A SORTILIN ANTAGONIST FOR USE IN THE PREVENTION OR TREATMENT OF HEARING LOSS

FIELD OF THE INVENTION

The present invention relates to the use of sortilin modulators, in particular compounds of Formula (I), (II), (III) and (IV), in the treatment or prevention of hearing loss.

BACKGROUND

Hearing loss refers to either the partial, or total, acquired inability of an individual to hear. Accordingly, hearing loss can result in a severely reduced quality of life. Hearing loss may have a variety of causes, for example, the natural aging process, repeated exposure to loud noises or as a consequence of exposure to various toxins or pharmaceutical substances. Hearing loss is most commonly associated with damage to the hair cells, structures in the inner ear, which once damaged are unable to divide or regenerate themselves. Currently, there are no approved drugs for treating hearing loss. Current, symptomatic, treatment options for individuals suffering from hearing loss primarily revolve around the use of hearing aids or cochlear implants. Whilst these devices are able to improve the hearing of some individuals, not all individuals experience the desired effect. Additionally, such devices are deemed to be aesthetically displeasing and only treat the symptoms of hearing loss, and not the underlying disease pathology. Further, the available treatments are not useful for preventing or restoring the initial hearing loss nor preventing progression of hearing loss.

Sortilin (encoded by SORT1) is a type 1 membrane receptor in the vacuolar protein sorting 10 protein (VPS10P) family of sorting receptors, and is abundantly expressed in the central nervous system, the inner ear, and in some peripheral tissues involved in metabolic control ¹ ’ ² ’ ³ ’ ⁴ . Sortilin has an amino acid sequence according to SEQ ID NO: 1 and comprises a signal peptide, a propeptide, the Vps1 Op domain, a 10cc domain (1 OCCa + 10CCb), a transmembrane domain and a large cytoplasmic tail. The luminal domain of sortilin has 6 potential /V-linked glycosylation sites, whilst the cytoplasmic tail enables for the recruitment of various adapter proteins.

Sorti I in binds to a vast number of ligands and membrane receptors and as a result engages in functions known to be important in cellular signalling and sorting. For example, sortilin is involved in signalling by proneurotrophins: the proforms of nerve growth factor (proNGF), brain derived neurotrophic factor (proBDNF), and neurotrophin-3 (proNT3), respectively. In complex with the protein p75NTR (p75 neurotrophin receptor), sortilin has been reported to form the receptor for pro- neurotrophin-mediated apoptotic effects leading to degeneration and cell death in cellular and animal models ⁵ ’ ⁶⁷ .

There is an unmet need for new substances useful in the treatment, e.g. by means of delaying progression of symptoms, and prevention of hearing loss, which address the underlying causes of the disease and not just the symptoms.

DESCRIPTION OF THE FIGURES

Figure 1 shows the mean maximal perilymph concentration of the compound exemplified in Example 8 in male Hartley guinea-pigs following trans-tympanic administration at either 0.1 mg/mL or 1 mg/mL at 1 h, 8 h and 72 h post dosing. Data is shown as Mean ± SD.

Figure 2 shows the mean maximal perilymph concentration of the compound exemplified in Example 8 in male Harley guinea-pigs following oral administration at 1 mg/mL, at 8 h and 24 h following three days of daily dosing. Data is shown as Mean ± SD.

DISCLOSURE OF THE INVENTION

In a first aspect, the present invention provides a sortilin antagonist for use in the prevention or treatment of hearing loss. Without wishing to be bound by theory, it is thought that the protein sortilin is upregulated in the ear of hearing loss patients. Sortilin can form a trimeric complex with the low-affinity nerve growth factor receptor (p75NTR) and a pro- neurotrophin, such as proNGF, proBDNF or proNT3, resulting in cell death of inner ear cells while at the same time preventing the conversion of the proform into its mature and trophic form ¹ . The present invention provides substances which interfere with the binding of sortilin to pro-neurotrophins, which: 1) prevents the trimeric apoptotic complex of sortilin, p75NTR and the pro-neurotrophin from forming, and 2) prevents sortilin mediated clearance of the pro-neurotrophins, providing a pool of substrate to be converted into the mature and trophic neurotrophin. Thus, sortilin-proneurotrophin interference can be used in protecting the ear from damage resulting from the aforementioned cell death.

It has surprisingly been found that sortilin antagonists, in particular compounds of Formula (I), (la), (II), (III), and (IV) may be useful in the treatment or prevention of hearing loss.

According to another aspect, the invention provides the use of a sortilin antagonist in the manufacture of a medicament for the prevention or treatment of hearing loss.

According to another aspect of the invention, there is provided a method of preventing or treating hearing loss by administering a therapeutically effective amount of a sortilin antagonist to a subject in need thereof.

The invention also provides a pharmaceutical formulation comprising the sortilin antagonist of the invention.

As used herein, the term “sortilin” may refer to full length sortilin (also referred to as immature sortilin), comprising a signal peptide, a propeptide, a Vps1 Op domain, a 10CC domain, a transmembrane domain and a large cytoplasmic tail, having an amino acid sequence according to SEQ ID NO: 1 or SEQ ID NO: 2, or it may refer to mature sortilin, comprising a Vps10p domain, a 10CC domain, a transmembrane domain and a large cytoplasmic tail, having an amino acid sequence according to SEQ ID NO: 3, or a naturally occurring fragment, homologue or variant thereof. The term “sortilin” or “sortilin molecule” (used interchangeably herein) as used herein may also include any protein that is functionally related to sortilin, i.e. a sortil in-related molecule, for example, SorSCI , SorCS2, SorCS3 or SorLA/SORL1. Therefore, for the avoidance of doubt, any reference to the term “sortilin” in a use, method or composition of the invention, may not only refer to sortilin as defined above but may also be taken to optionally alternatively refer to a functionally related protein, such as SorCSI , SorCS2, SorCS3 or SorLA/SORL1 . It is understood that the sortilin is capable of interacting with a pro-neurotrophin molecule to form a sortilin/pro-neurotrophin complex. This sortilin/pro-neurotrophin complex may or may not be capable of interacting with a p75NTR molecule to form a trimeric complex comprising sortilin, pro-neurotrophin and p75NTR. It is understood that this trimeric complex may be responsible for adverse biological responses, such as stimulating apoptosis in retinal and ganglion cells.

As used herein, the term “pro-neurotrophin” refers to the larger precursors of neurotrophins, which undergo proteolytic cleavage to yield the mature form of the neurotrophin. Neurotrophins are a family of proteins that induce the survival, development and function of neurons, and are commonly referred to as growth factors. Pro-neurotrophins are biologically active and have distinct roles compared to their neurotrophin counterparts, such as induction of apoptosis. Examples of pro-neurotrophins include proNGF, proBDNF, proNT3, proNT4, preferably the pro-neurotrophin is proNGF, proBDNF or proNT3, even more preferably the pro-neurotrophin is proNGF.

As used herein, the term “sortilin antagonist” refers to a substance (such as an antibody, protein, or other molecule) that interferes with, blocks, or otherwise attenuates the effect of, a sortilin protein binding to a pro-neurotrophin (e.g proNGF, proNT3, proBDNF) and preventing the formation of the trimeric complex between sortilin, p75NTR and the pro-neurotrophin. The term “sortilin antagonist” also includes a substance or agent that interferes with the formation of a high affinity trimeric complex. In the latter scenario, it is recognised that a trimeric complex may be formed in that sortilin can bind to p75NTR (but not proNGF) and p75NTR can simultaneously bind the NGF domain of proNGF. However, the resulting trimeric complex may be of lower affinity for its receptor and as a result have significantly reduced capacity to stimulate apoptosis via the mechanism describe above. The term “sortilin antagonist” also includes a substance or agent that interferes with, blocks, or otherwise attenuates the effect of, a sortilin protein interacting with p75NTR. This interaction may be completely prevented, in which case the trimeric complex is prevented from forming, or only partially prevented, in which case the trimeric complex may be formed but may have reduced biological potency.

The sortilin antagonist herein disclosed is intended for the use in the prevention or treatment of hearing loss. The hearing loss may for example be noise-induced hearing loss, ototoxicity-induced hearing loss, age-induced hearing loss, idiopathic hearing loss, tinnitus and/or sudden hearing loss. It is envisaged that the sortilin antagonist herein disclosed may be particularly useful for the prevention or treatment of noise-induced hearing loss and ototoxicity-induced hearing loss.

Thus, in an embodiment, the sortilin antagonist for use according to the invention may disrupt interaction between a sortilin molecule and a pro-neurotrophin molecule, or disrupt the interaction between a sortilin molecule and a p75NTR molecule. Said sortilin molecule may be mature sortilin or a sortilin-related molecule such as SorCS1 , SorCS2, SorCS3 or SorLA.

Preferably, the sortilin antagonist is a sortilin inhibitor. As used herein, the term “sortilin inhibitor” refers to a substance that binds to a sortilin protein, thereby preventing it from binding to a pro-neurotrophin and preventing the formation of the aforementioned trimeric complex, or resulting in the formation of a trimeric complex that is less active or inactive. The term “sortilin inhibitor” may also refer to a soluble fragment of sortilin, or sortilin-related molecule, which can bind competitively to biologically available pro-neurotrophins but prevent formation of an effective trimeric complex. As a result, the sortilin inhibitor results in an inhibition of receptor activation, which would otherwise occur as a result of ligand binding and/or allows for a shift in unbound/biologically available pro-neurotrophin, which in turn can act through a secondary route/receptor either in its pro- or mature form, specifically, but not limited to Trk family receptors.

