 2.47-2.39 (4 H, m, 2 x CH2), 2.24 (2 H, t, J 7.5, CH2), 1.67-1.33 (38 H, m, 19 x CH2), 1.28 (6 H, s, 2 x Me); 6c (125 MHz, D2O) 175.7 (C=O), 173.9 (4 x C=O), 172.1 (2 x C=O), 102.1 (4 x CH), 99.7 (4 x CH), 77.0 (4 x CH), 76.3 (4 x CH),

[0220] 74.6 (4 x CH), 73.1 (4 x CH), 72.9 (4 x CH), 72.8 (4 x CH), 72.2 (2 x CH2), 72.1 (2 x CH2),

[0221] 71.7 (4 x CH), 70.5 (4 x CH2), 69.4 (4 x CH), 67.6 (4 x CH2), 60.8 (4 x CH2), 60.6 (4 x CH2), 57.1 (2 x C), 45.0 (CH), 41.3 (2 x CH2), 40.2 (CH2) 39.4 (4 x CH2), 36.3 (4 x CH2), 35.9 (CH2), 30.2 (CH2), 28.8 (4 x CH2), 28.4 (4 x CH2), 26.0 (4 x CH2), 25.7 (CH2), 25.3 (CH2),

[0222] 24.8 (4 x CH2), 18.7 (2 x Me).

[0223]

[0224] Maltose tetramer azide 15 (211 mg, 86.3 pmol) was dissolved in anhydrous DMF (4 mL), and the reaction mixture was stirred under argon at r.t. Sulfur trioxide trimethylamine complex (1.7 g, 12 mmol, 140 eq.) was added, and the reaction mixture was stirred at 60°C for 72 hours. MeOH (1 mL) was added, the mixture stirred for 30 min and concentrated in vacuo. Chromatography (acetonitrile:water:aq. ammonia, 6:2: 1 -> 5:2: l-> 4:2: 1 -> 3:2: 1) afforded a product as an ammonium salt. This was dissolved in water and passed through a Dowex 50WX8-200 (Na+) resin column. The desired product was eluted with water to furnish sulfated maltose tetramer azide 17 (365 mg, 86 pmol, 90%) as a sodium salt.XH NMR (500 MHz, D2O) 6 5.58 (d, J = 3.5 Hz, 4H), 4.96 (d, J = 4.3 Hz, 4H), 4.82 (q, J = 7.5 Hz, 4H), 4.76 (t, J = 4.6 Hz, 4H), 4.95 - 4.8 (m, 4 H), 4.55 (dd, J = 13.1, 3.9 Hz, 4H), 4.52 - 4.39 (m, 10H), 4.31 - 4.09 (m, 28H), 3.89 (dd, J = 10.1, 6.8 Hz, 4H), 3.81-3.73 (m, 8H), 3.78 - 3.59 (m, 12H), 3.50 (d, J = 9.7 Hz, 4H), 3.33 (t, J = 6.8 Hz, 8H), 3.18 (t, J = 7.0 Hz, 2H), 2.48 (t, J = 5.9 Hz, 8H), 2.52 - 2.36 (m, 4H), 2.28 - 2.22 (t, J = 7.4 Hz, 4H), 2.07 (s, 4H), 1.56 - 1.36 (m, 50H), 1.25 (s, 6H).13C NMR (126 MHz, D2O) 6 177.1, 172.9, 171.2, 99.1, 93.2,

[0225] 76.3, 75.3, 74.1, 72.6, 72.3, 71.2, 71.0, 69.2, 69.2, 66.9, 66.6, 65.3, 56.2, 50.0, 40.6, 38.6,

[0226] 35.3, 34.9, 27.6, 27.4, 26.9, 25.0, 24.8, 24.1, 23.9, 17.8, 0.0. HRMS (ESI) calcd. for Cii2Hi6oF6N8Na280i4oS28 [M- 8Na]8“ m / z 635.4516; found: 635.4538.

[0227] Example 18: Sulfated maltose tetramer amine 18

[0228] To sulfated maltose tetramer azide 17 (40 mg, 0.008 mmol) in water:THF (1: 1, 6 mL) was added aqueous ammonium hydroxide (28% w / w, 0.6 mL) followed by Pd(0H)2 / C (20% Pd w / w, 10 mg) and the resulting mixture stirred under an atmosphere of hydrogen for 25 h. The reaction mixture was then filtered over celite, the cake washed with water:THF (10 mL) and the filtrate concentrated in vacuo to provide the sulfated maltose tetramer amine 18 as a fluffy white solid (38 mg, 0.007 mmol, 95%). HR.MS [ESI, M - 8Na]8Found : [Ci03Hi58N8Na28Oi39S28 - 8Na]8~ requires; 6H (500 MHz, D2O) 5.63 (4 H, d, J 3.4, 4 x CH), 5.00 (4 H, d, J 4.3, 4 x CH), 4.90-4.87 (4 H, m, 4 x CH), 4.82-4.79 (4 H, 4 x CH), 4.62 (4 H, dd, J 8.1, 3.3, 4 x CH), 4.59-4.17 (37 H, m, 21 x CH, 8 x CH2), 3.97-3.92 (4 H, m, 2 x CH2), 3.79-3.55 (20 H, m, 10 x CH2), 3.23 (8 H, t, J 6.9, 4 x CH2), 3.07 (2 H, t, J 1.1, CH2), 2.54- 2.41 (12 H, m, 6 x CH2), 2.28 (2 H, t, J 7.7, CH2), 1.77-1.34 (38 H, m, 19 x CH2), 1.29 (6 H, s, 2 x Me); 6c (125 MHz, D2O) 175.4 (C=O), 173.9 (4 x C=O), 172.2 (2 x C=O), 100.0 (4 x CH), 94.1 (4 x CH), 77.4 (4 x CH), 76.3 (4 x CH), 75.0 (4 x CH), 73.5 (4 x CH), 73.3 (4 x CH), 72.2 (4 x CH), 72.1 (2 x CH2), 72.0 (2 x CH2), 71.9 (4 x CH), 70.2 (4 x CH, 4 x CH2), 67.8 (4 x CH2), 67.6 (4 x CH2), 66.2 (4 x CH2), 57.1 (2 x C), 44.9 (CH), 41.5 (2 x CH2), 39.52 (4 x CH2), 39.47 (CH2), 36.2 (4 x CH2), 35.6 (CH2), 28.6 (4 x CH2), 28.3 (4 x CH2), 26.5 (CH2), 26.0 (4 x CH2), 25.3 (CH2), 25.0 (CH2), 24.8 (4 x CH2), 18.7 (2 x Me).

[0229] - 53 -

[0230] SUBSTITUTE SHEET (RULE 26)

[0231] To a solution of commercially available 3,5-bis(trifluoromethyl)benzoic acid (10.2 mg, 0.040 mmol) in DMF (0.5 mL) was added HATU (13.4 mg, 0.035 mmol) and DIPEA (9 pL, 0.051 mmol) and the resulting mixture stirred at r.t. for 10 mins before the dropwise addition of maltose tetramer amine 16 (47 mg, 0.019 mmol) as a solution in DMF (1 mL). After stirring at r.t. for 4 h, DMF was removed in vacuo and the resulting crude material purified by flash column chromatography on silica gel eluting with acetonitrile - water (0->50% water) affording maltose tetramer bis(trifluoromethyl)benzamide 19 as a fluffy white solid (12.0 mg, 0.005 mmol, 23%). HRMS [ESI, M + 2H]2+Found: 1328.6074 [CmHissFeNsOse + 2H]2+requires 1328.6085; 6H (600 MHz, D2O) 8.29 (2 H, s, 2 x ArH), 8.27 (1 H, s, ArH), 5.35 (4 H, d, J 3.9, 4 x CH), 4.44-4.40 (1 H, m, CH), 4.39 (4 H, d, J 8.0, 4 x CH), 3.89-3.44 (64 H, m, 24 x CH + 20 x CH2), 3.39 (2 H, t, J 7.0, CH2), 3.37 (4 H, t, J 9.4, 4 x CH), 3.24 (4 H, dd, J 9.4, 8.1, 4 x CH), 3.21 (8 H, td, J 6.9, 2.2, 4 x CH2), 2.42 (8 H, t, J 5.8, 4 x CH2), 2.39-2.32 (4 H, m, 2 x CH2), 2.19 (2 H, t, J 7.3, CH2), 1.64-1.29 (38 H, m, 19 x CH2), 1.19 (6 H, s, 2 x Me); 6c (150 MHz, D2O) 175.6 (C=O), 173.8 (4 x C=O), 172.0 (2 x C=O), 167.2 (C=O),

[0232] 135.8 (C), 131.5 (q, J 33.5, 2 x C), 127.8 (2 x CH), 125.8 (CH), 123.1 (q, J 272.3, 2 x C) 102.1 (4 x CH), 99.6 (4 x CH), 76.8 (4 x CH), 76.3 (4 x CH), 74.5 (4 x CH), 73.0 (4 x CH),

[0233] 72.8 (4 x CH), 72.7 (4 x CH), 72.1 (2 x CH2), 72.0 (2 x CH2), 71.7 (4 x CH), 70.5 (4 x CH2), 69.3 (4 x CH), 67.5 (4 x CH2), 60.7 (4 x CH2), 60.5 (4 x CH2), 57.0 (2 x C), 45.0 (CH), 41.3 (2 x CH2), 40.0 (CH2), 39.4 (4 x CH2), 36.2 (4 x CH2), 35.8 (CH2), 28.7 (4 x CH2), 28.4 (4 x CH2), 28.2 (CH2), 25.9 (4 x CH2), 25.8 (CH2), 25.2 (CH2), 24.8 (4 x CH2), 18.6 (2 x Me); 6F (564 MHz, D2O) -62.6 (2 x CF3).

