WO2012089606A1

WO2012089606A1 - Azabicyclo [4.1.0] hept - 4 - yl derivatives as human orexin receptor antagonists

Info

Publication number: WO2012089606A1
Application number: PCT/EP2011/073736
Authority: WO
Inventors: Romano Di Fabio
Original assignee: Glaxo Group Limited
Priority date: 2010-12-28
Filing date: 2011-12-22
Publication date: 2012-07-05

Abstract

This invention relates to azabicyclo[4.1.0]hept-4-yl derivatives and their use as pharmaceuticals.

Description

AZABICYCLO [4.1.0] HEPT - 4 - YL DERIVATIVES AS HUMAN OREXIN RECEPTOR ANTAGONISTS

Many medically significant biological processes are mediated by proteins participating in signal transduction pathways that involve G-proteins and/or second messengers.

Polypeptides and polynucleotides encoding the human 7-transmembrane G-protein coupled neuropeptide receptor, orexin-1 (HFGAN72), have been identified and are disclosed in EP875565, EP875566 and WO 96/34877. Polypeptides and polynucleotides encoding a second human orexin receptor, orexin-2 (HFGANP), have been identified and are disclosed in EP893498.

Polypeptides and polynucleotides encoding polypeptides which are ligands for the orexin-1 receptor, e.g. orexin-A (Lig72A) are disclosed in EP849361.

The orexin ligand and receptor system has been well characterised since its discovery (see for example Sakurai, T. et al (1998) Cell, 92 pp 573 to 585; Smart et al (1999) British Journal of Pharmacology 128 pp 1 to 3; Willie et al (2001) Ann. Rev.

Neurosciences 24 pp 429 to 458; Sakurai (2007) Nature Reviews Neuroscience 8 pp 171 to 181; Ohno and Sakurai (2008) Front. Neuroendocrinology 29 pp 70 to 87). From these studies it has become clear that orexins and orexin receptors play a number of important physiological roles in mammals and open up the possibility of the development of new therapeutic treatments for a variety of diseases and disorders as described hereinbelow.

Experiments have shown that central administration of the ligand orexin-A stimulated food intake in freely-feeding rats during a 4 hour time period. This increase was approximately four-fold over control rats receiving vehicle. These data suggest that orexin- A may be an endogenous regulator of appetite (Sakurai, T. et al (1998) Cell, 92 pp 573 to 585; Peyron et al (1998) J. Neurosciences 18 pp 9996 to 10015; Willie et al (2001) Ann. Rev. Neurosciences 24 pp 429 to 458). Therefore, antagonists of the orexin-A receptor(s) may be useful in the treatment of obesity and diabetes. In support of this it has been shown that orexin receptor antagonist SB334867 potently reduced hedonic eating in rats (White et al (2005) Peptides 26 pp 2231 to 2238) and also attenuated high-fat pellet self- administration in rats (Nair et al (2008) British Journal of Pharmacology, published online 28 January 2008).

The search for new therapies to treat obesity and other eating disorders is an important challenge. According to WHO definitions a mean of 35% of subjects in 39 studies were overweight and a further 22% clinically obese in westernised societies. It has been estimated that 5.7% of all healthcare costs in the USA are a consequence of obesity. About 85%) of Type 2 diabetics are obese. Diet and exercise are of value in all diabetics. The incidence of diagnosed diabetes in westernised countries is typically 5% and there are estimated to be an equal number undiagnosed. The incidence of obesity and Type 2 diabetes is rising, demonstrating the inadequacy of current treatments which may be either ineffective or have toxicity risks including cardiovascular effects. Treatment of diabetes with sulfonylureas or insulin can cause hypoglycaemia, whilst metformin causes GI side- effects. No drug treatment for Type 2 diabetes has been shown to reduce the long-term complications of the disease. Insulin sensitisers will be useful for many diabetics, however they do not have an anti-obesity effect.