Preferably, the sortilin antagonist of the present invention prevents the proteinprotein interaction between a sortilin molecule and a pro-neurotrophin, further preventing the formation of the apoptotic trimeric complex usually formed between sortilin, pro-neurotrophin and the p75NTR receptor, or resulting in the formation of a low affinity trimeric complex, which is biologically less active or inactive or has minimal activity.

The term “sortilin antagonist” is intended to include any molecule which has the desired effect herein described. Accordingly, the sortilin antagonist may for example be a small molecule pharmaceutical compound or a larger molecule, such as a biologic. Examples of suitable larger molecule biologies include polypeptides, such as immunoglobulins (i.e. antibodies) or fragments thereof. As would be understood by a person of skill in the art, a small molecule would typically be produced by means of a chemical process whereas a biologic may be produced by means of a biological process, such as recombinant expression in a living cell or in cell free expression system. Where the sortilin antagonist is a small molecule compound, the sortilin antagonist may have a molecular weight of <2000 Da, preferably the sortilin antagonist has a molecular weight of <1000 Da.

In a preferred aspect of the invention, the sortilin antagonist is a compound of Formula (I): or a pharmaceutically acceptable salt, solvate, hydrate, geometrical isomer, tautomer, N-oxide, and/or prodrug thereof, wherein

Y is selected from the group consisting of -O-, -NR ⁵ -, and -S-;

Z is selected from the group consisting of optionally substituted C ₁ -C ₆ alkyl, optionally substituted C ₆ -C ₁₀ aryl, and optionally substituted C ₁ -C ₉ heteroaryl;

A, B, C and D are each independently selected from the group consisting of H, optionally substituted C ₁ -C ₆ alkyl, halo, NO2, optionally substituted C ₆ -C ₁₀ aryl, optionally substituted C ₁ -C ₉ heteroaryl, -OR ⁶ , NR ⁷ R ⁸ , -SR ⁹ , -C(O)OR ¹⁰ , -C(O)NR ¹¹ R ¹² , -C(O)SR ¹³ , C(O ₂ )SR ¹⁴ ;

R ¹ , R ⁴ , R ⁵ , R ⁶ , R ⁹ , R ¹⁰ , R ¹³ , and R ¹⁴ are each independently selected from the group consisting of H and C ₁ -C ₆ alkyl group, wherein the C ₁ -C ₆ alkyl is optionally substituted with one or more halo atoms; and

R ² , R ³ , R ⁷ , R ⁸ , R ¹¹ , and R ¹² are each independently selected from the group consisting of H and C ₁ -C ₆ alkyl, wherein the C ₁ -C ₆ alkyl is optionally substituted with one or more halo atoms, or R ² and R ³ , R ⁷ and R ⁸ , and/or R ¹¹ and R ¹² can be taken together with the nitrogen atom to which they are attached to from a 5- or 6-membered heterocycle, optionally substituted with one or more halo atoms;

Compounds of Formula (I) act as sortilin antagonists by binding to sortilin. They are therefore useful in the treatment or prevention of diabetic retinopathy.

It is highly preferred that YY is -NR ⁵ -, mmoorree preferably Y is

-NH-. In a particularly preferred aspect of the invention, the compound of Formula

(I) is a compound of Formula (la): Preferred compounds of Formula (I) and (la), as detailed below, may exhibit increased binding affinity to sortilin and therefore increased efficacy in the treatment or prevention of hearing loss.

In a preferred aspect of the invention, A and D are H.

In another preferred aspect of the invention, B and C are independently selected from the group consisting of H, halo, CF ₃ , NO ₂ , and C ₁ -C ₄ alkyl.

It is more preferred that C is H.

It is also more preferred that B is selected from the group consisting of halo, t-Bu, and CF ₃ , more preferably B is Br, I, t-Bu or CF ₃ .

In another preferred aspect of the invention, Z is an optionally substituted Ce-Cw aryl or an optionally substituted C ₁ -C ₉ heteroaryl.

It is preferred that the optionally substituted C ₆ -C ₁₀ aryl is an optionally substituted phenyl.

Particularly preferred optionally substituted C ₁ -C ₉ heteroaryl groups are optionally substituted 5- or 6-membered heteroaryl groups in which 1 to 5 of the ring atoms are carbon and one or more of the ring atoms are a heteroatom, more preferably optionally substituted 6-membered heteroaryl groups in which 1 to 5 of the ring atoms are carbon and one or more of the ring atoms are a heteroatom.

It is preferred that the ring atoms of the optionally substituted 6-membered heteroaryl group are selected from the group consisting of carbon and nitrogen. In which case, particularly preferred examples of optionally substituted C ₁ -C ₉ heteroaryl groups are optionally substituted pyridyl, pyridazinyl, pyrimidinyl, pyrazinyl, and triazinyl. More preferred optionally substituted C ₁ -C ₉ heteroaryl groups are optionally substituted pyridyl and pyrimidinyl. Therefore, in a particularly preferred aspect of the invention, Z is an optionally substituted phenyl, an optionally substituted pyridyl, or an optionally substituted pyrimidinyl.

Z may be substituted with one or more substituents independently selected from the group consisting of -OR ¹⁵ , halo, and C ₁ -C ₄ alkyl, wherein R ¹⁵ is H, halo, and C ₁ -C ₃ alkyl. More preferably, Z is substituted with one or more substituents selected from the group consisting of Cl, -OMe, and Me.

In addition to the above-mentioned preferred optional substituents, an alkylene group may be attached to two adjacent atoms of group Z to form a 5- or 6-membered partially saturated or saturated ring, optionally wherein the alkylene group is substituted with one or more halo atoms. Examples of such fused bicyclic ring systems are shown below in the following two examples of group Z:

Particular combinations of Z and its substituents may be particularly preferred, as detailed below.

In a particularly preferred aspect of the invention, Z is C ₆ -C ₁₀ aryl or C ₁ -C ₉ heteroaryl. Each of C ₆ -C ₁₀ aryl and C ₁ -C ₉ heteroaryl are independently substituted with one or more substituents independently selected from the group consisting of -OR ¹⁵ , halo, and C ₁ -C ₄ alkyl, wherein R ¹⁵ is H, halo, or C ₁ -C ₃ alkyl, preferably one or more substituents independently selected from the group consisting of Cl, -OMe, and Me; and/or an alkylene group is attached to two adjacent atoms of group Z to form a 5- or 6-membered partially saturated or saturated ring, optionally wherein the alkylene group is substituted with one or more halo atoms.

In an even more preferred aspect of the invention, Z is phenyl, pyridyl, or pyrimidinyl. Each of phenyl, pyridyl, and pyrimidinyl are independently substituted with one or more substituents independently selected from the group consisting of -OR ¹⁵ , halo, and C ₁ -C ₄ alkyl, wherein R ¹⁵ is H, halo, or C ₁ -C ₃ alkyl, preferably one or more substituents independently selected from the group consisting of Cl, -OMe, and Me; and/or an alkylene group is attached to two adjacent atoms of group Z to form a 5- or 6-membered partially saturated or saturated ring, optionally wherein the alkylene group is substituted with one or more halo atoms.

Particular compounds of Formula (I) for use according to the invention are:

• 2-[(6-methylpyridin-2-yl)carbamoyl]benzoic acid;

• 2-methyl-6-[(6-methylpyridin-2-yl)carbamoyl]benzoic acid;

• 5-bromo-2-[(6-methylpyridin-2-yl)carbamoyl]benzoic acid;

• 5-chloro-2-[(6-methylpyridin-2-yl)carbamoyl]benzoic acid;

• 5-methyl-2-[(6-methylpyridin-2-yl)carbamoyl]benzoic acid;

• 2-[(6-methylpyridin-2-yl)carbamoyl]-5-(propan-2-yl)benzoic acid;

• 2-[(6-methylpyridin-2-yl)carbamoyl]-5-(trifluoromethyl)benzo ic acid;

• 2-[(6-methylpyridin-2-yl)carbamoyl]-5-nitrobenzoic acid;

• 4-[(6-methylpyridin-2-yl)carbamoyl]-[1 , 1 '-biphenyl]-3-carboxylic acid;

• 4-bromo-2-[(6-methylpyridin-2-yl)carbamoyl]benzoic acid;

• 4-chloro-2-[(6-methylpyridin-2-yl)carbamoyl]benzoic acid;

• 4-methyl-2-[(6-methylpyridin-2-yl)carbamoyl]benzoic acid;

• 2-[(6-methylpyridin-2-yl)carbamoyl]-4-(trifluoromethyl)benzo ic acid;

• 4,5-dichloro-2-[(6-methylpyridin-2-yl)carbamoyl]benzoic acid;

• 4,5-dimethyl-2-[(6-methylpyridin-2-yl)carbamoyl]benzoic acid;

• 3-methyl-2-[(6-methylpyridin-2-yl)carbamoyl]benzoic acid;