[0234] To maltose tetramer bis(trifluoromethyl)benzamide 19 (9.9 mg, 0.004 mmol) and sulfur trioxide trimethylamine complex (76 mg, 0.546 mmol) was added DMF (0.6 mL) and toluene (0.4 mL) and the resulting mixture heated to 60 °C and stirred for 24 h. After cooling to r.t., the reaction mixture was diluted with aqueous ammonium hydroxide (5% w / w, 2 mL) and concentrated in vacuo. The crude material was subsequently solved in water (~5 mL) and transferred into a dialysis cassette (MWCO 1000 Da). The cassette was placed in 1 L dialysate solution of ammonium bicarbonate (7 pM) and the dialysate was changed at the following time intervals: 1, 2, 3, 16 and 2 hours. The sample was then removed from the cassette, passed through an Amberlyst™ exchange resin (Na+form), and lyophilised to afford sulfated maltose tetramer bis(trifluoromethyl)benzamide 20 as a fluffy white solid (18.2 mg, 0.003 mmol, 89%). HRMS [ESI, M - 8Na]8’ Found: 666.1967 [CmHieoFeNsNazsO^oSzs - 8Na]8“ requires 666.1961; 6H (600 MHz, D2O) 8.31 (2 H, s, 2 x ArH), 8.30 (1 H, s, ArH), 5.56 (4 H, d, J 3.4, 4 x CH), 4.92 (4 H, d, J 4.5, 4 x CH), 4.84-4.76 (4 H, 4 x CH)*, 4.74 (4 H, t, J 4.9, 4 x CH), 4.56 (4 H, dd, J 7.9, 3.4, 4 x CH), 4.50-4.46 (9 H, m, 9 x CH), 4.39 (4 H, dd, J 11.2, 2.7, 4 x CH), 4.33-4.09 (24 H, m, 8 x CH + 8 x CH2), 3.88-3.84 (4 H, m, 2 x CH2), 3.72-3.45 (20 H, m, 10 x CH2), 3.42 (2 H, t, J 6.9, CH2), 3.13 (8 H, td, J 7.0, 3.0, 4 x CH2), 2.44 (8 H, t, J 5.8, 4 x CH2), 2.41-2.33 (4 H, m, 2 x CH2), 2.21 (2 H, t, J 7.4, CH2), 1.66-1.26 (38 H, m, 19 x CH2), 1.20 (6 H, s, 2 x Me); 6c (150 MHz, D2O) 175.7 (C=O), 173.8 (4 x C=O), 172.2 (2 x C=O), 167.6 (C=O), 135.9 (C), 131.5 (q, J 33.7, 2 x C), 127.9 (2 x CH), 125.9 (CH), 123.1 (q, J 272.4, 2 x C), 100.0 (4 x CH), 94.1 (4 x CH), 77.5 (4 x CH), 76.3 (4 x CH), 75.0 (4 x CH), 73.4 (4 x CH), 73.2 (4 x CH), 72.13 (4 x CH), 72.05 (2 x CH2), 71.93 (2 x CH2), 71.86 (4 x CH), 70.23 (4 x CH), 70.18 (4 x CH2), 67.8 (4 x CH2), 67.5 (4 x CH2), 66.2 (4 x CH2), 57.1 (2 x C), 44.9 (CH), 41.5 (2 x CH2), 40.0 (CH2), 39.5 (4 x CH2), 36.2 (4 x CH2), 35.8 (CH2), 28.6 (4 x CH2), 28.3 (4 x CH2), 28.1 (CH2), 26.0 (4 x CH2), 25.8 (CH2), 25.2 (CH2), 24.8 (4 x CH2), 18.6 (2 x Me); 6F (564 MHz, D2O) -62.6 (2 x CF3).

[0235] *chemical shift obscured by residual solvent peak, determined via HSQC.

[0236] Example 21: Tris(perfluoro-t-butoxy) carboxylic acid 22

[0237] To tris(perfluoro-t-butoxy) alcohol 21 (Jiang, Z-X. and Yu, Y.B., Tetrahedron 2007, 63, 3982-3988) (120 mg, 0.152 mmol) in acetone (4.8 mL) at 0 °C was added Jones reagent (2.5 M, 0.15 mL) and the resulting mixture slowly left to warm to r.t. over 18 h. Ethyl acetate (10 mL) and water (10 ml) were then added and the phases separated. The aqueous phase was further extracted with ethyl acetate, the combined organic extracts dried over MgSCh, and concentrated in vacuo to afford the carboxylic acid 22 as a white solid (115 mg, 0.143 mmol, 94%). HRMS [ESI, M + CF3O2Na - H]“ Found: 938.9540 [CizHyF^Os + CF3O2Na - H]“ requires 938.9532; 6H (500 MHz, acetone-de) 4.47 (2 H, s, CH2); 6c (125 MHz, acetone-de) 169.8 (C=O), 121.2 (q, J 292.3, 9 x C), 81.2-80.2 (m, 3 x C), 66.6 (3 x CH2), 54.1 (C); 6F (470 MHz, D2O) -71.1 (9 x CF3). To a solution of tris(perfluoro-t-butoxy) carboxylic acid 22 (17.9 mg, 0.022 mmol) in DMF:THF (1 : 1, 1 mL) was added HATU (23.8 mg, 0.024 mmol) and DIPEA (7 pL, 0.037 mmol) and the resulting mixture stirred at r.t. for 10 mins. This mixture was then added dropwise to a solution of maltose tetramer amine 16 (36.0 mg, 0.015 mmol) in DMF:THF (2: 1, 1.5 mL). After stirring at 40 °C for 4 h, concentration in vacuo afforded the crude material, which was purified by flash column chromatography on silica gel eluting with acetonitrile - water (0->50% water) affording the amide 23 as a fluffy white solid (30.0 mg, 0.009 mmol, 63%). HRMS [ESI, M + 2Na]2+Found : 1624.0745 [C120H191F27N8O59 + 2Na]2+requires 1624.0795; 6H (500 MHz, methanol-d4) 5.17 (4 H, d, J 3.8, 4 x CH), 4.44 (1 H, quint, J 6.8, CH), 4.34 (6 H, s, 3 x CH2), 4.28 (4 H, d, J 7.8, 4 x CH), 3.92-3.50 (56 H, m, 16 x CH, 20 x CH2), 3.45 (4 H, dd, J 9.7, 3.8, 4 x CH), 3.37 (4 H, ddd, J 9.5, 4.5, 2.0, 4 x CH), 3.30-3.15 (18 H, m, 8 x CH, 5 x CH2), 2.45-2.40 (12 H, m, 6 x CH2), 2.18 (2 H, t, J 7.6, CH2), 1.67-1.29 (38 H, m, 19 X CH2), 1.29 (6 H, s, 2 X Me); 6c (125 MHz, methanol-d4), 175.1 (C=O), 173.8 (4 x C=O), 172.6 (2 x C=O), 169.1 (C=O), 121.5 (q, J 292.8, 9 x C), 104.3 (4 x CH), 102.9 (4 x CH), 81.3 (4 x CH), 80.9 (3 x C)*, 77.9 (4 x CH), 76.6 (4 x CH), 75.1 (4 x CH), 74.8 (4 x CH), 74.7 (4 x CH), 74.2 (4 x CH), 73.6 (4 x CH2), 71.5 (4 x CH), 70.8 (4 x CH2), 68.7 (4 x CH2), 66.7 (3 x CH2), 62.8 (4 x CH2), 62.2 (4 x CH2), 58.2 (2 x C), 54.5 (C), 46.5 (CH), 42.2 (2 x CH2), 41.0 (CH2), 40.4 (4 x CH2), 37.6 (4 x CH2), 37.2 (CH2), 30.7 (4 x CH2), 30.5 (4 x CH2), 30.2 (CH2), 27.8 (5 x CH2), 26.8 (4 x CH2), 26.7 (CH2), 19.7 (2 x Me); 6F (470 MHz, methanol-d4) -71.4 (9 x CF3). *chemical shift determined via HMBC

[0238] To maltose tetramer tris(perfluoro-t-butoxy)amide 23 (30.0 mg, 0.009 mmol) and sulfur trioxide trimethylamine complex (192 mg, 1.31 mmol) was added DMF (1.5 mL) and toluene (1.5 mL) and the resulting mixture heated to 60 °C and stirred for 50 h. After cooling to r.t., the reaction mixture was diluted with aqueous ammonium hydroxide (5% w / w, 2 mL) and concentrated in vacuo. The crude material was subsequently dissolved in water (~10 mL) and transferred into a dialysis cassette (MWCO 1000 Da). The cassette was placed in 1 L dialysate solution of ammonium bicarbonate (7 pM) and the dialysate was changed at the following time intervals: 1, 2, 3, 16 and 2 hours. The sample was then removed from the cassette, passed through an Amberlyst™ exchange resin (Na+form), and lyophilised to afford the sulfated amide 24 as a fluffy white solid (52.7 mg, 0.009 mmol, 93%). HRMS [ESI, M - 8Na]8“ Found: 734.4438 [Ci2oHi63F27N8Na280i43S28 - 8Na]8’ requires 734.4430; 6H (500 MHz, D2O) 5.63 (4 H, d, J 3.4, 4 x CH), 5.00 (4 H, d, J 4.4, 4 x CH), 4.91-4.88 (4 H, m, 4 x CH), 4.82-4.79 (4 H, 4 x CH)*, 4.63 (4 H, dd, J 8.0, 3.4, 4 x CH), 4.58-4.52 (9 H, m, 9 x CH), 4.48-4.15 (34 H, m, 12 x CH + 11 x CH2), 3.97-3.91 (4 H, m, 2 x CH2), 3.77 (8 H, t, J 6.0, 4 x CH2), 3.74-3.53 (12 H, m, 6 x CH2), 3.27-3.22 (2 H, m, CH2), 3.22 (8 H, t, J 6.9, 4 x CH2), 2.53 (8 H, t, J 5.8, 4 x CH2), 2.49-2.40 (4 H, m, 2 x CH2), 2.23 (2 H, t, J 7.6, CH2), 1.69-1.32 (38 H, m, 19 x CH2), 1.28 (6 H, s, 2 x Me); 6c (125 MHz, D2O) 175.6 (C=O), 173.7 (4 x C=O),

