This invention relates to covalently bonded conjugates of cyclodextrins and deferasirox or an analogue thereof, pharmaceutical compositions comprising such conjugates and methods to make such conjugates. The conjugates may be useful in the treatment of various medical conditions including metal overload, such as iron overload and copper overload.

Background

Deferasirox, 4-[bis(2-hydroxyphenyl)-1/-/-1 ,2,4-triazol-1-yl]benzoic acid (compound 1), is an iron chelator which is used clinically to treat iron overload (haemochromatosis) resulting from frequent blood transfusions, for example in patients with beta thalassaemia. Iron overload can lead to reactive oxygen species generation, disorders related to metal dyshomeostasis, oxidative stress, organ damage and myelodysplastic syndromes (MDS), and cancers such as blood cancers, which can lead to diseases such as acute myeloid leukaemia (AML).

Deferasirox is sold by Novartis under the name Exjade®. It is available commercially as film-coated tablets for oral administration.

Compound 1

Whilst deferasirox is an effective treatment for iron overload, it can lead to undesirable side effects including gastrointestinal disorders, impaired renal function and impaired hepatic function. In addition, some patients have been shown to experience drug-related toxicity (hepatotoxicity) from deferasirox, and so cannot tolerate it (Lee et al., PLoS ONE 8(5), 2013). It would therefore be desirable to provide alternative forms of deferasirox which are still effective for the treatment of iron overload but which have the potential to give rise to fewer or less severe side effects.

Cyclodextrins are cyclic molecules formed from 5 or more, usually 6 or more, 1 4 linked oD-glucopyranoside units. Cyclodextrins are extensively used in the pharmaceutical industry as formulating agents, for taste masking, for forming host-guest complexes or for encapsulating hydrophobic molecules (such as cholesterol and lipids) such as in the treatment of lysosomal disorder diseases, Niemann-Pick disease type C, or hypervitaminosis. For example, hydroxypropyl beta-cyclodextrin is used in the treatment of Niemann-Pick disease type C.

Deferiprone is another iron chelator which is used clinically to treat iron overload. The iron binding and antioxidant activity of two conjugates of deferiprone with b-cyclodextrins have been investigated (Puglisi et al, Dalton Trans., 2012, 41 , 2877-2883).

It is known to increase the solubility of drugs such as deferasirox by complexing the drug to a cyclodextrin such as hydroxypropyl beta-cyclodextrin (Dandagi et al., International Journal of Pharmacy and Pharmaceutical Sciences, Vol. 6, Suppl 2, 2014; and WO 2012/042224). However, in this case the drug (e.g. deferasirox) and the cyclodextrin (e.g. hydroxypropyl beta- cyclodextrin) are not covalently bonded together, but rather are held together by intermolecular forces.

Deferasirox has been covalently linked to calixarene macrocycles for use as an iron sensor (Rouge et al., Tetrahedron, 67 (2011), p2916-2924). However, compounds containing deferasirox covalently bonded to a cyclodextrin are not disclosed.

In contrast, the present invention provides covalently bonded conjugates of cyclodextrins and deferasirox or an analogue thereof. The compounds of the invention can be used in the treatment of various medical conditions including metal overload, such as iron overload and copper overload. In addition, the undesirable side effects that can occur from the use of deferasirox are minimised.

The compounds of the invention can also be used in the simultaneous treatment of (i) diseases which may benefit from treatment with a metal chelator (e.g. metal dyshomeostasis), and (ii) diseases which may benefit from treatment with a cyclodextrin or cyclodetrin derivative (e.g. lipid imbalance). Summary of Invention

The invention provides compounds of Formula I:

Formula I and deuterated or isotopically labelled derivatives, stereoisomers, geometric isomers, tautomers and pharmaceutically acceptable salts thereof, wherein CD, L, y and x are as hereinafter defined. Also provided are pharmaceutical compositions comprising such compounds, therapeutic methods using such compounds and methods to make such compounds.

The invention further provides a compound of Formula I, or a deuterated or isotopically labelled derivative thereof, or a stereoisomer, a geometric isomer, a tautomer or a

pharmaceutically acceptable salt thereof, for use in therapy.

The invention also provides a compound of Formula I, or a deuterated or isotopically labelled derivative thereof, or a stereoisomer, a geometric isomer, a tautomer or a

pharmaceutically acceptable salt thereof, for use in the treatment of metal dyshomeostasis, for example iron overload or copper overload, particularly iron overload.

The invention also provides a compound of Formula I, or a deuterated or isotopically labelled derivative thereof, or a stereoisomer, a geometric isomer, a tautomer or a

pharmaceutically acceptable salt thereof, for use in the treatment of cancer, preferably wherein the cancer is blood cancer such as myeiodyspiastic syndromes or acute myeloid leukemia.

The invention also provides a compound of Formula I, or a deuterated or isotopically labelled derivative thereof, or a stereoisomer, a geometric isomer, a tautomer or a

pharmaceutically acceptable salt thereof, for use in the treatment of lysosomal storage diseases. The invention also provides a compound of Formula I, or a deuterated or isotopically labelled derivative thereof, or a stereoisomer, a geometric isomer, a tautomer or a

pharmaceutically acceptable salt thereof, for use in the treatment of oxidative stress.

The invention also provides a compound of Formula I, or a deuterated or isotopically labelled derivative thereof, or a stereoisomer, a geometric isomer, a tautomer or a

pharmaceutically acceptable salt thereof, for use in the treatment of lipid imbalance.

The invention also provides a compound of Formula I, or a deuterated or isotopically labelled derivative thereof, or a stereoisomer, a geometric isomer, a tautomer or a

pharmaceutically acceptable salt thereof, for use in the simultaneous treatment of (i) metal dyshomeostasis, for example iron overload or copper overload, particularly iron overload; and (ii) lipid imbalance.

The invention also provides a compound of Formula I, or a deuterated or isotopically labelled derivative thereof, or a stereoisomer, a geometric isomer, a tautomer or a

pharmaceutically acceptable salt thereof, for use in the detection or diagnosis of metal dyshomeostasis, for example iron overload or copper overload, particularly iron overload.

The invention further provides a method of treatment of metal dyshomeostasis, for example iron overload or copper overload, particularly iron overload, the method comprising administering a therapeutically effective dose of a compound of Formula I, or a deuterated or isotopically labelled derivative thereof, or a stereoisomer, a geometric isomer, a tautomer or a pharmaceutically acceptable salt thereof, to a patient in need thereof.

The invention further provides a method of treatment of cancer, for example blood cancer such as myelodyspiastic syndromes or acute myeloid leukemia, the method comprising administering a therapeutically effective dose of a compound of Formula I, or a deuterated or isotopically labelled derivative thereof, or a stereoisomer, a geometric isomer, a tautomer or a pharmaceutically acceptable salt thereof, to a patient in need thereof.

The invention further provides a method of treatment of lysosomal storage diseases, the method comprising administering a therapeutically effective dose of a compound of Formula I, or a deuterated or isotopically labelled derivative thereof, or a stereoisomer, a geometric isomer, a tautomer or a pharmaceutically acceptable salt thereof, to a patient in need thereof.

The invention further provides a method of treatment of oxidative stress, the method comprising administering a therapeutically effective dose of a compound of Formula I, or a deuterated or isotopically labelled derivative thereof, or a stereoisomer, a geometric isomer, a tautomer or a pharmaceutically acceptable salt thereof, to a patient in need thereof.

The invention further provides a method of treatment of lipid imbalance, the method comprising administering a therapeutically effective dose of a compound of Formula I, or a deuterated or isotopically labelled derivative thereof, or a stereoisomer, a geometric isomer, a tautomer or a pharmaceutically acceptable salt thereof, to a patient in need thereof. The invention further provides a method of simultaneous treatment of (i) metal dyshomeostasis, for example iron overload or copper overload, particularly iron overload; and (ii) lipid imbalance, the method comprising administering a therapeutically effective dose of a compound of Formula I, or a deuterated or isotopically labelled derivative thereof, or a stereoisomer, a geometric isomer, a tautomer or a pharmaceutically acceptable salt thereof, to a patient in need thereof.

The invention further provides the use of a compound of Formula I, or a deuterated or isotopically labelled derivative thereof, or a stereoisomer, a geometric isomer, a tautomer or a pharmaceutically acceptable salt thereof, in the manufacture of a medicament for the treatment of metal dyshomeostasis, for example iron overload or copper overload, particularly iron overload; or for the treatment of cancer, preferably wherein the cancer is blood cancer such as myelodysplastic syndromes or acute myeloid leukemia; or for the treatment of lysosomal storage diseases, oxidative stress, or lipid imbalance; or for the simultaneous treatment of (i) metal dyshomeostasis, for example iron overload or copper overload, particularly iron overload; and (ii) lipid imbalance.

Brief Description of the Drawings

Figure 1 shows the results of a dose-response study comparing compounds of the invention with deferasirox.

Figure 2 shows the results of a toxicity study comparing compounds of the invention with deferasirox.

Figure 3 shows the results of an anti-aggregation assay.

Figure 4 shows the results of an antioxidant capacity assay.

Detailed Description

The invention provides compounds of Formula I:

Formula I or a deuterated or isotopically labelled derivative thereof, or a stereoisomer, a geometric isomer, a tautomer or a pharmaceutically acceptable salt thereof wherein:

CD denotes an optionally substituted cyclodextrin moiety or an optionally substituted cyclodextrin polymer moiety;

each L is independently a bond or a covalent linker attached to a carbon atom in CD; and

x and y independently denote integers greater than or equal to 1.

The linker L is preferably stable in vivo, such that the cyclodextrin moiety is not removed from the rest of the compound of Formula I by cleavage of the linker in vivo.

The value of x will depend on the number of sites available in the cyclodextrin moiety for attachment to a linker. For non-polymeric cyclodextrin moieties, x preferably denotes 1-8, more preferably x denotes 1 or x denotes 6, 7 or 8. For polymeric cyclodextrin moieties, the value of x will also depend on the degree of polymerisation, for example x may denote 1-8, preferably 1 , 6, 7 or 8, times the number of CD monomer units making up the polymer.

The value of y will depend on the number of sites in each linker L which are available for attachment to a deferasirox moiety. Preferably, y denotes 1-3, more preferably y denotes 1 or 2, even more preferably 1.

Preferred compounds of the invention include those of formula II:

Formula II

wherein L, x, y and CD are as defined above.

Preferred are compounds of the invention wherein:

L denotes -Z ¹ -W ¹ -Q-W ² -Z ² -, wherein Z ² is attached to the CD moiety;

Z ¹ denotes C=0, C(0)NR ¹ , CHR ² , CH ₂ NR ¹ , or C(0)0, preferably C=0 or C(0)NH, more preferably C(0)NH;

W ¹ is absent or denotes CM ₆ alkylene or CM ₆ alkenylene, wherein the Ci_i ₆ alkylene or C1-16 alkenylene may optionally include one or more heteroatoms independently selected from O, S and NR ¹ and/or may optionally include 1 or more, preferably 1 or 2, C=0 or C=S groups in the carbon chain;

Q is absent or denotes -CºC-, a 5 to 10 membered carbocyclic ring system, or a 5 to 10 membered heterocyclic ring system;

W ² is absent or denotes C1.16 alkylene or Ci_i _S alkenylene, wherein the Ci_i _S alkylene or C1-16 alkenylene may optionally include one or more heteroatoms independently selected from O, S and NR ¹ in the carbon chain and/or may optionally include 1 or more, preferably 1 or 2, C=0 or C=S groups in the carbon chain;

Z ² is absent or denotes S, O, C(0)NR ¹ , NR ¹ C(0) or NR ¹ , preferably S or NR ¹ ; each R ¹ independently denotes H, Ci _-20 alkyl, Ci_i ₂ alkenyl, C _3.i0 cycloalkyl, phenyl or benzyl, wherein the Ci_ ₂₀ alkyl, C _M0 alkenyl, C _3-i0 cycloalkyl, phenyl or benzyl may be optionally substituted by one or more substituents selected from halo, -OH, -N ₃ or NR ⁴ R ⁴ and wherein the C _1-20 alkyl group may include one or more S, C=S. C=0, NR ³ or O groups in the carbon chain; each R ² independently denotes H or OH; each R ³ independently denotes H or Ci.io alkyl, optionally substituted by NR ⁴ R ⁴ ; and each R ⁴ independently denotes H or Ci_io alkyl, wherein the C _MO alkyl may be optionally substituted by guanidino; and wherein the linker L may optionally be substituted at any available position with one or two substituents selected from groups of formula III or formula IV, preferably formula IV, or guanidino-Ci-io alkyl:

Formula III or

Formula IV

Preferred substitution positions for the groups of formula I I I or formula IV include any N atoms in primary or secondary amino groups in the linker L.