As well as having a role in food intake, the orexin system is also involved in sleep and wakefulness. Rat sleep/EEG studies have shown that central administration of orexin- A, an agonist of the orexin receptors, causes a dose-related increase in arousal, largely at the expense of a reduction in paradoxical sleep and slow wave sleep 2, when administered at the onset of the normal sleep period (Hagan et al (1999) Proc.Natl.Acad.Sci. 96 pp 10911 to 10916). The role of the orexin system in sleep and wakefulness is now well established (Sakurai (2007) Nature Reviews Neuroscience 8 pp 171 to 181 ; Ohno and Sakurai (2008) Front. Neuroendocrinology 29 pp 70 to 87; Chemelli et al (1999) Cell 98 pp 437 to 451; Lee et al (2005) J. Neuroscience 25 pp 6716 to 6720; Piper et al (2000) European J

Neuroscience 12 pp 726-730 and Smart and Jerman (2002) Pharmacology and Therapeutics 94 pp 51 to 61). Antagonists of the orexin receptors may therefore be useful in the treatment of sleep disorders including insomnia. Studies with orexin receptor antagonists, for example SB334867, in rats (see for example Smith et al (2003) Neuroscience Letters 341 pp 256 to 258) and more recently dogs and humans (Brisbare-Roch et al (2007) Nature Medicine 13(2) pp 150 to 155) further support this.

In addition, recent studies have suggested a role for orexin antagonists in the treatment of motivational disorders, such as disorders related to reward seeking behaviours for example drug addiction and substance abuse (Borgland et al (2006) Neuron 49(4) pp 589-601; Boutrel et al (2005) Proc.Natl.Acad.Sci. 102(52) pp 19168 to 19173; Harris et al (2005) Nature 437 pp 556 to 559).

International Patent Applications WO99/09024, W099/58533, WO00/47577 and WO00/47580 disclose phenyl urea derivatives and WO00/47576 discloses quinolinyl cinnamide derivatives as orexin receptor antagonists. WO05/118548 discloses substituted 1,2,3,4-tetrahydroisoquinoline derivatives as orexin antagonists.

WO01/96302, WO02/44172, WO02/89800, WO03/002559, WO03/002561, WO03/032991, WO03/037847, WO03/041711, WO08/038251, WO09/003993,

WO09/003997 and WO09/124956 all disclose cyclic amine derivatives.

WO08/038251 discloses 3-aza-bicyclo[3.1.0]hexane derivatives as orexin antagonists. We have now found that certain azabicyclo[4.1.0]hept-4-yl derivatives have beneficial properties including, for example, high potency, good brain penetration and good bioavailability. Such properties make these azabicyclo[4.1.0]hept-4-yl derivatives very attractive as potential pharmaceutical agents which may be useful in the prevention or treatment of obesity, including obesity observed in Type 2 (non-insulin-dependent) diabetes patients, sleep disorders, anxiety, depression, schizophrenia, drug dependency or compulsive behaviour. Additionally these compounds may be useful in the treatment of stroke, particularly ischemic or haemorrhagic stroke, and/or blocking the emetic response, i.e. useful in the treatment of nausea and vomiting.

Accordingly the present invention provides a compound of formula (I)

(I)

wherein:

Het is a heteroaryl group selected from thiazolyl and thiadiazolyl said heteroaryl group being optionally substituted with 1, 2 or 3 substituents independently selected from the group consisting of: Ci_-4alkyl, halo, Ci_-4alkoxy, haloCi_-4alkyl, haloCi_-4alkoxy and cyano; X is C or N;

Y is C or N;

Ri is Ci_-4alkyl, halo, Ci_-4alkoxy, haloCi_-4alkyl, haloCi_-4alkoxy, cyano, phenyl or a 5 or 6 membered heterocyclyl group containing 1, 2 or 3 atoms selected from N, O or S, which phenyl or heterocyclyl group is optionally substituted with Ci_-4alkyl, halo, Ci_-4alkoxy, haloCi_-4alkyl, haloCi_-4alkoxy or cyano;

R-2 is Ci_-4alkyl, halo, Ci_-4alkoxy, haloCi_-4alkyl, haloCi_-4alkoxy, cyano, phenyl or a 5 or 6 membered heterocyclyl group containing 1, 2 or 3 atoms selected from N, O or S, which phenyl or heterocyclyl group is optionally substituted with Ci_-4alkyl, halo, Ci_-4alkoxy, haloCi_-4alkyl, haloCi_-4alkoxy or cyano;

R₃ is Ci_-4alkyl, halo, Ci_-4alkoxy, haloCi_-4alkyl, haloCi_-4alkoxy or cyano;

m is 0 or 1; and

n is 0 or 1;

or a pharmaceutically acceptable salt thereof.