[0239] 172.2 (2 x C=O), 169.8 (C=O), 119.9 (q, J 292.6, 9 x C), 100.0 (4 x CH), 94.1 (4 x CH), 79.0 (3 x C)**, 77.4 (4 x CH), 76.3 (4 x CH), 75.0 (4 x CH), 73.5 (4 x CH), 73.2 (4 x CH),

[0240] 72.2 (4 x CH), 72.1 (2 x CH2), 71.9 (4 x CH, 2 x CH2), 70.2 (4 x CH, 4 x CH2), 67.8 (4 x CH2), 67.6 (4 x CH2), 66.2 (4 x CH2), 65.0 (3 x CH2), 57.1 (2 x C), 53.1 (C), 44.9 (CH), 41.6 (2 x CH2), 39.5 (5 x CH2), 36.2 (4 x CH2), 35.9 (CH2) , 28.6 (4 x CH2), 28.3 (4 x CH2), 28.2 (CH2), 26.0 (4 x CH2), 25.9 (CH2), 25.3 (CH2), 24.8 (4 x CH2), 18.6 (2 x Me); 6F (470 MHz, methanoldi -70.4 (9 x CF3).

[0241] *chemical shift obscured by residual solvent peak, determined via HSQC.

[0242] **chemical shift determined via HMBC.

[0243] Example 24: Maltose tetramer undecanamide 25

[0244] To a solution of undecanoic acid (1.9 mg, 0.011 mmol) in DMF (0.5 mL) was added HATU (4.2 mg, 0.011 mmol) and DIPEA (3 pL, 0.018 mmol) and the resulting mixture stirred at r.t. for 10 mins. This mixture was then added dropwise to a solution of maltose tetramer amine 16 (17.0 mg, 0.007 mmol) in DMF (0.5 mL). After stirring at r.t. for 4 h, concentration in vacuo afforded the crude material, which was purified by flash column chromatography on silica gel eluting with acetonitrile - water (0->40% water) affording the maltose tetramer undecanamide 25 as a fluffy white solid (12.4 mg, 0.005 mmol, 68%). HRMS [ESI, M + 2Na]2+Found: 1314.6675 [C114H206N8O56 + 2Na]2+requires 1314.6650; 6H (500 MHz, D2O) 5.42 (4 H, d, J 3.9, 4 x CH), 4.52-4.47 (1 H, m, CH), 4.47 (4 H, d, J 8.0, 4 x CH), 3.96-3.54 (64 H, m, 24 x CH + 20 x CH2), 3.45 (4 H, t, J 9.5, 4 x CH), 3.32 (4 H, dd, J 9.4, 8.1, 4 x CH), 3.22 (8 H, t, J 7.0, 4 x CH2), 3.22-3.19 (2 H, m, CH2), 2.51 (8 H, t, J 5.7, 4 x CH2), 2.47-2.41 (4 H, m, 2 x CH2), 2.27-2.22 (4 H, m, 2 x CH2), 1.69-1.30 (54 H, m, 27 x CH2), 1.29 (6 H, s, 2 x Me), 0.91 (3 H, t, J 6.9, Me); 6c (125 MHz, D2O) 176.6 (C=O), 175.4 (C=O), 173.7 (4 x C=O), 172.0 (2 x C=O), 102.1 (4 x CH), 99.7 (4 x CH), 77.0 (4 x CH), 76.3 (4 x CH), 74.6 (4 x CH), 73.0 (4 x CH), 72.9 (4 x CH), 72.7 (4 x CH), 72.2 (2 x CH2), 72.1 (2 x CH2), 71.7 (4 x CH), 70.5 (4 x CH2), 69.3 (4 x CH), 67.5 (4 x CH2), 60.8 (4 x CH2), 60.5 (4 x CH2), 57.1 (2 x C) , 45.0 (CH), 41.3 (2 x CH2), 39.4 (4 x CH2), 39.0 (CH2), 36.3 (4 x CH2), 35.9 (2 x CH2), 31.4 (CH2), 28.9 (2 x CH2), 28.8 (4 x CH2), 28.7 (CH2), 28.5 (CH2), 28.4 (4 x CH2), 28.3 (CH2), 28.2 (CH2), 26.0 (4 x CH2), 25.8 (CH2), 25.5 (CH2), 25.2 (CH2), 24.9 (4 x CH2), 22.2 (CH2), 18.7 (2 x Me), 13.7 (Me).

[0245] To maltose tetramer undecanamide 25 (10.8 mg, 0.004 mmol) and sulfur trioxide trimethylamine complex (81 mg, 0.585 mmol) was added DMF (0.5 mL) and toluene (0.5 mL) and the resulting mixture heated to 60 °C and stirred for 24 h. After cooling to r.t., the reaction mixture was diluted with aqueous ammonium hydroxide (5% w / w, 2 mL) and concentrated in vacuo. The crude material was subsequently dissolved in water (~5 mL) and transferred into a dialysis cassette (MWCO 1000 Da). The cassette was placed in 1 L dialysate solution of ammonium bicarbonate (7 pM) and the dialysate was changed at the following time intervals: 1, 2, 3, 16 and 2 hours. The sample was then removed from the cassette, passed through an Amberlyst™ exchange resin (Na+form), and lyophilised to afford sulfated maltose tetramer undecanamide 26 as a fluffy white solid (19.6 mg, 0.004 mmol, 86%). HRMS [ESI, M - 9Na]9-Found: 581.4135 [Cii4Hi78N8Na280i4oS28 - 9Na]9~ requires 581.4144; 6H (500 MHz, D2O) 5.62 (4 H, d, J 3.4, 4 x CH), 4.98 (4 H, d, J 4.5, 4 x CH), 4.90 (4 H, t, J 7.3, 4 x CH), 4.83-4.79 (4 H, 4 x CH)*, 4.63 (4 H, dd, J 8.0, 3.3, 4 x CH), 4.57-4.15 (37 H, m, 21 x CH + 8 x CH2), 3.97-3.92 (4 H, m, 2 x CH2), 3.77 (8 H, t, J 5.9, 4 x CH2), 3.73-3.54 (12 H, m, 6 x CH2), 3.22 (8 H, t, J 7.0, 4 x CH2), 3.21 (2 H, m, CH2), 2.53 (8 H, t, J 5.9, 4 x CH2), 2.49-2.40 (4 H, m, 2 x CH2), 2.28-2.23 (4 H, m, 2 x CH2), 1.69-1.31 (54 H, m, 27 x CH2), 1.29 (6 H, s, 2 x Me), 0.91 (3 H, t, J 6.7, Me); 6c (125 MHz, D2O) 177.0 (C=O), 175.6 (C=O), 173.8 (4 x C=O), 172.2 (2 x C=O), 100.0 (4 x CH), 94.1 (4 x CH), 77.5 (4 x CH), 76.4 (4 x CH), 75.0 (4 x CH), 73.4 (4 x CH), 73.2 (4 x CH), 72.2 (4 x CH), 72.1 (2 x CH2), 72.0 (2 x CH2), 71.9 (4 x CH), 70.2 (4 x CH, 4 x CH2), 67.8 (4 x CH2), 67.6 (4 x CH2), 66.2 (4 x CH2), 57.1 (2 x C), 44.9 (CH), 41.5 (2 x CH2), 39.6 (4 x CH2), 39.0 (CH2), 36.2 (4 x CH2), 35.9 (CH2), 35.8 (CH2), 31.3 (CH2), 28.7 (2 x CH2), 28.60 (4 x CH2), 28.55 (CH2), 28.3 (5 x CH2), 28.13 (CH2), 28.08 (CH2), 26.0 (4 x CH2), 25.8 (CH2), 25.5 (CH2), 25.2 (CH2), 24.8 (4 x CH2), 22.1 (CH2), 18.7 (2 x Me), 13.6 (Me).

