WO2017152237A1

WO2017152237A1 - Novel analogues of desferrioxamine b (dfob)

Info

Publication number: WO2017152237A1
Application number: PCT/AU2017/050215
Authority: WO
Inventors: Rachel Codd; Thomas James TELFER
Original assignee: The University Of Sydney
Priority date: 2016-03-10
Filing date: 2017-03-10
Publication date: 2017-09-14

Abstract

The present invention relates to new Desferrioxamine B-based compounds that are useful for iron-chelation therapy, to their preparation, and to compositions including the compounds. The present invention also relates to the use of the compounds, as well as compositions including the compounds, in the treatment of conditions of iron dyshomeostasis.

Description

Novel analogues of Desferrioxamine B (DFOB)

Field of the invention

The present invention relates to new desferrioxamine B-based compounds that are useful for iron-chelation therapy, to their preparation, and to compositions including the compounds. The present invention also relates to the use of the compounds, as well as compositions including the compounds, in the treatment of conditions of iron

dyshomeostasis (such as iron overload).

Background of the invention

Desferrioxamine B (DFOB) is an iron(lll) chelating molecule produced by the bacterium Streptomyces pilosus (S. pilosus) and other Actinomycetes. The bacteria produce

DFOB to bind iron in the local environment, as an essential requirement for growth. The structure of DFOB (1 ) is given below.

DFOB has been used in the clinic to treat patients with secondary iron overload, which can occur as a complication of the treatment of transfusion-dependent blood disorders, including beta-thalassaemia, sickle cell anaemia and myelodysplastic syndromes. Transfusion-dependent iron overload is the most common condition of metal toxicity worldwide, with the highest mortality. More generally, derangement of iron homeostasis leading to excessive iron intake and storage is deleterious to several tissues, and has been implicated in cardiac dysfunction and failure, liver dysfunction and cirrhosis, and endocrine abnormalities including hypothyroidism, hypogonadism, and diabetes mellitus.

DFOB is effective at removing iron from plasma and it is non-toxic. However, the administration regimen of the drug is arduous, requiring slow sub-cutaneous infusion over 60 to 70 hours per week. This regimen is necessary due to the short plasma half- life of DFOB (10 to 15 minutes), which requires slow administration to maintain steady- state concentrations of the drug. DFOB has a second shortcoming: it is inefficient at removing iron stored inside cells, which is problematic in the context of the accumulation of iron in major organs, including heart, liver and pancreas.

Two synthetic iron chelators, desferasirox and deferiprone, are orally active, but have a lower iron binding affinity, are less selective towards iron and have some toxicities, compared to DFOB.

It would be useful if agents, which provide an improvement over the current iron- chelating therapies, could be developed.

Reference to any prior art in the specification is not an acknowledgment or suggestion that this prior art forms part of the common general knowledge in any jurisdiction or that this prior art could reasonably be expected to be understood, regarded as relevant, and/or combined with other pieces of prior art by a skilled person in the art.

Summary of the invention

The present inventors have proposed that the issues with current agents used in iron- chelation therapy could be overcome by developing an agent that has improved pharmacokinetic properties. Specifically, the present inventors have sought to develop an agent that has a longer plasma half-life, is less prone to degradation by plasma amidases, and has an increased ability to cross cell membranes to access intracellular iron.

In a first aspect, the present invention relates to a compound of formula (I):

(D

or a pharmaceutically acceptable salt or prodrug thereof, wherein: each n is independently selected from 4 or 5; m is 2; and one or more of the CH2 groups are substituted with at least one fluorine atom. The primary amine group can be subsequently converted using semi-synthetic reactions to an amide group, a secondary amine group or an azide group. Therefore, the primary amine group may be replaced with a group selected from an amide group, a secondary amine group and an azide group.

In one embodiment, one CH2 group is substituted with at least one fluorine atom. In another embodiment, two CH2 groups are each substituted with at least one fluorine atom. In another embodiment, three CH2 groups are each substituted with at least one fluorine atom.

In a second aspect, the present invention relates to a pharmaceutical composition including a compound of formula (I) (according to the first aspect of the invention) together with a pharmaceutically acceptable carrier, diluent or excipient. Compounds and pharmaceutical compositions according to the present invention may be suitable for iron chelation therapy. Accordingly, in another aspect, the present invention relates to a method of treating a condition of iron dyshomeostasis in a subject, the method including administering to the subject an effective amount of a compound of formula (I) according to the first aspect of the invention or a pharmaceutical composition according to the second aspect of the invention.

