The present invention relates to a composition comprising a compound of formula (I) as defined herein and therapeutic uses thereof such as use in a method of treating cough.

Background of invention

Suplatast tosilate ((±)-3-{[4-(3-ethoxy-2-hydroxypropoxy)phenyl]amino}-3- oxopropyl)(dimethyl)sulfonium; 4-methylbenzenesulfonate) (abbreviated ST herein) is a drug marketed in Japan for oral treatment of atopic dermatitis, asthma and allergy (rhinitis). Suplatast tosilate has been described in the patent literature; see e.g. US 4,670,583 and US 4,556,737.

After oral administration suplatast tosilate is metabolized into M-1 ((±)-4-(3-ethoxy-2 hydroxypropoxy)acrylanilide) as well as metabolites derived from glutathione conjugation thereof e.g. M-2. These were identified in the plasma, urine, and bile.

Cough is an airway defensive reflex facilitating clearance of accumulated secretions and protecting airways and lungs from aspiration, inhaled particulates and irritants. However, when associated with disease, coughing can be a distressing symptom significantly affecting patient's lifestyle and wellbeing The marked decrease in health- related quality of life is responsible for cough being the most common symptom bringing patients to medical attention and indeed is one of the primary causes for patients with as yet undiagnosed IPF (idiopathic pulmonary fibrosis) to seek medical assistance.

Cough can be subdivided into acute cough lasting for less than 3 weeks, sub-acute cough lasting between 3 and 8 weeks and chronic cough lasting for more than 8 weeks. Acute cough is most frequently associated with upper respiratory infection and although usually self-limiting, both prescription and over the counter (OTC) medication are commonly used to treat it with limited success.

Chronic cough is a common symptom of respiratory conditions such as chronic obstructive pulmonary disease (COPD), asthma, upper airways cough syndrome, idiopathic pulmonary fibrosis and some non-respiratory conditions such as gastro oesophageal reflux disease. If the underlying disease is identified and appropriately treated, the cough will often disappear; however, there remains a significant cohort of patients for whom no specific cause of the cough can be identified. Indeed it has been reported that chronic non-productive cough having no identifiable cause can account for up to 40% of patients presenting to special cough clinics. In those patients heightened cough reflex sensitivity is persistent and their condition falls into a category of unexplained (idiopathic) cough, more recently defined as cough hypersensitivity syndrome. Since persistent cough is often debilitating and embarrassing, there is a clear need for an effective antitussive agent.

Current therapies for the management of cough are of limited benefit to many patients, and involve undesirable side effects or dose-limiting toxicities. Thus, there is still a need for improved therapies for the management and treatment of cough, especially those that have the capacity to treat the chronic disease without having adverse effects.

Summary of invention

The inventors have surprisingly found that one particular metabolite of suplatast tosilate, namely M-1 , is effective per se in reducing cough.

It is thus an aspect to provide a composition comprising a compound of formula (I)

wherein A, B and C independently is selected from hydrogen (H) and deuterium (D), wherein R is selected from the group consisting of H, a prodrug moiety, a protecting group and a functional group selected from alkyl, substituted alkyl, alkenyl, alkoxy, alkoxyalkyl, carbonylalkyl, carbonylaminoalkyl, alkoxyphosphonic acid,

alkylcarbonylalkoxy, cycloalkyl, cycloalkenyl, heterocycle, sulfur group, halogen, nitrogen group, -CN, -CH ₂ F, -CF ₃ ,-CF ₂ CF ₃ , and -CF ₂ CF ₂ CF ₃ ,

for use in a method of treating cough. When R is H and A, B and C each are hydrogen the compound is M-1.

In a particular embodiment R is H. In a particular embodiment one, two or three of A, B and C are substituted with deuterium (D).

M-1 has a high clearance rate, leading to a short elimination half-life. To fully benefit from the surprising pharmacological profile offered by M-1 , there is a need to improve the pharmacokinetic properties and/or the stability of the drug.

It is thus a further aspect to provide a deuterated compound of formula (I)

wherein A, B and C independently is selected from hydrogen (H) and deuterium (D), with the proviso that at least one of A, B and C is deuterium,

wherein R is selected from the group consisting of H, a prodrug moiety, a protecting group and a functional group selected from alkyl, substituted alkyl, alkenyl, alkoxy, alkoxyalkyl, carbonylalkyl, carbonylaminoalkyl, alkoxyphosphonic acid,

alkylcarbonylalkoxy, cycloalkyl, cycloalkenyl, heterocycle, sulfur group, halogen, nitrogen group, -CN, -CH ₂ F, -CF ₃ ,-CF ₂ CF ₃ , and -CF ₂ CF ₂ CF ₃ .

In a particular embodiment R is H and each of A, B and C are deuterium (M-1 3D). In one embodiment said compound has an altered pharmacokinetic profile compared to a compound of wherein R, A, B and C each are hydrogen (M-1). Also provided is a deuterated compound as disclosed herein for use in the treatment of cough.

In one embodiment the composition for use comprises, separately or together, one or more additional active pharmaceutical ingredients, such as ingredients used for the treatment of cough. All types of cough are encompassed, including acute cough, sub-acute cough and chronic cough, cough associated with respiratory conditions, and cough associated with non-respiratory conditions.

Also provided is a deuterated compound as disclosed herein for use in the treatment of inflammatory disorders such as allergy.

Definitions

The term "pharmaceutically acceptable derivative" in present context includes pharmaceutically acceptable salts, which indicate a salt which is not harmful to the patient. Such salts include pharmaceutically acceptable basic or acid addition salts as well as pharmaceutically acceptable metal salts, ammonium salts and alkylated ammonium salts. A pharmaceutically acceptable derivative further includes esters and prodrugs, or other precursors of a compound which may be biologically metabolized into the active compound, or crystal forms of a compound.

The term "acid addition salt" is intended to include "pharmaceutically acceptable acid addition salt" which indicates salts which are not harmful to the patient. Acid addition salts include salts of inorganic acids as well as organic acids. Representative examples of suitable inorganic acids include hydrochloric, hydrobromic, hydroiodic, phosphoric, sulfuric, nitric acids and the like. Representative examples of suitable organic acids include formic, acetic, trichloroacetic, trifluoroacetic, propionic, benzoic, cinnamic, citric, fumaric, glycolic, lactic, maleic, malic, malonic, mandelic, oxalic, picric, pyruvic, salicylic, succinic, methanesulfonic, ethanesulfonic, tartaric, ascorbic, pamoic, bismethylene salicylic, ethanedisulfonic, gluconic, citraconic, aspartic, stearic, palmitic, EDTA, glycolic, p-aminobenzoic, glutamic, benzenesulfonic, p-toluenesulfonic acids and the like. Further examples of pharmaceutically acceptable inorganic or organic acid addition salts include the pharmaceutically acceptable salts listed in J. Pharm. Sci. 66, 2, (1977) which is incorporated herein by reference.

The term "therapeutically effective amount" of a compound as used herein refers to an amount sufficient to cure, alleviate, prevent, reduce the risk of, or partially arrest the clinical manifestations of a given disease or disorder and its complications. An amount adequate to accomplish this is defined as "therapeutically effective amount". Effective amounts for each purpose will depend on the severity of the disease or injury as well as the weight and general state of the subject. It will be understood that determining an appropriate dosage may be achieved using routine experimentation, by constructing a matrix of values and testing different points in the matrix, which is all within the ordinary skills of a trained physician or veterinary.

The terms "treatment" and "treating" as used herein refer to the management and care of a patient for the purpose of combating a condition, disease or disorder. The term is intended to include the full spectrum of treatments for a given condition from which the patient is suffering, such as administration of the active compound for the purpose of: alleviating or relieving symptoms or complications; delaying the progression of the condition, disease or disorder; curing or eliminating the condition, disease or disorder; and/or preventing the condition, disease or disorder, wherein "preventing" or

"prevention" is to be understood to refer to the management and care of a patient for the purpose of hindering the development of the condition, disease or disorder, and includes the administration of the active compounds to prevent or reduce the risk of the onset of symptoms or complications. The individual or patient to be treated is preferably a mammal, in particular a human being. Treatment of animals, such as mice, rats, dogs, cats, cows, horses, sheep and pigs, is, however, also within the scope of the present invention. The patients to be treated according to the present invention can be of various ages.

The term "compound" as used herein, refers to a collection of molecules having an identical structure, except that there may be isotopic variation among the constituent atoms of the molecules. Thus, it will be clear to those of skill in the art that a compound represented by a particular chemical structure containing indicated deuterium atoms, will also contain lesser amounts of isotopologues having hydrogen atoms at one or more of the designated deuterium positions in that structure.

The term "isotopologue" refers to a species that differs from specific compounds of this invention only in the isotopic composition thereof.

The term "impure isotopologue" refers to a species that differs from specific

compounds of this invention only in the isotopic composition thereof. It will be recognized that some variations of the natural isotopic abundance occurs in a synthesized compound depending upon the origin of chemical materials used in the synthesis. Thus, a preparation of deuterated compound according to formula I will inherently contain small amounts of impure isotopologues.

The term "isotopic enrichment factor" as used herein means the ratio between the isotopic abundance and the natural abundance of a specific isotope. When a position is designated specifically as "D" or "deuterium", the position is understood to have deuterium at an abundance that is substantially greater than natural abundance of deuterium which is 0.015%. All percentages given for the amount of deuterium present are mole percentages. It is thus understood that pharmaceutical compositions according to the present invention comprise compounds which have isotopic enrichment factors significantly above 1.

The term "composition" or "pharmaceutical composition" as used herein, refers to compositions comprising compounds according to formula (I), which have identical structure, except that there may be isotopic variation among the constituent atoms of the molecules. Thus, it will be clear to those of skill in the art that a compound represented by a particular chemical structure containing indicated deuterium atoms, will also contain lesser amounts of impure isotopologues having hydrogen atoms at one or more of the designated deuterium positions in that structure.

As used herein, the term "prodrug" includes derivatives of compounds such as biohydrolyzable amides and biohydrolyzable esters thereof, compounds in which the biohydrolyzable functionality is encompassed in the compound; and compounds which may be oxidized or reduced biologically at a given functional group to yield drug substances. Examples of the latter type of functional group include 1 ,4-dihydropyridine, N-alkylcarbonyl-1 ,4-dihydropyridine, 1 ,4-cyclohexadiene, tert-butyl and the like.

Description of Drawings

Fig. 1.: Metabolic pathway of Suplatast Tosilat (ST).

Fig. 2.: Synthesis of Compound (III) (M-1).

Fig. 3.: Synthesis of Compound (IV) (M-1 3D).

Detailed description of the invention

Suplatast tosilate ((±)-3-{[4-(3-ethoxy-2-hydroxypropoxy)phenyl]amino}-3- oxopropyl)(dimethyl)sulfonium; 4-methylbenzenesulfonate) (abbreviated ST herein) is a drug marketed in Japan for oral treatment of atopic dermatitis, asthma and allergy (rhinitis). It is characterized by its ability to inhibit Th2 cytokine production and by its high degree of safety. It is approved for treatment of children and has during its 15 years on the market only been associated with very few serious adverse effects. (Suplatast tosilate)

Suplatast tosilate is a racemic mixture. There are no significant differences between the two enantiomers with respect to pharmacology (Tada et al: J. Med. Chem. 1998, 41 , 3330-3336)

Suplatast tosilate was developed as a derivative of S-methylmethionine in the attempt to identify sulfonium compounds with immunological activities with the ultimate goal to find a suitable clinical candidate for the treatment of allergic disorders (Tada et al: J. Med. Chem. 1998, 41 , 3330-3336). The potential therapeutic effects of S- methylmethionine in cytoprotection and wound healing have been described (Kim et al: Pharmacology 2010; 85: 68-76).