[0246] Example 26: Triundecanyl ester 28

[0247] To undecanoic acid (165 mg, 0.886 mmol) was added thionyl chloride (1 mL) and the resulting mixture heated to 75 °C and stirred for 2 h. After cooling to r.t., thionyl chloride was removed in vacuo affording undecanoyl chloride which was dissolved in dichloromethane (2 mL) and cooled to 0 °C. A solution of benzyl ether 27 (52 mg, 0.215 mmol) in dichloromethane (2 mL) was then added followed by the dropwise addition of triethylamine (0.14 mL, 0.972 mmol). 4-Dimethylaminopyridine (10.7 mg, 0.088 mmol) was finally added, and the reaction mixture warmed to r.t. After stirring for 16 h, concentration in vacuo was followed by the addition of hexanes (20 mL) and water (20 mL) and separation of the resulting phases. The organic phase was further washed with brine (20 mL), dried over MgSCh and concentrated in vacuo providing the triundecanyl ester benzyl ether 28 as a pale-yellow oil (157 mg, 0.215 mmol, 93%). HRMS [ESI, M + Na]+Found: 753.5652 [C45H78O7 + Na]+requires 753.5640; 6H (500 MHz, CDCI3) 7.34-7.27 (5 H, m, 5 x ArH), 4.47 (2 H, s, CH2), 4.13 (6 H, s, 3 x CH2), 3.43 (2 H, s, CH2), 2.25 (6 H, t, J 7.6, 3 x CH2), 1.59-1.54 (6 H, m, 3 x CH2), 1.31-1.26 (42 H, m, 21 x CH2), 0.88 (9 H, t, J 7.0, 3 x Me); 6c (125 MHz, CDCI3) 173.5 (3 x C=O), 138.1 (C), 128.5 (2 x CH), 127.8 (CH), 127.7 (2 x CH), 73.6 (CH2), 68.4 (CH2), 62.8 (3 x CH2), 42.8 (C), 34.3 (3 x CH2), 32.1 (3 x CH2), 29.7 (3 x CH2), 29.6 (3 x CH2), 29.5 (3 x CH2), 29.4 (3 x CH2), 29.3 (3 x CH2), 25.1 (3 x CH2), 22.8 (3 x CH2), 14.3 (3 x Me).

[0248] Example 27: Triundecanyl ester alcohol 29

[0249] To triundecanyl ester benzyl ether 28 (23.0 mg, 0.031 mmol) in THF (3 mL) was added Pd(OH)2 / C (20% Pd w / w, 2.4 mg) and the resulting mixture stirred under an atmosphere of hydrogen for 17 h. The reaction mixture was then filtered over celite™, the cake washed with dichloromethane (5 mL) and the filtrate concentrated in vacuo. The crude residue was purified by flash column chromatography on silica gel eluting with hexane - ethyl acetate (0->40% ethyl acetate) to give triundecanyl alcohol 29 as a colourless oil (13.0 mg, 0.020 mmol, 65%). HRMS [ESI, M + Na]+Found: 663.5170 [C38H72O7 + Na]+requires 663.5170; 6H (500 MHz, CDCI3) 4.11 (6 H, s, 3 x CH2), 3.49 (2 H, d, J 6.9, CH2), 2.52 (1 H, t, J 6.9, OH), 2.32 (6 H, t, J 7.6, 3 x CH2), 1.63-1.58 (6 H, m, 3 x CH2), 1.31-1.26 (42 H, m, 21 x CH2), 0.88 (9 H, t, J 7.0, 3 x Me); 6c (125 MHz, CDCI3) 174.0 (3 x C=O), 62.2 (3 x CH2), 60.9 (CH2), 44.1 (C), 34.3 (3 x CH2), 32.0 (3 x CH2), 29.7 (3 x CH2), 29.6 (3 x CH2), 29.44 (3 x CH2), 29.39 (3 x CH2), 29.3 (3 x CH2), 25.1 (3 x CH2), 22.8 (3 x CH2), 14.2 (3 x Me).

[0250] 30

[0251] To triundecanyl alcohol 29 (12.1 mg, 0.019 mmol) in acetone (2 mL) at 0 °C was added Jones reagent (2.5 M, 50 pL) and the resulting mixture slowly left to warm to r.t. over 17 h. Ethyl acetate (5 mL) and water (5 ml) were then added and the phases separated. The organic phase was further washed with water (5 mL), dried over MgS04 and concentrated in vacuo to afford triundecanyl carboxylic acid 30 as a white solid (10.4 mg, 0.016 mmol, 84%). HRMS [ESI, M - H]- Found: 653.4996 [C38H70O8 - H]“ requires 653.4998; 6H (500 MHz, CDCI3) 4.31 (6 H, s, 3 x CH2), 2.30 (6 H, t, J 7.6, 3 x CH2), 1.60-1.57 (6 H, m, 3 x CH2), 1.33-1.25 (42 H, m, 21 x CH2), 0.88 (9 H, t, J 7.0, 3 x Me); 6c (125 MHz, CDCI3) 174.4 (C=O), 173.2 (3 x C=O), 61.5 (3 x CH2), 50.2 (C), 34.2 (3 x CH2), 32.0 (3 x CH2), 29.7 (3 x CH2), 29.6 (3 x CH2), 29.5 (3 x CH2), 29.4 (3 x CH2), 29.3 (3 x CH2), 25.0 (3 x CH2), 22.8 (3 x CH2), 14.3 (3 x Me).

[0252]

[0253] To a solution of triundecyl carboxylic acid 30 (6.2 mg, 0.010 mmol) in DMF (0.5 mL) was added HATU (3.8 mg, 0.010 mmol) and DIPEA (3 pL, 0.018 mmol) and the resulting mixture stirred at r.t. for 10 mins. This mixture was then added dropwise to a solution maltose tetramer amide 16 (15.3 mg, 0.006 mmol) in DMF (0.5 mL). After stirring at r.t. for 16 h, concentration in vacuo afforded the crude material, which was purified by flash column chromatography on silica gel eluting with acetonitrile - water (0->40% water) affording maltose tetramer undecyl amide 31 as a fluffy white solid (11.6 mg, 0.004 mmol, 60%). HRMS [ESI, M + CH2O2 - 2H]2-Found: 1548.3481 [C141H254N8O62 + CH2O2 - 2H]2~ requires 1548.3450; 6H (500 MHz, methanol-d4) 5.17 (4 H, d, J 3.8, 4 x CH), 4.44 (1 H, quint, J 6.9, CH), 4.31 (6 H, s, 3 x CH2), 4.28 (4 H, d, J 7.8, 4 x CH), 3.92-3.80 (16 H, m, 8 x CH2), 3.70- 3.51 (40 H, m, 16 x CH, 12 x CH2), 3.45 (4 H, dd, J 9.7, 3.7, 4 x CH), 3.37 (4 H, ddd, J 9.6, 4.5, 1.9, 4 x CH), 3.29-3.18 (18 H, m, 8 x CH, 5 x CH2), 2.45-2.40 (12 H, m, 6 x CH2), 2.33 (6 H, t, J 7.3, 3 x CH2), 2.18 (2 H, t, J 7.5, CH2), 1.65-1.30 (92 H, m, 43 x CH2, 2 x Me), 0.90 (9 H, t, J 6.9, 3 x Me); 6c (125 MHz, methanol-d4) 176.3 (3 x C=O), 176.1 (C=O), 175.3 (C=O), 173.9 (4 x C=O), 172.7 (2 x C=O), 104.3 (4 x CH), 102.9 (4 x CH), 81.3 (4 x CH), 77.9 (4 x CH), 76.6 (4 x CH), 75.1 (4 x CH), 74.8 (4 x CH), 74.7 (4 x CH), 74.2 (4 x CH),

[0254] 73.6 (4 x CH2), 71.5 (4 x CH), 70.8 (4 x CH2), 68.7 (4 x CH2), 63.3 (3 x CH2), 62.8 (4 x CH2), 62.2 (4 x CH2), 58.2 (2 x C), 54.8 (C), 46.5 (CH), 42.3 (2 x CH2), 40.4 (4 x CH2), 40.1 (CH2),

[0255] 37.6 (4 x CH2), 37.2 (CH2), 34.8 (3 x CH2), 33.0 (3 x CH2), 30.70 (4 x CH2), 30.65 (3 x CH2), 30.57 (3 x CH2), 30.5 (4 x CH2), 30.41 (3 x CH2), 30.36 (3 x CH2), 30.2 (3 x CH2), 30.1 (CH2), 27.8 (4 x CH2), 27.6 (CH2), 26.8 (4 x CH2), 26.7 (CH2), 26.0 (3 x CH2), 23.7 (3 x CH2), 19.7 (2 x Me), 14.4 (3 x Me).

[0256] To maltose tetramer triundecyl amide 31 (5.6 mg, 0.002 mmol) and sulfur trioxide trimethylamine complex (36 mg, 0.259 mmol) was added DMF (0.3 mL) and toluene (0.3 mL) and the resulting mixture heated to 60 °C and stirred for 46 h. After cooling to r.t., the reaction mixture was diluted with aqueous ammonium hydroxide (5% w / w, 1 mL) and concentrated in vacuo. The crude material was subsequently dissolved in water (~5 mL) and transferred into a dialysis cassette (MWCO 1000 Da). The cassette was placed in 1 L dialysate solution of ammonium bicarbonate (7 pM) and the dialysate was changed at the following time intervals: 1, 2, 3, 16 and 2 hours. The sample was then removed from the cassette, passed through an Amberlyst™ exchange resin (Na+form), and lyophilised to afford sulfated maltose tetramer triundecylsulfonate 32 as a fluffy white solid (9.9 mg, 0.002 mmol, 95%). HRMS [ESI, M - 6Na]6-Found: 929.0663 [Cio8Hi63N8Na3iOi49S3i - 6Na]6’ requires 929.0676; 6H (500 MHz, D2O) 5.61 (4 H, d, J 3.4, 4 x CH), 4.98 (4 H, d, J 4.6, 4 x CH), 4.91-4.89 (4 H, m, 4 x CH), 4.80 (4 H, 4 x CH)*, 4.63 (4 H, dd, J 7.8, 3.5, 4 x CH), 4.55-4.14 (43 H, m, 21 x CH + 11 x CH2), 3.96-3.91 (4 H, m, 2 x CH2), 3.77 (8 H, t, J 6.0, 4 x CH2), 3.73-3.54 (12 H, m, 6 x CH2), 3.30 (2 H, t, J 6.8, CH2), 3.22 (8 H, t, J 7.0, 4 x CH2), 2.53 (8 H, it, J 5.9, 4 x CH2), 2.48-2.40 (4 H, m, 2 x CH2), 2.24 (2 H, t, J 7.6, CH2), 1.68-1.53 (20 H, m, 10 x CH2), 1.43-1.38 (18 H, m, 9 x CH2), 1.28 (6 H, s, 2 x Me); 6c (125 MHz, D2O) 175.9 (C=O), 173.8 (4 x C=O), 172.2 (2 x C=O), 170.8 (C=O), 100.1 (4 x CH), 94.0 (4 x CH), 77.6 (4 x CH),