• 5-bromo-2-[(butan-2-yl)carbamoyl]benzoic acid;

• 5-bromo-2-[(propan-2-yl)carbamoyl]benzoic acid;

• 5-bromo-2-(phenylcarbamoyl)benzoic acid;

• 5-bromo-2-[(3-methylphenyl)carbamoyl]benzoic acid;

• 5-bromo-2-[(pyridin-2-yl)carbamoyl]benzoic acid;

• 5-bromo-2-[(6-chloropyridin-2-yl)carbamoyl]benzoic acid;

• 5-bromo-2-[(4-methylpyridin-2-yl)carbamoyl]benzoic acid; • 5-bromo-2-[(3-methylpyridin-2-yl)carbamoyl]benzoic acid;

• 5-bromo-2-[(6-methylpyridin-3-yl)carbamoyl]benzoic acid;

• 5-bromo-2-[(2-methylpyridin-4-yl)carbamoyl]benzoic acid;

• 5-bromo-2-[(2-methylpyrimidin-4-yl)carbamoyl]benzoic acid;

• 2-[(6-methoxypyridin-2-yl)carbamoyl]-5-(trifluoromethyl)benz oic acid;

• 2-[(5,6-dimethylpyridin-2-yl)carbamoyl]-5-(trifluoromethyl)b enzoic acid;

• 2-[(5,6,7,8-tetrahydroquinolin-2-yl)carbamoyl]-5-(trifluorom ethyl)benzoic acid, or a pharmaceutically acceptable salt, solvate, hydrate, tautomer, optical isomer, N-oxide, and/or prodrug thereof.

Particularly preferred compounds of Formula (I) for use according to the invention are:

• 5-bromo-2-[(6-methylpyridin-2-yl)carbamoyl]benzoic acid;

• 5-chloro-2-[(6-methylpyridin-2-yl)carbamoyl]benzoic acid;

• 5-methyl-2-[(6-methylpyridin-2-yl)carbamoyl]benzoic acid;

• 2-[(6-methylpyridin-2-yl)carbamoyl]-5-(propan-2-yl)benzoic acid;

• 2-[(6-methylpyridin-2-yl)carbamoyl]-5-(trifluoromethyl)benzo ic acid;

• 2-[(6-methylpyridin-2-yl)carbamoyl]-5-nitrobenzoic acid;

• 4-bromo-2-[(6-methylpyridin-2-yl)carbamoyl]benzoic acid;

• 2-[(6-methylpyridin-2-yl)carbamoyl]-4-(trifluoromethyl)benzo ic acid;

• 4,5-dichloro-2-[(6-methylpyridin-2-yl)carbamoyl]benzoic acid;

• 4,5-dimethyl-2-[(6-methylpyridin-2-yl)carbamoyl]benzoic acid;

• 5-bromo-2-[(3-methylphenyl)carbamoyl]benzoic acid;

• 5-bromo-2-[(pyridin-2-yl)carbamoyl]benzoic acid;

• 5-bromo-2-[(6-chloropyridin-2-yl)carbamoyl]benzoic acid;

• 5-bromo-2-[(4-methylpyridin-2-yl)carbamoyl]benzoic acid;

• 5-bromo-2-[(2-methylpyridin-4-yl)carbamoyl]benzoic acid;

• 5-bromo-2-[(2-methylpyrimidin-4-yl)carbamoyl]benzoic acid;

• 2-[(6-methoxypyridin-2-yl)carbamoyl]-5-(trifluoromethyl)benz oic acid;

• 2-[(5,6-dimethylpyridin-2-yl)carbamoyl]-5-(trifluoromethyl)b enzoic acid; • 2-[(5,6,7,8-tetrahydroquinolin-2-yl)carbamoyl]-5-(trifluorom ethyl)benzoic acid; or a pharmaceutically acceptable salt, solvate, hydrate, tautomer, optical isomer, N-oxide, and/or prodrug thereof.

In another preferred aspect of the invention, the sortilin antagonist is a compound of Formula (II): or a pharmaceutically acceptable salt, solvate, hydrate, tautomer, optical isomer, N-oxide, and/or prodrug thereof, wherein

R ¹⁶ is C1-C6 alkyl, optionally substituted with one or more halo atoms; n is 0, 1 , or 2;

E is selected from the group consisting of H, optionally substituted 6- to 10-membered aryl, and optionally substituted 5- to 10-membered heteroaryl;

Preferred compounds of Formula (II), as detailed below, may exhibit increased binding affinity to sortilin and therefore increased efficacy in the treatment or prevention of hearing loss according to the invention.

It is preferred that R ¹⁶ is t-butyl or neopentyl, most preferably t-butyl.

It is also preferred that n is 1 or 2, most preferably n is 1 .

In preferred compounds of Formula (II), E is selected from the group consisting of H, optionally substituted phenyl, and optionally substituted 5- or 6-membered heteroaryl. Preferably, phenyl and 5- or 6-membered heteroaryl are each independently optionally substituted with one or more substituents independently selected from the group consisting of:

(i) halo, C ₁ -C ₃ alkyl, and C ₁ -C ₃ alkoxy; and

(ii) phenyl, -O-phenyl, -CH ₂ -phenyl, 5- or 6-membered heteroaryl, -O-(5- or 6-membered heteroaryl), and -CH ₂ -(5- or 6-membered heteroaryl), each of which is independently optionally substituted with one or more halo atoms.

In a more preferred aspect of the invention, E is selected from the group consisting of:

(a) H;

(b) phenyl and 5- or 6-membered heteroaryl, optionally substituted with one or more substituents independently selected from the group consisting of halo, phenyl, phenoxyl, and 5- or 6-membered heteroaryl.

Preferably, each 5-membered heteroaryl group defined above is independently selected from the group consisting of pyrrolyl, pyrazolyl, imidazolyl, triazolyl, furanyl, oxazolyl, isoxazolyl, oxadiazolyl.

Preferably, each 6-membered heteroaryl group defined above is independently selected from the group consisting of pyridyl, pyrimidinyl, pyridazinyl, pyrazinyl, triazinyl, most preferably pyridyl.

In an even more preferred aspect of the invention, E is selected from the group consisting of:

(a) H;

(b) phenyl, optionally substituted with one or more halo atoms; and (c) pyridyl and oxadiazolyl, optionally substituted with one or more substituents independently selected from halo, phenyl, phenoxyl, and pyrazolyl.

Particular compounds of Formula (II) for use according to the invention are:

• 1 -benzyl-3-tert-butyl-1 H-pyrazole-5-carboxylic acid;

• 3-tert-butyl-1 -[(3,5-dichlorophenyl)methyl]-1 H-pyrazole-5-carboxylic acid;

• 3-tert-buty I- 1 -{[6-(1 H-pyrazol-1 -y l)py ridin-3-y l]methy I}- 1 H-pyrazole-5- carboxylic acid;

• 3-tert-buty I- 1 -[(6-phenoxypyridin-3-yl)methyl]-1 H-pyrazole-5-carboxylic acid;

• 3-tert-butyl-1 -methyl-1 H-pyrazole-5-carboxylic acid;

• 3-tert-buty I- 1 -phenyl-1 H-pyrazole-5-carboxylic acid;

• 3-tert-buty I- 1 -(2-phenylethyl)-1 H-pyrazole-5-carboxylic acid;

• 3-tert-buty I- 1 -[(5-phenyl-1 ,3,4-oxadiazol-2-yl)methyl]-1 H-pyrazole-5- carboxylic acid;

• 1 -benzyl-3-(2,2-dimethylpropyl)-1 H-pyrazole-5-carboxylic acid;

• 1 -(2-phenylethyl)-3-(propan-2-yl)-1 H-pyrazole-5-carboxylic acid;

• 1 -benzyl-3-(3,3-dimethylbutyl)-1 H-pyrazole-5-carboxylic acid; or a pharmaceutically acceptable salt, solvate, hydrate, tautomer, optical isomer, N-oxide, and/or prodrug thereof.

Particularly preferred compounds of Formula (II) for use according to the invention are:

• 1 -benzyl-3-tert-butyl-1 H-pyrazole-5-carboxylic acid;

• 3-tert-buty I - 1 -[(3,5-dichlorophenyl)methyl]-1 H-pyrazole-5-carboxylic acid;

• 3-tert-buty I - 1 -[(6-phenoxypyridin-3-yl)methyl]-1 H-pyrazole-5-carboxylic acid;

• 3-tert-buty I- 1 -[(5-phenyl-1 ,3,4-oxadiazol-2-yl)methyl]-1 H-pyrazole-5- carboxylic acid; or a pharmaceutically acceptable salt, solvate, hydrate, tautomer, optical isomer, N-oxide, and/or prodrug thereof. In another preferred aspect of the invention, the sortilin antagonist is a compound of Formula (III) or (IV): or a pharmaceutically acceptable salt, solvate, hydrate, geometrical isomer, tautomer, N-oxide, and/or prodrug thereof, wherein

W is selected from the group consisting of optionally substituted C ₁ -C ₆ alkyl, optionally substituted 6- to 10-membered aryl, optionally substituted 5- to 10-membered heteroaryl, optionally substituted 3- to 7-membered cycloalkyl, and optionally substituted 5- or 6-membered heterocyclyl.