For reasons of stability, linker L is preferably not -C(0)S-.

When Z ¹ denotes CH ₂ , it is preferred that at least one of the W ¹ , Q, W ² or Z ² groups in the linker L is also present.

When Q denotes a 5 to 10 membered carbocyclic ring system or a 5 to 10 membered heterocyclic ring system, suitable such systems include phenylene, triazole, and 1 ,4-piperidine.

Preferred compounds of the invention include those wherein

Z ¹ denotes C=0 or C(0)NH, preferably C(0)NH; and/or

Q denotes phenylene, particularly 1 ,4-phenylene, triazole or 1 ,4-piperidine; and/or Z ² denotes S or NR ¹ .

Examples of suitable linkers L include the following:

C=0;

-C(0)NH-;

-C(0)NH(CH ₂ ) _n NH(CH ₂ ) _n -Z ² , preferably wherein Z ² denotes S or NR ¹ ;

-C(0)NH[(CH ₂ ) _n NH] _m (CH ₂ ) _n -Z ² , preferably wherein Z ² denotes S or NR ¹ ;

-C(0)NH(CH ₂ ) _n 0(CH ₂ ) _n -Z ² preferably wherein Z ² denotes S or NR ¹ ;

-C(0)NH[(CH ₂ )n0] _m (CH ₂ )n-Z ² , preferably wherein Z ² denotes S or NR ¹ ;

-C(0)NH(CH ₂ ) _n S(CH ₂ ) _ir Z ² , preferably wherein Z ² denotes S or NR ¹ ;

-C(0)NH[(CH ₂ )nS]m(CH ₂ ) _n -Z ² , preferably wherein Z ² denotes S or NR ¹ ;

-C(0)NH(CH ₂ ) _n S-;

-C(0)NH(CH ₂ ) _n NH-; -C(0)S(CH ₂ ) _n NH; and

-C(0)0(CH ₂ ) _n -Q-, preferably wherein Q denotes 1 ,2,3-triazole-;

wherein each n independently denotes 1-10 and each m independently denotes 1-10.

Examples of linkers comprising Q groups include -C(0)Q-, -C(0)NH(CH ₂ ) _n Q(CH ₂ ) _n -Z ² and -C(0)0(CH ₂ ) _n Q-, wherein n and Z ² are as previously defined. In such linkers, Q preferably denotes -CºC-, phenylene, particularly 1 ,4-phenylene, triazole (preferably 1 ,2,3-triazole) or 1 ,4- piperidine and/or Z ² denotes S or NR ¹ . Examples of such linkers include the following:

wherein each n independently denotes 1-10, preferably 1 or 2.

Preferred linkers L are those comprising at least one amide, ether or thioether linkage, more preferably at least one amide linkage.

More preferred linkers L include: -C(0)NH-; -C(0)S(CH ₂ ) _n NH; -C(0)0(CH ₂ ) _n -Q-, preferably wherein Q denotes 1 ,2,3-triazole-, -C(0)NH(CH ₂ ) _n S-; and -C(0)NH(CH ₂ ) _n NH-. Even more preferably, L is -C(0)NH-; -C(0)S(CH ₂ ) ₂ NH; -C(0)0(CH ₂ ) ₃ -Q-, wherein Q denotes 1 ,2,3- triazole-. Most preferably, L is -C(0)NH-.

The invention includes any and all combinations of preferred features disclosed herein, whether or not such combinations are explicitly disclosed.

Definitions

References herein to the compounds of the invention include deuterated or isotopically labelled derivatives, stereoisomers, geometric isomers, tautomers and pharmaceutically acceptable salts thereof.

Any alkyl, alkylene, alkenyl, alkenylene, alkynyl or alkynylene groups in the compounds of the invention can be linear (straight chained) or branched. Preferred alkyl, alkylene, alkenyl, alkenylene, alkynyl and alkynylene groups in any of the above compound definitions have 1 to 10 carbon atoms, preferably 1 to 6 carbon atoms, more preferably 1 to 4 carbon atoms.

As used herein, any carbocyclic groups may be saturated, partially unsaturated or fully unsaturated (aromatic). Such groups may be mono- or bicyclic.

Preferred aromatic carbocyclic groups are phenyl and naphthyl.

As used herein, any heterocyclic groups contain carbon atoms and from 1 to 4 hetero atoms selected from N, S and O. They may be mono- or bicyclic and saturated, partially unsaturated or fully unsaturated (heteroaromatic). Preferred saturated heterocyclic rings include piperidine. Preferred heteroaromatic groups include pyridyl and triazole.

As used herein, halogen or halo denotes F, Cl, Br or I, preferably Br or I

Tosylate means para-toluenesulphonate (P-CH ₃ C ₆ H ₄ SO ₃ ), meslyate means methanesulphonate (CH ₃ SO ₃ ) and triflate means trifluoromethanesulphonate (CF ₃ SO ₃ )·

Guanidino means -NHC(=NH)NH ₂ .

As used herein,“deferasirox moiety” refers to the portion of the compounds of the invention shown below:

The compounds of the invention may include atoms in any isotopic form. They may be enriched in non-natural isotopic forms (i.e. labelled). For example, deuterated compounds may have one or more deuterium atoms, D, substituted for any or all of the non-labile hydrogen atoms present in the compounds of formula I. Preferably, the non-labile hydrogen atoms are attached to carbon atoms in the compounds of the invention. Other isotopic labels which may be incorporated into the compounds of the invention include ³ H, ¹⁴ C, ¹³ C, ¹⁵ N, ¹²⁵ l, ¹⁸ F, and ³⁵ S.

Some compounds of the invention may form salts. The invention therefore includes pharmaceutically acceptable salts of the compounds of formula I. As used herein,

“pharmaceutically acceptable salts” refer to derivatives of the disclosed compounds wherein the parent compound is modified by making an acid or base salt thereof. Suitable pharmaceutically acceptable salts are known in the art. Lists of suitable such salts may be found for example in Remington: The Science and Practice of Pharmacy, 22 ^nd Edition, 2013, Pharmaceutical Press. Cvclodextrins

Cyclodextrins are cyclic molecules formed from 5 or more, usually 6 or more, 1 4 linked oD-glucopyranoside units. Two alternative depictions of alpha- (m is 0 and n is 6), beta- (m is 1 and n is 7) and gamma- (m is 2 and n is 8) cyclodextrin are shown below.

As used herein, cyclodextrin moiety or cyclodextrin polymer moiety means a cyclodextrin or cyclodextrin polymer attached to the linker L via a carbon atom in the cyclodextrin moiety or cyclodextrin polymer moiety. References herein to cyclodextrins or cyclodextrin moieties apply equally to cyclodextrin polymers or cyclodextrin polymer moieties.

Any cyclodextrin which is pharmaceutically acceptable and which has at least one site available for reaction to attach it to a linker or directly to a deferasirox moiety can potentially be used in the present invention. Suitable cyclodextrins, including pharmaceutical grade cyclodextrins, are available commercially, for example from Sigma-Aldrich, CycloLab, CTD Holdings, Inc. and Tokyo Chemical Industry UK Ltd., or may be synthesised using known methods.

The properties of the compounds of the invention such as solubility can be tailored through choice of the cyclodextrin moiety, including the type of cyclodextrin (for example alpha, beta or gamma) and the nature of any substituents on the cyclodextrin. For example, polar substituents such as guanidino may increase the aqueous solubility of the cyclodextrin, and hence of the compounds of the invention.

The cyclodextrin is preferably attached to the linker L via the carbon(s) at one or more of the 6-positions in the glucosyl units of the cyclodextrin. However, attachment at the 2 and/or 3 positions of one or more of the glucosyl units, instead of or as well as attachment at one or more of the 6-positions, is also possible. Alpha-cyclodextrin contains 6 glucosyl units, beta- cyclodextrin contains 7 glucosyl units and gamma-cyclodextrin contains 8 glucosyl units. If a linker L is attached to the 6-position of each of the glucosyl units in alpha-, beta- or gamma- cyclodextrin, x in Formula I will denote 6, 7 or 8 respectively.

Suitable cyclodextrins include alpha-, beta- and gamma-cyclodextrin and substituted derivatives thereof. Substituted derivatives can have one or more substituents, which may be the same or different. Suitable optional substituents for the cyclodextrin include amino, guanidino, Ci-C ₁₀ alkylamino, di-( CrC _{1 0} alkyl)amino, CrC _{1 0} alkyl, CrC _{1 0} hydroxyalkyl, CrC _{1 0} carboxyalkyl, C _r Ci ₀ alkylcarbonyl, benzoyl, succinyl, piperizinyl, amino-Ci.i ₀ alkyl, amino-Ci. ₁₀ alkylpiperizinyl, trimethylsilyl, sulfate and phosphate. Preferred C _r Ci ₀ alkyl groups include methyl, ethyl, butyl, pentyl and octyl. Preferred CrC _{1 0} hydroxyalkyl groups include hydroxyethyl and hydroxypropyl (preferably 2-hydroxypropyl). Preferred C -C carboxyalkyl groups include carboxymethyl and carboxyethyl. Preferred C -C alkylcarbonyl groups include acetyl.

Preferred cyclodextrin substituents include amino, guanidino, C -C alkylamino, piperizinyl, amino-Ci-i ₀ alkyl, and amino-Ci_ioalkylpiperizinyl.

Cyclodextrin polymers and copolymers may also be used to form the compounds of the invention, including beta-cyclodextrin polymer, beta-cyclodextrin/epichlorohydrin copolymer and cross-linked CD polymers such as those according to Li et al in Chem. Sci., 2016, 7, 905.

Amino-substituted cyclodextrins may be linked to the rest of the compound of formula I via the nitrogen atom of the amino group. Particularly preferred amino-substituted cyclodextrins for use to form the compounds of the present invention are 6 ^A -amino-6 ^A -deoxy-p-cyclodextrin and 3 ^A -amino-3 ^A -deoxy-2 ^A (S),3 ^A (R)^-cyclodextrin.

Unsubstituted cyclodextrins may be linked to the rest of the compound of formula I via any OH group.

Preferred compounds of the invention include those of Formula I wherein the optionally substituted cyclodextrin moiety or optionally substituted cyclodextrin polymer moiety is attached to the rest of the compound via an amide linkage.

Particularly preferred compounds of the invention are shown below:

Compound 3 (CD2)

CD4 Compound 2 (CD1) and compound 3 (CD2) are particularly preferred.

Conjugate formation

The compounds of the invention are formed by covalently linking one or more molecules of deferasirox or an analogue thereof to a cyclodextrin moiety, optionally via a linker.

Deferasirox is commercially available or may be synthesised according to known methods, such as those disclosed in W097/49395 and US8,772,503.

Deferasirox itself has a carboxylic acid group which can react with a functional group in the cyclodextrin moiety or a linker attached to the cyclodextrin moiety to form a compound of the invention. For example, reaction of the carboxyl group in deferasirox with an amino functional group will lead to an amide linkage whilst reaction with a hydroxyl functional group will lead to an ester linkage. Ester linkages may be more prone to hydrolysis in vivo than amide linkages. However, compounds comprising sterically hindered ester linkages may be resistant enough to hydrolysis to be used in vivo. In addition, compounds comprising ester linkages may be useful in in vitro applications.