[0257] 76.4 (4 x CH), 74.9 (4 x CH), 73.3 (4 x CH), 73.2 (4 x CH), 72.2 (4 x CH), 72.1 (2 x CH2), 71.9 (4 x CH, 2 x CH2), 70.34 (4 x CH2), 70.27 (4 x CH), 67.8 (4 x CH2), 67.6 (4 x CH2), 66.2 (4 x CH2), 65.8 (3 x CH2), 57.1 (2 x C), 50.4 (C), 44.8 (CH), 41.5 (2 x CH2), 39.6 (4 x CH2),

[0258] 39.4 (CH2), 36.2 (4 x CH2), 35.9 (CH2), 28.6 (4 x CH2), 28.3 (4 x CH2), 28.0 (CH2), 26.0 (4 x CH2), 25.7 (CH2), 25.4 (CH2), 24.8 (4 x CH2), 18.7 (2 x Me). chemical shift obscured by residual solvent peak, determined via HSQC.

[0259] To a mixture of maltose tetramer amine 16 (14.5 mg, 0.006 mmol) and commercially available biotin- / V-hydroxysuccinimide ester (3.1 mg, 0.009 mmol) in DMF (1 mL) was added DIPEA (3 pL, 0.0150 mmol). After stirring at r.t. for 4 h, concentration in vacuo gave the crude material, which was purified by flash column chromatography on silica gel eluting with acetonitrile - water (0^55% water) affording the maltose tetramer biotinylamide 33 as a fluffy white solid (15.2 mg, 0.006 mmol, 96%). HRMS [ESI, M + 2H]2+Found: 1322.1489 [C113H200N10O57S + 2H]2+requires 1322.1484; 6H (500 MHz, D2O) 5.42 (4 H, d, J 3.9, 4 x CH), 4.65 (1 H, dd, J 7.8, 4.9, CH), 4.51-4.44 (2 H, m, 2 x CH), 4.48 (4 H, d, J 8.0, 4 x CH), 3.97- 3.54 (64 H, m, 24 x CH + 20 x CH2), 3.45 (4 H, t, J 9.5, 4 x CH), 3.38-3.30 (5 H, m, 5 x CH),

[0260] 3.22 (8 H, t, J 6.9, 4 x CH2), 3.22-3.19 (2 H, m, CH2), 3.03 (1 H, dd, J 13.0, 4.9,1 / 2CH2), 2.82 (1 H, d, J 13.0,1 / 2CH2), 2.51 (8 H, t, J 5.6, 4 x CH2), 2.48-2.41 (4 H, m, 2 x CH2), 2.29-

[0261] 2.23 (4 H, m, 2 x CH2), 1.78-1.39 (44 H, m, 23 x CH2), 1.29 (6 H, s, 2 x Me); 6c (125 MHz, D2O) 176.4 (C=O), 175.5 (C=O), 173.8 (4 x C=O), 172.0 (2 x C=O), 165.3 (C=O), 102.1 (4 x CH), 99.7 (4 x CH), 76.9 (4 x CH), 76.3 (4 x CH), 74.6 (4 x CH), 73.1 (4 x CH), 72.9 (4 x CH), 72.7 (4 x CH), 72.2 (2 x CH2), 72.1 (2 x CH2), 71.7 (4 x CH), 70.5 (4 x CH2), 69.3 (4 x CH), 67.5 (4 x CH2), 62.1 (CH), 60.8 (4 x CH2), 60.5 (4 x CH2), 60.3 (CH), 57.1 (2 x C), 55.5 (CH), 45.0 (CH), 41.3 (2 x CH2), 39.8 (CH2), 39.4 (4 x CH2), 39.1 (CH2), 36.3 (4 x CH2), 35.8 (CH2), 35.6 (CH2), 28.8 (4 x CH2), 28.4 (4 x CH2), 28.2 (CH2), 28.0 (CH2), 27.8 (CH2), 25.9 (4 x CH2), 25.8 (CH2), 25.3 (CH2), 25.2 (CH2), 24.8 (4 x CH2), 18.7 (2 x Me).

[0262] To maltose tetramer biotinylamide 33 (10.5 mg, 0.004 mmol) and sulfur trioxide trimethylamine complex (82 mg, 0.556 mmol) was added DMF (0.5 mL) and toluene (0.5 mL) and the resulting mixture heated to 60 °C and stirred for 48 h. After cooling to r.t., the reaction mixture was diluted with aqueous ammonium hydroxide (5% w / w, 1 mL) and concentrated in vacuo. The crude material was subsequently solved in water (~5 mL) and transferred into a dialysis cassette (MWCO 1000 Da). The cassette was placed in 1 L dialysate solution of ammonium bicarbonate (7 pM) and the dialysate was changed at the following time intervals: 1, 2, 3, 16 and 2 hours. The sample was then removed from the cassette, passed through an Amberlyst™ exchange resin (Na+form), and lyophilised to afford the biotinylamide sulfonate 34 as a fluffy white solid (16.9 mg, 0.003 mmol, 76%). HRMS [ESI, M - 8Na]8-Found: 677.1984 [Cn3Hi7iNioNa29Oi44S3o - 8Na]8’ requires 677.1985; 6H (500 MHz, D2O) 5.62 (4 H, d, J 3.4, 4 x CH), 4.99 (4 H, d, J 4.5, 4 x CH), 4.93-4.88 (1 H, m, CH), 4.90 (4 H, t, J 7.2, 4 x CH), 4.81-4.79 (4 H, 4 x CH)*, 4.63 (4 H, dd, J 7.8, 3.2, 4 x CH), 4.57-4.16 (38 H, m, 22 x CH, 8 x CH2), 3.96-3.92 (4 H, m, 2 x CH2), 3.77 (8 H, t, J 6.0, 4 x CH2), 3.73-3.54 (12 H, m, 6 x CH2), 3.41-3.37 (1 H, m, CH), 3.26-3.09 (12 H, m, 6 x CH2), 2.53 (8 H, t, J 5.7, 4 x CH2), 2.49-2.41 (4 H, m, 2 x CH2), 2.29 (2 H, t, J 7.2, CH2), 2.25 (2 H, t, J 7.4, CH2), 1.83-1.35 (44 H, m, 22 x CH2), 1.29 (6 H, s, 2 x Me); 6c (125 MHz, D2O) 176.5 (C=O), 175.6 (C=O), 173.8 (4 x C=O), 172.2 (2 x C=O), 159.8 (C=O), 99.9 (4 x CH),

[0263] 94.1 (4 x CH), 77.6 (4 x CH), 76.4 (4 x CH), 75.0 (4 x CH), 73.4 (4 x CH), 73.2 (4 x CH),

[0264] 72.2 (4 x CH), 72.1 (2 x CH2), 72.0 (2 x CH2), 71.9 (4 x CH), 70.2 (4 x CH, 4 x CH2), 67.8 (4 x CH2), 67.6 (4 x CH2), 66.2 (4 x CH2), 65.2 (CH), 59.3 (CH), 57.1 (2 x C), 54.8 (CH), 44.9 (CH), 41.5 (2 x CH2), 39.6 (4 x CH2), 39.2 (CH2), 38.7 (CH2), 36.2 (4 x CH2), 35.9 (CH2), 35.6 (CH2), 28.6 (4 x CH2), 28.3 (4 x CH2), 28.2 (CH2), 27.9 (CH2), 27.8 (CH2), 26.0 (4 x CH2), 25.9 (CH2), 25.3 (CH2), 25.2 (CH2), 24.8 (4 x CH2), 18.7 (2 x Me).

[0265] *chemical shift obscured by residual solvent peak, determined via HSQC.