R ¹⁷ is oxo or CF ₃ ;

R ¹⁸ is H or Me;

R ¹⁹ is selected from the group consisting of C ₁ -C ₆ alkyl, C ₃ -C ₇ cycloalkyl, C ₂ -C ₆ alkenyl, C ₁ -C ₆ alkoxy, and phenyl.

Preferred compounds of Formula (III) or (IV), as detailed below, may exhibit increased binding affinity to sortilin and therefore increased efficacy in the treatment or prevention of hearing loss according to the invention.

It is preferred that R ¹⁷ is oxo.

It is also preferred that R ¹⁸ is H.

It is also preferred that R ¹⁹ is selected from the group consisting of C ₁ -C ₆ alkyl, C ₃ - C ₇ cycloalkyl, C ₂ -C ₆ alkenyl, and C ₁ -C ₆ alkoxy, more preferably isopropyl, cyclohexyl, t-butyl, neopentyl, and t-butoxyl, more preferably t-butyl and neopentyl, most preferably neopentyl.

W is optionally substituted with one or more substituents. It is preferred that W is optionally substituted with one or more substituents independently selected from the group consisting of:

(i) halo, C ₁ -C ₄ alkyl, and C ₁ -C ₃ alkoxy; and

(ii) phenyl, -O-phenyl, -CH ₂ -phenyl, 5- or 6-membered heteroaryl, -O-(5- or 6- membered heteroaryl), and -CH ₂ -(5- or 6-membered heteroaryl), each of which is independently optionally substituted with one or more halo atoms;

In a more preferred aspect of the invention, W is selected from the group consisting of phenyl, -( C ₁ -C ₄ alkyl)-phenyl, 5- or 6-membered heteroaryl, 5- or 6-membered heterocyclyl, C ₃ -C ₇ cycloalkyl, and 9- or 10-membered bicyclic aryl or heteroaryl, each of which is independently optionally substituted with one or more substituents independently selected from the group consisting of:

(i) halo, C ₁ -C ₄ alkyl, and C ₁ -C ₃ alkoxy; and

(ii) benzyl and phenoxyl, each of which is independently optionally substituted with one or more halo atoms;

In an even more preferred aspect of the invention, W is selected from the group consisting of:

(a) phenyl and -(C ₁ -C ₄ alkyl)-phenyl, wherein each phenyl group is optionally substituted with one or more substituents independently selected from halo and C ₁ -C ₄ alkoxy;

(b) 5-membered heteroaryl, optionally substituted with one or more substituents independently selected from the group consisting of halo and C ₁ -C ₄ alkyl; (c) 6-membered heteroaryl and 6-membered heterocyclyl, optionally substituted with one or more substituents independently selected from the group consisting of halo, benzyl, and phenoxyl, wherein benzyl and phenoxyl are each independently optionally substituted with one or more halo atoms;

(d) C ₃ -C ₇ cycloalkyl, optionally substituted with one or more halo atoms; and

(e) 10-membered bicyclic heteroaryl, optionally substituted with one or more halo atoms.

Preferably, the 5-membered heteroaryl of group W is selected from the group consisting of pyrrolyl, pyrazolyl, imidazolyl, triazolyl, furanyl, oxazolyl, isoxazolyl, oxadiazolyl, most preferably pyrrolyl and pyrazolyl.

Preferably, the 6-membered heteroaryl of group W is selected from the group consisting of pyridyl, pyrimidinyl, pyridazinyl, pyrazinyl, triazinyl, most preferably pyridyl.

Preferably, the 5-membered heterocyclyl of group W is selected from the group consisting of pyrrolidinyl, pyrrolinyl, pyrazolidinyl, imidazolidinyl, pyrazolinyl, imidazolinyl, tetrahydrofuranyl, and dioxolanyl.

Preferably, the 6-membered heterocyclyl of group W is selected from the group consisting of morpholinyl, piperidinyl, piperazinyl, dioxanyl, and tetrahydropyranyl, most preferably morpholinyl.

Preferably, the 9- or 10-membered bicyclic aryl of heteroaryl of group W is selected from the group consisting of indenyl, naphthalenyl, indolyl, isoindolyl, indazolyl, benzimidazolyl, azaindolyl, azaindazolyl, benzofuranyl, isobenzofuranyl, benzoisoxazolyl, benzoxazolyl, quinoxalinyl, phthalazinyl, cinnolinyl, quinazolinyl, naphthyridinyl, pyridopyrimidinyl, pyridopyrazinyl, quinolinyl, and isoquinolinyl, most preferably quinoxalinyl.

Preferably, the 10-membered bicyclic heteroaryl of group W is selected from the group consisting of quinoxalinyl, phthalazinyl, cinnolinyl, quinazolinyl, naphthyridinyl, pyridopyrimidinyl, pyridopyrazinyl, quinolinyl, isoquinolinyl, most preferably quinoxalinyl.

Particular compounds of Formula (III) and (IV) for use according to the invention are:

• (2S)-2-[(3,5-dichlorophenyl)formamido]-5,5-dimethylhexanoic acid;

• (2S)-2-[(4-chloro-1 H-pyrrol-2-yl)formamido]-5,5-dimethylhexanoic acid;

• (2S)-5,5-dimethyl-2-[(6-phenoxypyridin-3-yl)formamido]hexano ic acid;

• (2S)-2-[(3,4-dichlorophenyl)formamido]-4-methylpentanoic acid;

• (2S)-2-[(3,5-dichlorophenyl)formamido]-4-methylpentanoic acid;

• (2S)-2-[(3-chlorophenyl)formamido]-4-methylpentanoic acid;

• (2S)-2-[(3-methoxyphenyl)formamido]-4-methylpentanoic acid;

• (2S)-2-{2-[(4-chlorophenyl)methyl]-2-methylpropanamido}-4-me thylpentanoic acid;

• (2S)-4-methyl-2-[(1 -methyl-1 H-pyrazol-4-yl)formamido]pentanoic acid;

• (2S)-4-methyl-2-[(quinoxalin-2-yl)formamido]pentanoic acid;

• (2S)-2-(cyclopentylformamido)-4-methylpentanoic acid;

• (2S)-2-({4-[(3,5-dichlorophenyl)methyl]morpholin-2-yl}formam ido)-4- methylpentanoic acid;

• (2S)-2-[(3,5-dichlorophenyl)formamido]-4-methylpent-4-enoic acid;

• (2S)-3-cyclopropyl-2-[(3,5-dichlorophenyl)formamido]propanoi c acid;

• (2S)-3-cyclohexyl-2-[(3,5-dichlorophenyl)formamido]propanoic acid;

• (2S)-2-[(3,5-dichlorophenyl)formamido]-4,4-dimethylpentanoic acid;

• (2S) -3-{bi cy clo[ 1 .1.1 ]pentan-1 -y l}-2-[(3, 5- dichlorophenyl)formamido]propanoic acid;

• (2S)-2-[(3,5-dichlorophenyl)formamido]-3-phenylpropanoic acid;

• (2S)-3-(tert-butoxy)-2-[(3,5-dichlorophenyl)formamido]propan oic acid;

• (2S)-3-(tert-butoxy)-2-[(3,5-dichlorophenyl)formamido]butano ic acid;

• 3,5-dichloro-N-[3,3-dimethyl-1 -(2H-1 ,2,3,4-tetrazol-5-yl)butyl]benzamide;

• (2S)-2-{[1-(3,5-dichlorophenyl)-2,2,2-trifluoroethyl]amino}- 4-methylpentanoic acid; • (2S)-4,4-dimethyl-2-[(6-phenoxypyridin-3-yl)formamido]pentan oic acid, or a pharmaceutically acceptable salt, solvate, hydrate, tautomer, N-oxide, and/or prodrug thereof.

Particularly preferred compounds of Formula (III) and (IV) for use according to the invention are:

• (2S)-2-[(3,5-dichlorophenyl)formamido]-5,5-dimethylhexanoic acid;

• (2S)-2-[(4-chloro-1 H-pyrrol-2-yl)formamido]-5,5-dimethylhexanoic acid;

• (2S)-5,5-dimethyl-2-[(6-phenoxypyridin-3-yl)formamido]hexano ic acid;

• (2S)-2-[(3,5-dichlorophenyl)formamido]-4-methylpentanoic acid;

• (2S)-2-({4-[(3,5-dichlorophenyl)methyl]morpholin-2-yl}formam ido)-4- methylpentanoic acid;

• (2S)-3-cyclohexyl-2-[(3,5-dichlorophenyl)formamido]propanoic acid;

• (2S)-2-[(3,5-dichlorophenyl)formamido]-4,4-dimethylpentanoic acid;

• (2S)-3-(tert-butoxy)-2-[(3,5-dichlorophenyl)formamido]propan oic acid;

• (2S)-4,4-dimethyl-2-[(6-phenoxypyridin-3-yl)formamido]pentan oic acid; or a pharmaceutically acceptable salt, solvate, hydrate, tautomer, N-oxide, and/or prodrug thereof.

According to another aspect of the invention, there is provided a pharmaceutical formulation comprising a sortilin antagonist according to the invention and one or more pharmaceutically acceptable carriers or excipients, for use in the treatment or prevention of hearing loss.