Suplatast tosilate has also been found effective in reduction of cough in pre-clinical and clinical investigations:

ST inhibits airway cough hypersensitivity underlying allergic eosinophilic inflammation (Myou et al: Clinical and Experimental Allergy, 2001 ,31 , 1939- 1944).

ST effectively reduce citric acid + enalapril induced cough (Zhou et al:

Pharmacology. 2015; 95(1-2): 36-41).

ST improved cough in a capsaicin challenge in patients with Cough Variant Asthma without significant side effects (Shioya et al: Eur J Clin Pharmacol (2002) 58: 171-176).

The cough threshold measured after four weeks of treatment with ST was significantly increased in Atopic cough (Ishiura et al: Arzneimittel-Forschung (Drug Research) 2008; 58(6):297-302).

ST benefits refractory chronic dry cough following lung cancer surgery

(Miyamoto et al: Gen Thorac Cardiovasc Surg. 2009; 57(9): 463-6). Suplatast tosilate metabolism

The metabolism of suplatast tosilate (ST) is schematized in figure 1 (Kuwata et al: Drug Metabolism and Pharmacokinetics Vol.7, No.4 (1992) pp.399-421 ; Masuda et al: Drug Metabolism and Pharmacokinetics Vol.7, No.4 (1992) pp. 423-439; Shindo et al: Drug Metabolism and Pharmacokinetics Vol.7 , No.4 (1992) pp. 441-459).

After oral suplatast tosilate administration to mice, rats, guinea pig, dog and monkeys, suplatast (base), M-1 ((±)-4-(3-ethoxy-2 hydroxypropoxy)acrylanilide), metabolites derived from glutathione conjugation as well as M-2 were identified in the plasma, urine, and bile.

Some suplatast tosilate is metabolized into M-1 which is in turn subject to rapid metabolism producing a glutathione conjugate (M-1-GSH), followed by formation of M- 1-Cys by deglutamation and deglycination, resulting in a mercapturic acid conjugate (M-1-Ac-Cys) and its S-oxide form (M-1-Ac-Cys(S-0. There is little information available on the pharmacokinetics properties of metabolites of suplatast tosilate.

M-1 metabolite and cough

M-1 is an example of a metabolite from suplatast tosilate metabolism. The inventors have surprisingly found that administration of M-1 per se is effective in reducing cough.

Use of M-1 has the potential to improve pharmacokinetic properties, including faster onset of action, dose frequency and/or duration of action.

Thus it is an aspect of the present invention to provide a composition comprising a compound of formula (I)

wherein A, B and C independently is selected from hydrogen (H) and deuterium (D), and wherein R is selected from the group consisting of H, a prodrug moiety, a protecting group and a functional group selected from alkyl, substituted alkyl, alkenyl, alkoxy, alkoxyalkyl, carbonylalkyl, carbonylaminoalkyl, alkoxyphosphonic acid,

alkylcarbonylalkoxy, cycloalkyl, cycloalkenyl, heterocycle, sulfur group, halogen, nitrogen group, -CN, -CH ₂ F, -CF ₃ ,-CF ₂ CF ₃ , and -CF ₂ CF ₂ CF ₃ ,

for use in a method of treating cough.

As used herein throughout, a compound of formula (I) is meant to encompass any combination of variable positions R, A, B and C, including (but not limited to) compounds (I), (II), (III) and (IV).

As used herein, "prodrug" refers to a derivative of a biologically active compound that may independently have pharmaceutical activity or may lack pharmaceutical activity but is converted to an active agent. A prodrug, according to the present invention, may be converted into an active compound through one or more steps.

In prodrug versions of M-1 , R can be substituted with other groups. In order to increase stability, one or more of the hydrogens at positions A, B and C can be substituted with deuterium.

As used herein, "alkyl" refers to linear, branched or cyclic hydrocarbon structures preferably having from 1 to 20 carbon atoms (a "C1-C20 alkyl") e.g., 1 to 10 carbon atoms or 1 to 6 carbon atoms. This term is exemplified by groups such as methyl, t- butyl, n-heptyl, octyl, cyclobutylmethyl, cyclopropylmethyl and the like. "Unsubstituted alkyl" refers to an alkyl group that is not substituted with any additional substituents. When an alkyl residue having a specific number of carbons is named, all geometric isomers having that number of carbons are intended to be encompassed; thus, for example, "butyl" is meant to include n-butyl, sec-butyl, isobutyl and t-butyl. As used herein, "substituted alkyl" refers to an alkyl group of from 1 to 10 carbon atoms, having from 1 to 5 substituents, including but not limited to, groups such as halogen, alkoxy, acyl, acylamino, acyloxy, amino, hydroxyl, mercapto, carboxyl, aryl, cyano, nitro and the like. For instance, an alkaryl group (alkyl-aryl) is a substituted alkyl and includes moieties such as propylbenzene where the moiety is attached to the parent structure via the aryl or the alkyi portion, most preferably via the alkyi portion of the substituent.

As used herein, "alkenyl" refers to linear, branched or cyclic hydrocarbon structures preferably having from 2 to 20 carbon atoms (a "C1-C20 alkenyl") and more preferably 2 to 10 carbon atoms or 2 to 6 carbon atoms and having at least 1 site of alkenyl unsaturation.

As used herein, "unsubstituted alkenyl" refers to an alkenyl group that is not substituted with any additional substituents. When an alkenyl residue having a specific number of carbons is named, all geometric isomers having that number of carbons are intended to be encompassed. This term is exemplified by groups such as propen-3-yl (— CH2— CH=CH ₂ ), 3-methyl-but-2-enyl and (=CH ₂ ). The group represented by =CH2 indicates connectivity from, e.g., an sp2 hybridized carbon atom of a parent structure to CH ₂ via a double bond.

As used herein, "substituted alkenyl" refers to an alkenyl group, preferably a C2- CiOalkenyl, having from 1 to 5 substituents, including but not limited to, substituents such as halogen, alkoxy, acyl, acylamino, acyloxy, amino, hydroxyl, mercapto, carboxyl, aryl, cyano, nitro and the like.

As used herein, "alkoxy" refers to the group "alkyl-O-" which includes, by way of example, methoxy, ethoxy, n-propoxy, iso-propoxy, n-butoxy, tert-butoxy, sec-butoxy, n-pentoxy, n-hexoxy, 1 ,2- dimethylbutoxy, and the like.

As used herein, "substituted alkoxy" refers to the group "substituted alkyl-O-".

As used herein, "alkoxyalkyl" refers to the group "alkyl-O-alkyl-" which includes, by way of example, methoxy methyl and the like.

As used herein, "alkanoate" refers to "alkyl-C(=0)-0- " which includes, by way of example, ethanoate and pentanoate. "Alkyl-alkanoate" refers to "-alkyl-0-C(=0)alkyl" such as in -CH(CH ₂ CH ₃ )-0-C(=0)-CH ₃ .

As used herein, "carbonylalkyl" refers to -C(=0)-alkyl, which includes, by way of example, C(=0)-CH ₂ CH ₃ .

As used herein "carbonylaminoalkyl" refers to CO(NH)alkyl or CON(alkyl) ₂ . As used herein, "alkoxyphosphonic acid" refers to "alkyl-0-P(=0)(OH) ₂ " or when referred to or implied as a moiety attached to a parent structure, the radical "-alkyl-O- P(=0)(OH) ₂ " such that the alkoxyphosphonic acid is attached to a parent structure via the alkyl moiety. This term is exemplified by groups such as methoxyphosphonic acid and ethoxyphosphonic acid and their radicals -CH2-0-P(=0)(OH) ₂ - CH(CH ₃ )OP(0)(OH) ₂ and -CH ₂ CH ₂ -0-P(=0)(OH) ₂ .

As used herein, "alkylcarbonylalkoxy" refers to alkyl-C(=0)-0-alkyl. In one variation, the alkylcarbonylalkoxy refers to a moiety C1-C4 alkyl-C(=0)-0-C1-C6 alkyl. An exemplary alkylcarbonylalkoxy is -CH ₂ CH ₂ C(=0)OCH ₃ .

As used herein, "C3-C8 monocyclic cycloalkyl" is a 3-, 4-, 5-, 6-, 7- or 8-membered saturated non-aromatic monocyclic cycloalkyl ring. Representative C3-C8 monocyclic cycloalkyl groups include, but are not limited to, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl and cyclooctyl. In one embodiment, the C3-C8 monocyclic cycloalkyl group is substituted with one or more of the following groups: -halo, -0-(C1- C6 alkyl), -OH, -ON, -COOR', -OC(0)R', -N(R) ₂ , -NHC(0)R' or -C(0)NHR' groups wherein each R' is independently -H or unsubstituted -C1-C6 alkyl. Unless indicated, the C3-C8 monocyclic cycloalkyl is unsubstituted. As used herein, "C3-C8 monocyclic cycloalkenyl" is a 3-, 4-, 5-, 6-, 7- or 8-membered non- aromatic monocyclic carbocyclic ring having at least one endocyclic double bond, but which is not aromatic. It is to be understood that when any two groups, together with the carbon atom to which they are attached form a C3-C8 monocyclic cycloalkenyl group, the carbon atom to which the two groups are attached remains tetravalent. Representative C3-C8 monocyclic cycloalkenyl groups include, but are not limited to, cyclopropenyl, cyclobutenyl, 1 ,3-cyclobutadienyl, cyclopentenyl, 1 ,3-cyclopentadienyl, cyclohexenyl, 1 ,3-cyclohexadienyl, cycloheptenyl, 1 ,3-cycloheptadienyl, 1 ,4- cycloheptadienyl, 1 ,3,5- cycloheptatrienyl, cyclooctenyl, 1 ,3-cyclooctadienyl, 1 ,4- cyclooctadienyl, or 1 ,3,5-cyclooctatrienyl. In one embodiment, the C3-C8 monocyclic cycloalkenyl group is substituted with one or more of the following groups: -halo, -O- (C1-C6 alkyl), -OH, -CN, -COOR', -OC(0)R\ -N(R') ₂ , -NHC(0)R or -C(0)NHR' groups wherein each R is independently -H or unsubstituted -C1-C6 alkyl. Unless indicated, the C3-C8 monocyclic cycloalkenyl is unsubstituted. The term "halo" as used herein refers to -F, -CI, -Br or -I.