[0266] Sulfated maltose tetramer amine 18 (Na+salt) was first converted to the triethylamine salt form by passing through Amberlyst™ exchange resin (NH4+form) followed by repeated evaporation from water: methanol :triethylamine (2: 1:0.1, 4 x 3 mL). To the Et3HN+salt of amine 18 (21.9 mg, 0.003 mmol) and biotin- / V-hydroxysuccinimide ester (9.9 mg, 0.029 mmol) in DMF (1 mL) was added triethylamine (4 pL, 0.029 mmol) and the resulting mixture stirred at r.t. for 42 h. The reaction mixture was then diluted with aqueous ammonium hydroxide (5% w / w, 1 mL) and concentrated in vacuo. The crude material was subsequently solved in water (~5 mL) and transferred into a dialysis cassette (MWCO 1000 Da). The cassette was placed in 1 L dialysate solution of ammonium bicarbonate (7 pM) and the dialysate was changed at the following time intervals: 1, 2, 3, 16 and 2 hours (cycle repeated twice). The sample was then removed from the cassette, passed through an Amberlyst exchange resin (Na+form), and lyophilised to afford the biotinylamide 35 as a fluffy white solid (14.5 mg, 0.003 mmol, 90%). HRMS [ESI, M - 6Na]6’ Found: 893.6034 [Cii3Hi72NioNa28Oi4iS29 - 6Na]6“ requires 893.6042; 6H (400 MHz, D2O) 5.61 (4 H, d, J 3.4, 4 x CH), 4.98 (4 H, d, J 4.6, 4 x CH), 4.91-4.88 (4 H, m, 4 x CH), 4.82-4.79 (4 H, 4 x CH)*, 4.67 (1 H, dd, J 8.0, 4.9, CH), 4.63 (4 H, dd, J 7.8, 3.4, 4 x CH), 4.57-4.15 (38 H, m, 22 x CH + 8 x CH2), 3.96-3.91 (4 H, m, 2 x CH2), 3.77 (8 H, t, J 6.0, 4 x CH2), 3.73-3.53 (12 H, m, 6 x CH2), 3.41-3.36 (1 H, m, CH), 3.23-3.20 (10 H, m, 5 x CH2), 3.05 (1 H, dd, J 13.1, 4.9,1 / 2CH2), 2.83 (1 H, d, J 13.1,1 / 2CH2), 2.53 (8 H, t, J 5.7, 4 x CH2), 2.49-2.40 (4 H, m, 2 x CH2), 2.34-2.23 (4 H, m, 2 x CH2), 1.80-1.38 (44 H, m, 22 x CH2), 1.28 (6 H, s, 2 x Me); 6H (100 MHz, D2O) 176.5 (C=O), 175.6 (C=O), 173.8 (4 x C=O), 172.2 (2 x C=O), 165.3 (C=O), 99.9 (4 x CH), 94.1 (4 x CH), 77.6 (4 x CH), 76.4 (4 x CH), 75.0 (4 x CH), 73.4 (4 x CH), 73.2 (4 x CH), 72.2 (4 x CH), 72.1 (2 x CH2), 72.0 (2 x CH2), 71.9 (4 x CH), 70.2 (4 x CH, 4 x CH2), 67.8 (4 x CH2), 67.6 (4 x CH2), 66.2 (4 x CH2), 62.2 (CH), 60.3 (CH), 57.1 (2 x C), 55.5 (CH), 44.9 (CH), 41.5 (2 x CH2), 39.8 (CH2), 39.6 (4 x CH2), 39.1 (CH2), 36.2 (4 x CH2), 35.9 (CH2), 35.6 (CH2), 28.6 (4 x CH2), 28.3 (4 x CH2), 28.2 (CH2), 28.0 (CH2), 27.8 (CH2), 26.0 (4 x CH2), 25.8 (CH2), 25.3 (CH2), 25.2 (CH2), 24.8 (4 x CH2), 18.7 (2 x Me). *chemical shift obscured by residual solvent peak, determined via HSQC.

[0267] Maltose tetramer amine 16 (113 mg, 0.047 mmol), 4,4-difluoro-l,3-dimethyl-4-bora- 5-carboxyethyl-3a,4a-diaza-s-indacene (19 mg, 0.051 mmol, 1.1 eq.) and HATU (21 mg, 0.055 mmol, 1.2 eq.) were dried together under vacuum before being brought under argon atmosphere. Then anhydrous DMF (2.5 mL) and DIPEA (10 pL, 0.058 mmol, 1.2 eq.) were added and the reaction mixture was stirred at r.t. for 1.5 hours. The mixture was then transferred to a falcon tube in minimal MeOH before 30 mL of EtOAc was added. The mixture was centrifuged at 4000 rpm for 4 minutes which gave a dark orange residue at the bottom of the tube. The supernatant was discarded and the residue was resuspended in 50% H2O in ACN before being purified via flash chromatography (0%-50% H2O in ACN) to give maltose tetramer BODIPY amide 36 (108 mg, 0.040 mmol, 85% yield) as a fluffy orange solid after lyophilisation. TLC Rf = 0.2 (25% H2O in ACN). 1H NMR (500 MHz, D2O) 6 = 7.48 (s, 1H, CH), 7.08 (d, J = 4.3 Hz, 1H CH), 6.39 (d, J = 3.9 Hz, 1H CH), 6.33 (s, 1H, CH), 5.39 (d, J = 3.9 Hz, 4H, CH), 4.48 (p, J = 7.3 Hz, 1H, CH), 4.42 (d, J = 8.0 Hz, 4H, CH), 3.96 - 3.48 (m, 64H), 3.45 (t, J = 9.5 Hz, 4H), 3.30 (dd, J = 9.4, 8.0 Hz, 4H, CH), 3.18 (m, 12H, CH2, CH2 and CH2), 2.68 (t, J = 7.5 Hz, 2H, CH2), 2.54 (s, 3H, CH3), 2.51 - 2.37 (m, 12H, CH2 and CH2), 2.28 (s, 3H, CH3), 2.18 (t, J = 7.5 Hz, 2H, CH2), 1.68 - 1.43 (m, 20H), 1.41 - 1.29 (m, 18H), 1.27 (s, 6H, CH3). 13C{1H} NMR (125 MHz, D2O) 6 = 175.3, 174.3, 173.73, 173.70, 172.0, 161.3, 156.1, 146.2, 135.3, 33.2, 128.9, 124.9, 121.2, 116.8, 102.1, 99.8, 77.1, 76.3, 74.6, 73.0, 72.9, 72.7, 72.1, 72.0, 71.7, 70.5, 69.3, 67.5, 60.7, 60.5, 57.0, 45.0, 41.3, 39.4, 39.1, 36.2, 35.8, 34.7, 28.8, 28.4, 28.1, 26.0, 25.7, 25.2, 24.8, 24.5, 18.6, 14.4, 10.7. HRMS (ESI-TOF) m / z calcd for [CnyHissBFzNioNaOs?]2* [M + H + Na]2+: 1365.1531, observed 1365.1464.

[0268] Maltose tetramer BODIPY amide 36 (32 mg, 0.012 mmol) and sulfur trioxide trimethylamine (232 mg, 1.67 mmol, 140 eq.) were dried together under vacuum before being brought under argon atmosphere. Then anhydrous DMF (1 mL) and toluene (1.5 mL) were added before the reaction mixture was stirred at 60°C for 16 hours. The reaction mixture was then cooled to r.t., diluted with NH4OH (5% w / iv) and concentrated in vacuo. The material was co-evaporated with NH4OH (5% w / iv) four times before being dissolved in H2O and transferred into a 1000 g / mol MWCO dialysis cassette. The cassette was placed in 1 L dialysate solution of (NH^CCh (7 pM) and the dialysate was changed at the following time intervals: 1, 2, 3, 10 and 2 hours. Then the sample was removed from the cassette and lyophilised to give sulfated maltose tetramer BODIPY amide 37 (57 mg, 0.010 mmol, 87% yield) as a fluffy orange solid. 1HNMR (600 MHz, D2O) 6 = 7.54 (s, 1H, C(7)H), 7.11 (d, J = 4.0 Hz, 1H, CH), 6.40-6.32 (m, 2H, CH and CH), 5.56 (d, J = 3.5 Hz, 4H, CH), 4.88 (d, J = 4.6 Hz, 4H, CH), 4.84 (t, J = 8.1 Hz, 4H, CH), 4.73 (t, J = 5.0 Hz, 4H, CH), 4.57 (dd, J = 8.1, 3.3 Hz, 4H, CH), 4.53-4.42 (m, 9H, CH, CH and CH), 4.42-4.22 (m, 16H, CH and CH), 4.22- 4.13 (m, 8H, CH), 4.13-4.04 (m, 4H, CH and CH), 3.84 (dt, J = 9.8, 6.9 Hz, 4H, 1 / 2 CH2), 3.68 (q, J = 7.8, 5.6 Hz, 8H, CH2), 3.63-3.41 (m, 12H, CH2and 1 / 2 CH2), 3.22-3.05 (m, 12H, CH2, CH2and CH2), 2.65 (t, J = 7.3 Hz, 2H, CH2), 2.52 (s, 3H,CH3), 2.48-2.32 (m, 12H, CH2and CH2), 2.30 (s, 3H, CH3), 2.11 (t, J = 7.4 Hz, 2H, CH2), 1.64-1.37 (m, 20H, CH2, CH2, CH2, CH2), 1.37-1.23 (m, 16H, C(32)H2, CH2), 1.20 (s, 8H, CH3and CH3). 13C{1H} NMR (150 MHz, D2O) 6 = 175.6, 174.6, 173.7, 172.2, 161.7, 155.8, 146.7, 135.5, 133.2, 129.0, 125.1, 121.4, 116.8, 99.9, 94.1, 77.5, 76.3, 74.9, 73.4, 73.2, 72.14, 72.05, 71.95, 71.87, 70.2, 70.1, 67.7, 67.5, 66.2, 57.1, 44.9, 41.5, 39.6, 39.1, 36.1, 35.8, 34.7, 28.6, 28.3, 28.0, 26.0, 25.6, 25.2, 24.8, 24.5, 18.6, 14.4, 10.8.