Scheme 1 illustrates two alternative methodologies for linking the deferasirox moiety and the CD moiety via an amide linkage. In the first method (Method 1), deferasirox is reacted with an amino-substituted cyclodextrin moiety (when the (linker) in compound 4 is absent) or with an amino-terminated linker attached to the cyclodextrin moiety (when the (linker) in compound 4 is present). The -C(0)NR ¹ -(linker)- in compound 5 corresponds to L in the compounds of Formula I.

Alternatively, a carboxyl-terminated linker can be attached to the deferasirox moiety which is then reacted with an amino functional group in a cyclodextrin moiety (Method 2, when the (linker ² ) in compound 6 is absent) or a linker attached to a cyclodextrin moiety (when the (linker ² ) in compound 6 is present) to form an amide linkage. The linker ¹ -C(0)NR ¹ -(linker ² )- in compound 7 corresponds to L in the compounds of Formula I.

If there is more than one amino group in the NHR ¹ -(linker)-CD or NHR^Iinker^-CD compounds available for reaction to form an amide bond, then multiple deferasirox moieties may be attached to each CD moiety.

Method 1

Method 2

Scheme 1

The same methodologies may be adapted for the formation of different types of linkage by suitable selection of the functional groups on the deferasirox moiety and/or the CD moiety.

Primary amines are preferred over secondary amines for the formation of amide linkages. Methods for the formation of amide bonds are known in the art. For example, the coupling reaction may take place in the presence of an activating agent and/or a coupling reagent. Suitable activating and coupling agents for amide bond formation are known in the art -and include those normally used in peptide synthesis. Such activating agents are normally designed to react with a carboxylic acid to form an activated compound which then reacts with an amine to form the desired amide bond. The carboxylic group in the deferasirox moiety can also be converted into an acid chloride prior to reaction with a functionalised cyclodextrin or attachment to a linker. The activating or coupling agent may be present on a solid support.

Suitable activating agents include 1-hydroxybenzotriazole (HOBt), HOBt-6- sulfonamidomethyl resin HCI, HOOBt (hydroxy-3, 4-dihydro-4-oxo-1 , 2, 3-benzotriazine), HOSu (N-hydrosxysuccinimide), HOAt (1-hydroxy-7-aza-1 H-benzotriazole), ethyl 2-cyano-2- (hydroxyimino)acetate and propylphosphonic anhydride (T3P), all of which may optionally be present on a solid support.

Suitable coupling agents include carbodiimides such as DCC (dicyclohexylcarbodiimide), DIC (diisopropylcarbodiimide) and EDC HCI (N-(3-dimethylaminopropyl)-N’- ethylcarbodiimideHCI). Other suitable coupling reagents include BOP ((benzotriazol-1-yloxy)- tris(dimethylamino)phosphonium hexafluorophosphate), PyPOB (benzotriazol-1-yl- oxytripyrrolidinophosphonium hexafluorophosphate), PyBrOP (bromo-tripyrrolidino- phosphonium hexafluorophosphate), PyAOP ((7-azabenzotriazol-1- yloxy)tripyrrolidinophosphonium hexafluorophosphate), PyOxim ((ethyl

cyano(hydroxyimino)acetato-0 ² )tri-1-pyrrolidinylphosphonium hexafluorophosphate), DEPBT (3- (diethoxyphosphoryloxy)-l ,2,3-benzotriazin-4(3H)-one), TBTU (2-(1 H-benzotriazole-1-yl)- 1 ,1 ,3,3-tetramethyluronium tetrafluoroborate), HBTU (2-(1 H-benzotriazole-1-yl)-1 , 1 ,3,3- tetramethyluronium hexafluorophosphate), HCTU (0-(6-chlorobenzotriazol-1-yl)-N,N,N',N'- tetramethyluronium hexafluorophosphate), HDMC (N-[(5-chloro-3-oxido-1 H-benzotriazol-1-yl)-4- morpholinylmethylene]-N-methylmethanaminium hexafluorophosphate), HATU (1- [bis(dimethylamino)methylene]-1 H-1 ,2,3-triazolo[4,5-b]pyridinium 3-oxide hexafluorophosphate), COMU (1-cyano-2-ethoxy-2-oxoethylidenaminooxy)dimethylamino-morph olino-carbenium hexafluorophosphate), TOTT (S-(1-oxido-2-pyridyl)-N,N,N',N'-tetramethylthiuronium

tetrafluoroborate), TFFH (fluoro-N,N,N',N'-tetramethylformamidinium hexafluorophosphate), EEDQ (N-ethoxycarbonyl-2-ethoxy-1 ,2-dihydroquinoline), T3P (propanephosphonic acid anhydride), DMTMM (4-(4,6-dimethoxy-1 ,3,5-triazin-2-yl)-4-methylmorpholinium salts), BTC (bis-trichloromethylcarbonate), and CDI (1 ,Tcarbonyldiimidazole).

Amide bonds can also be formed by microwave condensation without the need for any coupling agent.

Scheme 2 illustrates another general methodology for synthesis of the compounds of the invention via nucleophilic displacement. In option a, a deferasirox moiety is attached, optionally via a linker ¹ , to a nucleophilic group, for example an amino group (-NH ₂ ) or a thiol group (-SH). The functionalised deferasirox moiety 24 can then react with a leaving group LG attached directly or via a linker (linker ² ) to a cyclodextrin moiety (compound 8) in a nucleophilic displacement reaction to attach the deferasirox moiety to the cyclodextrin moiety either directly or via a linker. In option b, a deferasirox moiety is attached to a leaving group, optionally via a linker (compound 25). The functionalised deferasirox moiety 25 can then react with a nucleophilic group attached directly or via a linker (linker ² ) to a cyclodextrin moiety (compound 9) in a nucleophilic displacement reaction to attach the deferasirox moiety to the cyclodextrin moiety either directly or via a linker. Suitable leaving groups LG for both options are known in the art and include Cl, Br, I, tosylate, mesylate, and triflate.

Option a

wherein Nu denotes a nucleophilic group

and LG denotes a leaving group

Option b

p g p

and LG denotes a leaving group

Scheme 2 Amino functionalised cyclodextrins for use in forming the compounds of the invention may be prepared by known methods. For example, the corresponding iodides or bromides 11 may be reacted with a primary amine NH ₂ R ¹ as illustrated in Scheme 3 below. 6-Deoxy-6-halo- substituted cyclodextrins are available commercially, for example from Cycloab, or can be synthesised using known methods (see for example A. Gadelle et al, J Agnew Chem., Int. Ed. Engl., 1991 , 30, 78-80). Alternatively, the iodide or bromide 11 may be reacted with sodium azide to introduce an azide group (C. Roehri-Stoeckel et al, Tetrahedron Letters, 1997, 38(9), 1551-1554) which can then be reduced to provide the corresponding amino compound 14.

Scheme 3

Azido-functionalised cyclodextrins such as 13 can also be further functionalised via Cu ¹ - catalyzed azide-alkyne cycloaddition (so-called“click” chemistry) to form triazine rings (Diaz- Moscoso et al, ChemMedChem, 6(1 ), 2011 , pages 181-192; C. Roehri-Stoeckel et al,

Tetrahedron Letters, 1997, 38(9), 1551-1554; Chui et al, Org. Biomol. Chem., 2014, 12, 3622).

If the alkyne 15 used in the reaction has an optionally protected amino functional group, as illustrated in Scheme 4, this can be used in a subsequent amide bond formation reaction to link it to a deferasirox moiety. Suitable amine protecting groups are known in the art (see for example Greene's Protective Groups in Organic Synthesis: Fifth Edition, Ed. Peter G. M. Wuts, John Wiley & Sons, 2014) and include fe/f-butoxycarbonyl (BOC), carbobenzyloxy (Cbz), benzyl (Bn), p-methoxybenzylcarbonyl (Moz), 9-fluorenylmethyloxycarbonyl (FMOC) and benzoyl (Bz).

P denotes an optional protecting group

Scheme 4

Alternatively, an alkyne can be coupled to a deferasirox moiety first, before the

Cu ¹ -catalyzed azide-alkyne cycloaddition is carried out. This is illustrated in Scheme 5 below. The alkyne can be coupled to the deferasirox moiety by reaction of a suitable amino-substituted alkyne, for example propargylamine, with deferasirox to form an amide linkage. Propargylamine is commercially available.

Scheme 5

If a halo-substituted aminoalkyne such as 1-amino-4-chloro-2-butyne is coupled to a deferasirox moiety via amide bond formation, the resulting compound can be linked to a CD moiety via nucleophilic displacement of the halo group as in Scheme 2, option b.

Halo-substituted cyclodextrins such as 11 can be reacted with other nucleophilic functional groups, in addition to amines, present in either a linker or a deferasirox moiety according to Scheme 2. For example, reaction with a thiol can lead to formation of a thioether linkage and reaction with an alkoxide can lead to formation of an ether linkage. Alternatively, at least one of the free hydroxyl groups in a cyclodextrin may be converted directly to a leaving group such as triflate or tosylate and then reacted with a suitable nucleophile.

As shown in Scheme 6 below, deferasirox can be converted to the halo-substituted compound 19, for example via reduction of the acid group to the corresponding alcohol followed by treatment with a halogenating agent. Suitable reagents for reduction of acids to alcohols are known in the art and include boranes such as borane tetrahydrofuran (BH ₃ -THF). Suitable halogenating agents are also known in the art and include HBr, PPh ₃ /Br ₂ , PPh ₃ /CBr ₄ , PPh ₃ /CI ₄ , PPh ₃ /l ₂ , and PPh ₃ /CH ₃ l. Reaction with a nucleophilic functional group in a linker or a functionalised cyclodextrin, for example an amino-cyclodextrin, will then allow attachment of the linker or the cyclodextrin to the deferasirox moiety to form compound 20.

Scheme 6

Reduction of the carboxyl group on deferasirox to the corresponding alcohol followed by conversion of the alcohol to a leaving group such as tosylate or triflate would provide an alternative route for attachment of a linker or a cyclodextrin moiety via reaction with a nucleophilic functional group in the linker or cyclodextrin moiety. The alcohol could also react with a carboxylic acid group in a linker or a cyclodextrin moiety to form an ester bond.

In another method for the formation of the compounds of the invention, the carboxyl group in deferasirox 1 can be reduced to give the corresponding aldehyde 21 which may then be subjected to a reductive animation. The amine used in the reductive animation reaction could be part of a linker, optionally already attached to a CD moiety, or could be an

amino-functionalised CD. This reaction is shown in schematic form in Scheme 7 below:

Scheme 7

Ester linkages may be formed by methods known in the art. For example,

halo-substituted alkynes 23 may be reacted with deferasirox under basic conditions as shown in Scheme 8 and Example 38. The alkyne group in the resulting ester 24 can then be coupled to an azido-functionalised cyclodextrin via Cu ¹ -catalyzed azide-alkyne cycloaddition.

Scheme 8

Thioester linkages may also be formed by methods known in the art. For example, suitably protected amino-substituted thiols may be reacted with deferasirox as illustrated in

Scheme 9 and Example 37 to form amino-functionalised thioesters. Following deprotection, the amino group can then be coupled with a suitably functionalised cyclodextrin moiety using the methodology discussed previously.

where P denotes a protecting group

Scheme 9

Cyclodextrins can also be modified by attachment of alkene-containing functional groups, using methodology analogous to that discussed above for alkyne functionalisation. The alkene bonds can then be attached to a deferasirox moiety attached to a linker containing an alkene group via a cross-metathesis reaction, or to a halo-substituted deferasirox moiety via a Heck reaction.

The Tables below shows various functionalised cyclodextrins which can be used to form a compound of the invention by attachment to one or more deferasirox moieties, optionally via an additional linker.

n = 6-8

Ref 1: Diaz-Moscoso etal, ChemMedChem, 6(1), 2011, pages 181-192

Ref.2: L. Gallego-Yerga et al, Org. Biol. Chem, 2015, 13, 1708-1723

Ref 3: Karginov et el, Antimicrobial Agents and Chemotherapy, Nov.2006, 3740-3753

n = 6-8

Depending on the reactions used to form the compounds of the invention, it may be necessary to protect some or all of the other functional groups present in the deferasirox moiety, the cyclodextrin moiety and/or the linker L. Suitable protecting groups are known in the art, for example from Greene's Protective Groups in Organic Synthesis: Fifth Edition, Ed. Peter G. M. Wuts, John Wiley & Sons, 2014.