The term "3- to 7-membered monocyclic heterocycle" refers to: a 3-, 4-, 5-, 6-, or 7- membered aromatic or non-aromatic monocyclic cycloalkyl in which 1-4 of the ring carbon atoms have been independently replaced with an NH, an O, or an S moiety. The non-aromatic 3- to 7-membered monocyclic heterocycles can be attached via a ring nitrogen, sulfur, or carbon atom. The aromatic 3- to 7-membered monocyclic heterocycles are attached via a ring carbon atom. Representative examples of a 3- to 7-membered monocyclic heterocycle group include, but are not limited to furanyl, furazanyl, imidazolidinyl, imidazolinyl, imidazolyl, isothiazolyl, isoxazolyl, morpholinyl, oxadiazolyl, oxazolidinyl, oxazolyl, oxazolidinyl, pyrimidinyl, phenanthridinyl, phenanthrolinyl, piperazinyl, piperidinyl, pyranyl, pyrazinyl, pyrazolidinyl, pyrazolinyl, pyrazolyl, pyridazinyl, pyridooxazole, pyridoimidazole, pyridothiazole, pyridinyl, pyrimidinyl, pyrrolidinyl, pyrrolinyl, quinuclidinyl, tetrahydrofuranyl, thiadiazinyl, thiadiazolyl, thienyl, thienothiazolyl, thienooxazolyl, thienoimidazolyl, thiomorpholinyl, thiophenyl, triazinyl, triazolyl, In one embodiment, the 3- to 7-membered monocyclic heterocycle group is substituted with one or more of the following groups: -halo, -0-(C- 1-C6 alkyl), -OH, -CN, -COOR', -OC(0)R', -N(R') ₂ , -NHC(0)R or -C(0)NHR groups wherein each R' is independently -H or unsubstituted -C1-C6 alkyl. Unless indicated, the 3- to 7-membered monocyclic heterocycle is unsubstituted.

It is also an aspect to provide a method for treating cough in an individual in need thereof, said method comprising administering an effective amount of a composition comprising a compound of formula (I)

wherein A, B and C independently is selected from hydrogen (H) and deuterium (D), and

wherein R is selected from the group consisting of H, a prodrug moiety, a protecting group and a functional group selected from alkyl, substituted alkyl, alkenyl, alkoxy, alkoxyalkyl, carbonylalkyl, carbonylaminoalkyl, alkoxyphosphonic acid,

alkylcarbonylalkoxy, cycloalkyl, cycloalkenyl, heterocycle, sulfur group, halogen, nitrogen group, -CN, -CH ₂ F, -CF ₃ ,-CF ₂ CF ₃ , and -CF ₂ CF ₂ CF ₃ .

Methods for treatment according to the present invention may further comprise one or more steps of administration of one or more additional active ingredients as defined herein.

In a further aspect there is provide the use of a composition comprising a compound of formula (I)

wherein A, B and C independently is selected from hydrogen (H) and deuterium (D), and

wherein R is selected from the group consisting of H, a prodrug moiety, a protecting group and a functional group selected from alkyl, substituted alkyl, alkenyl, alkoxy, alkoxyalkyl, carbonylalkyl, carbonylaminoalkyl, alkoxyphosphonic acid,

alkylcarbonylalkoxy, cycloalkyl, cycloalkenyl, heterocycle, sulfur group, halogen, nitrogen group, -CN, -CH ₂ F, -CF ₃ ,-CF ₂ CF ₃ , and -CF ₂ CF ₂ CF ₃ ,

for the manufacture or preparation of a medicament for the treatment of cough.

A compound of formula (I) may also be referred to as Compound (I) herein, and is meant to make reference also to the definitions of R, A, B and C provided herein. In the 'base' compound; M-1 , R is H and each of A, B and C are H. In prodrug versions of M-1 , R can be substituted with other groups. In order to increase stability, one or more of the hydrogens at positions A, B and C can be substituted with deuterium. As used herein, "prodrug" refers to a derivative of a biologically active compound that may independently have pharmaceutical activity or may lack pharmaceutical activity but is converted to an active agent. A prodrug, according to the present invention, may be converted into an active compound through one or more steps. It follows that any one of positions A, B and C each can be selected from hydrogen (H) and deuterium (D).

In one embodiment A is deuterium and B+C each are hydrogen.

In one embodiment B is deuterium and A+C each are hydrogen.

In one embodiment C is deuterium and A+B each are hydrogen.

In one embodiment A+B each are deuterium and C is hydrogen.

In one embodiment A+C each are deuterium and B is hydrogen.

In one embodiment B+C each are deuterium and A is hydrogen.

In one embodiment A+B+C each are deuterium.

In one embodiment A+B+C each are hydrogen.

In one embodiment the composition according to the present invention comprises a compound wherein R is H and A, B and C independently is selected from hydrogen (H) and deuterium (D) (formula II):

When R is H, the compound is not a prodrug. A compound of formula (II) may also be referred to as Compound (II) herein, and is meant to make reference also to the definitions of A, B and C provided herein.

In another embodiment the composition according to the present invention comprises compound wherein R is H and A, B and C each are hydrogen (M-1 ; formula III)

A compound of formula (III) may also be referred to as Compound (III) herein.

In yet another embodiment the composition according to the present invention comprises a compound wherein R is H and A, B and C each are deuterium (formula IV)

(IV) A compound of formula (IV) may also be referred to as Compound (IV) herein.

In one embodiment the compound is further modified in order to increase stability of the compound. Such increase of stability may be achieved by any means known to the skilled person.

Increased stability may be determined by a number of ways known to the skilled person. This includes determination of the stability of the compounds in aqueous buffer, in presence of microsomes derived from human or non-human liver, during incubations in presence of mammalian cells or human or non-human origin or upon administration to humans or other mammals. In one embodiment the compound of the invention has increased stability and/or decreased rate of conversion (e.g. to M-1-GSH) compared to a compound of formula (I) wherein R, A, B and C each are hydrogen; i.e. increased stability and/or decreased rate of conversion (to e.g. M-1-GSH) compared to the compound of formula (III) (M-1).

It is understood that reference to a compound of the invention, such as defined by formula (I), (II), (III) and (IV) herein, such reference is intended to include all forms of the compound such as amorphous, crystalline, or polymorphs, unless specified otherwise or contradicted by the description. The term is also intended to include isotopes and pro-drugs as well as salts thereof, such as the acid addition salts thereof, e.g. pharmaceutically acceptable acid addition salts.

Cough

It follows that any type of cough with any given aetiology is meant to be encompassed by the present invention.

In one embodiment the present invention provides a composition comprising a compound of formula (I) as defined herein for use in a method of treating cough selected from the group consisting of chronic cough, sub-acute cough and acute cough.

When making reference to a composition comprising a compound of formula (I) as defined herein for use in a method of treating cough associated with a certain condition this is meant to encompass also e.g. cough caused by a certain condition, cough accompanying a certain condition, cough induced by a certain condition.

In another embodiment the present invention provides a composition comprising a compound of formula (I) as defined herein for use in a method of treating cough selected from the group consisting of chronic cough associated with respiratory conditions, chronic cough associated with non-respiratory conditions, cough related respiratory disorder, cough resulting from acute disease, and urge to cough associated with any respiratory disease.

In yet another embodiment the present invention provides a composition comprising a compound of formula (I) as defined herein for use in a method of treating cough selected from the group consisting of idiopathic chronic cough, refractory cough and treatment resistant cough.

In a further embodiment the present invention provides a composition comprising a compound of formula (I) as defined herein for use in a method of treating cough selected from the group consisting of chronic cough variant asthma, cough variant asthma, cough associated with asthma, upper airway cough syndrome (UACS) (also known as post nasal drip cough), cough associated with bronchitis such as chronic bronchitis, cough associated with eosinophilic bronchitis, atopic cough, cough associated with bronchial hyperresponsiveness, cough hypersensitivity syndrome and cough associated with bronchospasm.

In one embodiment the present invention provides a composition comprising a compound of formula (I) as defined herein for use in a method of treating post nasal drip cough caused by one or more from the group consisting of colds, flu, allergies (allergic postnasal drip), sinus infection (sinusitis), objects stuck in the nose, pregnancy, certain medications, deviated septum, changing weather fronts, cold temperatures, excess dryness in the air, certain foods and fumes from chemicals, perfumes, cleaning products, smoke or other irritants.

In one embodiment the present invention provides a composition comprising a compound of formula (I) as defined herein for use in a method of treating cough selected from the group consisting of cough associated with lung cancer, cough associated with chronic obstructive pulmonary disease (COPD), cough associated with interstitial lung disease including idiopathic pulmonary fibrosis (IPF), congestive heart disease and sarcoidosis, cough associated with thoracic tumor and cough associated with gastro esophageal reflux disease (GERD) (both acid and non-acid reflux).

In one embodiment the present invention provides a composition comprising a compound of formula (I) as defined herein for use in a method of treating cough associated with interstitial lung disease including idiopathic pulmonary fibrosis (IPF).

In one embodiment the present invention provides a composition comprising a compound of formula (I) as defined herein for use in a method of treating cough associated with idiopathic pulmonary fibrosis (IPF) (or IPF-cough). In one embodiment the present invention provides a composition comprising a compound of formula (I) as defined herein for use in a method of treating cough selected from the group consisting of viral and post-viral cough, bacterial and post- bacterial cough, cough associated with bacterial infection including whooping cough (pertussis), cough associated with upper respiratory infection, cough resulting from a cold and cough resulting from the flu.

In one embodiment the present invention provides a composition comprising a compound of formula (I) as defined herein for use in a method of treating cough selected from the group consisting of iatrogenic cough and drug-induced cough.

In one embodiment said drug-induced cough is selected from ACE inhibitor-induced cough, ACE-2 inhibitor-induced cough (Angiotensin 2 receptor blockers), beta-blocker- induced cough, NSAID-induced cough and calcium antagonist induced cough.

Pathologic cough, particularly if chronic, can become life-altering for patients, affecting overall quality of life, and ability to maintain relationships or jobs. Social isolation and depression often become significant factors for these patients, as pathologic coughers are ostracized in public venues such as the work place, restaurants, public transport, theaters, etc. Many chronic cough patients cough persistently especially during the active daytime period. Patients with IPF are among those with the most severe cough, the most debilitating symptom of their disease. Currently available antitussive drugs have shown little objective evidence that they are effective for cough in any disorder. Further, safety and abuse liability concerns have restricted use of certain narcotic antitussives. The last new cough therapy to be approved was dextromethorphan nearly 50 years ago. To date, no other agent in development has shown effectiveness in the treatment of pathologic cough, especially using objective measures of cough frequency.

The prevalence of chronic cough is estimated to be over 10% of adults in the U.S. While much of these patients' underlying aetiology (such as GERD, asthma, COPD, etc.) may contribute to cough, 20-40% of these are not responsive to treatment for such underlying aetiology or have no known trigger and many treatment responsive patients are still not well controlled for their cough. Additionally, although pertussis is thought to be significantly underdiagnosed, the incidence of this infection has been increasing due to a less potent and waning effectiveness of the vaccine as well as lower vaccination rates. Pertussis often leads to particularly severe and violent coughing bouts lasting for 3 months or more. Acute cough following an upper respiratory infection, another pathologic cough, is estimated to represent nearly 3 billion cough days annually in the U.S. and significantly impacts patient productivity.