[0269] Example 36: Detection of cell-bound sulfated maltose tetramer BODIPY amide 37 on immune cells in blood and brain after EAE induction

[0270] Female C57BI / 6J mice were immunised for EAE with 50-100 pg / mouse MOG3s-55 (Genscript, Piscataway, NJ USA) in complete Freund's adjuvant (Sigma, St. Louis, MO, USA), containing 500 pg / mouse heat-killed Mycobacterium tuberculosis (Difco Laboratories, Detroit, USA) and 200 ng / mouse of pertussis toxin (List Biological Laboratories, Campbell, CA USA). Two days later, mice received a second dose of pertussis toxin at 200 ng / mouse. Twenty-four days after EAE immunisation mice received a single IP dose of sulfated BODIPY amide 37 or non-sulfated BODIPY amide 36 (60 pg in 100 pl PBS). The compound was allowed to circulate for 45 minutes before mice were sacrificed.

[0271] Following euthanasia, blood was collected from the heart, and red blood cells were lysed by incubation in 2mL RBC lysis buffer (Sigma-Aldrich, MA). Remaining blood cells were washed then pelleted by centrifugation before resuspension in flow cytometry staining (FACs) buffer and assessed by flow cytometry. Residual blood in brain blood vessels was then flushed out by cardiac perfusion with 20 mL of PBS before brains were collected. Brains were cut in half sagittally and prepared for both flow cytometry and confocal microscopy analysis. For flow cytometry analysis, half-brains were processed into a single-cell suspension. CNS cells were separated from myelin by resuspending the samples in a 37% Percoll solution (Sigma- Aldrich, MA) and centrifuging at low acceleration. Following the removal of myelin, cells were resuspended in FACs buffer. Flow cytometric data were collected on a BD FACS Canto II (BD Biosciences, NJ) and analysed with FlowJo software (Treestar, Ashland, OR USA), immune cell types were identified as CD45+ (all immune cells), CD45+CD4+ (CD4 T helper cells), CD45+CD8+ (CD8 cytotoxic T cells, CD45+Ly6C+ (monocytes), and CD45+Ly6G+ (neutrophils). BODIPY expression is displayed as BODIPY mean fluorescent intensity (MFI) as a percentage of BODIPY MFI detected in a sham injection control animal or raw MFI. Data were pooled from two independent experiments (n=2-15 per group) and displayed as mean ± SEM. *p<0.05, **p<0.01 by two-way ANOVA with Holm-Sidak's multiple comparisons test. All in vivo BODIPY amide 37 treatment experiments included non-injected control animals. The results are shown in Figures 2 and 3.

[0272] Example 37: In vitro analysis of sulfated maltose tetramer BODIPY amide 37 adhesion to immune cells

[0273] Spleens were collected from healthy C57BL / 6J mice and processed into single-cell suspensions. Splenocytes were cultured in the presence of BODIPY amide 37 (0, 1.2, 4, 12 ng / ml) for 12 or 24 hours at 37°C and 5% CO2. Following incubation, BODIPY amide 37 was rinsed away and the cells were stained with fluorescently tagged antibodies for flow cytometry analysis. CD4 T helper cells were identified as CD45+CD4+. Flow cytometry of splenocytes was performed on a BD FACS Canto II (BD Biosciences, NJ) and analysed using FlowJo software (Treestar Inc., Ashland, OR, USA) to determine the % of immune cells positive for BODIPY amide 37. ***p<0.001 for both time and concentration by 2-way ANOVA. The results are shown in Figure 1.

[0274] Example 38: Detection of sulfated maltose tetramer BODIPY amide 37 in tissues following in vivo treatment

[0275] Female C57BI / 6J mice were immunised for EAE (as in Example 36) or were healthy. Twenty-four to twenty-eight days after immunisation, BODIPY amide 37 was administered to healthy or EAE mice either i.p. or p.o. at 10 pg / mouse or 60 pg / mouse in 100 pl of PBS and allowed to circulate for 45 minutes, 4 hours or 12 hours before animals were euthanised. Following euthanasia, blood was collected from the heart before animals were perfused. Residual blood in CNS blood vessels was then flushed out by cardiac perfusion with 20 mL of PBS before brains were collected. Red blood cells in the blood were lysed by incubation in 2mL RBC lysis buffer (Sigma-Aldrich, MA). Remaining blood cells were washed then pelleted by centrifugation before resuspension in flow cytometry staining (FACs) buffer. For flow cytometry analysis, half-brains were processed into a single-cell suspension. CNS cells were separated from myelin by resuspending the samples in a 37% Percoll solution (Sigma-Aldrich, MA) and centrifuging at low acceleration. Following the removal of myelin, cells were resuspended in FACs buffer. Brain and blood single-cell suspensions were stained with fluorescently labelled antibodies for flow cytometry analysis as described in Example 36.

[0276] In some experiments, blood was collected and processed to obtain plasma for analysis of free BODIPY amide 37 levels. Here, blood was collected into tubes with EDTA to prevent clotting and red blood cells were lysed 2x by incubation with RBC lysis buffer as above. Afterwards, samples were centrifuged at 10,000 x g for 10 minutes and plasma was carefully collected without disturbing cell pellets. BODIPY amide 37 levels in plasma were estimated by relative fluorescence units collected on an EnSpire plate reader (PerkinElmer). Data are displayed as mean ± SEM. All in vivo BODIPY amide 37 treatment experiments included non-injected control animals. The results are shown in Figures 8 and 9.

[0277] Example 39: Visualisation of sulfated maltose tetramer BODIPY amide 37 in brain parenchyma following in vivo treatment.

[0278] Female C57BI / 6J mice were immunised for EAE (as in Example 36) or were healthy. Twenty-four to twenty-eight days after immunisation, sulfated BODIPY amide 37 or nonsulfated BODIPY amide 36 was administered to healthy or EAE mice at 60 pg / mouse in 100 pl of PBS i.p. and allowed to circulate for 45 minutes before animals were euthanised. Following euthanasia, blood was collected from the heart before animals were perfused. Residual blood in CNS blood vessels was then flushed out by cardiac perfusion with 20 mL of PBS before brains were collected. Brains were cut in half sag itta lly and prepared for both flow cytometry and confocal microscopy analysis. The brain hemispheres were snap- frozen in Polyfreeze OCT compound (Sigma, USA) and stored at -80°C until sectioning.

[0279] Brain hemispheres were cut into 20 pm sagittal sections using a Leica CM3050 S cryostat microtome (Leica Biosystems, Wetzlar, Germany) and mounted on Superfrost plus slides (Thermo Fisher Scientific, MA). Immediately prior to staining, slides were thawed at 37°C for 5 minutes and sections were fixed in 4% paraformaldehyde (PFA) for 10 minutes at room temperature. Tissue autofluorescence was quenched by immersion in fresh sodium borohydride solution (1 mg / ml in ddHzO) for 30 minutes. Tissue was then blocked in 5% donkey serum (Sigma-Aldrich, MA) for 2 hours at room temperature. After blocking, tissue was incubated with rabbit-anti-mouse collagen IV antibody (Abeam, Cambridge, UK) in Trisbuffered saline (TBS) overnight at 4°C. Sections were washed thoroughly in 2.5xl0'4% Triton-X-100 in TBS and then incubated with a donkey-anti-rabbit AF647 (Abeam, Cambridge, UK) antibody for 2 hours at room temperature. Finally, sections were washed twice in PBS and in ddHzO before they were mounted with an anti-fade glycerol mounting medium containing DAPI (Abeam, Cambridge, UK) and 0.17 nm coverslips. BODIPY amide 37 infiltration into the brain parenchyma was visualised by confocal microscopy. The results are shown in Figure 4. Example 40: Sulfated maltose tetramer BODIPY amide 37 adhesion to brain sections

[0280] Whole brains were collected from healthy C57BL / 6J mice as described in Example 39 and fixed overnight in 4% PFA and then cryopreserved in sucrose solution for 48 hours. Tissue was embedded in Polyfreeze OCT compound (Sigma, USA), snap frozen, and then cut into 20 pm sagittal sections. Sections underwent antigen retrieval in sodium citrate buffer at 80-95°C for 15 minutes before quenching, blocking and primary antibody incubation. Sulfated BODIPY amide 37 or non-sulfated BODIPY amide 36 was added to the secondary antibody solution at 1 pg / mL and left on sections for 3 hours at room temperature. Sections were then washed in PBS and ddHzO before mounting with anti-fade glycerol mounting medium containing DAPI (Abeam, Cambridge, UK) as stated above. Sections were analysed by confocal microscopy. The results are shown in Figures 6 and 7.

[0281] Example 41: Sulfated maltose tetramer BODIPY amide 37 adhesion to brain endothelial (bEnd.3) cells

[0282] Coverslips were rinsed in 70% ethanol and left to dry in a 6-well plate while a 0.5 mg / ml solution of Collagen I (Sigma-Aldrich, USA) was prepared. The collagen solution was then applied to coverslips in the 6-well plate and left on a shaker for 2 hours at room temperature. Excess collagen was aspirated, and wells were washed twice with IX dPBS (Gibco, USA). Following washes, 3xl05bEnd.3 cells (ATCC, USA) were seeded on each coverslip. Cells were left to become confluent in incubator for 48-72 hours. Once confluent, media was aspirated and replaced with pre-warmed media containing different concentrations of sulfated BODIPY amide 37 or non-sulfated BODIPY amide 36. After 1 hour, the media was removed, and cells washed twice with IX dPBS. The bEnd.3 cells were then fixed with 4% PFA (Sigma-Aldrich, USA) for 15 minutes at room temperature. PFA was aspirated, coverslips washed twice with IX dPBS, and mounted on a Superfrost microscope slide (Fisherbrand, USA) using DAPI containing mounting medium (Abeam, UK). The results are shown in Figure 5. ****p<0.0001 and *p<0.05 by 2-way ANOVA compared to non-sulfated BODIPY amide 36.