In a further aspect the present invention relates to the use of a compound of formula (I) according to the first aspect of the invention or a pharmaceutical composition according to the second aspect of the invention in the manufacture of a medicament for treating a condition of iron dyshomeostasis. In a further aspect the present invention relates to the use of a compound of formula (I) according to the first aspect of the invention or a pharmaceutical composition according to the second aspect of the invention for the treatment of a condition of iron

dyshomeostasis in a subject.

In a further aspect the present invention relates to a compound of formula (I) according to the first aspect of the invention or a pharmaceutical composition according to the second aspect of the invention for use in the treatment of a condition of iron

dyshomeostasis in a subject.

The condition of iron dyshomeostasis may be selected from primary iron overload and secondary iron overload. The compounds of formula (I) may be used in therapy alone or in combination with one or more other therapeutic agents, for example, as part of a combination therapy. In a further aspect, the present invention relates to a process for making a compound of formula (I) according to the first aspect of the invention, the process including:

- culturing Streptomyces pilosus in a medium supplemented with a fluorinated diamine substrate at a non-toxic concentration. The fluorinated diamine substrate may be selected from 1 ,4-diamino-2-fluorobutane, or a 1 ,4-diaminobutane or 1 ,5-diaminopentane substituted with at least one fluorine atom, or a salt thereof.

The 1 ,4-diamino-2-fluorobutane may be a racemic mixture (i.e. rac-1 ,4-diamino-2- fluorobutane), (R)-1 ,4-diamino-2-fluorobutane or (S)-1 ,4-diamino-2-fluorobutane. The salt may be a hydrochloride salt (e.g. 1 ,4-diamino-2-fluorobutane.2HCI).

The concentration of the fluorinated diamine substrate may be about 35 mM or less.

The medium may include a further compound that inhibits production of 1 ,4- diaminobutane and/or 1 ,5-diaminopentane.

Further aspects of the present invention and further embodiments of the aspects described in the preceding paragraphs will become apparent from the following description, given by way of example and with reference to the accompanying drawings.

Brief description of the drawings

Figure 1. LC trace with total ion current (TIC) detection mode from semi-purified supernatant of S. pilosus cultured in: (a) base medium, or (b) medium supplemented with rac-1 ,4-diamino-2-fluorobutane (rac-DFB), analyzed as isolated or Fe(lll)-loaded solutions of the base medium system (c) or the rac-DFB supplemented system (d). The gradient in (a) or (c) was the same in (b) or (d), respectively. Peak numbering refers to the structures of the compounds 1 -4 or the equivalent 1 : 1 complexes formed with Fe(lll) (number underlined). Figure 2(a). MS signals from LC peaks from the as isolated extract consistent with: DFOB at f_R 34.85 min (1 ); or F1-DFOA1 at f_R 33.80 min (2), or F₂- DFOA₂ at 31.37 min (3); or F3-DFOA3 at 28.92 min (4). Data shown as experiment (lower x axis) or calculated (upper x axis: [M+H]⁺ [M+2H]⁺ adducts),

Figure 2(b). MS signals from LC peaks from the Fe(lll)-loaded extract consistent with: Fe(lll)-DFOB at f_R 28.49 min (1_Fe); or Fe(lll)-Fi-DFOAi at 27.85 min (2_Fe); or Fe(lll)- F2-DFOA2 at 26.84 min (3_Fe); or Fe(lll)-F₃-DFOA3 at 24.20 min (4_Fe). Data shown as experiment (lower x axis) or calculated (upper x axis: [M+H]⁺ and [M+2H]²⁺ adducts).

Figure 3. The production of iron-binding compounds over time (monitored using an Fe(lll) addition assay) using as a substrate the racemic form, as well as the (R)- and (SJ-isomers, of 1 ,4-diamino-2-fluorobutane to supplement a S. pilosus bacteriological medium.

Detailed description of the embodiments

Compounds are generally described herein using standard nomenclature. For compounds having asymmetric centres, it will be understood that, unless otherwise specified, all of the optical isomers and mixtures thereof are encompassed. Compounds with two or more asymmetric elements can also be present as mixtures of

diastereomers. Where a compound exists in various tautomeric forms, a recited compound is not limited to any one specific tautomer, but rather is intended to encompass all tautomeric forms. Compounds according to the formula provided herein, which have one or more stereogenic centres, have an enantiomeric excess of at least 50%. For example, such compounds may have an enantiomeric excess of at least 60%, 70%, 80%, 85%, 90%, 95%, or 98%. Some embodiments of the compounds have an enantiomeric excess of at least 99%. It will be apparent that single enantiomers (optically active forms) can be obtained by asymmetric synthesis, synthesis from optically pure precursors,

biosynthesis or by resolution of the racemates, for example, enzymatic resolution or resolution by conventional methods such as crystallization in the presence of a resolving agent, or chromatography, using, for example, a chiral HPLC column.