Pharmaceutical Formulations

The compounds of the present invention are preferably administered in a pharmaceutical formulation along with pharmaceutically acceptable excipients. Suitable pharmaceutically acceptable excipients are known in the art (see, for example, Handbook of Pharmaceutical Excipients, Sheskey, Paul J; Cook, Walter G; Cable, Colin G, Eighth Edition, Pharmaceutical Press, 2017). Methods for making pharmaceutical formulations are also known in the art.

Preferably, compounds of the present invention are formulated for oral administration. Suitable formulations for oral administration include tablets, capsules, dispersible powders and suspensions, preferably tablets.

Tablets for oral administration may comprise diluents or fillers, binders, disintegrants, lubricants, coloring agents and/or preservatives in addition to the active ingredient. Suitable binders include alginate, candelilla wax, carnuba wax, corn starch, ethyl cellulose, and hydroxypropylcellulose. Suitable disintegrants include croscarmellose sodium, microcrystalline cellulose and povidone. Suitable lubricants include talc, calcium stearate, magnesium stearate, poloxamers and polyethylene glycols. Suitable diluents include crospovidone, lactose, mannitol, sorbitol, and sucrose,

Preferred excipients for tablet formulation include microcrystalline cellulose,

crospovidone, povidone, magnesium stearate, silica and poloxamers

Tablets for oral administration are preferably film-coated. Suitable coating formulations comprise a film-forming polymer, a plasticiser and a colorant.

The dosage of the compounds of the invention will depend on the medical condition and the patient being treated. Doses in the range of 10 and 120 mg/kg, preferably 20-80 mg/kg. Preferably, each dosage unit, for example each tablet, will contain a weight of the compound of the invention which provides a dosage of active ingredient which is equivalent to 90, 180 or 360 mg of deferasirox.

Uses

The compounds of the invention may be useful for the treatment of metal

dyshomeostasis, for example iron overload or copper overload, particularly iron overload.

The compounds of the invention may also be useful for the detection of metal dyshomeostasis, for example iron overload or copper overload, particularly iron overload. The compounds of the invention may be useful for the treatment of iron overload (haemochromatosis) including haemosiderosis. Iron overload can be caused by the genetic disorder hereditary haemochromatosis (HHC) or be the result of frequent blood transfusions during the treatment of anaemias such as beta thalassaemia, sickle cell anaemia, leukaemia, aplastic anaemia or myelodysplastic syndromes (MDS). Haemosiderosis is an iron overload disorder resulting in the formation of haemosiderin. It can result from diseases such as Goodpasture's syndrome, granulomatosis with polyangiitis, and idiopathic pulmonary haemosiderosis.

The compounds of the invention show similar solubility and iron-binding properties compared to deferasirox but show surprisingly lower toxicity. The compounds of the invention may therefore give rise to fewer or less severe side effects compared to deferasirox itself.

The compounds of the invention may also be useful in the treatment of other diseases which may benefit from treatment with a metal chelator, including Niemann Pick C disease, Parkinson’s disease (PD), Alzheimer’s disease (AD), Wilson’s disease and dementia with Lewy bodies (DLB). The compounds of the invention may also be useful in the treatment of cancer, preferably wherein the cancer is blood cancer such as myelodysplastic syndromes or acute myeloid leukemia.

Similarly, the compounds of the invention may also be useful in the treatment of other diseases which may benefit from treatment with a cyclodextrin or cyclodetrin derivative, including lysosomal storage diseases, oxidative stress, and lipid imbalance.

The compounds of the invention may also be used in the simultaneous treatment of (i) diseases which may benefit from treatment with a metal chelator, and (ii) diseases which may benefit from treatment with a cyclodextrin or cyclodetrin derivative.

Thus, the compounds of the invention may also be useful in the simultaneous treatment of: (i) metal dyshomeostasis (such as iron or copper overload, preferably iron overload), cancer (preferably blood cancer such as myelodysplastic syndromes or acute myeloid leukemia), Niemann Pick C disease, Parkinson’s disease (PD), Alzheimer’s disease (AD), Wilson’s disease or dementia with Lewy bodies (DLB); and

(ii) lysosomal storage diseases, oxidative stress, or lipid imbalance.

For example, the compounds of the invention may be useful in the treatment of (i) metal dyshomeostasis (such as iron or copper overload, preferably iron overload); and (ii) lipid imbalance.

The aggregation of proteins into toxic conformations plays a critical role in the development of different neurodegenerative disorders such as PD, AD and DLB. DLB and PD are amongst a group of diseases referred to as a-synucleinopathies that are characterized by o synuclein (aSyn) accumulation in cortical and subcortical regions. Therefore, compounds that can delay and/or prevent the aggregation process of aSyn could potentially lead to a new therapeutic strategy for treating PD and other a-synucleinopathies. aSyn consists of 140 amino acid residues and in its monomeric form is an intrinsically disordered protein with no persistent secondary structure. The interaction of proteins with metals often plays a crucial role in the aggregation. In particular, copper and iron are the most effective ions in promoting aSyn aggregation. The compounds of the invention can chelate iron and copper and are therefore potentially useful in the of treatment of a-synucleinopathies.

Mounting evidence suggests a pivotal role of oxidative stress in neurodegenerative diseases such as PD. Antioxidants such as polyphenols and vitamins A and E have been proposed as therapeutic agents for preventing and reducing the rate of progression of PD. The compounds of the invention demonstrate antioxidant activity, and may therefore be useful for the treatment of PD for this reason.

Compounds of the invention may also be of use in vitro, for example in assays including diagnostic assays, or in sensors including electrochemical iron sensors.

The present invention covers all combinations of preferred features disclosed herein, even if not explicitly disclosed. All references referred to herein are incorporated by reference.

The invention will be illustrated further by the following non-limiting examples.

6 ^A -Amino-6 ^A -deoxy-p-cyclodextrin was synthesized by a microwave-assisted procedure starting from the corresponding 6-tosylate derivative using a literature method (Puglisi, A., Spencer, J., Clarke, J., & Milton, J. (2012), Microwave-assisted synthesis of 6-amino-p- cyclodextrins, Journal of Inclusion Phenomena and Macrocyclic Chemistry, 73(1-4), 475-478). 3 ^A -Amino-3 ^A -deoxy-2 ^A (S),3 ^A (R)-p-cyclodextrin was obtained from Tokyo Chemical Industry UK Ltd.. Deferasirox was purchased from Molekula, UK, and human aSyn was purchased from Sigma-Aldrich. Cu ²⁺ stock solutions were prepared by dissolving the corresponding perchlorate in water and titrating the resulting solutions with standardized EDTA by using murexide.

The sugar units in b-CyD derivatives are labelled A-G counter-clockwise starting from the modified ring (denoted as A) and viewing from the upper rim. The deferasirox moiety numbering is as shown above for compound 1.

Proton NMR spectra were measured on a Varian VNMRS solution-state spectrometer at 500 MHz at 30°C using residual isotopic solvent (CHCI ₃ , d _H = 7.27 ppm, DMSO, d _H = 2.50 ppm, MeOH, d _H = 3.31 ppm, H ₂ 0, d _H = 4.79 ppm) as internal reference. Chemical shifts are quoted in parts per million (ppm). Coupling constants (J) are recorded in Hertz (Hz).

Carbon NMR spectra were recorded at 125 MHz on a Varian 500 MHz spectrometer and are proton decoupled, using residual isotopic solvent (CHCI ₃ , d ₀ = 77.00 ppm, DMSO, d ₀ =

39.52 ppm, MeOH, d ₀ = 49.00 ppm, except D ₂ 0 internal standard was DSS-d ₆ (4, 4-di methyl-4- silapentane-1 -sulfonic acid). Proton and carbon spectra assignments are supported by variable temp. NMR, DEPT, HSQC and gCOSY.

Infrared spectra were recorded on a Perkin Elmer FT-IR spectrometer as either an evaporated film or liquid film or as a solid. Absorption maxima are reported in wave numbers (cm ^-1 ). Only significant absorptions are presented in the data, with key stretches identified in brackets.

High resolution mass spectra (HRMS) were acquired by electrospray ionization (ESI) on MAT95 XP. The mass to charge ratio (m/z) are given as the molecular ion [M+H] Low resolution mass spectrometry data were recorded on Fission Instrument VG autospec at 70 eV. HRMS data (ESI) were recorded in-house on the Bruker Daltonics, Apex III, ESI source: Apollo ESI with methanol as spray solvent. Only molecular ions, fractions from molecular ions and other major peaks are reported as mass/charge (m/z) ratios.

Dynamic light-scattering (DLS) measurements were carried out using a Zetasizer Nano ZS (Malvern Instruments, UK) equipped for backscattering at 173° with a 633 nm He-Ne laser. Each DLS measurement was run using automated, optimal measurement times and laser attenuation settings according to literature (Oliveri, V., Sgarlata, C., & Vecchio, G. (2016), Cyclodextrins 3-Functionalized with 8-Hydroxyquinolines: Copper-Binding Ability and Inhibition of Synuclein Aggregation, Chemistry-An Asian Journal, 1 1 (17), 2436-2442).

All reactions were conducted under an atmosphere of nitrogen unless otherwise stated. Anhydrous solvents and reagents were used as purchased e.g. DMF ACS reagent, >99.8% or were purified under nitrogen as follows: dichloromethane from calcium hydride, THF from sodium wire/benzophenone and methanol from 3A molecular sieves overnight.

Melting point measurements are recorded on a Griffen Gallenkamp melting point apparatus.

General cyclodextrin synthesis was performed using standard methods:

Abbreviations:

HOBt hydroxybenzotriazole

DCC L/,L/'-dicyclohexylcarbodiimide

DMF L/,L/'-dimethylformamide

TLC thin layer chromatography

WT wild type

NPC1 Niemann Pick C-1

CHO Chinese Hamster ovary

THF tetrahydrofuran

DMAP 4 H > tr¾ »t h V I ** i n o ovr i <1 i n s

DCU dicyclohexylurea

Example 1 : Synthesis of 4-r3.5bis(2-hvdroxyphenyl)-1 H-1.2.4-triazol-1 -yH-N-r6 ^A -amino-6 ^A - deoxy-B-cyclodextrinlbenzamide (compound 2 - CPU

HOBt (24 mg, 0.175 mmol), DCC (36 mg, 0.175 mmol) and triethylamine (50 mI_, 0.53 mmol) were added to a solution of deferasirox (65.5 mg, 0.175 mmol) in dry DMF (5 ml_). After 20 min, 6 ^A -amino-6 ^A -deoxy-p-cyclodextrin (205 mg, 0.175 mmol) was added. The reaction was stirred at room temperature under argon for 48 h. The solvent was evaporated to dryness in vacuo. The crude product was then triturated with diethyl ether (3 x 3 ml_), followed by an acetone trituration cold (3 ^c 4 mL) and hot acetone trituration (3 « 3 mL) and the crude solids were purified by reverse phase chromatography using a 30g Biotage KP-RC-18 column (eluent: H ₂ 0 CI-l ₃ 0l-l) to give the title product (white solid). Yield: 64 % (168 mg, 0.11 mmol); TLC: Rf=0.28 (iPr0H/Ac0Et/H ₂ 0/NH ₃ 4:3:2:1); ¹ H NMR (500 MHz, D ₂ 0): d 7.95 (d, J13.14 = 7.7 Hz, 1 H, H - 13) , 7.48 (t, J = 7.8 Hz, 1 H, H-15), 7.39 (m, 1 H, H-21), 7.31 (d, J8,7 = J10.11 = 8.0 Hz, 2H, H-8 and H-10), 7.26 (d, J23.22 = 7.5 Hz, 1 H, H-23), 7.09 (d, J16.15 = 8.2 Hz, 1 H, H-16), 7.04 (m, 4H, H-7, H-1 1 , H-14 and H-22), 6.79 (d, J20.21 = 7.8 Hz, 1 H, H-20), 5.02 (d, J1G,2G = 3.6 Hz, 1 H, H-1G of CyD), 4.96 (d, J = 3.5 Hz, 3H, Hs-1 of CyD), 4.92 (d, J = 3.6 Hz, 1 H, H-1A of CyD), 4.82 (d, J = 3.6 Hz, 2H, Hs-1 of CyD), 4.05 (d, J6A.6A = 13.7 Hz, 1 H, H-6A of CyD), 4.01 - 3.16 (m, 39H, Hs-2, Hs-3, Hs-4 and Hs-5 of CyD), 3.16 - 2.96 ppm (m, 2H, H-6A and H- 6X of CyD). ESI-MS: m/z=1490.45 [M+H] ⁺ Ret time: 13.0 min. Purity >95% .