[0283] Example 42: Detection of fluorescently labelled mimetics

[0284] MC38 murine colon adenocarcinoma cells were suspended in DMEM media (Gibco, Waltham, MA, USA) supplemented with 10% heat-inactivated FCS (Moregate Biotech, Hamilton, NZ), 100 U / mL penicillin and 100 mg / mL streptomycin (Gibco) at a density of 2 x 105cells / mL. 1.5 mL cell suspension (300,000 cells) was seeded into wells of a 12 well plate containing poly-D-lysine coated coverslips (12-13mm diameter, 1.5H) and incubated for 16 hours at 37°C + 5% CO2 to allow cells to adhere. 5 pM of sulfated BODIPY amide 37 was added to the cells and incubated for 1 hour before removing the media from the wells and washing the cells two times in PBS. Cells were then fixed in 4% PFA w / v solution in PBS for 15 minutes at room temperature, washed three times with PBS, and then permeabilized with 0.1% Triton™ X-100 in PBS for 5 minutes. Cells were washed three times with PBS and then stained with Alexa Fluor Plus 647 Phalloidin (Invitrogen, Carlsbad, CA, USA) in PBS containing 1% bovine serum albumin (BSA) for 60 minutes at room temperature. After this incubation, cells were stained with 2 pg / mL Hoechst 33342 for 30 minutes at room temperature, washed three times in PBS, and then mounted on glass slides with Prolong™ Glass Antifade Mountant (Invitrogen). Slides were dried for 48 hours before imaging on a Nikon Ni-E Fluorescence microscope (Nikon, Tokyo, Japan). The results are shown in Figure 10. These data show that heparin sulphate mimetics selectively associate with tumour cells in vitro and demonstrates their successful detection using fluorescence-based imaging.

[0285] Example 43: Detection of19F-labelled maltose tetramers by Magnetic Resonance Imaging (MRI)

[0286] The F6 compound 20 (sulfated maltose tetramer bis(trifluoromethyl)benzamide) and the F27 compound 24 (sulfated maltose tetramer tris(perfluoro-t-butoxy)amide) were serially diluted in PBS to prepare 40 mM, 20 mM, 10 mM, and 5 mM solutions with respective19F concentrations of 1080 mM, 540 mM, 270 mM, and 135 mM for compound 24, and 240 mM, 120 mM, 60 mM, and 30 mM for compound 20. 150 pL volumes of each concentration were transferred into 0.5 mL polypropylene tubes for phantom imaging.19F images of phantoms were acquired on a MRS DRYMAG 7.0T MRI system (MRS 7024, MR Solutions, Guildford, UK) using a 2D FLASH sequence with the following parameters: field-of-view (FOV) = 50 x 50 mm; matrix = 64 x 64; slice thickness = 5 mm; TR = 200 ms; TE = 2.8 ms; flip angle (FA) = 20°; bandwidth (BW) = 33 KHz; number of averages (NA) = 480; acquisition time = 102 min. DICOM images were exported using OsiriX Lite software (Pixmeo SARL, Bernex, Switzerland) and processed using ImageJ. Signal-to-noise ratios were calculated by normalising the region-of-interest mean intensity values by standard deviation of signal noise. Both19F-labelled compounds could be detected down to 5 mM concentrations with linear signal-to-noise ratios (SNR). As expected, the densely19F-labelled compound 24 is more sensitive than compound 20. The results are shown in Figures 11-13. These data confirm19F-labelled maltose tetramers can be successfully detected using MRI.

[0287] Example 44: Heparanase inhibition assay

[0288] A colorimetric heparanase assay was used to assess the compounds for heparanase inhibition in vitro. The assay used fondaparinux as the enzyme substrate and WST-1 for colour development. Recombinant human heparanase was purchased from R&D Systems. Assays were carried out in 96-well plates, with assay solutions having a total volume in each well of 100 pL. Final concentrations in the wells were as follows: 40 mM sodium acetate buffer (pH 5.0), pMlOO fondaparinux (MW = 1726.77 g / mol, 12.5 mg / ml), 1 nM heparanase and sulfated maltose tetramer 7a or sulfated maltose tetramer 7c at the desired concentration. Values provided for each inhibitor concentration are from the average (n=3) of independent experiments ± SD, each independent experiment contained 3 technical replicates. The wells were pre-treated with BSA (4% w / v) in 0.05% Tween 20 phosphate buffered saline (PBST) for 2 h at 37°C, after which the plates were washed 3 times with PBST and shaken dry. The unused wells of the plate were filled with water in order to avoid excessive evaporation during the incubation. Heparanase was added as the last component of the solution. The plates were then sealed with tape and aluminium foil before being incubated at 37°C for 6 h. The assays were stopped by the addition of 100 pL of 1.69 mM (WST-1) (4-[3-(4-iodophenyl)-2-(4-nitrophenyl)-2H-5-tetrazolio]-l,3-benzene disulfonate) in 0.1 M NaOH (1.1 mg / ml). Plates were covered and incubated at 60°C for 1 h and the absorbance was measured at 584 nm. IC50 values were calculated using Prism 9.0.1 software (GraphPad Software, La Jolla, CA, USA). The results are shown in Figures 14 and 15.

[0289] INDUSTRIAL APPLICABILITY

[0290] The invention relates to compounds that are useful for the treatment or prevention of diseases including cancer, inflammation, diabetic nephropathy, neurodegenerative disorders, and for certain cosmeceutical and dermatological uses.

Claims

CLAIMS1. A compound of formula (I) :wherein:X is (CH2)Pand p is an integer from 1 to 20;Ri is H or CH3; andwherein• is the point of attachment of R2 to the O atom;Y is (CH2)nand n is an integer from 1 to 3;R3 is H or C1-C3 alkyl;Z is (CH2)m and m is an integer from 1 to 6; andwherein R4 is H or SO3H, and • is the point of attachment to Z; or a salt thereof.

2. A compound as claimed in claim 1, wherein p is an integer from 5 to 12.

3. A compound as claimed in claim 1 or claim 2, wherein p is 5, 6 or 12.

4. A compound as claimed in any one of claims 1 to 3, wherein n is 2.

5. A compound as claimed in any one of claims 1 to 4, wherein R3is H.

6. A compound as claimed in any one of claims 1 to 4, wherein R3 is methyl or ethyl.

7. A compound as claimed in any one of claims 1 to 6, wherein m is 4, 5 or 6.

8. A compound as claimed in any one of claims 1 to 7, wherein one or more of the FU groups are SO3H or SChNa.

9. A compound as claimed in claim 1 selected from the group comprising:

10. A compound of formula (II):wherein: Ri is H or CH3; andwherein• is the point of attachment of R2 to the O atom;Y is (CH2)n and n is an integer from 1 to 3;R3 is H or C1-C3 alkyl;Z is (CH2)m and m is an integer from 1 to 6; andwherein R4 is H or SO3H, and • is the point of attachment to Z; andA is a Ci-Ce alkyl group substituted with -NH2, -N3, or -NH(C=O)Rs wherein Rs is: a) a C1-C12 alkyl group; b) a biotinyl substituent; c) a group comprising a fluorescent label; d) a group comprising a fluorine-18 label; e) a group comprising a fluorine-19 label; f) a group comprising a crown ether-based caged ligand for rhodium, iridium, actinium-225 or thorium-227; or g) a group comprising an N-acetate or a C-14 radiolabelled N-acetate; or a salt thereof.

11. A compound as claimed in claim 10, wherein n is 2.A compound as claimed in claim 10 or claim 11, wherein Ri is H.

13. A compound as claimed in claim 10 or claim 11, wherein Ri is methyl.

14. A compound as claimed in any one of claims 10 to 13, wherein R3is H.

15. A compound as claimed in any one of claims 10 to 13, wherein R3 is methyl or ethyl.

16. A compound as claimed in any one of claims 10 to 15, wherein m is 4, 5 or 6.

17. A compound as claimed in any one of claims 10 to 16, wherein A is a Ci-Ce alkylamine.

18. A compound as claimed in any one of claims 10 to 16, wherein A is a Ci-Ce alkylazide.

19. A compound as claimed in any one of claims 10 to 16, wherein A is Ci-Ce alkyl group substituted with NH(C=0)R4 wherein FU is a C6-C12 alkyl group, a C6-C12 alkyl group comprising a biotin substituent, a C6-C12 alkyl group comprising a fluorine labelled BODIPY group, a C6-C12 alkyl group comprising a fluorescent label, a C6-C12 alkyl group comprising a fluorine-18 label, or a C6-C12 alkyl group comprising a fluorine-19 label.

20. A compound as claimed in any one of claims 10 to 19, wherein one or more of the R4 groups are SO3H or SChNa.

21. A compound as claimed in claim 10 selected from the group comprising:5 and22. A pharmaceutical composition or cosmeceutical composition comprising an effective amount of a compound of any one of claims 1 to 21 and a suitable carrier, diluent or excipient.

23. A method of treating or preventing any one or more of cancer, inflammation, diabetic nephropathy, a neurodegenerative disorder, multiple sclerosis, and a dermatological condition comprising administering a pharmaceutically effective amount of a compound of any one of claims 1 to 21 to a patient requiring treatment.

24. A method as claimed in claim 23, wherein the neurodegenerative disorder is senile dementia, pre-senile dementia, multi-infarct dementia or Alzheimer's disease.

25. A method of rejuvenating skin or preventing skin-aging comprising administering an effective amount of a compound of any one of claims 1 to 21 to human skin.

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