The compounds for use according to the invention may include isotopically- labelled and/or isotopically-enriched forms of the compounds. The compounds for use according to the invention herein may contain unnatural proportions of atomic isotopes at one or more of the atoms that constitute such compounds. Examples of isotopes that can be incorporated into the disclosed compounds include isotopes of hydrogen, carbon, nitrogen, oxygen, phosphorus, sulfur, chlorine, such as ² H, ³ H, ¹¹ C, ¹³ C, ¹⁴ C, ¹³ N, ¹⁵ O, ¹⁷ 0, ³² P, ³⁵ S, ¹⁸ F, ³⁶ CI. The compounds for use according to the invention may be used as such or, where appropriate, as pharmacologically acceptable salts (acid or base addition salts) thereof. The pharmacologically acceptable addition salts mentioned below are meant to comprise the therapeutically active non-toxic acid and base addition salt forms that the compounds are able to form. Compounds that have basic properties can be converted to their pharmaceutically acceptable acid addition salts by treating the base form with an appropriate acid. Exemplary acids include inorganic acids, such as hydrogen chloride, hydrogen bromide, hydrogen iodide, sulphuric acid, phosphoric acid; and organic acids such as formic acid, acetic acid, propanoic acid, hydroxyacetic acid, lactic acid, pyruvic acid, glycolic acid, maleic acid, malonic acid, oxalic acid, benzenesulphonic acid, toluenesulphonic acid, methanesulphonic acid, trifluoroacetic acid, fumaric acid, succinic acid, malic acid, tartaric acid, citric acid, salicylic acid, p-aminosalicylic acid, pamoic acid, benzoic acid, ascorbic acid and the like. Exemplary base addition salt forms are the sodium, potassium, calcium salts, and salts with pharmaceutically acceptable amines such as, for example, ammonia, alkylamines, benzathine, and amino acids, such as, e.g. arginine and lysine. The term addition salt as used herein also comprises solvates which the compounds and salts thereof are able to form, such as, for example, hydrates, alcoholates and the like.

Throughout the present disclosure, a given chemical formula or name shall also encompass all pharmaceutically acceptable salts, solvates, hydrates, geometrical isomers, tautomers, optical isomers, N-oxides, and/or prodrug forms thereof. It is to be understood that the compounds for use according to the invention include any and all hydrates and/or solvates of the compound formulas. It is appreciated that certain functional groups, such as the hydroxy, amino, and like groups form complexes and/or coordination compounds with water and/or various solvents, in the various physical forms of the compounds. Accordingly, the above formulas are to be understood to include and represent those various hydrates and/or solvates.

Compounds for use according to the invention also include tautomeric forms. Tautomeric forms result from the swapping of a single bond with an adjacent double bond together with the concomitant migration of a proton. Tautomeric forms include prototropic tautomers which are isomeric protonation states having the same empirical formula and total charge. Example prototropic tautomers include ketone - enol pairs, amide - imidic acid pairs, lactam - lactim pairs, amide - imidic acid pairs, enamine - imine pairs, and annular forms where a proton can occupy two or more positions of a heterocyclic system, for example, 1 H- and 3H- imidazole, 1 H, 2H- and 4H- 1 ,2,4-triazole, 1 H- and 2H- isoindole, and 1 H- and 2H- pyrazole. Tautomeric forms can be in equilibrium or sterically locked into one form by appropriate substitution.

The compounds described herein can be asymmetric (e.g. having one or more stereocenters). All stereoisomers, such as enantiomers and diastereomers, are intended unless otherwise indicated. Compounds for use according to the present invention that contain asymmetrically substituted carbon atoms can be isolated in optically active or racemic forms. Methods on how to prepare optically active forms from optically active starting materials are known in the art, such as by resolution of racemic mixtures or by stereoselective synthesis. Many geometric isomers of olefins, C=N double bonds, and the like can also be present in the compounds described herein, and all such stable isomers are contemplated in the present invention. Cis- and trans-geometric isomers of the compounds for use according to the present invention are described and may be isolated as a mixture of isomers or as separated isomeric forms.

In the case of the compounds, which contain an asymmetric carbon atom, the invention relates to the D form, the L form, and D, L mixtures and also, where more than one asymmetric carbon atom is present, to the diastereomeric forms. Those compounds for use according to the invention which contain asymmetric carbon atoms, and which as a rule accrue as racemates, can be separated into the optically active isomers in a known manner, for example using an optically active acid. However, it is also possible to use an optically active starting substance from the outset, with a corresponding optically active or diastereomeric compound then being obtained as the end product.

The term "prodrugs" refers to compounds that may be converted under physiological conditions or by solvolysis to a biologically active compound for use according to the invention. A prodrug may be inactive when administered to a subject in need thereof, but is converted in vivo to an active compound of the invention. Prodrugs are typically rapidly transformed in vivo to yield the parent compound of the invention, e.g. by hydrolysis in the blood. The prodrug compound usually offers advantages of solubility, tissue compatibility or delayed release in a mammalian organism (see Silverman, R. B., The Organic Chemistry of Drug Design and Drug Action, 2nd Ed., Elsevier Academic Press (2004), page 498 to 549). Prodrugs of a compound for use according to the invention may be prepared by modifying functional groups, such as a hydroxy, amino or mercapto groups, present in a compound for use according to the invention in such a way that the modifications are cleaved, either in routine manipulation or in vivo, to the parent compound for use according to the invention. Examples of prodrugs include, but are not limited to, acetate, formate and succinate derivatives of hydroxy functional groups or phenyl carbamate derivatives of amino functional groups.

The term “treatment” as used herein may include prophylaxis of the named disorder or condition, or amelioration or elimination of the disorder once it has been established. The term “prevention” refers to prophylaxis of the named disorder or condition.

Methods delineated herein include those wherein the subject is identified as in need of a particular stated treatment. Identifying a subject in need of such treatment can be in the judgment of a subject or a health care professional and can be subjective (e.g. opinion) or objective (e.g. measurable by a test or diagnostic method).

In other aspects, the methods herein include those further comprising monitoring subject response to the treatment administrations. Such monitoring may include periodic imaging or sampling of subject tissue, fluids, specimens, cells, proteins, chemical markers, genetic materials, etc. as markers or indicators of the treatment regimen. In other methods, the subject is pre-screened or identified as in need of such treatment by assessment for a relevant marker or indicator of suitability for such treatment.

The invention provides a method of monitoring treatment progress. The method includes the step of determining a level of diagnostic marker (Marker) (e.g. any target or cell type delineated herein modulated by a compound herein) or diagnostic measurement (e.g., screen, assay) in a subject suffering from or susceptible to a disorder or symptoms thereof delineated herein, in which the subject has been administered a therapeutic amount of a compound herein sufficient to treat the disease or symptoms thereof. The level of Marker determined in the method can be compared to known levels of Marker in either healthy normal controls or in other afflicted patients to establish the subject's disease status. In preferred embodiments, a second level of Marker in the subject is determined at a time point later than the determination of the first level, and the two levels are compared to monitor the course of disease or the efficacy of the therapy. In certain preferred embodiments, a pre-treatment level of Marker in the subject is determined prior to beginning treatment according to this invention; this pretreatment level of Marker can then be compared to the level of Marker in the subject after the treatment commences, to determine the efficacy of the treatment.

A level of Marker or Marker activity in a subject may be determined at least once. Comparison of Marker levels, e.g., to another measurement of Marker level obtained previously or subsequently from the same patient, another patient, or a normal subject, may be useful in determining whether therapy according to the invention is having the desired effect, and thereby permitting adjustment of dosage levels as appropriate. Determination of Marker levels may be performed using any suitable sampling/expression assay method known in the art or described herein. Preferably, a tissue or fluid sample is first removed from a subject. Examples of suitable samples include blood, urine, tissue, mouth or cheek cells, and hair samples containing roots. Other suitable samples would be known to the person skilled in the art. Determination of protein levels and/or mRNA levels (e.g., Marker levels) in the sample can be performed using any suitable technique known in the art, including, but not limited to, enzyme immunoassay, is ELISA, radiolabeling/assay techniques, blotting/chemiluminescence methods, real-time PCR, and the like.

For clinical use, the compounds disclosed herein are formulated into pharmaceutical compositions (or formulations) for various modes of administration. It will be appreciated that compounds for use according to the invention may be administered together with a physiologically acceptable carrier, excipient, and/or diluent (i.e. one, two, or all three of these). The pharmaceutical compositions disclosed herein may be administered by any suitable route, preferably by oral, ocular (including intravitreal), rectal, nasal, topical (including buccal and sublingual), sublingual, transdermal, intrathecal, transtympanic, transmucosal or parenteral (including subcutaneous, intramuscular, intravenous and intradermal) administration. Preferably, the pharmaceutical compositions are administered orally or transtympanically. Other formulations may conveniently be presented in unit dosage form, e.g., tablets and sustained release capsules, and in liposomes, and may be prepared by any methods well known in the art of pharmacy. Pharmaceutical formulations are usually prepared by mixing the active substance, or a pharmaceutically acceptable salt thereof, with conventional pharmaceutically acceptable carriers, diluents or excipients. Examples of excipients are water, gelatin, gum arabicum, lactose, microcrystalline cellulose, starch, sodium starch glycolate, calcium hydrogen phosphate, magnesium stearate, talcum, colloidal silicon dioxide, and the like. Such formulations may also contain other pharmacologically active agents, and conventional additives, such as stabilizers, wetting agents, emulsifiers, flavouring agents, buffers, and the like. Usually, the amount of active compounds is between 0.1-95% by weight of the preparation, preferably between 0.2-20% by weight in preparations for parenteral use and more preferably between 1-50% by weight in preparations for oral administration. The formulations can be further prepared by known methods such as granulation, compression, microencapsulation, spray coating, etc. The formulations may be prepared by conventional methods in the dosage form of tablets, eye drops, creams, capsules, granules, powders, syrups, suspensions, suppositories or injections. Liquid formulations may be prepared by dissolving or suspending the active substance in water or other suitable vehicles. Tablets and granules may be coated in a conventional manner. To maintain therapeutically effective plasma concentrations for extended periods of time, compounds disclosed herein may be incorporated into slow release formulations.