In one embodiment the stereogenic centres of the compounds of formula (I) are in a cis (IS, 4S,6S)- configuration.

In one embodiment the stereogenic centres of the compounds of formula (I) are in a trans (\R,4S,6R)- configuration.

In one embodiment Het is thiazolyl.

In one embodiment Het is thiadiazolyl.

In one embodiment Het is substituted with trifluorom ethyl.

In one embodiment X and Y are both carbon.

In one embodiment X and Y are both nitrogen.

In one embodiment X is nitrogen and Y is carbon.

In one embodiment X is carbon and Y is nitrogen.

In one embodiment Ri is triazolyl.

In one embodiment Ri is pyrazolyl.

In one embodiment Ri is pyrimidinyl.

In one embodiment m is 1 and n is 0.

In one embodiment m is 1, n is 0 and R₂ is methyl. In one embodiment the invention provides the compound of formula (I) selected from the group consisting of:

N-[((lR,4^6R)-3-{[6-methyl-3-(2H-l,2,3-triazol-2-yl)-2-pyridinyl]carbonyl}-3- azabicyclo[4.1.0]hept-4-yl)methyl]-5-(trifluoromethyl)-l,3,4-thiadiazol-2-amine;

N-[((lR,4S,6R)-3-{[6-methyl-3-(lH-pyrazol-l-yl)-2-pyridinyl]carbonyl}-3- azabicyclo[4.1.0]hept-4-yl)methyl]-5-(trifluoromethyl)-l ,3,4-thiadiazol-2-amine;

N-[((lR,4^6R)-3-{[5-methyl-2-(2H-l,2,3-triazol-2-yl)phenyl]carbonyl}-3- azabicyclo[4.1.0]hept-4-yl)methyl]-5-(trifluoromethyl)-l,3,4-thiadiazol-2-amine;

N-[((lR,4S,6R)-3-{[6-methyl-3-(lH-l,2,3-triazol-l-yl)-2-pyridinyl]carbonyl}-3- azabicyclo[4.1.0]hept-4-yl)methyl]-5-(trifluoromethyl)-l,3,4-thiadiazol-2-amine;

N-[((lR,4^6R)-3-{[2-(2H-l,2,3-triazol-2-yl)phenyl]carbonyl}-3-azabicyclo[4.1.0]hept-4- yl)methyl]-5-(trifluorom ethyl)- 1 ,3 ,4-thiadiazol-2-amine;

N-[((lR,4^,6R)-3-{[6-methyl-3-(2-pyrimidinyl)-2-pyridinyl]carbonyl}-3- azabicyclo[4.1.0]hept-4-yl)methyl]-5-(trifluoromethyl)-l,3-thiazol-2-amine;

N-[((lR,4^6R)-3-{[6-methyl-3-(2-pyrimidinyl)-2-pyridinyl]carbonyl}-3- azabicyclo[4.1.0]hept-4-yl)methyl]-5-(trifluoromethyl)-l ,3,4-thiadiazol-2-amine;

N-[((lR,4^,6R)-3-{[6-methyl-3-(2-pyrimidinyl)-2-pyridinyl]carbonyl}-3- azabicyclo[4.1.0]hept-4-yl)methyl]-4-(trifluoromethyl)-l,3-thiazol-2-amine; and

N-[((l,S',45',65)-3-{[6-methyl-3-(2-pyrimidinyl)-2-pyridinyl]carbonyl}-3- azabicyclo[4.1.0]hept-4-yl)methyl]-5-(trifluoromethyl)-l,3,4-thiadiazol-2-amine, or a pharmaceutically acceptable salt thereof.

The Het group may be attached to the aminomethyl linker by means of a bond between the nitrogen atom in said linker and any carbon or nitrogen atom in said Het group.. Preferably the Het group is attached to the linker by means of a bond between the nitrogen atom in the linker and a carbon atom in the Het group ring.

When Ri or R₂ is a heterocyclic group it can be any 5 or 6 membered heterocyclyl group containing 1, 2 or 3 atoms selected from N, O or S. Examples of such heterocyclic groups include pyrimidinyl, oxadiazolyl, oxazolyl, isoxazolyl, thiazolyl, triazolyl, imidazolyl, pyrazolinyl, pyridazinyl, pyrazinyl, pyridinyl, furanyl, thienyl, pyrrolyl, thiadiazolyl, triazinyl and isothiazolyl.