Cough is the symptom for which patients most often seek medical attention, and chronic cough due to any cause affects an estimated 5-18% of the general population. A significant set of these patients (as many as approximately 40% of chronic coughers) is estimated to have cough refractory to treatment of associated conditions, such as gastroesophageal reflux or post-nasal drip. Post nasal drip cough, also referred to as upper airway cough syndrome (UACS), can have a number of causes, including colds, flu, allergies (allergic postnasal drip), sinus infection (sinusitis), objects stuck in the nose, pregnancy, certain medications, deviated septum, changing weather fronts, cold temperatures, excess dryness in the air, certain foods, fumes from chemicals, perfumes, cleaning products, smoke or other irritants,

Individuals with treatment-refractory chronic cough often report being distressed, depressed, angry and/or anxious. Nearly 80% feel cough interferes with social activities. In Afferent's Phase 2 study in chronic cough, patients enrolled were, on average, coughing approximately 40 times per hour and for over 10 years.

The last new cough therapy to be approved was dextromethorphan more than 50 years ago. There is little evidence suggesting that currently available antitussive drugs are effective for cough in any disorder. Further, safety and abuse liability concerns have restricted use of certain antitussives.

"Cough related respiratory disorder" or "- respiratory disease" refers to, without limitation, cough hypersensitivity syndrome, chronic obstructive pulmonary disease (COPD), asthma, bronchospasm, and the like. Respiratory disorders include, for example, sub-acute or chronic cough, treatment resistant cough, idiopathic chronic cough, cough associated with upper respiratory infection, post-viral cough, iatrogenic cough (e.g., as induced by ACE-inhibitors), idiopathic pulmonary fibrosis or cough associated with smoking or a form of bronchitis. Respiratory disorders can include urge to cough associated with any respiratory disease, for example urge to cough associated with chronic obstructive pulmonary disease (COPD), cough-variant asthma, interstitial lung disease, or whooping cough.

"Acute cough" is understood to mean a cough lasting up to two weeks in duration. For instance, acute cough can be the result of an acute disease, such as a cold or flu. An acute cough will disappear when the underlying cause (e.g., cold or flu) is eliminated.

"Sub-acute cough" is understood to mean a cough lasting between two and eight weeks. In some cases, a sub-acute cough follows a period in which a subject is infected with a disease (e.g., cold or flu). A sub-acute cough is one that often remains after the underlying cause has been removed. For instance, a sub-acute cough is found post-infection (e.g., post-viral infection).

"Chronic cough" refers to a persistent or refractory cough lasting longer than eight weeks that may not have an obvious underlying cause and is may not be associated with other respiratory diseases, such as asthma or COPD (i.e., idiopathic). Chronic cough is also characterized in that there are no hallmarks to define and diagnose it, in contrast to other respiratory diseases (e.g., COPD). Another characteristic of chronic cough is that a subject suffering from chronic cough may be apparently normal in most other respects. Chronic cough is characterized by frequent coughing (e.g., at least 5-10 coughs per hour during daytime), and bothersome coughing during sleep. Chronic cough can last for a period of years, including over a decade.

In order to determine if a subject is afflicted by a chronic cough, a practitioner or clinician can perform a three step test. First, the subject can be treated for putative post nasal drips. In some cases, such treatment takes the form of an antihistamine. Second, the subject can be treated with a proton-pump inhibitor (e.g., to treat putative gastroesophageal disease such as reflux disease). Third, a subject can be treated with steroids (e.g., to treat a putative case of asthma).

If a subject continues to display a chronic cough after the above three-step treatment regimen, the cough is said to be chronic cough and is likely refractory. It is understood that patients suffering from refractory cough often have suffered both acute and subacute cough before being diagnosed with chronic cough.

In one embodiment the composition comprising a compound of formula (I) as defined herein is for use in a method of treating cough in an individual or subject in need thereof. The subject is preferably a mammal, such as a human.

In one embodiment the composition comprising a compound of formula (I) as defined herein is for use in a method of treating cough in a non-human mammal. In one embodiment the composition is for veterinary use. In one embodiment the composition comprising a compound of formula (I) as defined herein is for use in a method of treating cough in an animal, such as an animal selected from the group consisting of mice, rats, dogs, cats, cows, horses, sheep, goats and pigs. Use for treating other animals is also encompassed within this invention.

Deuterated M-1 metabolite

It is also an aspect to provide compounds and pharmaceutical compositions comprising novel M-1 analogues wherein one or more protons are substituted with deuterium.

The inventors have surprisingly found that deuterated M-1 wherein one or more protons in specific positions are substituted with deuterium have beneficial properties.

The inventors have surprisingly found that administration of deuterated M-1 per se is effective in reducing cough.

Thus it is also an aspect to provide a deuterated compound of formula (I)

wherein A, B and C independently is selected from hydrogen (H) and deuterium (D), with the proviso that at least one of A, B and C is deuterium, wherein R is selected from the group consisting of H, a prodrug moiety, a protecting group and a functional group selected from alkyl, substituted alkyl, alkenyl, alkoxy, alkoxyalkyl, carbonylalkyl, carbonylaminoalkyl, alkoxyphosphonic acid,

alkylcarbonylalkoxy, cycloalkyl, cycloalkenyl, heterocycle, sulfur group, halogen, nitrogen group, -CN, -CH ₂ F, -CF ₃ ,-CF ₂ CF ₃ , and -CF ₂ CF ₂ CF ₃ .

In this embodiment a compound of formula (I) is meant to encompass any combination of variable positions R, A, B and C, with the proviso that at least one of A, B and C is deuterium.

It follows that any one of positions A, B and C each can be selected from hydrogen (H) and deuterium (D), with the proviso that at least one of A, B and C is deuterium.

A compound of formula (I) may also be referred to as Compound (I) herein, and is meant to make reference also to the definitions of R, A, B and C provided herein.

In one embodiment there is provided a compound of formula (I) wherein R is H.

In one embodiment there is provided a compound of formula (I) wherein A, B and C each are deuterium.

In one embodiment there is provided a compound of formula (I) wherein A is deuterium and B and C each are hydrogen. In one embodiment there is provided a compound of formula (I) wherein B is deuterium and A and C each are hydrogen.

In one embodiment there is provided a compound of formula (I) wherein C is deuterium and A and B each are hydrogen.

In one embodiment there is provided a compound of formula (I) wherein A and B each are deuterium and C is hydrogen.

In one embodiment there is provided a compound of formula (I) wherein A and C each are deuterium and B is hydrogen. In one embodiment there is provided a compound of formula (I) wherein B and C each are deuterium and A is hydrogen. In one embodiment there is provided a deuterated compound wherein R is H and A, B and C independently is selected from hydrogen (H) and deuterium (D), with the proviso that at least one of A, B and C is deuterium (formula II)

A compound of formula (II) may also be referred to as Compound (II) herein, and meant to make reference also to the definitions of A, B and C provided herein.

In another embodiment there is provided a deuterated compound wherein R is H and A, B and C each are deuterium (formula IV)

A compound of formula (IV) may also be referred to as Compound (IV) herein.

It is understood that reference to a compound of the invention, such as defined by formula (I), (II) and (IV) herein, such reference is intended to include all forms of the compound such as amorphous, crystalline, or polymorphs, unless specified otherwise or contradicted by the description. The term is also intended to include isotopes and pro-drugs as well as salts thereof, such as the acid addition salts thereof, e.g.

pharmaceutically acceptable acid addition salts.

It is understood that the present invention also relates to chiral analogues of deuterated compounds, such as enantiomers or optical isomers of the compounds and pharmaceutical compositions of the present invention. Such enantiomers or optical isomers may have different pharmacokinetic properties.

Degree of deuterium labelling

It will be recognized that some variations of the natural isotopic abundance occurs in a synthesized compound depending upon the origin of chemical materials used in the synthesis. Thus, a preparation of any compound will inherently contain small amounts of deuterated isotopologues. The concentration of naturally abundant stable hydrogen and carbon isotopes, notwithstanding this variation, is small and immaterial as compared to the degree of stable isotopic substitution of compounds of this invention. See for instance: Gannes LZ et al. 1998.

In a compound of this invention, when a particular position is designated as including deuterium, it is understood that the abundance of deuterium at that position is substantially greater than the natural abundance of deuterium, which is 0.015%. Thus, in compounds of larger deuterium abundance, the isotopic enrichment factor is significantly above 1. A position designated as having deuterium according to the present invention typically has a minimum isotopic enrichment factor of at least 3000 (45% deuterium incorporated) at each atom designated as deuterium in said compound. Unless otherwise stated, when a position is designated specifically as "H" or "hydrogen", the position is understood to have hydrogen at its natural abundance isotopic composition. In the compounds of the present invention any atom not specifically designated as a particular isotope is meant to represent any stable isotope of that atom.

In one embodiment of the present invention, the composition of the invention comprising the compounds disclosed herein have an isotopic enrichment factor for each designated deuterium atom of at least 3333 (50 % deuterium incorporated at each position designated as a deuterium atom), such as at least 3500 (52.5 % deuterium incorporated at each position designated as a deuterium atom), such as at least 3666 (55% deuterium incorporated at each position designated as a deuterium atom), such as at least 4000 (60% deuterium incorporation at each position designated as a deuterium atom), such as at least 4333 (65% deuterium incorporation at each position designated as a deuterium atom), such as at least 4500 (67.5% deuterium incorporation at each position designated as a deuterium atom), such as at least 4666.6 (70% deuterium incorporation at each position designated as a deuterium atom), such as at least 5000 (75% deuterium incorporation at each position designated as a deuterium atom), such as at least 5333 (80% deuterium incorporation at each position designated as a deuterium atom), such as at least 5500 (82.5% deuterium incorporation at each position designated as a deuterium atom), such as at least 5666 (85% deuterium incorporation at each position designated as a deuterium atom), such as at least 6000 (90% deuterium incorporation at each position designated as a deuterium atom), such as at least 6333.3 (95% deuterium incorporation at each position designated as a deuterium atom), such as at least 6400 (96% deuterium incorporation at each position designated as a deuterium atom), such as at least

6466.7 (97% deuterium incorporation at each position designated as a deuterium atom), such as at least 6533.3 (98% deuterium incorporation at each position designated as a deuterium atom), such as at least 6600 (99% deuterium incorporation at each position designated as a deuterium atom), such as at least 6633 (99.5 % deuterium incorporation at each position designated as a deuterium atom), such as at least 6660 (99.9% deuterium incorporation at each position designated as a deuterium atom).

In a preferred embodiment of the present invention, the isotopic enrichment factor is at least 6000 (90% deuterium incorporation at each position designated as a deuterium atom), such as at least 6333.3 (95% deuterium incorporation at each position designated as a deuterium atom), such as at least 6600 (99% deuterium incorporation at each position designated as a deuterium atom). In another embodiment of the present invention, the composition of the invention comprises the compounds disclosed herein wherein the abundance of hydrogen in the positions designated as deuterium is less than 49.9%, such as less than 45%, such as less than 40%, such as less than 35%, such as less than 30%, such as less than 25%, such as less than 20%, such as less than 15%, such as less than 12,5%, such as less than 10%, such as less than 9%, such as less than 8%, such as less than 7%, such as less than 6%, such as less than 5%, such as less than 4%,, such as less than 3%, such as less than 2%, such as less than 1 %, such as less than 0.5%, such as less than 0.01 %. The composition according to the present invention may also comprise compounds according to formula (I) wherein deuterium is incorporated to a varying degree in any of the positions A, B and C. In one embodiment of the present invention at least 50% of the compounds are isotopically enriched with deuterium, such as in at least 55% of the compounds isotopical iy enriched with deuterium, such as at least 60% of the compounds isotopical iy enriched with deuterium, such as at least 65% of the compounds isotopical iy enriched with deuterium, such as at least 70% of the compounds isotopical iy enriched with deuterium, such as at least 75% of the compounds isotopical iy enriched with deuterium, such as at least 80% of the compounds isotopical iy enriched with deuterium, such as at least 85% of the compounds isotopical iy enriched with deuterium, such as at least 90% of the compounds isotopical iy enriched with deuterium, such as at least 95% of the compounds isotopical iy enriched with deuterium, such as at least 96% of the compounds isotopical iy enriched with deuterium, such as at least 97% of the compounds isotopical iy enriched with deuterium, such as at least 98% of the compounds isotopical iy enriched with deuterium, such as at least 99% of the compounds isotopical iy enriched with deuterium, such as at least 99.5% of the compounds isotopical iy enriched with deuterium, such as at least 99.9% of the compounds isotopical iy enriched with deuterium.