A "pharmaceutically acceptable salt" of a compound disclosed herein is an acid or base salt that is generally considered in the art to be suitable for use in contact with the tissues of human beings or animals without excessive toxicity or carcinogenicity, and preferably without irritation, allergic response, or other problem or complication. In particular, pharmaceutically acceptable salts in accordance with the present invention are those that do not adversely affect the ability of the compound to bind iron, and that do not adversely affect the ability of the compound to cross cell membranes to access intracellular iron. Such salts include mineral and organic acid salts of basic residues such as amines. Suitable pharmaceutically acceptable salts include, but are not limited to, salts of acids such as hydrochloric, phosphoric, hydrobromic, malic, glycolic, fumaric, sulfuric, sulfamic, sulfanilic, formic, toluenesulfonic, methanesulfonic, benzenesulfonic, ethane disulfonic, 2-hydroxyethylsulfonic, nitric, benzoic, 2-acetoxybenzoic, citric, tartaric, lactic, stearic, salicylic, glutamic, ascorbic, pamoic, succinic, fumaric, maleic, propionic, hydroxymaleic, hydroiodic, phenylacetic, alkanoic (such as acetic, HOOC-(CH₂)_n-COOH where n is any integer from 0 to 6, i.e. 0, 1 , 2, 3, 4, 5 or 6), and the like. Similarly, pharmaceutically acceptable cations include, but are not limited to sodium, potassium, calcium, aluminum, lithium and ammonium. A person skilled in the art will recognize further pharmaceutically acceptable salts for the compounds provided herein. In general, a pharmaceutically acceptable acid or base salt can be synthesized from a parent compound that contains a basic or acidic moiety by any conventional chemical method. Briefly, such salts can be prepared by reacting the free acid or base forms of these compounds with a stoichiometric amount of the appropriate base or acid in water or in an organic solvent (such as ether, ethyl acetate, ethanol, isopropanol or

acetonitrile), or in a mixture of the two.

It will be apparent that each compound of formula (I) may, but need not, be present as a hydrate, solvate or non-covalent complex. In addition, the various crystal forms and polymorphs are within the scope of the present invention, as are prodrugs of the compounds of formula (I) provided herein.

A "prodrug" is a compound that may not fully satisfy the structural requirements of the compounds provided herein, but is modified in vivo, following administration to a subject or patient, to produce a compound of formula (I) provided herein. For example, a prodrug may be an acylated derivative of a compound as provided herein. Prodrugs include compounds wherein hydroxy, carboxy, amine or sulfhydryl groups are bonded to any group that, when administered to a mammalian subject, cleaves to form a free hydroxy, carboxy, amino, or sulfhydryl group, respectively. Examples of prodrugs include, but are not limited to, acetate, formate, phosphate and benzoate derivatives of alcohol and amine functional groups within the compounds provided herein. Prodrugs of the compounds provided herein may be prepared by modifying functional groups present in the compounds in such a way that the modifications are cleaved in vivo to generate the parent compounds. A "substituent" as used herein, refers to a molecular moiety that is covalently bonded to an atom within a molecule of interest. For example, a "CH₂ substituent" is a moiety such as a halogen or an alkyl group that is covalently bonded to the carbon atom of the CH₂ group. The term "substituted," as used herein, means that any one or more hydrogens on the designated atom is replaced with a selection from the indicated substituents, provided that the designated atom's normal valence is not exceeded, and that the substitution results in a stable compound, i.e., a compound that can be isolated, characterized and tested for biological activity. Suitable substituents are halogen (for example, fluorine, chlorine, bromine or iodine atoms). Particularly preferred is fluorine. As used herein a wording defining the limits of a range of length such as, for example, "from 1 to 5" means any integer from 1 to 5, i.e. 1 , 2, 3, 4 and 5. In other words, any range defined by two integers explicitly mentioned is meant to comprise and disclose any integer defining said limits and any integer comprised in said range.

As discussed above, the present invention relates to a compound of formula (I):

(D

or a pharmaceutically acceptable salt or prodrug thereof, wherein: each n is independently selected from 4 or 5; m is 2; and one or more of the CH₂ groups are substituted with at least one fluorine atom. The primary amine group can be subsequently converted using semi-synthetic reactions to an amide group, a secondary amine group or an azide group. Therefore, in the compound of formula (I), the primary amine group may be replaced with a group selected from an amide group, a secondary amine group and an azide group.