When Ri or R₂ is a heterocyclic group, said group may be attached to the X/Y containing ring by means of a bond between a carbon atom of said X/Y containing ring and a carbon or a heteroatom of the heterocyclic group. For example where Ri is a triazolyl group the attachment to the X/Y containing ring may be by means of a bond between a carbon atom on the X/Y containing ring and a) one of the two carbon atoms or b) one of the three nitrogen atoms of the triazolyl group.

When the compound contains a Ci_-4alkyl group, whether alone or forming part of a larger group, e.g. Ci_-4alkoxy, the alkyl group may be straight chain, branched or cyclic, or combinations thereof. Examples of Ci_-4alkyl are methyl or ethyl.

Examples of haloCi_-4alkyl include trifluoromethyl (i.e. -CF₃).

Examples of Ci_-4alkoxy include methoxy and ethoxy.

Examples of haloCi_-4alkoxy include trifluoromethoxy (i.e. - OCF₃). Halogen or "halo" (when used, for example, in

means fluoro, chloro, bromo or iodo.

It is to be understood that the present invention covers all combinations of particularised groups and substituents described herein above.

It will be appreciated that for use in medicine the salts of the compounds of formula

(I) should be pharmaceutically acceptable. Suitable pharmaceutically acceptable salts will be apparent to those skilled in the art. Pharmaceutically acceptable salts include those described by Berge, Bighley and Monkhouse J.Pharm.Sci (1977) 66, pp 1-19. Such pharmaceutically acceptable salts include acid addition salts formed with inorganic acids e.g. hydrochloric, hydrobromic, sulphuric, nitric or phosphoric acid and organic acids e.g. succinic, maleic, acetic, fumaric, citric, tartaric, benzoic, p-toluenesulfonic, methanesulfonic or naphthalenesulfonic acid. Other salts e.g. oxalates or formates, may be used, for example in the isolation of compounds of formula (I) and represent another aspect of this invention.

Certain of the compounds of formula (I) may form acid addition salts with one or more equivalents of the acid. The present invention includes within its scope all possible stoichiometric and non-stoichiometric forms.

The compounds of formula (I) may be prepared in crystalline or non-crystalline form and, if crystalline, may optionally be solvated, eg. as the hydrate. This invention includes within its scope stoichiometric solvates (eg. hydrates) as well as compounds containing variable amounts of solvent (eg. water).

As used herein "pharmaceutically acceptable derivative" includes any

pharmaceutically acceptable ester or salt of such ester of a compound of formula (I) which, upon administration to the recipient is capable of providing (directly or indirectly) a compound of formula (I) or an active metabolite or residue thereof.

The stereogenic centres of the compounds of formula (I) are in a cis (IS, 4S,6S)- or a a trans (\R,4S,6R)- configuration. The invention also extends to any tautomeric forms or mixtures thereof.

The subject invention also includes isotopically-labeled compounds which are identical to those recited in formula (I) but for the fact that one or more atoms are replaced by an atom having an atomic mass or mass number different from the atomic mass or mass number most commonly found in nature. Examples of isotopes that can be incorporated into compounds of the invention include isotopes of hydrogen, carbon, nitrogen, oxygen, fluorine, iodine and chlorine such as ³H, ^UC, ¹⁴C, ¹⁸F, ¹²³I or ¹²⁵I.

Compounds of the present invention and pharmaceutically acceptable salts of said compounds that contain the aforementioned isotopes and/or other isotopes of other atoms are within the scope of the present invention. Isotopically labeled compounds of the present invention, for example those into which radioactive isotopes such as ³H or ¹⁴C have been incorporated, are useful in drug and/or substrate tissue distribution assays. Tritiated, ie. ³H, and carbon-14, ie. ¹⁴C, isotopes are particularly preferred for their ease of preparation and detectability. ^UC and ¹⁸F isotopes are particularly useful in PET (positron emission tomography).

Since the compounds of formula (I) are intended for use in pharmaceutical compositions it will readily be understood that they are each preferably provided in substantially pure form, for example at least 60% pure, more suitably at least 75% pure and preferably at least 85%>, especially at least 98%> pure (% are on a weight for weight basis). Impure preparations of the compounds may be used for preparing the more pure forms used in the pharmaceutical compositions.