Example 2: Synthesis of 4-r3,5-bis(2-hvdroxyphenyl)-1 H-1 ,2,4-triazol-1-yll-N-r3-deoxy-3-amino- beta-cyclodextrinl benzamide (compound 3 - CD2)

HOBt (24 mg, 0.175 mmol), DCC (36 mg, 0.175 mmol) were added to a solution of deferasirox (65.5 mg, 0.175 mmol) in dry DMF (5 mL). After 20 min, 3 ^A -amino-3 ^A -deoxy-p-cyclodextrin (199 mg, 0.175 mmol) was added. Reaction was stirred at room temperature under argon for 48 h. The solvent was evaporated to dryness in vacuo. The crude product was then triturated with diethyl ether (3 x 3 mL), followed by an acetone trituration cold (3 x 4 mL) and hot acetone trituration (3 x 3 mL) and the crude solids were purified by reverse phase chromatography using a 30g Biotage KP-RC-18 column (eluent: H ₂ 0®CH ₃ 0H) to give the title product (white solid). Yield: 57 % (149 mg, 0.10 mmol); TLC: Rf = 0.43 (iPr0H/Ac0Et/H ₂ 0/NH ₃ 4:3:2:1); 1 H NMR (500 MHz, D ₂ 0): d 7.94 (d, J = 7.7 Hz, 1 H, H-13), 7.80 (bs, 2H, H-8 and H-10), 7.64-7.26 (m, 5H, H-7, H-1 1 , H-15, H-21 and H-23), 7.14-6.95 (m, 3H, H-14, H-16 and H-22), 6.71 (s, 1 H, H- 20), 5.09 - 4.99 (m, 2H, Hs-1 of CyD), 4.97 (d, J = 3.4 Hz, 1 H, H-1 of CyD), 4.85 (s, 2H, Hs-1 of CyD), 4.81 (s, 1 H, H-1 of CyD), 4.59 (bs, 1 H, H-1A of CyD), 4.46 - 2.78 (m, 42H, Hs-2, Hs-3, Hs-4,Hs-5 and Hs-6 of CyD). ESI-MS: [P+H+K] ²⁺ , 1490.45 [M+H] Ret time: 13.3 min. Purity >95%.

Example 3: Solubility study

To determine solubility, a standard turbidimetric solubility assay (pH 7.8) was performed using 1 % DMSO solution diluted into buffer (HEPES 35 mM) with solubilities measured over a range from 3 - 100 mM. The plates were sealed and shaken for 3h at 25°C. The absorbance at 650 nm was measured using a SpectraMaxM5 (Molecular Devices). Aspirin was used as a positive control and pyrene as a negative control. The results are shown in the Table below:

The results showed that compounds 2 and 3 possessed similar solubility to deferasirox.

Example 4: Toxicity and dose-response studies

A dose-response study was performed on WT and NPC1 deficient CHO cells using 24 or 72 hours of treatment to determine efficacy and cytotoxicity profiles. Cell Lines: WT and Npd null CHO cells were used. Compounds 2 and 3 of the invention were compared with deferasirox.

LysoTracker Green staining: In vitro flow cytometry (FACS) experiments were performed in order to measure the relative acidic compartment volume of live CHO cells. Cells were stained with LysoTracker Green (250nM, 10min) in PBS at RT, centrifuged at 1200x, 10min and cells resuspended in FACS buffer (PBS, 1 % BSA, 0.1% sodium azide). Cells were stained with propidium iodide (20nM) immediately prior to analysis on the FACs machine to detect dead cells. FACS analyses were performed by recording 10,000 cells using a FACsCanto with FACSDiva BD software. Statistical analysis was done by using GraphPad Prism, Version 6. Oh. All experiments were statistically analyzed with One-Way Anova.

Figure 1 shows lysosomal volumes after 24 hour treatment at a compound concentration of 250 mM (CD1 is compound 2, CD2 is compound 3 and Ex-Jade is deferasirox). Figure 2 shows the results of the toxicity studies after 72 hours (CD1 is compound 2, CD2 is compound 3 and Ex-Jade is deferasirox).

Studies on WT cells showed that the compounds of the invention did not have any detrimental effect in terms of cell viability and did not affect relative lysosomal volume in concentrations up to 500mM. On the other hand, deferasirox increased the lysosomal volume at low concentrations (50mM, 100mM) in WT cells, while extremely decreasing total acidic compartment volume at high concentrations (250mM, 500 mM). In addition, 72 hours treatment with deferasirox reduced viability at all doses tested, compared to the untreated group and the compounds of the invention in a dose-dependent dependent manner after 100pM concentration.

In NPC1 null cells, treatment with the compounds of the invention for 72 hours significantly decreased the lysosomal volume at all concentrations, except for compound 2 at 500mM. Different concentrations showed similar responses with compound 2, while compound 3 showed the largest reduction in lysosomal volume at 100 and 250 mM. Deferasirox treatment increased the lysosomal volume at low concentrations, while was below WT levels at high concentrations. Toxicity experiments revealed that the compounds of the invention were well tolerated in NPC1 CHO cells. On the other hand, deferasirox caused 40% of cells to die, even at the low concentrations.

Compared to deferasirox, the compounds of the invention showed little toxicity at doses >10 mM in CHO cells. Indeed, little toxicity was seen for the compounds of the invention even at 500 mM concentration.

Example 5: Iron Binding Data:

An automated titration system was used in the study consisting of a Metrohm Dosimat 765 litre ml syringe, a Mettler Toledo MP230 pH meter with SENTEK pH electrode (P11), and an HP 8453 UV-visible spectrophotometer with a Hellem quartz flow cuvette being circulated through by a Gilson Mini-plus #3 pump— speed capability (20 mL/min). A potassium chloride electrolyte solution (0.1 mol/L) was used to maintain the ionic strength. The temperature of the test solutions was maintained in a thermostatic jacketed titration vessel at 25 °C (± 0.1 °C) using a Fisherbrand Isotemp water bath. The pH electrodes were calibrated using GLEE (Gans, P., & O’Sullivan, B. (2000). GLEE, a new computer program for glass electrode calibration, Talanta, 51(1), 33-37) with data obtained by titrating a volumetric standard HCI (0.1 mol/L) in KCI (0.1 mol/L) with KOH (0.1 mol/L) under an argon gas atmosphere in the vessel.

The solution under investigation was stirred vigorously during the experiment. For pKa determinations, a cuvette path length of 10 mm was used while for metal stability constant determinations, a cuvette path length of 50 mm was used. All instruments were interfaced to a computer and controlled by an in-house program. An automated titration adopted the following strategy: the pH of a solution was increased by 0.1 pH unit by the addition of potassium hydroxide solution (0.1 mol/L) from the autoburette. The pH readings were judged to be stable if their values varied by less than 0.01 pH unit after a set incubation period. For pKa

determinations, an incubation period of 1.5 mins was adopted; for metal stability constant determinations, an incubation period of 3 mins was adopted. The cycle was repeated until the defined end point pH value was achieved.

Titrations were carried out in the solution with molar ratio of DMSO : H ₂ 0 being 0.2 : 1. Under this condition, the pH meter readings are shifted, compared to the aqueous solution. All the titration data were analysed with the HypSpec2014 program (Gans, P., Sabatini, A., & Vacca, A. (1999), Determination of equilibrium constants from spectrophometric data obtained from solutions of known pH: the program pHab. Annali di chimica, 89(1-2), 45-49; and Gans, P., Sabatini, A., & Vacca, A. (1996), Investigation of equilibria in solution. Determination of equilibrium constants with the HYPERQUAD suite of programs, Talanta, 43(10), 1739-1753; http://www.hyperquad.co.uk/). The speciation plot was calculated with the HYSS program. Analytical grade reagent materials were used in the preparation of all solutions. The results are shown in the tables below:

Deferasirox (comparative)

Compound 2 (CD1)

Compound 3 (CD2)

‘Determined in the solution with molar ratio of DMSO : H ₂ 0 being 0.2 : 1.

[Fe]total: 1.000x1 O ⁶ M, [Ljtotal: 1.000x10 ⁵ M

The above results demonstrate that attachment of deferasirox to a cyclodextrin moiety does not adversely affect the Fe(lll) chelation properties of the drug since the stability constants (given as log units) are extremely high even in the presence of the cyclodextrin, showing that conjugation is not deleterious to iron binding.

Example 6: Antiaqqreqation assay

To investigate the influence of compounds 2 and 3 (CD1 and CD2) on aSyn aggregation, the copper-induced fibrillation of the protein in the absence and presence of each compound was monitored using dynamic light scattering (DLS). In order to determine the dimension of the aggregates and the extent of aSyn aggregation, the distributions of the diameter values in solution were recorded at different times (t=0 and t=24 h). The

hydrodynamic diameter of aSyn (mean diameter = 6.0 ± 0.3 nm) confirms the existence of the protein in a monomeric. Moreover, the size distribution of aSyn remains essentially unchanged after 24 h in these conditions. The presence of Cu ²⁺ promotes the aggregate formation, and the dimension of the aggregates significantly increases after 24 h (Figure 3).

aSyn solutions (0.5 mg/ml_) were buffered at pH 7.4 (MOPS, 20 mM). Copper-mediated aggregation of aSyn was studied using suitable Cu(CI0 ₄ ) ₂ solutions in order to have a final concentration of 265 mM in cuvette. CyD derivatives were added in equimolar amount to the copper ions (Oliveri, V., Sgarlata, C., & Vecchio, G. (2016), Cyclodextrins 3-Functionalized with 8-Hydroxyquinolines: Copper-Binding Ability and Inhibition of Synuclein Aggregation,

Chemistry-An Asian Journal, 11(17), 2436-2442). Example 7: Trolox Equivalent Antioxidant Capacity Assay

In vitro antioxidant assays were performed by 2,2’-azinobis(3-ethylbenzothiazoline-6- sulfonic acid) diammonium salt (ABTS’ ⁺ ) radical cation decolorization assay using 6-hydroxy- 2,5,7,8-tetramethylchroman-2-carboxylic acid (Trolox) according to the method disclosed in Oliveri, V., Bellia, F. and Vecchio, G. (2015), Cyclodextrin 3-Functionalized with 8- Hydroxyquinoline as an Antioxidant Inhibitor of Metal-Induced Amyloid Aggregation,

ChemPlusChem, 80: 762-770.

The capacity of compounds of the invention to scavenge ABTS' ⁺ at 1 , 3, and 6 minutes was compared to Trolox, an analogue of vitamin E, and was thus expressed as TEAC values (Figure 4). Both compounds 2 and 3 showed a greater ability to scavenge free radicals than Trolox.