In the compounds of formula (I), one, two or three CH₂ groups may be substituted with at least one fluorine atom. For example, one set of CH₂ groups may be substituted with at least one fluorine atom (e.g. as shown in compound 2 below), two sets of CH₂ groups may be substituted with at least one fluorine atom (e.g. as shown in compound 3 below), or three sets of CH₂ groups may be substituted with at least one fluorine atom (e.g. as shown in compound 4 below).

Examples of compounds of formula (I), in accordance with the present invention, are:

4

The compounds of the present invention can be synthesised by any suitable method known to a person skilled in the art. The present inventors have found that precursor- directed biosynthesis can be used to produce the compounds of the present invention. Briefly, a fluorinated diamine substrate (such as 1 ,4-diamino-2-fluorobutane) is introduced into bacteriological medium inoculated with S. pilosus, or another

Actinomycete. Conditions to prepare base medium as described in Chiani et a/¹ may be used. The present inventors have found that the exogenous substrate (1 ,4-diamino-2- fluorobutane) competes with the native substrate 1 ,5-diaminopentane in the

biosynthesis, which results in the production of a mixture of the compounds of the present invention. These compounds are able to be separated by liquid

chromatography. The compounds retain the ability to bind iron(lll), as demonstrated from mass spectrometry measurements of the compounds in the presence of iron(lll) (Figures 1 and 2). Fluorination chemistry can be difficult so it is particularly attractive that the compounds of the present invention can be produced by a bacterium by co-opting its native biosynthetic machinery. This also makes the synthesis amenable to industrial-scale production, using existing fermentation systems for DFOB production. Accordingly, the present invention also relates to a process for making a compound of formula (I):

(I)

or a pharmaceutically acceptable salt or prodrug thereof, wherein: each n is independently selected from 4 or 5; m is 2; and one or more of the CH₂ groups are substituted with at least one fluorine atom, the process including:

- culturing Streptomyces pilosus in a medium supplemented with a fluorinated diamine substrate at a non-toxic concentration.

The fluorinated diamine substrate may be selected from 1 ,4-diamino-2-fluorobutane, or a 1 ,4-diaminobutane or 1 ,5-diaminopentane substituted with one or more fluorine atoms, or a salt thereof. The fluorine atom(s) may be, for example, substituted on any of the two internal CH₂ groups penultimate to the amine groups of the 1 ,4-diaminobutane, or the three internal CH₂ groups penultimate to the amine groups of the 1 ,5- diaminopentane. Examples of suitable fluorinated diamine substrates include the following:

,4-diaminobutane-based 1 ,5-diaminopentane-based 1 5-dtaminopentane-based

The 1 ,4-diamino-2-fluorobutane (or any of the other fluorinated diamine substrates) may be used as a racemic mixture (e.g. rac-1 ,4-diamino-2-fluorobutane), as a mixture of enantiomers, or in optically-pure form (e.g. as (R)A ,4-diamino-2-fluorobutane or SJ-1 ,4- diamino-2-fluorobutane). The salt may be a hydrochloride salt (e.g. 1 ,4-diamino-2- fluorobutane.2HCI), or a hydrobromide salt.

Native substrates that are used by S. pilosus to prepare DFOB include 1 ,5- diaminopentane and 1 ,4-diaminobutane. These substrates are produced by the bacterium itself. It may be desirable in the process of the present invention to

supplement the medium with a mixture of fluorinated diamines. It may also be useful to supplement the mixture with a compound that attenuates the production of the native diamine substrates. The medium may therefore, for example, include a fluorinated diamine substrate (which is intended to be incorporated into the final product) and may also include a compound that inhibits the production of the native diamine substrate. Under these conditions, it would be expected that, since the bacterium has a reduced supply of the native substrate, it would be effectively forced to use whatever substrate is available (i.e. the fluorinated diamine substrate). This would lead to an increase in the yield of the fluorinated DFOB derivative. Examples of suitable inhibitors include 1 ,4- diamino-2-butanone and 5-hydroxylysine. The total concentration of all of the

exogenous compounds in the medium (the fluorinated diamine substrate and the inhibitor) may be about 35 mM or less. A person skilled in the art will understand that the actual concentration used will vary as a function of the culture conditions.

The process may also include the further step of converting the primary amine group to a group selected from an amide group, a secondary amine group and an azide group. A person skilled in the art will be aware of suitable methods for carrying out these conversions.