In a preferred embodiment, the sortilin antagonists herein described may be administered in a gel formulation, preferably a Poloxamer 407 gel formulation.

Said formulation may comprise the following excipients, reactants and solvents; Poloxamer 407, 5% glucose, Sodium dihydrogenophosphate, Di-sodium hydrogenophosphate, acetonitrile, trifluoroacetic acid and de-ionized water. Preferably, the gel formulation is a 14 % poloxamer solution in diluted pH 7.4 phosphate buffer (0.013M) in deionized water. Such a formulation has been shown to be liquid (fluid) at room temperature and quickly becomes a gel at 37°C (body temperature), see Table 1 below. Additionally, the advantageous properties of such a formulation have been shown to be unaltered following the administration of the compounds herein disclosed, for example, AF38469 (example 8) at the targeted 1 mg/mL concentration. Accordingly, the aforementioned gel formulation is an appropriate vehicle solution to administer said compounds.

Table 1 : Poloxamer gelling tests at 20°C and 37°C. The dose level and frequency of dosage of the specific compound will vary depending on a variety of factors including the potency of the specific compound employed, the metabolic stability and length of action of that compound, the patient's age, body weight, general health, sex, diet, mode and time of administration, rate of excretion, drug combination, the severity of the condition to be treated, and the patient undergoing therapy. The daily dosage may, for example, range from about 0.001 mg to about 100 mg per kilo of body weight, administered singly or multiply in doses, e.g. from about 0.01 mg to about 25 mg each. Normally, such a dosage is given orally but parenteral administration may also be chosen.

As described above, in addition to the small molecule compounds herein described, the sortilin antagonist of the present invention may also be a biologic molecule.

Accordingly, the sortilin antagonist for use according to the present invention may be an anti-sorti lin antibody or an antigen-binding fragment thereof.

By the term “an antibody or antigen-binding fragment thereof” we intend any substantially intact antibody molecules, as well as chimeric antibodies, humanised antibodies, isolated human antibodies, single chain antibodies, bispecific antibodies, antibody heavy chains, antibody light chains, homodimers and heterodimers of antibody heavy and/or light chains, and antigen-binding fragments and derivatives of the same. Suitable antigen-binding fragments and derivatives include, but are not limited to, Fv fragments, Fab-like fragments, single variable domains and domain antibodies. The skilled person will recognise that using antibody fragments can confer a number of advantages compared to the use of whole antibodies. For example, the smaller size of the fragments may lead to improved pharmacological properties, such as better penetration of solid tissue. Moreover, antigen-binding fragments such as Fab, Fv, ScFv and dAb antibody fragments can be expressed in and secreted from bacteria, such as E. coli, thus allowing the facile production of large amounts of the said fragments. The phrase “an antibody or an antigen-binding fragment thereof’ is also intended to encompass antibody mimics (for example, non-antibody scaffold structures that have a high degree of stability yet allow variability to be introduced at certain positions). Those skilled in the art of biochemistry will be familiar with many such molecules, as discussed in Gebauer & Skerra, 2009, Curr Opin Chem Biol 13(3): 245-255 (the disclosures of which are incorporated herein by reference). Exemplary antibody mimics include: affibodies (also called Trinectins; Nygren, 2008, FEBS J, 275, 2668-2676); CTLDs (also called Tetranectins; Innovations Pharmac. Technol. (2006), 27-30); adnectins (also called monobodies; Meth. Mol. Biol., 352 (2007), 95-109); anticalins (Drug Discovery Today (2005), 10, 23-33); DARPins (ankyrins; Nat. Biotechnol. (2004), 22, 575-582); avimers (Nat. Biotechnol. (2005), 23, 1556-1561); microbodies (FEBS J, (2007), 274, 86-95); peptide aptamers (Expert. Opin. Biol. Ther. (2005), 5, 783-797); Kunitz domains (J. Pharmacol. Exp. Ther. (2006) 318, 803-809); affilins (Trends. Biotechnol. (2005), 23, 514-522); affimers (Avacta Life Sciences, Wetherby, UK).

The skilled person will appreciate that the invention also encompasses modified versions of the antibodies and antigen-binding fragments herein disclosed. For example, via the attachment of polyethylene glycol or another suitable polymer.

To elicit the desired effect, the sortilin antibody or the antigen-binding fragment herein disclosed may bind against any sequence of sortilin that produces said desired effect, i.e. an inhibitory effect. Such an effect may result from the inhibition of the binding of ligands to their respective binding sites, the inhibition of the formation of complexes with co-receptors or alterations in the confirmation of the receptor.

In preferred embodiments, the anti-sortilin antibody or the antigen-binding fragment thereof binds to at least one amino acid of any of the amino acid sequences according to SEQ ID NO: 1 to 14, or at least one amino acid of the amino acid sequence TGL.

The sortilin antibody or the antigen-binding fragment thereof may be a polyclonal antibody or a monoclonal antibody. Those skilled in the art will readily understand the methods of production by which polyclonal and monoclonal antibodies directed against sortilin are produced. The use of a polyclonal antibody may affect the binding of several ligands, due to polyclonal antibodies binding to several binding sites, accordingly several downstream pathways may be inhibited/disrupted. The use of a monoclonal antibody results in the antibody binding to a specific site and therefore will inhibit access of other molecules to that specific site.

The sortilin antibody or the antigen-binding fragment thereof may be obtained from a variety of different species. In a preferred embodiment, the sortilin antibody or antigen-binding fragment thereof may be a human anti-sortilin antibody, a goat anti-sortilin antibody, a rabbit anti-sortilin antibody or an IgG antibody. Said antibody or antigen-binding fragment thereof may also be a humanised antibody. Whilst the preferred antibody isotype may be an IgG antibody, it is understood that the present invention also encompasses other antibody isotypes, for example, IgM, IgD, IgA or IgE.

The sortilin antibody or the antigen-binding fragment thereof may also be conjugated to a cell-penetrating peptide in order to aid the delivery of the antibody or antigen-binding fragment to the desired location. Such a cell-penetrating peptide may be a TAT peptide.

DEFINITIONS

“Optional” or “optionally” means that the subsequently described event or circumstance may, but need not, occur, and that the description includes instances where the event or circumstance occurs and instances in which it does not.

The term “C ₁ -C ₆ alkyl” denotes a straight or branched alkyl group having from 1 to 6 carbon atoms, i.e. 1 , 2, 3, 4, 5 or 6 carbon atoms. For parts of the range “C ₁ -C ₆ alkyl” all subgroups thereof are contemplated, such as C1-C5 alkyl, C ₁ -C ₄ alkyl, C ₁ - C ₃ alkyl, C ₁ -C ₂ alkyl, C ₁ alkyl, C ₂ -C ₆ alkyl, C ₂ -C ₅ alkyl, C ₂ -C ₄ alkyl, C ₂ -C ₃ alkyl, C ₂ alkyl, C ₃ -C ₆ alkyl, C ₃ -C ₅ alkyl, C ₃ -C ₄ alkyl, C ₃ alkyl, C ₄ -C ₆ alkyl, C ₄ -C ₅ alkyl, C ₄ alkyl, C ₅ -C ₆ alkyl, C ₅ alkyl, and C ₆ alkyl. Examples of “C ₁ -C ₆ alkyl” include methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, sec-butyl, and t-butyl.

When a term denotes a range, for instance “1 to 6 carbon atoms” in the definition of C ₁ -C ₆ alkyl, each integer is considered to be disclosed, i.e. 1 , 2, 3, 4, 5 and 6.

The term “C ₃ -C ₇ cycloalkyl” denotes a cyclic or partially cyclic alkyl group having from 3 to 7 carbon atoms, i.e. 3, 4, 5, 6, or 7 carbon atoms. Examples of “C ₃ -C ₇ cycloalkyl” include cyclopropyl, cyclobutyl, cyclopropylmethyl, cyclopentyl, and cyclohexyl. The term “cycloalkyl” includes bridged bicyclic cycloalkyl groups such as the following group:

The term “C ₂ -C ₆ alkenyl” denotes a straight or branched alkyl group having at least one carbon-carbon double bond, and having from 2 to 6 carbon atoms. Examples of “C ₂ -C ₆ alkenyl” include 2-propenyl, 2-butenyl, 3-butenyl, 2-methyl-2-propenyl, 2-hexenyl, 5-hexenyl, 2,3-dimethyl-2-butenyl.