Example 8: Synthesis of Heptakis (per6-2-allylamino-6-deoxy)-3-cyclodextrin

In a 100 mL carousel tube equipped with a magnetic stirrer /?epfa/c/s-6-iodo-6-deoxy-p- cyclodextrin (0.16 g, 0.084 mmol, 1 equiv) ( Angew . Chem. Int. Ed., 2005, 44, 7584 -7587; Chem. Commun., 2012, 48, 8063-8065; Angew. Chem. Int. Ed., in English, 1991 , 30, 78-80; J. Am. Chem. Soc., 1995, 117, 336-343) was dissolved in DMF (0.5 mL) and the warmed to 70°C before allylamine (0.34 mL, 5.88 mmol) was added. The reaction mixture was then heated for two days. The solution was cooled to room temperature before reagent grade acetone (50 mL) was added. The resulting solution was cooled overnight at -20°C, the precipitate was allowed to settle and the supernatant solution was decanted. The residue was dissolved in a minimum amount of methanol and crashed out of acetone (150 mL) then membrane filtered and further purified using reverse phase C-18 column (gradient water: methanol 10%). to give the product (32 mg, 27% yield) as a solid. ¹ H NMR (D ₂ 0) d = 5.85 (7H, m), 5.24-5.18 (14H, m), 3.86-3.76 (14H, m), 3.60 (7H, m), 3.47 (7H, m), 3.24 (14H, brs), 2.99-2.87 (14H, m). ¹³ C NMR (D ₂ 0) d = 134.3, 118.6, 101.0, 81.9, 75.4, 72.8, 69.4, 51.3, 48.3. HRMS: m/z calculated for CesF sO^Ny [M+H] ⁺ : 1408.5379. Found: 1408.7086.

Example 9: Synthesis of Heptakis (per6-2-cvclopropylamino-6-deoxy)-8-cyclodextrin

The title compound was synthesised by a method analogous to that of Example 8 using cyclopropylamine in place of the allylamine. Yield: 39 mg (26%). ¹ H NMR (DMSO-d ₆ ) d = 5.69 (14H, m), 4.80 (7H, s), 3.70 (7H, brs), 3.60 (7H, m), 3.34-3.31 (21 H, m), 2.94 (7H, m), 2.88 (7H, brs), 1.80 (7H, m), 0.33 (14H, brs), 0.23 (14H, brs). ¹³ C NMR (DMSO-d ₆ ) d = 102.4, 83.5, 73.4, 72.9, 70.6, 49.4, 30.5, 6.7, 6.3. HRMS: m/z calculated for CssH^sOssNy [M+H] ⁺ : 1408.5379. Found: 1408.7001. Calculated for ΰ ₆₃ H ₁₀ 5N ₇ O ₂₈ 1.2 HI 7.7 H ₂ 0 (C, 44.49; H, 7.21 ; N, 5.76), found: C, 44.74; H , 7.48; N, 5.52.

Example 10: Synthesis of Octakis (per6-n-butylamino-6-deoxy)-y-cyclodextrin

The title compound was synthesised by a method analogous to that of Example 8 using n-butylamine in place of the allylamine and a g-cyclodextrin in place of the b-cyclodextrin. Yield: 77 mg (45%). ¹ H NMR (DMSO-de) d = 5.78 (8H, m), 5.75 (8H, d , J = 5.0 Hz), 4.88 (8H, m), 3.64 (8H, brs), 3.56 (8H, t, J = 10.0 Hz), 3.34 (24H, m), 2.78 (8H, m), 1.39-1 .28 (48H, m), 0.85 (24H, t, J = 5.0 Hz). ¹³ C NMR (DMSO-d ₆ ) d = 102.3, 83.2, 75.4 (2C), 71.1 , 49.8, 49.6, 32.3, 20.4, 14.3. HRMS: m/z calculated for C ₈ oHi ₅₂ N ₈ 0 ₃₂ [M] ⁺ : 1739.0669. Found: 1739.0608. Calculated for C80H152N8O32 0.9 HI 0.2 H ₂ 0 (C, 51.75; H , 8.32; N, 6.03), found: C, 51.78; H, 8.37; N, 6.02.

Example 1 1 : Synthesis of Heptakis (per6-n-butylamino-6-deoxy)-3-cyclodextrin

The title compound was synthesised by a method analogous to that of Example 8 using n-butylamine in place of the allylamine. Yield: 97 mg (76%). ¹ H NMR (DMSO-d ₆ ) d = 5.71 (14H, m), 4.82 (7H, brs), 3.68 (7H, brs), 3.61 (7H, t, J = 10 Hz), 3.37-3.30 (21 H , m), 2.86-2.78 (14H, m), 2.50 (7H, brs), 1.38-1 .35 (28H , m), 1.30-1.25 (28H, m), 0.85 (21 H, t, J = 10 Hz). ¹³ C NMR (DMSO-d ₆ ) d = 102.5, 83.7, 73.4, 72.9, 71.0, 49.7, 32.3, 31 .1 , 20.4, 14.2. HRMS: m/z calculated for C ₇ oHi ₃₃ N ₇ 0 ₂₈ [M+H] ⁺ : 1520.9277, Found: 1520.9556. Calculated for C ₇₀ H ₁₃₃ N ₇ O ₂₈ (C, 55.28; H, 8.81 ; N, 6.45), found: C, 55.22; H, 8.89; N, 6.51.

Example 12: Synthesis of i-6-deoxy)-8-cyclodextrin

The title compound was synthesised by a method analogous to that of Example 8 using

1-methylpentylamine in place of the allylamine. Yield: 50 mg (35%). ¹ H NMR (DMSO-d _s ) d = 5.46 (14H, brs), 4.82 (7H, s), 3.65 (7H , m), 3.64 (7H, m), 3.31-3.29 (14H, m), 2.85, (7H, m), 2.48 (7H, m), 1.39 (7H, s), 1.25 (42H, brs), 0.85 (21 H, s). ¹³ C NMR (DMSO-d _e ) d = 102.6, 83.7, 73.4, 73.0, 71.0, 50.1 , 49.8, 31.8, 30.2, 27.1 , 22.6, 14.3. HRMS: m/z calculated for C ₈₄ H ₁₆ I0 ₂₈ N ₇ [M+H] ⁺ : 859.61 16. Found: 859.5776. Calculated for C ₈₄ H ₁₆₁ N ₇ 0 ₂₈ 9.7 CH ₄ 0 0.3 C ₃ H ₆ 0 (C, 55.55; H, 9.93; N, 4.79), found: C, 55.93; H , 10.33; N, 5.19.

Example 13: Synthesis of Heotakis (per6-2-r2-(2-hvdroxy-ethylamino)-ethylaminol-6-deoxy)-B- cvclodextrin The title compound was synthesised by a method analogous to that of Example 8 using 2-(2- hydroxyethylamino)-ethylamine in place of the allylamine. Yield: 44 mg (28%). ¹ H NMR (D ₂ 0) d = 8.05 (7H, brs), 5.04 (7H, brs), 3.88 (14H , brs), 3.65-3.40 (42H, m), 2.88-2.77 (28H, brs), 2.24 (7H, s). ¹³ C NMR (D ₂ 0) d = 166.0, 101.8, 75.4, 72.0 (2C), 70.6, 60.1 , 58.4, 49.8, 47.8. HRMS: m/z calculated for C7oHi ₄ oNi ₄ 0 ₃ 5 [M+2H+K] ²⁺ : 908.9384. Found: 908.7281. Calculated for C70H140N14O35 6.3 C ₃ HSO 3.1 HI (C, 42.70; H, 7.29; N, 7.84), found: C, 42.67; H, 7.35; N, 7.84.

|-6-deoxy)-3-

The title compound was synthesised by a method analogous to that of Example 8 using 2-(2- hydroxyethoxy)-ethylamine in place of the allylamine. Yield: 68 mg (47%). ¹ H NMR (D ₂ 0) d = 5.08 (7H, brs), 3.91 (14H, m), 3.68-3.50 (63H, m), 2.99-2.92 (14H, m) 2.84 (7H, brs), 2.93 (7H, brs). ¹³ C NMR (D ₂ 0) d = 101.2, 82.3, 72.7, 72.0, 71.7, 70.2, 68.7, 60.5, 49.1 , 48.5. HRMS: m/z calculated for C ₇₀ H ₁₃₃ N ₇ O ₄₂ [M+2H] ²⁺ : 872.9322. Found: 872.9364.

Example 15: Synthesis of Octakis (per6-2-[2-(2-hydroxy-ethoxy)-ethylaminol-6-deoxy-)-y-

The title compound was synthesised by a method analogous to that of Example 14. Yield: 82 mg (41 %). ¹ H NMR (D ₂ 0) d = 5.1 1 (8H, brs), 3.86 (16H, m), 3.72-3.47 (72H, m), 2.99-2.92 (16H, m) 2.84 (8H, brs), 2.92 (8H, brs). ¹³ C NMR (D ₂ 0) d = 100.7, 81 .3, 75.3, 72.8, 71 .6, 70.3, 69.0, 60.5, 49.0, 48.4. HRMS: m/z calculated for C ₈ oH ₁₅₂ 0 ₄₈ N ₈ [M+2H] ²⁺ : 997.4850. Found: 997.4923. Calculated for CsoH^C sNs (C, 48.19; H, 7.68; N , 5.62), found: C, 47.98; H, 7.61 ; N , 5.52

Example 16: Synthesis of l-2-(6-amino-l i-6-deoxy)-3-cyclodextrin

The title compound was synthesised by a method analogous to that of Example 8 using 1 ,6-hexanediamine in place of the allylamine. Yield: 60 mg (39%). ¹ H NMR (D ₂ 0 20% HCI v/w) d = 7.98 (7H, s), 5.04 (7H, brs), 3.84 (14H, brs), 3.59 (14H, brs), 3.16 (7H, brs), 2.95-2.79 (7H, m), 2.57 (14H, brs), 1.57 (42H, brs), 1.29 (42H, brs). ¹³ C NMR (D ₂ 0 20% HCI v/w) d = 169.0, 165.8, 162.4, 102.6, 85.5, 83.1 , 72.9 9, 70.7, 49.9, 37.5, 30.8, 26.7. HRMS: m/z calculated for Ce4Hi08Ni4O ₂ 8 [M ^— 2H] ² : 909.6077, Found: 909.5383. Calculated for C84Hi ₃ 8Ni40 ₂ 8 2 C7H8 2.8 HI (C, 49.78; H, 7.96; N, 8.29), found: C, 49.62; H, 8.27; N, 8.03.

The title compound was synthesised by a method analogous to that of Example 17. Yield: 42 mg (21 %). ¹ H NMR (D ₂ 0 20% HCI v/w) d = 7.97 (8H, s), 5.08 (8H, brs), 3.86 (16H, brs), 3.68 (16H, brs), 3.16 (8H, s), 2.93 (8H, s), 2.63 (16H, brs), 1.60-1.30 (64H, m). ¹³ C NMR (D ₂ 0) d = 101.2, 82.5, 72.1 (2C), 69.7, 49.4, 41.8, 37.9, 28.4 (2C), 26.1 (2C). HRMS: m/z calculated for 0 ₉₆ H _ΐ92 Ni ₆ q ₃₂ [(M-8cNH ₂ +2H) ²⁺ + H ₂ 0]: 996.2456. Found: 996.0742.

Example 18: Synthesis of Heptakis (per6-6-amino-1-hexanol-6-deoxy)-3-cyclodextrin

The title compound was synthesised by a method analogous to that of Example 8 using 6-amino-1-hexanol in place of the allylamine. Yield: 130 mg (85%). ¹ H NMR (D ₂ 0) d = 5.04 (7H, brs), 3.84 (14H, brs), 3.60-4.43 (21 H, m), 2.58 (7H, brs), 2.17 (7H, s), 1.50 (42H, brs), 1.31 (42 H, brs). ¹³ C NMR (D ₂ 0) d = 101.8, 83.3, 73.0, 72.0, 70.2, 61.8, 49.5, 45.7, 31.6, 28.6, 26.6, 25.3. HRMS: m/z calculated for C ₈₄ H _{1 61} N ₇ 035 [M+2H] ²⁺ : 915.0597, Found: 915.0584. Calculated for C ₈₄ H _{1 61} N ₇ 0 ₃ 5 8.6 H ₂ 0 2.4 C ₂ H ₆ 0 (C, 50.92; H, 9.27; N, 4.68), found: C, 50.72; H, 9.54; N, 4.95.