Without wishing to be bound by theory or mode of action, the present inventors hypothesise that the compounds of the present invention may have increased plasma protein binding, which in turn may increase the plasma half-life time of the compounds in the body, thereby reducing administration time. Perhaps more importantly, the compounds of the present invention may out-perform DFOB on the measure of accessing intracellular iron, due to the reduction in the p a value of the terminal amine group of the compounds of the present invention that arises from the electron

withdrawing nature of a proximal fluorine group. A reduced p a value translates to a higher proportion of drug present at physiological pH in neutral form, which is required for the effective passage across cell membranes. In addition, an attenuation of plasma degradation may arise due to the presence of the fluorine group proximal to the terminal amine (the region where degradation is thought to be initiated) and/or near the amide bonds for amidase-mediated degradation.

The therapeutic use of compounds of formula (I), their pharmaceutically acceptable salts, solvates, hydrates, prodrugs and also formulations and pharmaceutical

compositions (including mixtures of the compounds of formula (I)) are within the scope of the present invention. Accordingly, the present invention also relates to a

pharmaceutical composition including a compound of formula (I):

(I)

or a pharmaceutically acceptable salt or prodrug thereof, wherein: each n is independently selected from 4 or 5; m is 2; and one or more of the CH₂ groups are substituted with at least one fluorine atom, together with a pharmaceutically acceptable carrier, diluent or excipient. A "pharmaceutical carrier, diluent or excipient" includes, but is not limited to, any physiological buffered (i.e., about pH 7.0 to 7.4) medium including a suitable water soluble carrier, conventional solvents, dispersion media, fillers, solid carriers, coatings, antibacterial and antifungal agents, isotonic and absorption delaying agents. Suitable water soluble carriers include, but are not limited to saline, dextrose, corn oil,

dimethylsulfoxide, and gelatin capsules. Other conventional additives include lactose, mannitol, corn starch, potato starch, binders such as crystalline cellulose, cellulose derivatives, acacia, gelatins, disintegrators such as sodium carboxymethyl-cellulose, and lubricants such as talc or magnesium stearate. Pharmaceutical compositions may be formulated for any appropriate route of

administration including, for example, topical (for example, transdermal or ocular), oral, buccal, nasal, vaginal, rectal or parenteral administration. The term "parenteral" as used herein includes subcutaneous, intradermal, intravascular (for example, intravenous), intramuscular, spinal, intracranial, intrathecal, intraocular, periocular, intraorbital, intrasynovial and intraperitoneal injection, as well as any similar injection or infusion technique. In certain embodiments, compositions in a form suitable for oral use or parenteral use are preferred. Suitable oral forms include, for example, tablets, troches, lozenges, aqueous or oily suspensions, dispersible powders or granules, emulsions, hard or soft capsules, or syrups or elixirs. For intravenous, intramuscular,

subcutaneous, or intraperitoneal administration, one or more compounds may be combined with a sterile aqueous solution which is preferably isotonic with the blood of the recipient. Such formulations may be prepared by dissolving solid active ingredient in water containing physiologically compatible substances such as sodium chloride or glycine, and having a buffered pH compatible with physiological conditions to produce an aqueous solution, and rendering said solution sterile. The formulations may be present in unit or multi-dose containers such as sealed ampoules or vials. Examples of suitable components are described in Martindale - The Extra Pharmacopoeia

(Pharmaceutical Press, London 1993) and Martin (ed.), Remington's Pharmaceutical Sciences.

For the treatment of a condition of iron dyshomeostasis, the dose of the biologically- active compound according to the invention may vary within wide limits and may be adjusted to individual requirements. Active compounds according to the present invention are generally administered in a therapeutically effective amount. Preferred doses range from about 0.1 mg to about 140 mg per kilogram of body weight per day (e.g. about 0.5 mg to about 7 g per patient per day). The daily dose may be

administered as a single dose or in a plurality of doses. The amount of active ingredient that may be combined with the carrier materials to produce a single dosage form will vary depending upon the host treated and the particular mode of administration. Dosage unit forms will generally contain between about 1 mg to about 500 mg of an active ingredient.

It will be understood, however, that the specific dose level for any particular patient will depend upon a variety of factors including the activity of the specific compound employed, the age, body weight, general health, sex, diet, time of administration, route of administration, and rate of excretion, drug combination (i.e. other drugs being used to treat the patient), and the severity of the particular disorder undergoing therapy. Such treatments may be administered as often as necessary and for the period of time judged necessary by the treating physician. A person skilled in the art will appreciate that the dosage regime or therapeutically effective amount of the compound of formula (I) to be administered may need to be optimized for each individual.