Thus, the present invention relates to compositions comprising novel M-1 analogues for which one or more protons have been substituted with deuterium.

It can be quite difficult in the laboratory to achieve 100% deuteration at any one site of a lab scale amount of compound (e.g., milligram or greater). When 100% deuteration is recited or a deuterium atom is specifically shown in a structure, it is assumed that a small percentage of hydrogen may still be present. Deuterium-enrichment can be achieved by either exchanging protons with deuterium or by synthesizing the molecule with enriched starting materials.

Stability

The new analogues of M-1 as defined herein are deuterated in specific positions and can thus be metabolically stabilized in order to reduce or delay metabolism and change the pattern of metabolism. The deuterated compounds of the present invention can be beneficial by having pharmacological properties comparable to non-deuterated M-1 and additionally improved pharmacokinetic properties compared to the non-deuterated M-1. The improvement of pharmacokinetic properties is obtained because deuterated analogues of M-1 can have reduced formation of metabolites after administration and can therefore be associated with less risk of adverse effects compared to non- deuterated M-1. In particular the compounds of the present invention may be beneficial due to their pharmacokinetic properties, which include a fast onset of action, a long duration of action and increased exposure. Furthermore the compounds of the current invention can be beneficial due to their lower propensity to affect the metabolism of other drugs (drug-drug interaction).

The stability and metabolism of compounds can be measured by assays involving human liver microsomes. When different compounds are compared, the half-life (T ½) and the rate of intrinsic clearance can be computed and used for comparison. An altered, inhibited or delayed metabolism can be observed when the half-life of a deuterated compound is increased compared to non-deuterated compound, or when the rate of intrinsic clearance is decreased for deuterated compound compared to non- deuterated compound.

Increased stability may be determined by a number of ways known to the skilled person. This includes determination of the stability of the compounds in aqueous buffer, in presence of microsomes derived from human or non-human liver, during incubations in presence of mammalian cells or human or non-human origin or upon administration to humans or other mammals. In one embodiment the compound of the invention has increased stability and/or decreased rate of conversion (to M-1-GSH) compared M-1 wherein R, A, B and C each are hydrogen; i.e. increased stability and/or decreased rate of conversion (to M-1-GSH) compared to M-1. In one embodiment the rate of intrinsic clearance (Clint) is decreased for the deuterated compound compared to non-deuterated compound.

In one embodiment the compound of the invention has an altered pharmacokinetic profile compared to a compound of wherein R, A, B and C each are hydrogen (M-1). In one embodiment the rate of intrinsic clearance of the deuterated compound measured by incubation with human liver microsomes is reduced.

In one embodiment the compound of the invention the plasma protein binding of the deuterated compound is reduced compared to a compound of wherein R, A, B and C each are hydrogen (M-1).

In one embodiment the compound of the invention is further modified in order to increase stability of the compound. Such increase of stability may be achieved by any means known to the skilled person.

Deuterated M-1 as a medicament

It is also an aspect of the present invention to provide a composition comprising a deuterated compound of formula (I)

wherein A, B and C independently is selected from hydrogen (H) and deuterium (D), with the proviso that at least one of A, B and C is deuterium,

wherein R is selected from the group consisting of H, a prodrug moiety, a protecting group and a functional group selected from alkyl, substituted alkyl, alkenyl, alkoxy, alkoxyalkyl, carbonylalkyl, carbonylaminoalkyl, alkoxyphosphonic acid,

alkylcarbonylalkoxy, cycloalkyl, cycloalkenyl, heterocycle, sulfur group, halogen, nitrogen group, -CN, -CH ₂ F, -CF ₃ ,-CF ₂ CF ₃ , and -CF ₂ CF ₂ CF ₃ ,

for use as a medicament and/or for use in medicine. It is a further aspect to provide a deuterated compound as defined herein for use in the treatment of an inflammatory disorder.

Provided herein is a composition comprising a deuterated compound of formula (I)

wherein A, B and C independently is selected from hydrogen (H) and deuterium (D), with the proviso that at least one of A, B and C is deuterium,

wherein R is selected from the group consisting of H, a prodrug moiety, a protecting group and a functional group selected from alkyl, substituted alkyl, alkenyl, alkoxy, alkoxyalkyl, carbonylalkyl, carbonylaminoalkyl, alkoxyphosphonic acid,

alkylcarbonylalkoxy, cycloalkyl, cycloalkenyl, heterocycle, sulfur group, halogen, nitrogen group, -CN, -CH ₂ F, -CF ₃ ,-CF ₂ CF ₃ , and -CF ₂ CF ₂ CF ₃ ,

for use in the treatment of an inflammatory disorder.

It is also an aspect to provide a method for treating an inflammatory disorder in an individual in need thereof, said method comprising administering an effective amount of a composition comprising a deuterated compound of formula (I)

wherein A, B and C independently is selected from hydrogen (H) and deuterium (D), with the proviso that at least one of A, B and C is deuterium,

wherein R is selected from the group consisting of H, a prodrug moiety, a protecting group and a functional group selected from alkyl, substituted alkyl, alkenyl, alkoxy, alkoxyalkyl, carbonylalkyl, carbonylaminoalkyl, alkoxyphosphonic acid,

alkylcarbonylalkoxy, cycloalkyl, cycloalkenyl, heterocycle, sulfur group, halogen, nitrogen group, -CN, -CH ₂ F, -CF ₃ ,-CF ₂ CF ₃ , and -CF ₂ CF ₂ CF ₃ .

Methods for treatment according to the present invention may further comprise one or more steps of administration of one or more additional active ingredients as defined herein. In a further aspect there is provided the use of a composition comprising a deuterated compound of formula (I)

wherein A, B and C independently is selected from hydrogen (H) and deuterium (D), with the proviso that at least one of A, B and C is deuterium,

wherein R is selected from the group consisting of H, a prodrug moiety, a protecting group and a functional group selected from alkyl, substituted alkyl, alkenyl, alkoxy, alkoxyalkyl, carbonylalkyl, carbonylaminoalkyl, alkoxyphosphonic acid,

alkylcarbonylalkoxy, cycloalkyl, cycloalkenyl, heterocycle, sulfur group, halogen, nitrogen group, -CN, -CH ₂ F, -CF ₃ ,-CF ₂ CF ₃ , and -CF ₂ CF ₂ CF ₃ ,

for the manufacture or preparation of a medicament for the treatment of an

inflammatory disorder.

It follows that any type of an inflammatory disorder with any given aetiology is meant to be encompassed by the present invention.

In one embodiment the inflammatory disorder is atopic dermatitis.

In one embodiment the inflammatory disorder is asthma

In one embodiment the inflammatory disorder is allergy (rhinitis)

In a particular embodiment the deuterated compounds of the present invention are used for treatment of various types of skin disorders or conditions, such as atopic dermatitis, seborrhoeic dermatitis, diaper dermatitis, allergic contact dermatitis, irritant contact dermatitis, unspecified contact dermatitis, infective dermatitis, exfoliative dermatitis, lichen simplex chronicus, lichen planus, pruritus/itch, pityriasis rosea, rosacea, psoriasis, urticaria (allergic and unspecified), erythema, sunburn, pemphigus and other acantholytic disorders. In one embodiment the inflammatory disorder is selected from the group consisting of interstitial cystitis, cutaneous mastocytosis, bronchitis and COPD.

In one embodiment the inflammatory disorder is a disease associated with increased levels of IgE and/or Th2 vs Th1 cells; in one embodiment the inflammatory disorder is selected from the group consisting of Kimura's disease, angiolymphoid hyperplasia and eosinophilic vasculitis.

Also provided herein is a composition comprising a deuterated compound of formula (I)

wherein A, B and C independently is selected from hydrogen (H) and deuterium (D), with the proviso that at least one of A, B and C is deuterium,

wherein R is selected from the group consisting of H, a prodrug moiety, a protecting group and a functional group selected from alkyl, substituted alkyl, alkenyl, alkoxy, alkoxyalkyl, carbonylalkyl, carbonylaminoalkyl, alkoxyphosphonic acid,

alkylcarbonylalkoxy, cycloalkyl, cycloalkenyl, heterocycle, sulfur group, halogen, nitrogen group, -CN, -CH ₂ F, -CF ₃ ,-CF ₂ CF ₃ , and -CF ₂ CF ₂ CF ₃ ,

for use in the treatment of a disorder or disease selected from the group consisting of diabetic neuropathy; micturition disorders including dysuria, pollakiuria, miction pain, and urinary incontinence; idiopathic pulmonary fibrosis and idiopathic hypereosinophilic syndrome.

Combination therapies

The compounds or compositions of the present invention may be combined with or comprise one or more additional active ingredients which are understood as other therapeutically effective compounds or pharmaceutically acceptable derivatives thereof. In a further aspect there is provided a composition comprising, separately or together, i) a compound of formula (I)

wherein A, B and C independently is selected from hydrogen (H) and deuterium (D), and

wherein R is selected from the group consisting of H, a prodrug moiety, a protecting group and a functional group selected from alkyl, substituted alkyl, alkenyl, alkoxy, alkoxyalkyl, carbonylalkyl, carbonylaminoalkyl, alkoxyphosphonic acid,

alkylcarbonylalkoxy, cycloalkyl, cycloalkenyl, heterocycle, sulfur group, halogen, nitrogen group, -CN, -CH ₂ F, -CF ₃ ,-CF ₂ CF ₃ , and -CF ₂ CF ₂ CF ₃ ,

and ii) one or more additional active pharmaceutical ingredients,

for use in a method of treating cough. In yet another aspect there is provided a composition comprising, separately or together, i) a deuterated compound of formula (I)

wherein A, B and C independently is selected from hydrogen (H) and deuterium (D) with the proviso that at least one of A, B and C is deuterium (formula II),

wherein R is selected from the group consisting of H, a prodrug moiety, a protecting groups and a functional group selected from alkyl, substituted alkyl, alkenyl, alkoxy, alkoxyalkyl, carbonylalkyl, carbonylaminoalkyl, alkoxyphosphonic acid,

alkylcarbonylalkoxy, cycloalkyl, cycloalkenyl, heterocycle, sulfur group, halogen, nitrogen group, -CN, -CH ₂ F, -CF ₃ ,-CF ₂ CF ₃ , and -CF ₂ CF ₂ CF ₃ ,

and ii) one or more additional active pharmaceutical ingredients. In one embodiment said one or more additional active pharmaceutical ingredients comprise ingredients used for the treatment of cough (anti-tussives). Anti-tussives encompassed by the present invention include Expectorants (help thin mucus, making it easier to cough up, e.g. comprising guaifenesin), Suppressants (help cut the number of times you cough, often comprising dextromethorphan, other cough suppressants include camphor, eucalyptus oil, and menthol) and Combination cough products having more than one active ingredient (e.g. comprising for example both guaifenesin and dextromethorphan). Cough medicines may also contain ingredients to help coat and soothe the throat. Combination products may have medicines to ease other symptoms, that may include decongestants for stuffy nose, antihistamines for allergies or a runny nose, or painkillers. In one embodiment said one or more additional active pharmaceutical ingredients used for the treatment of cough (anti-tussives) is selected from the group consisting of codein, aspirin, dextromethorphan, guaifenesin, camphor, eucalyptus oil, menthol, oxymetazoline, levodropropizine, noscapine, theobromine, benzonatate, triprolidine, pseudoephedrine, chlorpheniramine, chlorpheniramine- hydrocodone- pseudoephedrine, hydrocodone and chlorpheniramine, colistimethate injection, homatropine, and potential new types of compounds (P2X3 antagonist, TRPV1 , TRPA1 , CB-2 and NK-1).