Example 19: of Octakis (per6-6-amino-1-hexanol-6-' -6-deoxy)-y-cyclodextrin

The title compound was synthesised by a method analogous to that of Example 18. Yield: 85 mg (43%). ¹ H NMR (D ₂ 0) d = D ₂ 0/DMSO d = 5.04 (8H, brs), 3.84 (16H, brs), 3.60-4.43 (24H, m), 2.58 (8H, brs), 2.17 (8H, s), 1.50 (48H, brs), 1.31 (48H, brs). ¹³ C NMR (D ₂ 0) d = 100.8, 82.4, 73.0, 72.1 , 69.7, 61.6, 49.1 , 31.7 (2C), 28.8, 26.7, 25.3. HRMS: m/z calculated for C ₉₆ H ₁₈₄ N ₃ 0 ₄ o [M+2H] ²⁺ : 1045.6383. Found: 1045.6260. Calculated for C ₉ 6H ₁₈₄ N ₈ 04o (C, 55.16; H, 8.87; N, 5.36), found: C, 55.09; H, 8.78; N, 5.25.

Example 20: Synthesis of i-6-deoxy)-3-cyclodextrin

The title compound was synthesised by a method analogous to that of Example 8 using 1-(2- aminoethyl)piperizine in place of the allylamine. Yield: 150 mg (62%). ¹ H NMR (D ₂ 0) d = 5.06 (7H, brs), 3.87 (7H, brs), 3.56 (7H, brs), 2.82 (56H, brs), 2.72 (7H, brs), 2.48 (63H, brs), 2.16 (7H, brs), 2.02 (7H, s), 1.86 (7H, s). ¹³ C NMR (D ₂ 0) d = 101.5, 72.0, 70.1 , 57.4, 57.0, 52.1 (14C), 45.7, 43.8 (14C), 36.5. HRMS: m/z calculated for C ₈₄ H _16I N ₂₁ 0 ₂₈ [M+H] ⁺ : 1914.3090, Found: 1914.2257. Calculated for C ₈₄ H ₁₆₁ N ₂₁ 0 ₂₈ 4.4 HBr 3.5 C ₂ H ₆ 0 (C, 44.97; H, 7.73; N, 12.10), found: C, 44.68; H, 7.96; N, 12.37. Example 21 : Synthesis of Octakis (per6-1-(2-aminoethyl)piperizine-6-deoxy-)-v-cyclodextrin

The title compound was synthesised by a method analogous to that of Example 21. Yield: 145 mg (65%). ¹ H NMR (D ₂ 0) d = 5.12 (8H, brs), 3.84 (8H, brs), 3.55 (8H, brs), 2.96-2.50 (136H, m). HRMS: m/z calculated for C ₉₆ H ₁₁₂ N ₂₄ O ₄₀ [M-H +H ₂ 0] ⁺ : 2257.7541 , Found: 2257.3044.

Example 22: -6-deoxy)-3-cyclodextrin

The title compound was synthesised by a method analogous to that of Example 8 using n-octylamine in place of the allylamine. Yield: 118 mg (71 %). ¹ H NMR (DMSO-d ₆ ) d = 5.70 (14H, brs), 4.80 (7H, brs), 3.69 (7H, brs), 3.60 (7H, brs), 3.30 (21 H, brs), 2.86 (7H, brs), 2.75 (7H, brs), 1.36 (14H, brs), 1.22 (87H, brs), 0.81 (21 H, brs). ¹³ C NMR (DMSO-d ₆ ) d = 102.7,

84.0, 73.4, 72.9, 71.0, 50.2, 31.9, 31.1 , 30.3, 29.7, 29.4, 27.6, 22.6, 14.0. HRMS: m/z calculated for C ₉₈ H ₁₈₉ N ₇ O ₂₈ [M+3H] ³⁺ : 638.4607, Found: 638.7935. Calculated for CgsH^gNyC^s 0.91 C ₆ H ₁₄ 0.02 HI (C, 62.30; H, 10.20; N, 4.92), found: C, 62.22; H, 10.29; N, 5.01.

Example 23: Synthesis of Heptakis (per6-2-decylamino-6-deoxy)-3-cyclodextrin

The title compound was synthesised by a method analogous to that of Example 8 using n-decylamine in place of the allylamine. Yield: 41 mg (23%). ¹ H NMR (DMSO-d ₆ ) d = 5.72 (brs, 14H), 4.80 (7H, brs), 3.63 (7H, brs), 3.61 (7H, brs), 3.31 (21 H, brs), 2.72 (7H, m), 2.15 (7H, s), 1.35 (14H, brs), 1.20 (1 12H, brs), 0.79 (21 H, m). ¹³ C NMR (DMSO-d ₆ ) d = 102.6, 83.5, 75.5, 72.9, 50.2, 46.9, 31.9, 30.4, 29.8 (4C), 29.4, 27.7, 22.6, 14.1. HRMS: m/z calculated for C ₁₁₂ H ₂₁₇ N ₇ 0 ₂₈ [M+2H] ²⁺ : 1055.2964. Found: 1055.8014.

Example 24: Synthesis of Heptakis (per6-2-{2-[3-(2-amino-ethylamino)-propylaminol- ethylamino}-6-deoxy)-3-cyclodextrin

The title compound was synthesised by a method analogous to that of Example 8 using [(2- amino-ethylamino)-propylamino]-ethylamine in place of the allylamine. Yield: 54 mg (30%). ¹ H NMR (D ₂ 0) d = (7H, brs, 7xN H), 5.07 (7H, brs, 7xH1), 3.85 (14H, brs, 7xH3, 7xH5), 3.57 (14H, brs, 7xH2, 7xH4), 2.25-3.00 (98H, m, 12CH ₂ x7, 7xH6, 7xH6’), absent (14H, brs, 7xNH ₂ ), absent (7H, brs, 7xN H), absent (7H, brs, I H). ¹³ C NMR (D ₂ 0) d = 101.5, 75.6, 72.9 (2C), 70.6, 49.6, 47.6, 46.5 (3C), 39.2 (2C), 28.0. HRMS: m/z calculated for C _9i Hiee0 ₂₈ N ₂₈ [M+3H] ³⁺ : 710.8336. Found: 710.8368. Calculated for Cgi H^NssOss 15.6 C ₂ H ₆ 0 3.5 HI (C, 44.52; H, 8.96; N, 11.89), found: C, 44.53; H, 8.89; N, 11.82. Example 25: Synthesis of Heptakis (per6-2-f3-[bis-(2-hvdroxy-ethyl)-aminol-propylamino)-6- deoxy)-B-cydodextrin

The title compound was synthesised by a method analogous to that of Example 8 using 3-[bis- (2-hydroxyethyl)-amino]-propylamine in place of the allylamine. Yield: 58 mg (32%). ¹ H NMR (D ₂ 0) d = 5.13 (7H, brs), 3.87 (14H, brs), 3.64 (28H, brs), 3.58 (14H, brs), 3.48 (7H, brs), 2.97 (7H, m), 2.67 (28H, brs), 2.59 (28H, brs), 1.67 (14H, brs HRMS: m/z calculated for

C ₉₁ H ₁₈₂ N ₁₄ O ₄₂ [M+2 H] ²⁺ : 1073.1365. Found: 1073.1488. Calculated for C ₉₁ H ₁₈₂ N ₁₄ 0 ₄₂ 1 .6 HI 10.1 CH ₄ 0 (C, 45.43; H, 8.45; N, 7.34), found: C, 45.47; H, 8.49; N, 7.30.

Example 26: Synthesis of Octakis (per6-2-f3-fbis-(2-hvdroxy-ethyl)-aminol-propylamino}-6- deoxy)-v-cvclodextrin

The title compound was synthesised by a method analogous to that of Example 25. Yield: 49 mg (24%). ¹ H NMR (D ₂ 0) d = 5.12 (8H, brs), 3.83 (16H, brs), 3.62 (32H, brs), 3.56 (16H, brs), 3.47 (8H, brs), 2.83 (8H, m), 2.64 (32H, brs), 2.59 (32H, brs), 1.65 (16H, brs). ¹³ C NMR (D ₂ 0) d = 100.7, 97.8, 72.8, 72.1 , 70.2, 58.8, 55.3, 52.5, 49.1 , 47.8, 25.3. HRMS: m/z calculated for CI ₀₄ H ₂₀₈ NI ₆ O ₄₈ [M-H ₂ O + 2K] ²⁺ : 1254.7087. Found: 1254.7581. Calculated for CIO ₄ H ₂ O ₈ N ₁₆ 0 ₄₈ 5 C ₃ H ₆ 0 12.75 H ₂ 0 (C, 48.11 ; H, 8.94; N, 7.54), found: C, 47.87; H, 8.70; N, 7.31.

27: of 1 2 :-(3-amino-i ino)-'

propylamino)-6-deoxy)-B-cyclodextrin

The title compound was synthesised by a method analogous to that of Example 8 using 3-[2-(3- amino-propylamino)-ethylamino]-propylamine in place of the allylamine. Yield: 37 mg (20%).

¹ H NMR (D ₂ 0) d = 7.98 (7H, brs), 5.04 (7H, brs), 3.85 (14H, brs), 3.56 (14H, brs), 3.05-3.55 (14H , m), 2.25-3.00 (84H, m), 2.57 (7H, brs), 2.00 (7H, brs), 1.31 (28H, brs). ¹³ C NMR (D ₂ 0) d = 165.4, 164.2, 101.8, 75.4, 72.8, 70.5, 47.3, 43.8 (2C), 38.2 (2C), 37.5, 28.1 (2C). HRMS: m/z calculated for C ₉₈ H ₂₁ O0 ₂₈ N ₂₈ [M-7XCH ₂ CH ₂ CH ₂ NH ₂ +2K] ²⁺ : 896.7820. Found: 896.0166.

Calculated for C ₉₈ H ₂₁₀ N ₂₈ O ₂₈ 10.9 C ₃ H _s O 4.5 HI (C, 45.67; H, 8.21 ; N, 11.41), found: C,

45.51 ; H, 8.37; N, 1 1.36.

:-(3-amino-i 6

The title compound was synthesised by a method analogous to that of Example 8 using 3-[2-(3- amino-propoxy)-ethoxy]-propylamine in place of the allylamine. Yield: 100 mg (53%). ¹ H NMR (D ₂ 0) d = 8.00 (7H, s), 5.04 (7H, brs), 3.84 (14H, brs), 3.59-3.53 (84H, brs), 3.26 (7H, brs), 2.96-2.64 (14H, m), 2.64 (7H, brs), 2.23 (14H, brs), 1.75 (28H, brs). ¹³ C NMR (D ₂ 0) d = 167.3, 101.6, 82.6, 73.0, 72.1 , 69.5, 69.3, 68.5, 68.3, 69.3, 49.0, 46.6, 37.7, 35.1 , 29.0. Calculated for C ₉₈ H ₁₉₆ N ₁₄ 0 ₄₂ (C, 52.48; H, 8.81 ; N, 8.74), found: C, 52.38; H, 8.72; N, 8.64. of Octakis (per6-2-{3-[2-(3-amino-i 6

The title compound was synthesised by a method analogous to that of Example 28. Yield: 97 mg (38%). ¹ H NMR (D ₂ 0 20% HCI v/w) d = 7.98 (8H, s), 5.07 (8H, brs), 3.83 (16H, brs), 3.58- 3.52 (88H, m), 2.94 (8H, s), 2.64 (16H, brs), 1.75 (32H, brs). ¹³ C NMR (D ₂ 0) d = 163.8, 101.6, 82.0, 72.9, 72.3, 69.4, 68.4 (2C), 49.1 , 46.7, 35.2, 28.4 (2C). HRMS: m/z calculated for

C ₁₁₂ H ₂₂₄ N ₁₆ 0 ₄₈ [(M-8xNH ₂ +2Na) ²⁺ + Na+H]: 1264.1707. Found: 1264.2082. Calculated for Cn ₂ H ₂₂₄ N ₁₆ 0 ₄₈ (C, 52.48; H, 8.81 ; N, 8.74), found: 52.35; H, 8.83; N, 8.64. i-2-f6-(6-amino-l |-6-deoxy-)-8-

The title compound was synthesised by a method analogous to that of Example 8 using (6- amino-hexylamino)-hexylamine in place of the allylamine. Yield: 48 mg (23%). ¹ H NMR (D ₂ 0) d = 7.96 (7H, brs), 7.90 (7H, s), 5.04 (7H, brs), 3.80 (14H, brs), 2.85 (7H, brs), 2.40-3.00 (35H, m), 2.22 (7H, s), 1.48 (56H, brs), 1.30 (1 12H, brs). ¹³ C NMR (D ₂ 0) d = 164.0, 101.7, 82.6, 75.4, 73.3, 48.1 , 47.6, 37.9 (2C), 28.6 (2C), 26.5 (2C), 25.9 (4C), 25.1. HRMS: m/z calculated for C ₁₂₆ H ₂ 590 ₂₈ N ₂₁ [(M-7XN H ₂ +2H) ²⁺ + H ₂ 0]: 1221.2085. Found: 1221.3658. Calculated for C ₁₂₆ H ₂₅₉ N ₂₁ 0 ₂₈ 10.7 C ₂ H _s O 3.6 HI (C, 51.02; H, 9.49; N, 8.48), found: C, 51.12; H, 9.38; N, 8.36.