It will be appreciated that different dosages may be required for treating different disorders. An effective amount of an agent is that amount which causes a statistically significant decrease in the level of iron burden in the patient. The patient may have a toxic amount of iron in their body as a result of a hereditary or primary hemochromatosis ("primary iron overload"). Iron overload in this condition results from increased intestinal iron absorption and a further derangement of iron metabolism, which occur as a result of a genetic mutation. The patient may have a toxic amount of iron in their body as a result of high parenteral iron administration ("secondary iron overload"), which is primarily observed in association with transfusion-dependent hereditary or acquired anaemias, or other transfusion-dependent conditions (such as chronic liver disease, Friedreich ataxia, aceruloplasminaemia, and congenital atransferrinaemia). Hereditary anaemias include inherited hemoglobinopathies (e.g. beta-thalassaemia and sickle cell disease), Blackfan-Diamond anaemia, congenital dyserythropoiesis anaemia, and sideroplastic anaemia. Acquired anaemias include myelodysplastic syndromes, myelofibrosis, aplastic anaemia, leukaemia, myeloproliferative disorders, stem cell transplantation, and chronic kidney disease. Increased dietary intake of iron may also contribute, or lead, to secondary iron overload. The terms "therapeutically effective amount" or "effective amount" refer to an amount of the compound of formula (I) that results in an improvement or remediation of the symptoms of iron overload. The dosage form and amount of the compounds or pharmaceutical compositions of the present invention can be readily established by reference to known treatment regimens. Preferred compounds of the invention will have certain pharmacological properties.

Such properties include, but are not limited to oral bioavailability and cell membrane permeability, such that the preferred oral dosage forms discussed above can provide therapeutically effective levels of the compound in vivo. The compounds of the present invention are preferably administered to a patient (for example, a human) orally or parenterally, and are present within at least one body fluid or tissue of the patient. Accordingly, the present invention further provides methods for treating patients suffering from a condition of iron dyshomeostasis. The terms "treating", "treatment" and "therapy" are used herein to refer to curative therapy. Therefore, in the context of the present disclosure, the term "treating" encompasses curing and ameliorating the severity of iron overload or its associated symptoms.

Patients may include but are not limited to primates, especially humans, domesticated companion animals such as dogs, cats, horses, and livestock such as cattle, pigs, sheep, with dosages as described herein.

Compounds and pharmaceutical compositions according to the present invention may be suitable for iron chelation therapy. Accordingly, the present invention also relates to a method of treating a condition of iron dyshomeostasis in a patient including

administration to the patient of a therapeutically effective amount of a compound of formula (I), or a pharmaceutically-acceptable salt, solvate, hydrate or prodrug thereof. The present invention also relates to the use of a therapeutically effective amount of a compound of formula (I), or a pharmaceutically-acceptable salt, solvate, hydrate or prodrug thereof, for treating a condition of iron dyshomeostasis. The present invention also provides a pharmaceutical composition for use in treating a condition of iron dyshomeostasis, in any of the embodiments described in the specification. The present invention also relates to the use of a therapeutically effective amount of a compound of formula (I), or a pharmaceutically acceptable salt, solvate, hydrate or prodrug thereof, for the manufacture of a medicament for treating a condition of iron dyshomeostasis. The present invention also relates to a compound of formula (I), or a pharmaceutically acceptable salt, solvate, hydrate or prodrug thereof, when used in a method of treating a condition of iron dyshomeostasis. The present invention also relates to a composition having an active ingredient for use in treating a condition of iron dyshomeostasis, wherein the active ingredient is a compound of formula (I), or a pharmaceutically acceptable salt, solvate, hydrate or prodrug thereof. The present invention also relates to the use of a pharmaceutical composition containing a compound of the formula (I), or a pharmaceutically acceptable salt, solvate, hydrate or prodrug thereof, in treating a condition of iron dyshomeostasis, such as described above. In one embodiment, the compound of formula (I) is essentially the only active ingredient of the composition. In one embodiment, the condition of iron dyshomeostasis is primary or secondary iron overload. Alternatively, or in addition to, the compounds may be administered in combination with other agents, for example, deferasirox, deferiprone and Desferal.

It will be understood that the invention disclosed and defined in this specification extends to all alternative combinations of two or more of the individual features mentioned or evident from the text or drawings. All of these different combinations constitute various alternative aspects of the invention.

The invention therefore provides, but is not limited to, the following embodiments:

1 . A compound of formula (I):

(I)

or a pharmaceutically acceptable salt or prodrug thereof, wherein: each n is independently selected from 4 or 5; m is 2; and one or more of the CH₂ groups are substituted with at least one fluorine atom.

2. The compound of embodiment 1 , wherein one CH₂ group is substituted with at least one fluorine atom. 3. The compound of embodiment 1 or embodiment 2, wherein two CH₂ groups are each substituted with at least one fluorine atom.

4. The compound of any one of the preceding embodiments, wherein three CH₂ groups are each substituted with at least one fluorine atom.