The term “C ₁ -C ₆ alkoxy” denotes -O-(C ₁ -C ₆ alkyl) in which a C ₁ -C ₆ alkyl group is as defined above and is attached to the remainder of the compound through an oxygen atom. Examples of “C ₁ -C ₆ alkoxy” include methoxy, ethoxy, n-propoxy, isopropoxy, n-butoxy, isobutoxy, sec-butoxy, t-butoxy and straight- and branched- chain pentoxy and hexoxy.

The term “halo atom” means a halogen atom, and is preferably F, Cl, Br or I.

The term “oxo” denotes a double bond to an oxygen atom (=O). This typically forms a ketone or aldehyde group.

The term “C ₆ -C ₁₀ aryl” or “6- to 10-membered aryl” denotes an aromatic monocyclic or fused bicyclic hydrocarbon ring system comprising 6 to 10 ring atoms. Examples of “C ₆ -C ₁₀ aryl” and “6- to 10-membered aryl” groups include phenyl, indenyl, naphthyl, and naphthalene.

The term “C ₁ -C ₉ heteroaryl” or “5- to 10-membered heteroaryl” denotes an aromatic monocyclic or fused bicyclic heteroaromatic ring system having 5 to 10 ring atoms in which 1 to 9 of the ring atoms are carbon and one or more of the ring atoms are selected from nitrogen, sulphur, and oxygen. Examples of “C ₁ -C ₉ heteroaryl” and “5- to 10-membered heteroaryl” include furyl, pyrrolyl, thienyl, oxazolyl, isoxazolyl, imidazolyl, thiazolyl, isothiazolyl, pyridinyl, pyrimidinyl, tetrazolyl, quinazolinyl, indolyl, indolinyl, isoindolyl, isoindolinyl, pyrazolyl, pyridazinyl, pyrazinyl, quinolinyl, quinoxalinyl, thiadiazolyl, benzofuranyl, 2,3- dihydrobenzofuranyl, 1 ,3-benzodioxolyl, 1 ,4-benzodioxinyl, 2,3-dihydro-1 ,4- benzodioxinyl, benzothiazolyl, benzimidazolyl, benzothiadiazolyl, benzotriazolyl and chromanyl.

The term “5-membered heterocycle” denotes a monocyclic ring system having 5 ring atoms in which 1 to 4 of the ring atoms are carbon and one or more of the ring atoms are selected from nitrogen, sulfur, and oxygen. Examples of “5- memebered heterocycle” include tetrahydrofuranyl, and pyrrolidinyl.

The term “6-membered heterocycle” denotes a monocyclic ring system having 6 ring atoms in which 1 to 5 of the ring atoms are carbon and one or more of the ring atoms are selected from nitrogen, sulfur, and oxygen. Examples of “5- memebered heterocycle” include tetrahydrofuranyl, and pyrrolidinyl.

“An effective amount” refers to an amount of a compound for use according to the invention that confers a therapeutic effect on the treated subject. The therapeutic effect may be objective (i.e. measurable by some test or marker) or subjective (i.e. subject gives an indication of or feels an effect).

As used herein, the terms “administration” or “administering” mean a route of administration for a compound disclosed herein. Exemplary routes of administration include, but are not limited to, oral, ocular (such as intravitreal), topical, intravenous, intraperitoneal, intraarterial, and intramuscular. The preferred route of administration can vary depending on various factors, e.g. the components of the pharmaceutical composition comprising a compound disclosed herein, site of the potential or actual disease and severity of disease.

The terms "subject" and "patient" are used herein interchangeably. They refer to a human or another mammal (e.g., mouse, rat, rabbit, dog, cat, cattle, swine, sheep, horse or primate) that can be afflicted with or is susceptible to a disease or disorder but may or may not have the disease or disorder. It is preferred that the subject is human.

Compounds for use according to the invention may be disclosed by the name or chemical structure. If a discrepancy exists between the name of a compound and its associated chemical structure, then the chemical structure prevails.

PREPARATION OF COMPOUNDS OF THE INVENTION

The compounds of Formulae (I), (II), (III), and (IV) disclosed herein may be prepared by, or in analogy with, conventional methods. Appropriate reaction conditions for the individual reaction steps are known to a person skilled in the art. The necessary starting materials for preparing the compounds of Formulae (I) and (II) are either commercially available, or may be prepared by methods known in the art.

The compounds of Formulae (I), (II), (III), and (IV) may possess one or more chiral carbon atoms, and they may therefore be obtained in the form of optical isomers, e.g., as a pure enantiomer, or as a mixture of enantiomers (racemate) or as a mixture containing diastereomers. The separation of mixtures of optical isomers to obtain pure enantiomers is well known in the art and may, for example, be achieved by fractional crystallization of salts with optically active (chiral) acids or by chromatographic separation on chiral columns.

Particular experimental procedures for examples of the invention are described below. The processes may be carried out to give a compound of the invention in the form of a free base or as an acid addition salt. A pharmaceutically acceptable acid addition salt may be obtained by dissolving the free base in a suitable organic solvent and treating the solution with an acid, in accordance with conventional procedures for preparing acid addition salts from base compounds. Examples of addition salt forming acids are mentioned above. The chemicals used in the synthetic routes delineated herein may include, for example, solvents, reagents, catalysts, and protecting group and deprotecting group reagents. Examples of protecting groups are t-butoxycarbonyl (Boc), benzyl and trityl(triphenylmethyl). The methods described below may also additionally include steps, either before or after the steps described specifically herein, to add or remove suitable protecting groups in order to ultimately allow synthesis of the compounds. In addition, various synthetic steps may be performed in an alternate sequence or order to give the desired compounds.

Synthetic chemistry transformations and protecting group methodologies (protection and deprotection) useful in synthesizing applicable compounds are known in the art and include, for example, those described in R. Larock, Comprehensive Organic Transformations, VCH Publishers (1989); T.W. Greene and P.G.M. Wuts, Protective Groups in Organic Synthesis, 3rd Ed., John Wiley and Sons (1999); L. Fieser and M. Fieser, Fieser and Fieser's Reagents for Organic Synthesis, John Wiley and Sons (1994); and L. Paquette, ed., Encyclopedia of Reagents for Organic Synthesis, John Wiley and Sons (1995) and subsequent editions thereof.

The invention will now be further illustrated by the following non-limiting examples. The specific examples below are to be construed as merely illustrative, and not limitative of the remainder of the disclosure in any way whatsoever. Without further elaboration, it is believed that one skilled in the art can, based on the description herein, utilize the present invention to its fullest extent. All references and publications cited herein are hereby incorporated by reference in their entirety. GENERAL SYNTHETIC PROCEDURE 1

In general synthetic procedure 1 , phthalic anhydride 1 undergoes condensation- cyc/o-dehydartion to produce phthalimide 2 followed by hydrolysis to form phthalamic acid 3. The regioisomers of phthalamic acid 3 are then separated via preparative HPLC. ⁸

GENEREAL SYNTHETIC PROCEDURE 2

In general, synthetic procedure 2, phthalic anhydride 4 is reacted with te/Y-butanol followed by separation of the regioisomeric mono esters by supercritical fluid chromatography (SFC) to produce phthalate mono-t-butyl ester 5. Phthalate mono-t-butyl ester 5 subsequently undergoes coupling deprotection resulting in phthalamic acid 6. ⁸

Palladium-catalysed coupling could be used to replace the bromine of phthalate mono-t-butyl ester 5 or phthalamic acid 6 with alternative substituents. ⁸

EXAMPLES Phthalamide 6 was degraded under assay conditions (see description of

Neurotensin (NTS) scintillation proximity assay (SPA) below) to afford the two corresponding phthalimide hydrolysis products, 5-bromo-2-[(6-methylpyridin-2- yl)carbamoyl]benzoic acid (Example 1) and 4-bromo-2-[(6-methylpyridin-2- yl)carbamoyl]benzoic acid (Example 2). ⁸ Alternatively, Examples 1 and 2 may be prepared in accordance with general synthetic procedures 1 and/or 2.

Examples 3 to 30

Examples 3 to 30 can be prepared in accordance with general synthetic procedures 1 and/or 2.