According to a further aspect of the present invention there is provided a process for the preparation of compounds of formula (I) and derivatives thereof. The following schemes detail some synthetic routes to compounds of the invention. In the following schemes reactive groups can be protected with protecting groups and deprotected according to well established techniques.

Schemes

According to a further aspect of the invention there is provided a process for the preparation of compounds of formula (I) or salts thereof. The following 4 schemes are examples of synthetic schemes that may be used to synthesise the compounds of the invention. In the schemes Het has the meaning given in formula (I), and the group Ar₁ denotes the 6 membered X/Y containing ring

wherein X, Y, Ri, R₂, R₃, m and n all have the meanings given in formula (I):

Scheme 1

Acetonitrile

i. Toluene, HCI cat. ii. Heptane

5

Scheme 2

Ts

 Scheme 4

toluene DCM

NaN0₂

DCM/ buffer solution pH=5 ate dimer

It will be understood by those skilled in the art that certain compounds of the invention can be converted into other compounds of the invention according to standard chemical methods.

The starting materials for use in the scheme are commercially available, known in the literature or can be prepared by known methods.

Pharmaceutically acceptable salts may be prepared conventionally by reaction with the appropriate acid or acid derivative.

The present invention provides compounds of formula (I) or a pharmaceutically acceptable salt thereof for use in human or veterinary medicine.

The compounds of formula (I) or their pharmaceutically acceptable salts may be of use for the treatment or prophylaxis of a disease or disorder where an antagonist of a human orexin receptor is required.

Coumpounds of formula (I) or their pharmaceutically acceptable salts may be of use for the treatment or prophylaxis of sleep disorders selected from the group consisting of Dyssomnias such as Primary Insomnia (307.42), Primary Hypersomnia (307.44),

Narcolepsy (347), Breathing-Related Sleep Disorders (780.59), Circadian Rhythm Sleep Disorder (307.45) and Dyssomnia Not Otherwise Specified (307.47); primary sleep disorders such as Parasomnias such as Nightmare Disorder (307.47), Sleep Terror Disorder (307.46), Sleepwalking Disorder (307.46) and Parasomnia Not Otherwise Specified (307.47); Sleep Disorders Related to Another Mental Disorder such as Insomnia Related to Another Mental Disorder (307.42) and Hypersomnia Related to Another Mental Disorder (307.44); Sleep Disorder Due to a General Medical Condition, in particular sleep disturbances associated with such diseases as neurological disorders, neuropathic pain, restless leg syndrome, heart and lung diseases; and Substance-Induced Sleep Disorder including the subtypes Insomnia Type, Hypersomnia Type, Parasomnia Type and Mixed Type; Sleep Apnea and Jet-Lag Syndrome.

In addition the compounds of formula (I) or their pharmaceutically acceptable salts may be of use for the treatment or prophylaxis of depression and mood disorders including Major Depressive Episode, Manic Episode, Mixed Episode and Hypomanic Episode;

Depressive Disorders including Major Depressive Disorder, Dysthymic Disorder (300.4), Depressive Disorder Not Otherwise Specified (311); Bipolar Disorders including Bipolar I Disorder, Bipolar Π Disorder (Recurrent Major Depressive Episodes with Hypomanic Episodes) (296.89), Cyclothymic Disorder (301.13) and Bipolar Disorder Not Otherwise Specified (296.80); Other Mood Disorders including Mood Disorder Due to a General Medical Condition (293.83) which includes the subtypes With Depressive Features, With Major Depressive-like Episode, With Manic Features and With Mixed Features),

Substance-Induced Mood Disorder (including the subtypes With Depressive Features, With Manic Features and With Mixed Features) and Mood Disorder Not Otherwise Specified (296.90).

Further, the compounds of formula (I) or their pharmaceutically acceptable salts may be of use for the treatment or prophylaxis of anxiety disorders including Panic Attack; Panic Disorder including Panic Disorder without Agoraphobia (300.01) and Panic Disorder with Agoraphobia (300.21); Agoraphobia; Agoraphobia Without History of Panic Disorder (300.22), Specific Phobia (300.29, formerly Simple Phobia) including the subtypes Animal Type, Natural Environment Type, Blood-Injection-Injury Type, Situational Type and Other Type), Social Phobia (Social Anxiety Disorder, 300.23), Obsessive-Compulsive Disorder (300.3), Posttraumatic Stress Disorder (309.81), Acute Stress Disorder (308.3), Generalized Anxiety Disorder (300.02), Anxiety Disorder Due to a General Medical Condition (293.84), Substance-Induced Anxiety Disorder, Separation Anxiety Disorder (309.21), Adjustment Disorders with Anxiety (309.24) and Anxiety Disorder Not Otherwise Specified (300.00).