In another embodiment said one or more additional active pharmaceutical ingredients comprise ingredients used for the treatment of the underlying cause of the cough.

In one embodiment the combination of additional medicaments has a dose-sparing effect of lowering the required dosage of the medication used in combination with the compound of the present invention.

In one embodiment of the present invention, the composition comprising a compound of formula (I) as defined herein, and the additional active ingredient, are administered simultaneously, sequentially or separately. In one embodiment of the present invention, the composition comprising a compound of formula (I) as defined herein is administered before the additional active ingredient. In another embodiment, the composition comprising a compound of formula (I) as defined herein is administered simultaneously with the additional active ingredient. In yet another embodiment, the composition comprising a compound of formula (I) as defined herein is administered after the additional active ingredient.

Routes of administration

It will be appreciated that the preferred route of administration will depend on the general condition and age of the subject to be treated, the nature of the condition to be treated, the location of the tissue to be treated in the body and the active ingredient chosen.

In one embodiment of the present invention, the composition comprising a compound of formula (I) as defined herein is administered by systemic administration, local administration, enteral administration or parenteral administration.

Appropriate dosage forms for such administration may be prepared by conventional techniques.

Systemic treatment

Systemic treatment according to the present invention is capable of introducing the agent into the blood stream to ultimately target the sites of desired action. Such routes of administration are any suitable routes, such as an enteral route, the oral, rectal, nasal, pulmonary, buccal, sublingual, transdermal, intracisternal, intraperitoneal, and parenteral (including subcutaneous, intramuscular, intrathecal, intravenous and intradermal) route. Oral administration

Oral administration is normally for enteral drug delivery, wherein the agent is delivered through the enteral mucosa. Syrups and solid oral dosage forms are commonly used. Parenteral administration

Parenteral administration is any administration route not being the oral/enteral route whereby the medicament avoids first-pass degradation in the liver. Accordingly, parenteral administration includes any injections and infusions, for example bolus injection or continuous infusion, such as intravenous administration, intramuscular administration, subcutaneous administration. Furthermore, parenteral administration includes inhalations and topical administration.

Accordingly, the agent may be administered topically to cross any mucosal membrane of an animal to which the biologically active substance is to be given, e.g. in the nose, vagina, eye, mouth, genital tract, lungs, gastrointestinal tract, or rectum, preferably the mucosa of the nose, or mouth, and accordingly, parenteral administration may also include buccal, sublingual, nasal, rectal, vaginal and intraperitoneal administration as well as pulmonal and bronchial administration by inhalation or installation. Also, the agent may be administered topically to cross the skin.

The subcutaneous and intramuscular forms of parenteral administration are generally preferred. Local treatment

The agent according to the invention may be used as a local treatment, i.e. be introduced directly to the site(s) of action. Accordingly, the agent may be applied to the skin or mucosa directly, or the agent may be injected into the site of action, for example into the diseased tissue or to an end artery leading directly to the diseased tissue.

Dosage

According to the present invention, the composition comprising a compound of formula (I) is administered to individuals in need of treatment in pharmaceutically effective doses. A therapeutically effective amount of a compound according to the present invention is an amount sufficient to cure, prevent, reduce the risk of, alleviate or partially arrest the clinical manifestations of a given disease and its complications. The amount that is effective for a particular therapeutic purpose will depend on the severity and the sort of the disease as well as on the weight and general state of the subject. The compounds of the present invention may be administered one or several times per day, such as from 1 to 8 times per day, such as from 1 to 6 times per day, such as from 1 to 5 times per day, such as from 1 to 4 times per day, such as from 1 to 3 times per day, such as from 1 to 2 times per day, such as from 2 to 4 times per day, such as from

2 to 3 times per day. Alternatively, the compounds may be administered less than once a day, for example once a day, such as once every second day, for example once every third day, such as once every fourth day, for example once every fifth day, such as once every sixth day, for example once every week.

In one embodiment the composition comprising a compound of formula (I) as defined herein is administered in a therapeutically effective amount, such as in an amount of 1 mg to 1000 mg compound of formula (I) (calculated as the free base) per day.

It follows that in one embodiment a compound according to the present invention is administered in an amount of 1 mg to 5 mg per day, such as 5 mg to 10 mg, 10 mg to 20 mg, 20 mg to 30 mg, 30 mg to 40 mg, 40 mg to 60 mg, 60 mg to 70 mg, 70 mg to 80 mg, 80 mg to 90 mg, 90 mg to 100 mg, 100 mg to 125 mg, 125 mg to 150 mg, 150 mg to 175 mg, 175 mg to 200 mg, 200 mg to 225 mg, 225 mg to 250 mg, 250 mg to 275 mg, 275 mg to 300 mg, 300 mg to 325 mg, 325 mg to 350 mg, 350 mg to 400 mg, 400 mg to 450 mg, or 450 mg to 500 mg, 500 mg to 600 mg, 600 mg to 700 mg, 700 mg to 800 mg, 800 mg to 900 mg, for example 900 mg to 1000 mg per day.

In another embodiment a compound according to the present invention is administered in an amount of 0.01 mg/kg bodyweight to 20 mg/ kg bodyweight, such as 0.01 mg/ kg bodyweight to 0.05 mg/ kg bodyweight, 0.05 to 0.1 mg/ kg bodyweight, 0.1 to 0.5 mg/ kg bodyweight, 0.5 mg to 1 mg/ kg bodyweight, 1 to 2 mg/ kg bodyweight, 2 to 3 mg/ kg bodyweight, 3 to 5 mg/ kg bodyweight, 5 to 10 mg/ kg bodyweight, for example 10 to 15 mg/ kg bodyweight.

Formulation

In one embodiment the composition comprising a compound of formula (I) as defined herein is a pharmaceutical composition, such as a pharmaceutically safe composition. According to the invention, the composition comprising a compound of formula (I) as defined herein may be administered in any suitable way e.g. orally, sublingually, or parenterally, and it may be presented in any suitable form for such administration, e.g. in the form of tablets, capsules, powders, syrups, solutions, implant or dispersions for injection. Typically, the composition comprising a compound of formula (I) as defined herein is administered as an oral dose form. In another embodiment the composition comprising a compound of formula (I) as defined herein is administered as an injectable dose form.

Preferably, and in accordance with the purpose of the present invention, the composition comprising a compound of formula (I) as defined herein is administered in the form of a solid pharmaceutical entity, suitably as a tablet or a capsule or in the form of a suspension, solution or implant or dispersion for injection.

Methods for the preparation of solid pharmaceutical preparations are well known in the art.

Tablets may thus be prepared by mixing the active ingredients with ordinary adjuvants and/or diluents and subsequently compressing the mixture in a convenient tabletting machine. Examples of adjuvants or diluents comprise: corn starch, lactose, talcum, magnesium stearate, gelatine, lactose, gums, and the like. Any other adjuvant or additive such as colourings, aroma, preservatives, etc. may also be used provided that they are compatible with the active ingredients. The compound (I) as used in accordance with the invention as the free base or the salt thereof may be administered alone or in combination with pharmaceutically acceptable carriers or excipients, in either single or multiple doses. The pharmaceutical compositions according to the invention may be formulated with pharmaceutically acceptable carriers or diluents as well as any other known adjuvants and excipients in accordance with conventional techniques such as those disclosed in Remington: The Science and Practice of Pharmacy, 19 Edition, Gennaro, Ed., Mack Publishing Co., Easton, Pa., 1995. Typically, the composition comprising a compound of formula (I) as defined herein is a solid oral dose form, such as tablets or capsules, or a liquid oral dose form. Examples

The present invention is further illustrated by the following examples which, however, are not to be construed as limiting the scope of protection. The features disclosed in the foregoing description and in the following examples may, both separately and in any combination thereof, be material for realizing the invention in diverse forms thereof.

Example 1

Preparation of Compound (III); M-1

The compounds of the invention can be prepared by different procedures. One example is provided by: Kizawa et al: Biol. Pharm. Bull. 28(6) 1061—1065 (2005).

(+/-)-4-(3-ethoxy-2-hydroxypropoxy) acrylanilide was synthesized as follows: 0.2 g of ST and 0.1 g of Na2C03 were dissolved in distilled water and heated at 50 °C for 45 min. After addition of 1 ml of 1 M HCI and 5 ml of extract solvent, dichloromethane : ethyl acetate (1 : 5), mixture was shaken vigorously and then centrifuged to obtain the organic phase. The aqueous phase was washed with 3 ml of the extract solvent twice. The organic phase was gathered into another tube and then evaporated to dryness under a dry nitrogen stream at 45 °C to obtain a residue of white powder. M-1 was confirmed by Μη peak at 265 by mass spectroscopy. Purity of M-1 was ^L 95% as measured by reverse phase HPLC with octadecylsilyl-column at 254 nm detection.

Another example is given below (see Fig. 2).

General procedure for preparation of compound intermediate 3

To a mixture of Compound intermediate 1 (10 g, 72.0 mmol) and Compound intermediate 2 (7.3 g, 72.0 mmol), was added pyridine (5.7 g, 72.0 mmol) in one portion at 15°C under N ₂ . The mixture was stirred at 15°C for 10 min. Then heated to 90°C and stirred for 15 hours. TLC (PE: EA=3: 1 , Rf=0.4) showed the reaction was completed. The mixture was cooled to 15°C. The residue was poured into HCI (1 N , 100 mL). The aqueous phase was extracted with EA (100 ml_*2). The combined organic phase was washed with NaHCO ₃ (100 ml_), dried with anhydrous Na ₂ S0 ₄ , filtered and

concentrated in vacuum to afford Compound intermediate 3 (10.0 g, 41.5 mmol, 57% yield) as brown solid.