In embodiments 1 to 4, the fluorine atoms may be in various substitution patterns such as mono, vicinal and geminal. In one embodiment, the fluorine atoms are vicinal. In another embodiment, the fluorine atoms are geminal.

5. The compound of any one of the preceding embodiments, wherein the compound is selected from the group consisting of:

OH O OH O OH

4

6. The compound of any one of the preceding embodiments, wherein the primary amine group is replaced with a group selected from an amide group, a secondary amine group and an azide group. 7. A process for making a compound of formula (I):

I H m I H m l

OH OH OH

(I)

- culturing Streptomyces pilosus in a medium supplemented with a fiuorinated diamine substrate at a non-toxic concentration.

8. The process of embodiment 7, wherein the fiuorinated diamine substrate is selected from 1 ,4-diamino-2-fluorobutane, or a 1 ,4-diaminobutane or 1 ,5- diaminopentane substituted with one or more fluorine atoms, or a salt thereof.

9. The process of embodiment 8, wherein the fiuorinated diamine substrate is 1 ,4- diamino-2-fluorobutane, or a salt thereof.

10. The process of embodiment 8 or embodiment 9, wherein the salt is a

hydrochloride salt.

1 1 . The process of any one of embodiments 6 to 10, wherein the medium is further supplemented with a compound that inhibits production of 1 ,4-diaminobutane and/or

1 ,5-diaminopentane. 12. The process of any one of embodiments 7 to 1 1 , wherein, in the compound of formula (I), one CH₂ group is substituted with fluorine.

13. The process of any one of embodiments 7 to 12, wherein two CH₂ groups are each substituted with fluorine. 14. The process of any one of embodiments 7 to 13, wherein three CH₂ groups are each substituted with fluorine.

15. The process of any one of embodiments 7 to 14, wherein the compound of formula (I) is selected from the group consisting of:

4

16. The process of any one of embodiments 7 to 15, wherein the process includes the further step of converting the primary amine group to a group selected from an amide group, a secondary amine group and an azide group. 17. A pharmaceutical composition including a compound of formula (I):

I H m I H m l

OH OH OH

(I)

or a pharmaceutically acceptable salt or prodrug thereof, wherein: each n is independently selected from 4 or 5; m is 2; and one or more of the CH₂ groups are substituted with at least one fluorine atom, together with a pharmaceutically acceptable carrier, diluent or excipient.

18. The pharmaceutical composition of embodiment 17, wherein the composition is suitable for parenteral or oral administration.

19. The pharmaceutical composition of embodiment 17 or 18, wherein, in the compound of formula (I), one CH₂ group is substituted with at least one fluorine atom.

20. The pharmaceutical composition of any one of embodiments 17 to 19, wherein two CH₂ groups are each substituted with at least one fluorine atom. 21 . The pharmaceutical composition of any one of embodiments 17 to 20, wherein three CH₂ groups are each substituted with at least one fluorine atom.

22. The pharmaceutical composition of any one of embodiments 17 to 21 , wherein the compound is selected from the group consisting of:

4

23. The pharmaceutical composition of any one of embodiments 17 to 22, wherein the primary amine group is replaced with a group selected from an amide group, a secondary amine group and an azide group. 24. A method of treating a condition of iron dyshomeostasis in a patient including administration to the patient of a therapeutically effective amount of a compound of formula (I):

I H m I H m I

OH OH OH

(I)

or a pharmaceutically-acceptable salt or prodrug thereof, wherein: each n is independently selected from 4 or 5; m is 2; and one or more of the CH₂ groups are substituted with at least one fluorine atom.

25. The method of embodiment 24, wherein the administration is selected from parenteral or oral administration.

26. The method of embodiment 24 or embodiment 25, wherein the condition of iron dyshomeostasis is primary iron overload or secondary iron overload.

27. The method of any one of embodiments 24 to 26, wherein, in the compound of formula (I), one CH₂ group is substituted with at least one fluorine atom. 28. The method of any one of embodiments 24 to 27, wherein two CH₂ groups are each substituted with at least one fluorine atom.

29. The method of any one of embodiments 24 to 28, wherein three CH₂ groups are each substituted with at least one fluorine atom.

30. The method of any one of embodiments 24 to 29, wherein the compound is selected from the group consisting of:

2

4

31 . The method of any one of embodiments 24 to 30, wherein the primary amine group is replaced with a group selected from an amide group, a secondary amine group and an azide group.