In addition the compounds of formula (I) or their pharmaceutically acceptable salts may be of use for the treatment or prophylaxis of substance-related disorders including Substance Use Disorders such as Substance Dependence, Substance Craving and Substance Abuse; Substance-Induced Disorders such as Substance Intoxication, Substance

Withdrawal, Substance-Induced Delirium, Substance-Induced Persisting Dementia, Substance-Induced Persisting Amnestic Disorder, Substance-Induced Psychotic Disorder, Substance-Induced Mood Disorder, Substance-Induced Anxiety Disorder, Substance- Induced Sexual Dysfunction, Substance-Induced Sleep Disorder and Hallucinogen

Persisting Perception Disorder (Flashbacks); Alcohol -Related Disorders such as Alcohol Dependence (303.90), Alcohol Abuse (305.00), Alcohol Intoxication (303.00), Alcohol Withdrawal (291.81), Alcohol Intoxication Delirium, Alcohol Withdrawal Delirium, Alcohol -Induced Persisting Dementia, Alcohol-Induced Persisting Amnestic Disorder, Alcohol -Induced Psychotic Disorder, Alcohol-Induced Mood Disorder, Alcohol-Induced Anxiety Disorder, Alcohol-Induced Sexual Dysfunction, Alcohol -Induced Sleep Disorder and Alcohol-Related Disorder Not Otherwise Specified (291.9); Amphetamine (or

Amphetamine-Like)-Related Disorders such as Amphetamine Dependence (304.40), Amphetamine Abuse (305.70), Amphetamine Intoxication (292.89), Amphetamine

Withdrawal (292.0), Amphetamine Intoxication Delirium, Amphetamine Induced Psychotic Disorder, Amphetamine-Induced Mood Disorder, Amphetamine-Induced Anxiety Disorder, Amphetamine-Induced Sexual Dysfunction, Amphetamine-Induced Sleep Disorder and Amphetamine-Related Disorder Not Otherwise Specified (292.9); Caffeine Related

Disorders such as Caffeine Intoxication (305.90), Caffeine-Induced Anxiety Disorder, Caffeine-Induced Sleep Disorder and Caffeine-Related Disorder Not Otherwise Specified (292.9); Cannabis-Related Disorders such as Cannabis Dependence (304.30), Cannabis Abuse (305.20), Cannabis Intoxication (292.89), Cannabis Intoxication Delirium, Cannabis- Induced Psychotic Disorder, Cannabis-Induced Anxiety Disorder and Cannabis-Related Disorder Not Otherwise Specified (292.9); Cocaine-Related Disorders such as Cocaine Dependence (304.20), Cocaine Abuse (305.60), Cocaine Intoxication (292.89), Cocaine Withdrawal (292.0), Cocaine Intoxication Delirium, Cocaine-Induced Psychotic Disorder, Cocaine-Induced Mood Disorder, Cocaine-Induced Anxiety Disorder, Cocaine-Induced Sexual Dysfunction, Cocaine-Induced Sleep Disorder and Cocaine-Related Disorder Not Otherwise Specified (292.9); Hallucinogen-Related Disorders such as Hallucinogen Dependence (304.50), Hallucinogen Abuse (305.30), Hallucinogen Intoxication (292.89), Hallucinogen Persisting Perception Disorder (Flashbacks) (292.89), Hallucinogen

Intoxication Delirium, Hallucinogen-Induced Psychotic Disorder, Hallucinogen-Induced Mood Disorder, Hallucinogen-Induced Anxiety Disorder and Hallucinogen-Related Disorder Not Otherwise Specified (292.9); Inhalant-Related Disorders such as Inhalant Dependence (304.60), Inhalant Abuse (305.90), Inhalant Intoxication (292.89), Inhalant Intoxication Delirium, Inhalant-Induced Persisting Dementia, Inhalant-Induced Psychotic Disorder, Inhalant-Induced Mood Disorder, Inhalant-Induced Anxiety Disorder and