General procedure for preparation of compound intermediate 4

3

To a mixture of Compound intermediate 3 (2.0 g, 8.3 mmol) and imidazole (1.1 g, 16.6 mmol) in THF (20 ml_), was added TBSCI (1.9 g, 12.4 mmol) in one portion at 0°C under N ₂ . The mixture was stirred at 15°C for 15 hours. TLC (PE:EA=3:1 , Rf=0.8) showed the reaction was completed. The aqueous phase was extracted with EA (100 ml_ x3). The combined organic phase was washed with saturated brine (50 ml_ x2), dried with anhydrous Na ₂ S0 ₄ , filtered and concentrated in vacuum. The residue was purified by silica gel chromatography (100-200 mesh silica gel, PE/EA=20/1 , 10/1) to afford Compound intermediate 4 (1.8 g, 5.1 mmol, 61 % yield) as yellow oil.

General procedure for preparation of compound intermediate 6

To a solution of Compound intermediate 4 (1.8 g, 5.0 mmol) in EtOH (25 ml_) was added Pd/C (10%, 0.5 g) under N ₂ . The suspension was degassed under vacuum and purged with H ₂ several times. The mixture was stirred under H ₂ (50 psi) at 15°C for 8 hours. TLC (PE:EA=3: 1 , Rf=0.3) showed the starting material was consumed completely. The reaction mixture was filtered and the filtrate was concentrated. The crude product was purified by silica gel chromatography eluted with (PE: EA=10: 1) to give Compound intermediate 6 (1.5 g, 4.6 mmol, 91 % yield) as brown solid. eneral procedure for preparation of compound intermediate 7

6

To a mixture of Compound intermediate 6 (1.5 g, 4.6 mmol) and Compound intermediate 5 (500 mg, 5.5 mmol) in DCM (20 mL), was added Et ₃ N (700 mg, 6.9 mmol) in one portion at 15°C under N ₂ . The mixture was stirred at 0°C for 10 min. Then stirred at 15°C and stirred for 3 hours. TLC (PE:EA=3:1 , Rf=0.5) showed the reaction was completed. The mixture was extracted with DCM (40 mL x3). The combined organic phase was washed with saturated brine (20 mL x2), dried with anhydrous Na ₂ S0 ₄ , filtered and concentrated in vacuum. The residue was purified by silica gel chromatography (PE/EA=20/1-10/1) to afford Compound intermediate 7 (1.5 g, 4.0 mmol, 85% yield) as white solid. LCMS: EW138-9-P1 M

General procedure for preparation of Compound (III) (M-1)

Cpd 1 , 253 mg

To a mixture of Compound intermediate 7 (1.2 g, 3.2 mmol), TBAF (1.2 g, 4.7 mmol) in MeOH (20 mL) was stirred at 15°C for 5 hours. TLC (PE:EA=1 :1 , Rf=0.3) showed the starting material was consumed completely. The reaction mixture was poured into H ₂ 0 (30 mL). The mixture was extracted with EA (3 x25 mL). The organic phase was washed with saturated brine (30 mL), dried over anhydrous Na ₂ S0 ₄ , concentrated in vacuum to give a residue. The residue was purified by prep-HPLC to afford Compound intermediate 7 (253.0 mg, 0.9 mmol, 30% yield) as white solid.

¹ H NMR: EW138-012-P1A, 400 MHz, CDCI ₃

δ 7.52-7.50 (d, J = 8.8 Hz, 2H), 7.19 (s, 1 H), 6.93-6.91 (d, J = 8.8 Hz, 2H), 6.47-6.42 (d, J = 17.2 Hz, 1 H), 6.28-6.22 (m, 1 H), 5.80-5.78 (d, J = 10 Hz, 1 H), 4.21-4.17 (m, 1 H), 4.05-4.03 (m, 2H), 3.64-3.58 (m, 4H), 1.27-1.23 (t, J = 6.8 Hz, 3H). Preparation of Compound (IV); M-1 3D

Synthesis of Compound (IV) (M-1 with deuterium at positions A, B and C); see Fig. 3

General procedure for preparation of compound intermediate 3

To a mixture of Compound intermediate 1 (10 g, 72.0 mmol) and Compound intermediate 2 (7.3 g, 72.0 mmol), was added pyridine (5.7 g, 72.0 mmol) in one portion at 15°C under N2. The mixture was stirred at 15°C for 10 min. Then heated to 90°C and stirred for 15 hours. TLC (PE:EA=3: 1 , Rf=0.4) showed the reaction was completed. The mixture was cooled to 15°C. The residue was poured into HCI (1 N, 100 ml_). The aqueous phase was extracted with EA (100 ml_x2). The combined organic phase was washed with NaHC0 ₃ (100 ml_), dried with anhydrous Na ₂ S0 ₄ , filtered and

concentrated in vacuum to afford Compound intermediate 3 (10.0 g, 41.5 mmol, 57% yield) as brown solid.

General procedure for preparation of compound intermediate 4

3

To a mixture of Compound intermediate 3 (2.0 g, 8.3 mmol) and imidazole (1.1 g, 16.6 mmol) in THF (20 ml_), was added TBSCI (1.9 g, 12.4 mmol) in one portion at

0°C under N ₂ . The mixture was stirred at 15°C for 15 hours. TLC (PE:EA=3: 1 , Rf=0.8) showed the reaction was completed. The aqueous phase was extracted with EA (100 ml_x3). The combined organic phase was washed with saturated brine (50 ml_x2), dried with anhydrous Na ₂ S0 ₄ , filtered and concentrated in vacuum. The residue was purified by silica gel chromatography (100-200 mesh silica gel, PE/EA=20/1 , 10/1) to afford Compound intermediate 4 (1.8 g, 5.1 mmol, 61 % yield) as yellow oil. eneral procedure for preparation of compound intermediate 6

To a solution of Compound intermediate 4 (1.8 g, 5.0 mmol) in EtOH (25 mL) was added Pd/C (10%, 0.5 g) under N2. The suspension was degassed under vacuum and purged with H ₂ several times. The mixture was stirred under H ₂ (50 psi) at 15°C for 8 hours. TLC (PE:EA=3: 1 , Rf=0.3) showed the starting material was consumed completely. The reaction mixture was filtered and the filtrate was concentrated. The crude product was purified by silica gel chromatography eluted with (PE: EA=10: 1) to give Compound intermediate 6 (1.5 g, 4.6 mmol, 91 % yield) as brown solid.

General procedure for preparation of compound intermediate 7

DCM solution used to next ste

To a solution of Compound intermediate 5 (200.00 mg, 2.66 mmol, 1.00 Eq, the deuterium staring material was commercially available in DCM (8 mL) was added a solution of (COCI) ₂ (405.16 mg, 3.19 mmol, 1.20 Eq) drop-wise at OoC under N ₂ . The reaction mixture was stirred at 15°C for 1 hour. TLC (added MeOH) showed the starting material was consumed completely. The DCM solution was added drop-wise into Compound intermediate 6 (1.3 g, 3.99 mmol, 1.50 Eq) and TEA (2 eq) in DCM (20 mL) at 0°C under N ₂ . The reaction mixture was stirred at 15°C for 2 hours. LCMS showed the starting material was consumed completely. The reaction was concentrated in vacuo at 30°C. The solution was washed with DCM (50 ml_x3) and brine (30 ml_x2), dried over anhydrous Na ₂ S0 ₄ , filtered and concentrated in vacuo. The residue was purified by HPLC to get Compound intermediate 7 (650.00 mg, 1.70 mmol, 64% yield) as yellow oil. LCMS: EW138-17-P1 M

General procedure for preparation of Compound (IV) (M-1 3D)

Cpd 1

To a mixture of Compound intermediate 7 (800.00 mg, 2.09 mmol, 1.00 Eq) in THF (10 ml_) was added TBAF (819.68 mg, 3.14 mmol, 1.50 Eq). The mixture was heated to 50°C for 5 hours. TLC (PE:EA=1 : 1) showed the starting material was consumed completely. The reaction mixture was concentrated. The residue was purified by column chromatography and prep-HPLC (twice) and prep-TLC to get Compound (IV) (209 mg, 780.00 umol, 37% yield) as a white solid.

¹ H NMR: EW138-20-P1 E, 400 MHz, CDCI3

δ 7.52-7.50 (d, J = 8.8 Hz, 2H), 7.15 (s, 1 H), 6.94-6.91 (d, J = 9.2 Hz, 2H), 4.18-4.05 (m, 1 H), 4.04-4.03 (m, 2H), 3.63-3.58 (m, 4H), 2.56-2.55 (d, J = 4.8 Hz, 1 H), 1.27-1.23 (t, J = 6.8 Hz, 3H).

Example 2

Evaluation of compounds in treatment of cough

The method is described in detail by Zhou et al: Pharmacology. 2015; 95(1-2): 36-41. In brief:

Male guinea pigs 300-350 g bodyweight (Harlan MD, distributed by INNOVO Ltd., Hungary) were used. The animals were housed in open cages in a temperature- controlled and ventilated environment (21-23°C) with a 12-hour light-dark cycle. Water and standard ascorbic acid containing guinea pig chow were provided ad libitum. The protocol has been approved by the Hungarian Food Chain Safety and Animal Health Directorate (PEI/001/3083-6/2014) and was carried out in accordance with European Directive 86/609/EEC. The enalapril maleate was dissolved in sterile saline (Salsol infusion, Teva) at 1.3 mg/mL concentration and was applied subcutaneously at 1.3 mg/kg dose (1 mL/kg b.d.w. volume from the 1.3 mg/mL solution) 30 min before vehicle, suplatast tosilat or Compound (IV) (M-1 3D) administration.

The vehicle for both suplatast tosilat and a Compound of formula (IV) (M-1 with deuterium at positions A, B and C) was 10% PEG400/saline mixture. Suplatast tosilat was well soluble in this mixture while Compound (IV) composed a suspension at 30 mg/mL concentration. Both compounds were administered intraperitoneally at 30 mg/kg dose (1 ml/kg b.d.w. volume from the 30 mg/mL solution/suspension) 30 min before citric acid challenge.

Three experimental groups were involved in this study: (1) vehicle, (2) suplatast tosilat and (3) Compound (IV) treated groups. N=6 guinea pigs were involved in each group.

The guinea pigs were exposed to 10% citric acid spray 30 min after the test item administration. The 10% citric acid spray was generated by a compressed air nebulizer (DeVilbiss Pulmoaid) connected to a transparent chamber (6.7 I volume). Compressed air with a flow of 0.16 l/s and pressure of 0.5 bar produced the spray. The citric acid vapour penetrated into the chamber through a short tube driven by constant air flow. The animals were put into a transparent chamber individually and exposed to the citric acid (10 w/v % citric acid dissolved in distilled water) aerosol for 3 minutes. The number of coughs during the 3 min inhalation period and the subsequent 5 min was counted. The coughing of the animals was defined as a strong contraction of abdomen which was followed by forced expiration through the opened mouth of the animals. The sound of the cough was monitored by a microphone connected to a loudspeaker placed into the cough chamber. A trained technician monitored the exposed animals and counted the cough numbers.

The average cough numbers in each group was expressed as mean ± S.E.M. The percentage antitussive effect was also calculated. Cough number in the treated group

Percentage antitussive effect = ¹ " " X 100

Cough number in the control group

The animals in the vehicle group produced 1 1.2 ± 2.6 coughs during the whole 8-min observational period. The cough number in the suplatast tosilat-treated group was lower (7.2 ± 2.8) than that of vehicle group (1 1.2 ± 2.6) indicating the antitussive effect of suplatast tosilat.