Example 31 1-benzyl-3-tert-butyl-1H-pyrazole-5-carboxylic acid

Ethyl 1-benzyl-3-(tert-butyl)-1 H-pyrazole-5-carboxylate (0.1 g, 0.349 mmol) was dissolved in 1 :1 THF/MeOH and heated to 50°C overnight. The reaction mixture was cooled, acidified using 1 N HCI and extracted 3 x EtOAc. The combined organic layers were dried over MgSO ₄ , filtered and concentrated. Purification was performed using RP Gilson HPLC 5-95% 0.01 % TFA-ACN over 20 min to provide the title compound (0.075 g, 83%). 1 H NMR (500 MHz, Chloroform-d) δ 7.30-7.17 (m, 5H),6.83 (s, 1 H), 5.71 (s, 2H), 1.34 (s, 9H). MS m/z: [M+H ]+ Calcd for

C15H18N2O2 258.1 ; Found 260.3. ⁹ Examples 32 to 41

Example 42

(2S)-2-[(3,5-dichlorophenyl)formamido]-5,5-dimethylhexano ic acid

To a solution of 2-amino-5,5-dimethylhexanoic acid (racemic) (100 mg, 0.628 mmol) in tetrahydrofuran (3140 pl) was added 3,5-dichlorobenzoyl chloride (132 mg, 0.628 mmol) and 1 N aqueous NaOH (1884 pl, 1.884 mmol). The resulting mixture was stirred overnight at room temperature. The reaction mixture was then concentrated and injected directly on to a reverse phase prep (5-95%MeCN/H2O TFA) providing 90 mg (43%) of a racemic white solid. The material was subsequently resolved using supercritical CO2 seperation on an IC (2 x 15 cm) column 15% isopropanol (0.1 % DEA)/CO2 at 100 bar 70 mL/min. Chiral separation gave 48 mg of peak A (20.3%) and 49 mg of peak B (19.9%) with peak B as the desired product with > 99% ee. 1 H NMR (500 MHz, Chloroform-d) δ 9.57 (s, 1 H), 7.66 (d, J = 1.7 Hz, 2H), 7.50 (s, 1 H), 6.76 (d, J = 7.7 Hz, 1 H), 4.79 (q, J = 7.3 Hz, 1 H), 2.08 - 1.90 (m, 1 H), 1 .89 - 1 .73 (m, 1 H), 1 .29 (m, 2H), 0.89 (s, 9H). MS m/z: [M+H ]+ Calcd for C15H19CI2NO3 331.07; Found 332.3. ⁹

Example 43

(2S)-2-[(4-chloro-1H-pyrrol-2-yl)formamido]-5,5-dimethylh exanoic acid A vial was charged with 2-amino-5,5-dimethylhexanoic acid (250 mg, 1.570 mmol), 4-chloro-1 H-pyrrole-2-carboxylic acid (286 mg, 1.963 mmol), DMF (5234 pl), DIEA (823 pl, 4.71 mmol), and HATU (746 mg, 1.963 mmol). The resulting mixture stirred was overnight at room temperature after which it was heated for 4 hours at 60°C. The reaction mixture was cooled to room temperature and purified by reverse phase prep (10-95%MeCN/H2O TFA) to provide 99.3 mg (22.1%) of racemic product as a brown solid. The material was subsequently resolved using supercritical CO2 separation on an AD-H (2 x 15 cm) column 15% isopropanol (0.1 % DEA)/CO2 at 100 bar 60 mL/min. Chiral separation gave 43 mg of peak A (6.7%) and 41 mg of peak B (5.4%) with peak B as the desired product with > 99% ee. 1 H NMR (500 MHz, DMSO-d6) δ 7.82 (s, 1 H), 6.93 (s, 1 H), 6.76 (s, 1 H), 4.05 (s, 1 H), 1.74 (m, 1 H), 1.61 (m, 1 H), 1.22 - 1.15 (m, 2H), 0.83 (s, 9H). MS m/z: [M+H ]+ Calcd for C13H19CIN2O3286.1 ; Found 287.4. ⁹

Example 44

(2S)-5,5-dimethyl-2-[(6-phenoxypyridin-3-yl)formamido]hex anoic acid

To a solution of 2-amino-5,5-dimethylhexanoic acid (100 mg, 0.628 mmol) in THF (3.1 mL) was added 6-phenoxynicotinoyl chloride (147 mg, 0.628 mmol) and 1 N NaOH (1884 pl, 1 .884 mmol). The resulting mixture was stirred overnight at room temperature. The reaction mixture was concentrated and purified by reverse phase prep (10-95%MeCN/H2O TFA) to provide 22.5 mg (10.1 %) of racemic product as a brown solid. The material was subsequently resolved using supercritical CO2 separation on an IA (2 x 15 cm) column 12% methanol (0.1% DEA)/CO2 at 100 bar 60 mL/min. Chiral separation gave 5.8 mg of peak A (2.6%) and 5.4 mg of peak B (2.4%) with peak B as the desired product with > 99% ee. 1 H NMR (500 MHz, DMSO-d6) δ 12.62 (s, 1 H), 8.67 (d, J = 7.6 Hz, 1 H), 8.63 (d, J = 2.3 Hz, 1 H), 8.29 (dd, J = 8.6, 2.4 Hz, 1 H), 7.45 (t, J = 7.9 Hz, 2H), 7.25 (t, J = 7.4 Hz, 1 H), 7.17 (d, J = 7.7 Hz, 2H), 7.11 (d, J = 8.5 Hz, 1 H), 4.32 (s, 1 H), 1.75 (dd, J = 32.9, 12.3 Hz, 2H), 1.41 - 1.13 (m, 2H), 0.87 (s, 9H). MS m/z: [M+H ]+ Calcd for C20H24N2O4 356.2; Found 357.5. ⁹ Examples 45 to 64

BIOLOGICAL DATA

Neurotensin scintillation proximity assay

The exemplified compounds 1 to 30 of the invention were tested in a Neurotensin (NTS) scintillation proximity assay (SPA) and the ICso data is shown in the table below. NTS, which is a 13 amino acid neuropeptide, is a sortilin ligand. The IC50 is a measure of the amount of the compound required to inhibit the binding of NTS to sortilin by 50%. The skilled person will recognise that the lower the IC50 value, the less of the compound needed to achieve the desired effect, and as a result, the chances of undesirable off-target effects are reduced.

Compound affinity was determined by measuring the displacement of 3H-neurotensin binding to hSortilin in SPA format. Total volume of 40 pl in 50 mM HEPES pH 7.4 assay buffer containing 100 mM NaCI, 2.0 mM CaCI2, 0.1 % BSA and 0.1% Tween-20. Compound pre-incubation for 30 min at RT with 150 nM of 6his-Sortilin before 5 nM [3/-/]-Neurotensin and Ni chelate imaging beads (Perkin Elmer) were added, after 6 h plate was read on a ViewLux with 360 s exposure time. Dose-response evaluation of compounds was performed with 10 concentrations of drugs (covering 3 decades). IC50 values were calculated by nonlinear regression using the sigmoid concentration-response (variable slope) using Xlfit 4 (I DBS, UK). All values reported are average of at least 4 determinations. ⁸

The data in the table below shows that the compounds disclosed herein are sortilin inhibitors.

* % inhibition at 50 pM.

The exemplified compounds 31 to 64 of the invention were tested in a progranulin (PGRN) peptide/SORT homogenous time-resolved fluorescence (HTRF) assay and the IC50 data shown in the table below. To determine the IC50S of compounds bound to SORT and thus preventing its interaction with the biotinylated 18-mer PGRN peptide a time-resolved fluorescence resonance energy transfer (TR- FRET) format was used. All test compounds were dissolved in DMSO; the final concentration of DMSO in the reaction buffer was 1 %. Compounds were prepared in a ten-point, three-fold serial dilution scheme using an acoustic dispenser. A solution of SORT protein in assay buffer (10 mM HEPES, pH 7.3, 150 mM NaCI, 0.005% Brij-35 and 0.01 % BSA, final concentration of SORT 1 nM) was added to each well and pre-incubated with compounds or DMSO vehicle at ambient temperature for 10 min with gentle shaking. Next, biotinylated 18-mer PGRN peptide was added to each well (final concentration 4 nM) and the binding reactions were incubated at ambient temperature for 30 min with gentle shaking. Solutions of SA/Dylight in assay buffer were added to each well, immediately followed by addition of Eu/anti-His in assay buffer. Reaction mixtures were incubated at ambient temperature for 120 min with gentle shaking, and the TR- FRET signal (the ratio of the 665 nm (Dylight) and 615 nm (Eu) fluorescence emissions using a 320 nm excitation wavelength) of each binding reaction was measured on an Envision plate reader (PerkinElmer) equipped with a LANCE/DELFIA Dual Enhanced mirror, a UV2(TRF) 320 nm excitation filter and APC 665 nm and Europium 615 nm emission filters, using the LANCE Eu/APC Dual 662 factory default protocol. Data were analyzed using ABase and a 4- parameter logistic fit based on the Levenberg-Marquardt algorithm. ⁹

The below ICsodata was obtained as described above for exemplified compounds 1 to 30, with the exception that the neurotensin employed in the SPA assay is

[125l]-Neurotensin.

*Racemate

As can be seen from Figure 1 , the compound according to Example 8 was present in the perilymph of male Hartley guinea-pigs up to the last time point of 72 hours following local dosing via the trans-tympanic route. This was evident at both doses tested (5 or 50 pg/ear; i.e. 0.1 or 1 mg/ml). Figure 2 also demonstrates that the compound according to Example 8 when administered orally (1 mg/mL) was also present in the perilymph of male Hartley guinea-pigs at the two time points tested (8 hours and 24 hours) following three days of daily dosing. The above data demonstrates how the compounds herein disclosed may be desirable for the prevention and/or treatment of hearing loss.

References

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8. T. J. Schroder et al. The identification of AF38469: An orally bioavailable inhibitor of the VPS10P family sorting receptor Sortilin, Bioorganic & Medicinal Chemistry Letters 24 (2014) 177-180.

9. Shawn J. Stachel, et al., Identification of potent inhibitors of the sortilin-progranulin interaction, Bioorganic & Medicinal Chemistry Letters, 30 (2020). Sequences referenced throughout the specification and forming part of the description.