Embodiments of the invention will now be discussed in more detail with reference to the examples which is provided for exemplification only and which should not be considered limiting on the scope of the invention in any way. Examples

Preparation of compounds

A pre-culture was established by adding an aliquot of a S. pilosus permanent to a 250- ml_ Erienmeyer flask filled with standard treated media (50 mL). This was shaken at 160 rpm and 28 °C for 96 h. The bacteria was withdrawn, spun down on a centrifuge, the supernatant was removed and replaced with fresh media, and were re-suspended before further use.

The following enriched medium was prepared that contained per 50 mL: YM broth (2.1 %), KH₂P0₄.3H₂0 (235 mM), Na₂HP0₄ (1 1 .6 mM), MgS0₄.7H₂0 (2.43 mM), CaCI₂ (13.6 mM), ZnS0₄.7H₂0 (13.9 μΜ), trizma base (35.0 mM), and threonine (0.84 mM). The enriched medium has been optimized for DFOB production.¹ The bacteria were split evenly between two cultures (50 mL) in 250 mL Erienmeyer flasks under the following conditions: enriched media control and racA ,4-diamino-2-fluorobutane.2HCI (10 mM, rac-DFB). The specific conditions were achieved by adding a concentrated, pH adjusted, syringe filtered standard solution of substrate to the media before bacteria addition. The bacteria were shaken at 160 rpm and 28 °C for 8 days, taking 700 μί aliquots of media from each culture on each day. A ferric assay was performed on these aliquot. After the 8 day growth cycle, the bacteria from each culture were collected and were lyophilised to dryness to approximate the bacterial load from each culture. The extracts were purified using conditions described previously.² Iron chelation

Extracts were analysed using liquid chromatography-mass spectrometry (LC-MS) as described previously.² Compared to the native system (Figure 1 (a)), the extract from the DFB-supplemented system (Figure 1 (b)) showed signals in the LC that analysed using MS as the fluorinated derivatives 2-4. Compounds were identified from MS patterns that represented the target fluorinated compounds (Figure 2). These compounds retained function as Fe(lll) chelating molecules (Figure 3), as shown from analysis in the presence of added Fe(lll). The compounds are racemic mixtures.

The (R)- and (SJ-isomers of 1 ,4-diamino-2-fluorobutane have also been used to supplement the S. pilosus bacteriological medium, and the production of iron-binding compounds over time monitored using an Fe(lll) addition assay (see Figure 3).

References

1 . M. Chiani, A. Akbarzadeh, A. Farhangi, M. Mazinani, Z. Saffari, K.

Emadzadeh, M.R. Mehrabi, Pak. J. Biol. Sci. 13 (2010) 546-550.

2. C.Z. Soe, R. Codd, ACS Chem. Biol. 9 (2014) 945-956. 3. T. J. Telfer, M. P. Gotsbacher, C. Z. Soe, R. Codd, ACS Chem. Biol. 11

(2016) 1452-1462.

Claims

1 . A compound of formula (I):

(I)

2. The compound of claim 1 , wherein one CH₂ group is substituted with at least one fluorine atom.

3. The compound of claim 1 or claim 2, wherein two CH₂ groups are each substituted with at least one fluorine atom.

4. The compound of any one of the preceding claims, wherein three CH₂ groups are each substituted with at least one fluorine atom.

5. The compound of any one of the preceding claims, wherein the compound is selected from the group consisting of:

3

4

6. The compound of any one of the preceding claims, wherein the primary amine group is replaced with a group selected from an amide group, a secondary amine group and an azide group.

7. A process for making a compound according to any one of claims 1 to 5, the process including:

8. The process of claim 7, wherein the fluorinated diamine substrate is selected from 1 ,4-diamino-2-fluorobutane, or a 1 ,4-diaminobutane or 1 ,5-diaminopentane substituted with one or more fluorine atoms, or a salt thereof.

9. The process of claim 8, wherein the fluorinated diamine substrate is 1 ,4- diamino-2-fluorobutane, or a salt thereof.

10. The process of claim 8 or claim 9, wherein the salt is a hydrochloride salt.

1 1 . The process of any one of claims 6 to 10, wherein the medium is further supplemented with a compound that inhibits production of 1 ,4-diaminobutane and/or 1 ,5-diaminopentane.

12. The process of any one of claims 7 to 12, wherein the process includes the further step of converting the primary amine group to a group selected from an amide group, a secondary amine group and an azide group.

13. A pharmaceutical composition according to any one of claims 1 to 6, together with a pharmaceutically acceptable carrier, diluent or excipient.

14. A method of treating a condition of iron dyshomeostasis in a patient including administration to the patient of a therapeutically effective amount of a compound of any one of claims 1 to 6.

15. The method of claim 14, wherein the condition of iron dyshomeostasis is primary iron overload or secondary iron overload.