Compound IV-treated animals produced lower number (4.8 ± 0.8) of cough during the whole observational period than that of vehicle (1 1.2 ± 2.6) group. Compound (IV) produced 56.7% antitussive effect. This antitussive effect of Compound (IV) was statistically significant (p<0.05). For statistical evaluation ANOVA followed by Dunnett's test was used (GraphPad Prism software).

Example 3

Pharmacological methods

Metabolic stability of the Compounds of the present invention can be evaluated by procedures know to persons skilled in the art. Several examples of such procedures can be found in Methling et al: Drug Metabolism and Disposition (2009), 37:479-493. Pharmacological methods for determining pharmacokinetic properties

A person skilled in the art will recognize that commercially available apparatus mentioned as part of the methods below are interchangeable with comparative apparatus for similar purposes obtained through other producers or vendors. Degradation by CYP450 isoenzymes

Incubation with recombinant human CYP isoforms (e.g. CYP2D6 and CYP3A4) is performed according to Suzuki et al. 1999, in brief:

The basic incubation medium contained 10 pmol/ml, 15 mM MgCI ₂ , 1.3 mM NADP, 3.3 mM glucose 6-phosphate, 0.4 I.U./ml glucose 6-phosphate dehydrogenase, 100 mM potassium phosphate buffer (pH 7.4), 0.1 mM EDTA, and 1 μΜ compound of the present invention, e.g .Compound (III) or Compound (IV), in a final volume of 200 ml. The mixture is incubated at 37°C in a shaking water bath for 60 min. The reaction is terminated by addition of 10 ml of perchloric acid and 50 ml of a methanolic solution of the internal standard. After termination of the incubation, the mixtures are centrifuged at 10,000 rpm for 1 min, and the supernatants are analyzed by H PLC- MS/MS. Stability in rat liver microsomes: Incubations of compounds or pharmacological compositions with Rat Liver Microsomes and S9

Microsomes from non-induced and from dexamethasone-, rifampicin-, phenobarbital-, and β-naphthoflavone-induced rat livers are used (Walter et al. 2003). The NADPH- regenerating system and NADPH negative controls are incubated for 5 min at 37 °C in open tubes. Next, the required volume of the substrates is added, and the solutions mixed and divided into 2-ml reaction vials. The reactions are started by the addition of the microsomal suspension to give a final concentration of 2 mg/ml protein (2-8 mg/ml protein for S9). The final volume of each incubation mixture is 1 ml containing 20 μΜ substrate, 0.5 mM NADP ⁺ , 5 mM glucose 6-phosphate, 10 mM MgCI2 ^■ 6H20, 5 mM EDTA, and 3.5 lU/ml glucose-6- phosphate dehydrogenase. Incubations are continued at 37 °C. At the appropriate times (0, 15, and 30 min) 180 μΙ is removed and added to 180 μΙ of ice-cold acetonitrile. After mixing, the samples are placed on ice for 30 min to facilitate protein precipitation. Finally, the samples are centrifuged at 14,000g at 0°C for 10 min. The centrifuged samples are stored at -32 °C. Immediately before HPLC analysis the samples are mixed and centrifuged once more. The supernatant are analyzed by quantitative HPLC analysis method.

HPLC/HRMS Analysis

All chromatographic separations for HRMS measurements and the isolation of metabolites are done with a HPLC system (e.g. Agilent 1 100) consisting of a quaternary gradient pump, an autosampler, and a solvent degasser. The column is connected to the BNMI-HP unit for beam splitting (20: 1) followed by the Bruker diode array UV detector (Bruker BioSpin GmbH, Rheinstetten, Germany) in parallel with the micrOTOF mass spectrometer (Bruker Daltonics, Bremen, Germany). The micrOTOF mass spectrometer is equipped with an electrospray ion source (temperature 180°C). Mass spectra are acquired with a scan range from 50 to 1500 m/z. All measurements are done in the positive mode. For all separations, a 125 x 4 mm LiChrospher 100 RP- 18e (5 μηι) column (Merck, Darmstadt, Germany) preceded by a pre-column of the same material are used. The flow rate is 0.5 ml/min. The chromatography is performed at 23 ± 2°C. Detection is done at λ = 204, 247, and 319 nm (maxima of absorption) and 362 nm (minimum of absorption) for analytes. Metabolite fractions for MS/MS analysis with an API 4000 mass spectrometer is collected manually. Eluents used in the gradients are acetonitrile (solvent B) and 50 mM ammonium acetate adjusted to pH 7.5 with 2.5% ammonia (solvent D). Solvent gradients for all chromatographic separations run from 10 to 100% solvent B in 25 min, with the shapes of the gradients optimized for separations. These methods are used in the analysis of incubations of compounds of the present invention in the presence of microsomes with UDP-GA or in the presence of HRP and H ₂ 0 ₂ with GSH. HPLC/MS/MS Analysis

The MS/MS analysis of metabolites of compounds of the invention can be done using the following procedure. The equipment can consist of an Agilent gradient pump 1 100, a column oven, an autosampler, and a linear ion trap quadrupole mass spectrometer 3200 Q TRAP (Applied Biosystems/MDS Sciex, Foster City, CA). The source type is Turbo Spray with a source temperature of 450°C. For all measurements the positive mode is used. A Phenomenex Synergi Hydro RP column, 150 x 2 mm (4 μηι), is used for the chromatography with a flow rate of 0.3 ml/min. Separations are performed using 95% A and 5% B for 30 s as a gradient, followed by a linear increase to 100% B over 15.5 min and then by 2 min of 100% B. Afterward the column is reconstituted to the starting conditions over 7 min. Solvent A used in the gradient is 5 mM ammonium acetate and solvent B is methanol containing 5 mM ammonium acetate. The column is heated to 30 °C.

MS/MS analysis

MS/MS analyses metabolites of compounds of the present invention also could be done with an API 4000 mass spectrometer (AB/MDS Sciex). Purified metabolite fractions are analyzed by flow injection analysis by using a solvent flow of acetonitrile- 50 mM ammonium acetate buffer (pH = 7.5) (solvent ratios resulting from the further separations) at flow rates of 10 and 20 μΙ/min, respectively. The mass spectrometer is equipped with an electrospray ion source (temperature 300°C). Collision-induced dissociation (CID) spectra are acquired for all metabolites with nitrogen as the collision gas. Collision energies used are in a range between 20 and 65 eV. Incubations of compounds with Human Liver Microsomes and S9

Incubations are done under the same conditions as described for rat liver microsomes. Modifications are as follows: final protein concentration is 4 mg/ml for S9; final volume of each incubation mixture is 320 μΙ; and sample volume 80 μΙ.

Stability in human hepatocytes

In vitro stability in the presence of hepatocytes is conducted as follows. Fresh or cryopreserved hepatocytes are thawed if necessary, isolated from shipping media and diluted to a viable cell density of 2 x 106 cells/mL according to the supplier's guidelines using Krebs-Henseleit buffer (KHB, pH 7.3, Sigma) supplemented with amikacin (84 μg/mL), calcium chloride (1 mM), gentamicin (84 μg/mL), HEPES (20 mM), heptanoic acid (4.2 μΜ) and sodium bicarbonate (2.2 mg/mL). Viability is determined by trypan blue exclusion using a hemacytometer (3500 Hausser, VWR). A 10-mM DMSO stock solution of each drug is diluted to 2 μΜ using supplemented KHB buffer to create the working standard. A 50-μΙ_ aliquot of test compound or control are added to each test well of a 96-well polypropylene plate (Costar) immediately followed by the addition of 50 μΙ_ of the hepatocyte suspension. One incubation plate is prepared for each timepoint (i.e., 0, 30, 60, and 120 minutes) with samples being prepared in duplicate. For these determinations, experiments are conducted in triplicate. Incubations are conducted at 37 °C, 5% C02 and 100% relative humidity in an incubator (Model 2300, VWR). At each timepoint, one incubation plate is removed from the incubator, and a solution containing internal standard (100 μΙ_, 2 μΜ labetalol) is added to each well. The plate is mixed at 700 rpm for 2 minutes on a plate shaker (IKA MTS 2/4 Digital Microtiter Shaker, VWR) and immediately centrifuged at 2,000 x g for 10 minutes using an Allegra benchtop centrifuge (Beckman Coulter). A 150-μΙ_ aliquot of the supernatant is transferred from each well to a 96-well shallow plate (Costar). The plates are sealed using reusable plate mats. Quantitation is performed using an ion trap LC-MS/MS method (Finnigan). Chromatographic separation is achieved using a YMC ODS AQ C18 column (2.1 x 30 mm, 3 μηι, 180 A) in conjunction with a 6-minute gradient using mobile phases A (aqueous 0.1 % formic acid containing 1 % isopropanol) and B (0.1 % formic acid in acetonitrile containing 1 % isopropanol). Mass spectrometric detection of the analytes is accomplished using ESI+ or APCI+ ionization modes. Analyte responses are measured using extracted ion chromatograms of characteristic fragments from the [M+H] ⁺ ion. Calculations are performed using Excel (Microsoft). Pharmacokinetic and bioavailability analysis of compounds following oral and intravenous administration to rats

Three male Sprague-Dawley rats (200-250 g each) are cannulated in the jugular vein and administered a single dose containing 2 mg/kg each of the compounds of the present invention. Three additional male Sprague-Dawley rats (200-250 g each) are administered a single dose containing 2mg/kg of the compounds of the present invention by oral gavage.

Blood (0.25ml) from intravenously treated rats is collected retro-orbitally at 2, 5, 15, and 30 minutes, and 1 , 2, 4, and 6 hours post-dosing. Blood (0.25ml) from orally treated rats is collected retro-orbitally at 5, 15, 30, and 45 minutes and 1 , 2, 4, and 6 hours post-dosing. Blood is collected into tubes containing K ₂ EDTA as coagulant at the above mentioned time points. Blood samples are stored on ice and then centrifuged to obtain plasma. The plasma (about 0.125 μΙ) is aliquoted into 96-well plats and stored at -80 °C until analysis by LC-MS/MS for example using an Applied Bio-system API 4000 mass spectrometer.

Metabolic stability of compounds of the present invention can also be evaluated by other procedures know to persons skilled in the art. Several examples of such procedures can be found in e.g. Konsoula et al: Int J Pharm. 2008 September 1 ; 361 (1- 2): 19-25 and Methling et al. 2009.

Stability in human plasma

The plasma is diluted to 80% with 0.05 M PBS (pH 7.4) at 37°C. The reactions are initiated by the addition of the compounds to 1 ml of preheated plasma solution to yield a final concentration of 200 μΜ. The assays are performed in a shaking water bath at 37°C and conducted in triplicate. Samples (50 μΙ) were taken at 0, 15, 30, 45, 60, 90 min and added to 200 μΙ acetonitrile in order to deproteinize the plasma. The samples are subjected to vortex mixing for 1 min and then centrifugation at 4°C for 15 min at 14,000 rpm. The clear supernatants are analyzed by HPLC. The values represent the mean of three independent experiments. The in vitro plasma half-life (t1/2) is calculated using the expression t _{1 2} =0.693/b, where b is the slope found in the linear fit of the natural logarithm of the fraction remaining of the parent compound vs. incubation time. References

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