Inhalant-Related Disorder Not Otherwise Specified (292.9); Nicotine-Related Disorders such as Nicotine Dependence (305.1), Nicotine Withdrawal (292.0) and Nicotine-Related Disorder Not Otherwise Specified (292.9); Opioid-Related Disorders such as Opioid Dependence (304.00), Opioid Abuse (305.50), Opioid Intoxication (292.89), Opioid Withdrawal (292.0), Opioid Intoxication Delirium, Opioid-Induced Psychotic Disorder, Opioid-Induced Mood Disorder, Opioid-Induced Sexual Dysfunction, Opioid-Induced Sleep Disorder and Opioid-Related Disorder Not Otherwise Specified (292.9); Phencyclidine (or Phencyclidine-Like)-Related Disorders such as Phencyclidine Dependence (304.60), Phencyclidine Abuse (305.90), Phencyclidine Intoxication (292.89), Phencyclidine

Intoxication Delirium, Phencyclidine-Induced Psychotic Disorder, Phencyclidine-Induced Mood Disorder, Phencyclidine-Induced Anxiety Disorder and Phencyclidine-Related Disorder Not Otherwise Specified (292.9); Sedative-, Hypnotic-, or Anxiolytic-Related Disorders such as Sedative, Hypnotic, or Anxiolytic Dependence (304.10), Sedative, Hypnotic, or Anxiolytic Abuse (305.40), Sedative, Hypnotic, or Anxiolytic Intoxication (292.89), Sedative, Hypnotic, or Anxiolytic Withdrawal (292.0), Sedative, Hypnotic, or Anxiolytic Intoxication Delirium, Sedative, Hypnotic, or Anxiolytic Withdrawal Delirium, Sedative-, Hypnotic-, or Anxiolytic-Persisting Dementia, Sedative-, Hypnotic-, or

Anxiolytic- Persisting Amnestic Disorder, Sedative-, Hypnotic-, or Anxiolytic-Induced Psychotic Disorder, Sedative-, Hypnotic-, or Anxiolytic-Induced Mood Disorder, Sedative-, Hypnotic-, or Anxiolytic-Induced Anxiety Disorder Sedative-, Hypnotic-, or Anxiolytic- Induced Sexual Dysfunction, Sedative-, Hypnotic-, or Anxiolytic-Induced Sleep Disorder and Sedative-, Hypnotic-, or Anxiolytic-Related Disorder Not Otherwise Specified (292.9); Poly sub stance-Related Disorder such as Polysubstance Dependence (304.80); and Other (or Unknown) Substance-Related Disorders such as Anabolic Steroids, Nitrate Inhalants and Nitrous Oxide.

In addition the compounds of formula (I) or their pharmaceutically acceptable salts may be of use for the treatment or prophylaxis of feeding disorders such as bulimia nervosa, binge eating, obesity, including obesity observed in Type 2 (non-insulin-dependent) diabetes patients. Further, the compounds of formula (I) or their pharmaceutically acceptable salts may be of use for the treatment or prophylaxis of stroke, particularly ischemic or haemorrhagic stroke and/or in blocking an emetic response i.e. nausea and vomiting.

The numbers in brackets after the listed diseases refer to the classification code in DSM-IV: Diagnostic and Statistical Manual of Mental Disorders, 4th Edition, published by the American Psychiatric Association. The various subtypes of the disorders mentioned herein are contemplated as part of the present invention.

The invention also provides a method for the treatment of a disease or disorder in a subject, for example those diseases and disorders mentioned hereinabove, comprising administering to said subject an effective amount of a compound of formula (I), or a pharmaceutically acceptable salt thereof. The invention also provides a compound of formula (I), or a pharmaceutically acceptable salt thereof, for use in the treatment or prophylaxis of a disease or disorder, for example those diseases and disorders mentioned hereinabove.

The invention also provides the use of a compound of formula (I), or a

pharmaceutically acceptable salt thereof, in the manufacture of a medicament for use in the treatment or prophylaxis of a disease or disorder, for example those diseases and disorders mentioned hereinabove.

For use in therapy the compounds of the invention are usually administered as a pharmaceutical composition. The invention also provides a pharmaceutical composition comprising a compound of formula (I), or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier.