Example 31 : Synthesis of Heptakis (per6-2-(3-{2-[2-(3-amino-propoxy)-ethoxyl-ethoxy>- propylamino)-6-deoxy)-8-cyclodextrin

The title compound was synthesised by a method analogous to that of Example 8 using 3-{2-[2- (3-amino-propoxy)-ethoxy]-ethoxy}-propylamine in place of the allylamine. Yield: 81 mg (41 %). ¹ H NMR (D ₂ 0) d = 8.02 (7H, s), 5.05 (7H, brs), 3.84 (14H, brs), 3.64-3.61 (56H, brs), 3.54 (56H, brs), 3.26 (7H, brs), 3.01-2.87 (7H, m), 2.66 (7H, brs), 2.26 (14H, brs), 1.77 (42H, brs). ¹³ C NMR (CDsOD) d = 166.1 , 162.5, 102.3, 82.9, 73.3, 72.9, 70.7, 70.2, 70.0, 69.9, 69.4, 68.8, 68.5, 67.5, 49.1 , 47.3, 47.2, 46.1 , 38.7, 35.1 , 29.3, 29.0. HRMS: m/z calculated for Cn ₂ H ₂₂₄ 0 ₄₉ N ₁₄ [M+3K] ³⁺ : 888.8126. Found: 888.1603. Calculated for C ₁₁₂ H ₂₂₄ N ₁₄ O ₄₉ 1.3 H ₂ 0 1.8 HI (C, 47.96; H, 8.21 ; N, 6.99), found: C, 48.05; H, 8.28; N, 6.89.

Example 32: Synthesis of Octakis (per6-2-(3-f2-r2-(3-amino-propoxy)-ethoxyl-ethoxy)- propylamino)-6-deoxy)-v-cvclodextrin

The title compound was synthesised by a method analogous to that of Example 31. Yield: 134 mg (46%). ¹ H NMR (D ₂ 0) d = 7.98 (8H, s), 5.06 (8H, brs), 3.82 (16H, brs), 3.60-3.53 (96, brs), 3.25 (16H, brs), 2.97-2.83 (32H, m), 2.65 (16H, brs), 2.24 (16H, brs), 1.84 (32H, brs).

Example 33: Synthesis of He ptakis (per6-11-azido-3.6,9-trioxandecan-1-amino-6-deoxy)-B- cvclodextrin

The title compound was synthesised by a method analogous to that of Example 8 using 11- azido-3,6,9-trioxandecan-1-amine in place of the allylamine. Yield: 47 mg (31 %). ¹ H NMR (D ₂ 0) d = 5.08 (7H, brs), 3.94 (7H, brs), 3.90 (7H, t, J= 10 Hz), 3.66-4.45 (126H, m), 3.16 (7H, s), 3.05 (7H, brs), 2.95 (7H, brs). ¹³ C NMR (D ₂ 0) d = 101.7, 82.6, 72.7, 72.0, 69.8, 69.7 (2C), 69.6 (2C), 69.5, 69.4, 69.3, 69.2, 68.1 , 66.5 (2C), 50.3, 48.9, 48.4, 39.2. HRMS: m/z calculated for C ₉₈ Hi ₈₂ N ₂₈ 0 ₄₉ [M+3H] ³⁺ : 846.0950, Found:846.4277.

Example 34: Synthesis of Heptakis (per6-2-(2-f2-[2-(2-amino-ethoxy)-ethoxyl-ethoxy)-ethoxy)- ethanol-6-deoxy)-3-cyclodextrin

The title compound was synthesised by a method analogous to that of Example 8 using 2-(2-{2- [2-(2-aminoethoxy)-ethoxy]-ethoxy}-ethoxy)-ethanol (HO-PEG5-NH ₂ ) in place of the allylamine. Yield: 27 mg (39%). ¹ H NMR (D ₂ 0) d = 5.08 (7H, brs), 4.12-3.85 (14H, m), 3.68-3.64 (161 H, brs), 2.81 (7H, brs), 2.34 (7H, brs). HRMS: m/z calculated for C ₁₁₂ H ₂₁₇ N ₇ 0 ₆₃ [M-2H] ²⁺ :

1333.19165, Found: 1333.5154.

Example 35: Synthesis of Heptakis (per6-2-[2-(2- [2-(2-amino-ethoxy)-ethoxyl-ethoxy|-

ethoxy)-ethoxyl-ethanol -6-deoxy)-8-cyclodextrin

The title compound was synthesised by a method analogous to that of Example 8 using 2-(2-{2- [2-(2-amino-ethoxy)-ethoxy]-ethoxy}-ethoxy)-ethoxy]-ethanol (HO-PEG6-NH ₂ ) in place of the allylamine. Yield: 40 mg (52%). ¹ H NMR (D ₂ 0) d = 5.05 (7H, brs), 4.12-3.85 (14H, m), 3.63- 3.58 (189H, brs), 2.80 (7H, brs), 2.32 (7H, brs). ¹³ C NMR (D ₂ 0) d = 101.7, 81.5, 72.6, 72.7 (2C), 69.7, 69.5 (3C), 68.86 (3C), 60.41 (2 C), 49.2, 48.3, 44.8. HRMS: m/z calculated for C ₁₂₆ H ₂₅₂ N ₇ O ₇₀ [M+3H+3Na] ³⁺ : 1018.5436, Found: 1018.2965. Calculated for C ₁₂₆ H ₂₄₅ N ₇ O ₇₀ · 3.05H Br (C, 46.92; H, 7.75; N, 3.04), found: C, 46.61 ; H, 7.77; N, 3.36.

Example 36: Synthesis of Heotakis (per6-2-(4.4.4-trifluoro-butylamino)-6-deoxy-)-B-cvclodextri n

The title compound was synthesised by a method analogous to that of Example 8 using 4,4,4- trifluoro-butylamine in place of the allylamine.

Yield: 40 mg (25%). ¹ H NMR (D ₂ 0) d = 5.07 (7H, brs), 3.91 (14H, brs), 3.48 (7H, brs), 3.16 (7H, brs), 3.04 (8H, brs), 2.83 (14H, brs), 2.65 (7H, brs), 2.22 (14H, m), 2.16 (7H, s), 1.80 (14H, brs). ¹³ C NMR (DMSO-d ₆ ) d = 128.2 (d, J = 273.8 Hz), 102.6, 83.4, 75.4, 73.4, 71.2, 49.2, 48.4, 46.7,

31.1 , 22.4. ¹⁹ F NMR (D ₂ 0) d = -66.09. HRMS: m/z calculated for 0 ₇₀ ^^ ₂ ^0 ₂₃ [M] ⁺ :

1898.6348. Found: 1898.8194. Calculated for C ₇₀ H ₁₁₂ F ₂₁ N ₇ O ₂₈ (C, 44.28; H, 5.95; N, 5.16), found: C, 44.19; H, 5.94; N, 5.21. Example 37: Synthesis of CD3

Step 1. Synthesis of S-(2-((tert-butoxycarbonyl)amino)ethyl) 4-(3,5-bis(2-hydroxyphenyl)-1 H- 1 ,2,4-triazol-1-yl)benzothioate

Deferasirox (250 mg, 1.0 eq) was dissolved in anhydrous DMF (6.0 ml) under inert conditions and cooled to 0°C. The reaction mixture was stirred for 10 minutes to ensure complete solvation before DCC (152.7 mg, 1.1 eq) and DMAP (9.00 mg, 0.1 eq) were then carefully added while maintaining the temperature at 0°C. After 5 minutes of vigorous stirring 2-(BOC- amino)ethanethiol (340 pi, 3.0 eq) was added and the resulting mixture was stirred for 12 hours at 0°C allowing to warm to room temperature.

After 12 hours, a white precipitate of DCU formed and was removed by filtration. The resulting filtrate was diluted using EtOAc (20 ml) and washed sequentially with saturated aqueous sodium hydrogen carbonate solution (2 x 20 ml), water (20 ml) and saturated aqueous sodium chloride solution (20 ml) before being dried using magnesium sulphate and concentrated to give a yellow, clear crude oil. The impure mixture was then purified using column chromatography (EtOAc: Hexanes, 1 : 1) to yield an off white, solid powder of the product (142.6mg, 40% yield, 100% purity, LC-MS; ret. Time = 3.45 min, MeCN H ₂ 0 gradient).

Steps 2 and 3. The compound formed in step 1 (S-(2-((tert-butoxycarbonyl)amino)ethyl) 4-(3,5- bis(2-hydroxyphenyl)-1/-/-1 ,2,4-triazol-1-yl)benzothioate) is deprotected by removal of the Boc group (step 2). The crude reaction product is then coupled to cyclodextrin using tosyl substituted beta-cyclodextrin (e.g. using the methodology set out in Puglisi et al., J Incl Phenom Macrocycle Chem (2012) 73:475-478) to give compound CD3.

Example 38: Synthesis of CD4

Inside an 8ml microwave reactor vessel Deferasirox (100 mg, 1.0 eq), K ₂ C0 ₃ (295.8 mg, 8.0 eq) and ZnCI ₂ (0.6 mg, 0.01 eq) were suspended in EtOH (4 ml) and stirred vigorously for 5 minutes. 5-Chloro-1-pentyne (57.2 pi, 2.0 eq) was then added and the reaction vessel underwent microwave irradiation at 200W, 125°C for 15 minutes. On completion, the reaction vessel was placed in the fridge for 12 hours leading to the formation of a white precipitate which was removed by filtration. The precipitate was washed with ice cold EtOH (10 ml) and the resultant filtrate was concentrated to give a white solid crude of a mixture of starting material and pent-4-yn-1-yl-4-(3,5-bis(2-hydroxyphenyl)-1/-/-1 ,2,4-triazol-1-yl)benzoate (91.0 mg, crude yield 77%). Step 2. Synthesis of CD4

Pent-4-yn-1-yl 4-(3,5-bis(2-hydroxyphenyl)-1 /-/-1 ,2,4-triazol-1-yl)benzoate (100 mg, 1.0 eq), from step 1 , 6A-azido-6A-deoxy-p-cyclodextrin (290.4 mg, 1.1 eq), CuS0 ₄ .5H ₂ 0 (3.2 mg, 0.1 eq) and sodium ascorbate (5.0 mg, 0.11 eq) are suspended in a mixture of H ₂ 0-CH ₂ Cl ₂ (1 :3, 4 ml). The reaction mixture is stirred vigorously at room temperature for 48 hours. Upon completion, the reaction mixture is extracted with CH ₂ CI ₂ (3 x 10 ml) and the resulting organic layers are combined, dried using magnesium sulphate and concentrated to dryness.

Example 39: Further toxicity study

A toxicity study in HepG2 (immortalized human liver carcinoma cells) was carried out. A typical protocol for such a study may be found in Cell Biol Toxicol (2012) 28:69-87. The results of the study are shown in the Table below:

This study confirmed that the conjugates of the invention are substantially less toxic in vivo than is deferasirox itself.