The compounds of formula (I) or their pharmaceutically acceptable salts may be administered by any convenient method, e.g. by oral, parenteral, buccal, sublingual, nasal, rectal or transdermal administration, and the pharmaceutical compositions adapted accordingly.

The compounds of formula (I) or their pharmaceutically acceptable salts which are active when given orally can be formulated as liquids or solids, e.g. as syrups, suspensions, emulsions, tablets, capsules or lozenges.

A liquid formulation will generally consist of a suspension or solution of the active ingredient in a suitable liquid carrier(s) e.g. an aqueous solvent such as water, ethanol or glycerine, or a non-aqueous solvent, such as polyethylene glycol or an oil. The formulation may also contain a suspending agent, preservative, flavouring and/or colouring agent.

A composition in the form of a tablet can be prepared using any suitable

pharmaceutical carrier(s) routinely used for preparing solid formulations, such as magnesium stearate, starch, lactose, sucrose and cellulose.

A composition in the form of a capsule can be prepared using routine encapsulation procedures, e.g. pellets containing the active ingredient can be prepared using standard carriers and then filled into a hard gelatin capsule; alternatively a dispersion or suspension can be prepared using any suitable pharmaceutical carrier(s), e.g. aqueous gums, celluloses, silicates or oils and the dispersion or suspension then filled into a soft gelatin capsule.

Typical parenteral compositions consist of a solution or suspension of the active ingredient in a sterile aqueous carrier or parenterally acceptable oil, e.g. polyethylene glycol, polyvinyl pyrrolidone, lecithin, arachis oil or sesame oil. Alternatively, the solution can be lyophilised and then reconstituted with a suitable solvent just prior to administration.

Compositions for nasal administration may conveniently be formulated as aerosols, drops, gels and powders. Aerosol formulations typically comprise a solution or fine suspension of the active ingredient in a pharmaceutically acceptable aqueous or nonaqueous solvent and are usually presented in single or multidose quantities in sterile form in a sealed container which can take the form of a cartridge or refill for use with an atomising device. Alternatively the sealed container may be a disposable dispensing device such as a single dose nasal inhaler or an aerosol dispenser fitted with a metering valve. Where the dosage form comprises an aerosol dispenser, it will contain a propellant which can be a compressed gas e.g. air, or an organic propellant such as a fluorochlorohydrocarbon or hydrofluorocarbon. Aerosol dosage forms can also take the form of pump-atomisers.

Compositions suitable for buccal or sublingual administration include tablets, lozenges and pastilles where the active ingredient is formulated with a carrier such as sugar and acacia, tragacanth, or gelatin and glycerin.

Compositions for rectal administration are conveniently in the form of suppositories containing a conventional suppository base such as cocoa butter.

Compositions suitable for transdermal administration include ointments, gels and patches.

In one embodiment the composition is in unit dose form such as a tablet, capsule or ampoule.

The composition may contain from 0.1% to 100% by weight, for example from 10 to 60% by weight, of the active material, depending on the method of administration. The composition may contain from 0% to 99% by weight, for example 40% to 90% by weight, of the carrier, depending on the method of administration. The composition may contain from 0.05mg to lOOOmg, for example from l .Omg to 500mg, of the active material, depending on the method of administration. The composition may contain from 50 mg to 1000 mg, for example from lOOmg to 400mg of the carrier, depending on the method of administration. The dose of the compound used in the treatment of the aforementioned disorders will vary in the usual way with the seriousness of the disorders, the weight of the sufferer, and other similar factors. However, as a general guide suitable unit doses may be 0.05 to 1000 mg, more suitably 1.0 to 500 mg, and such unit doses may be administered more than once a day, for example two or three a day. Such therapy may extend for a number of weeks or months.

Orexin-A (Sakurai, T. et al (1998) Cell, 92 pp 573-585) can be employed in screening procedures for compounds which inhibit the ligand' s activation of the orexin-1 or orexin-2 receptors.

In general, such screening procedures involve providing appropriate cells which express the orexin-1 or orexin-2 receptor on their surface. Such cells include cells from mammals, yeast, Drosophila or s. coli. In particular, a polynucleotide encoding the orexin- 1 or orexin-2 receptor is used to transfect cells to express the receptor. The expressed receptor is then contacted with a test compound and an orexin-1 or orexin-2 receptor ligand, as appropriate, to observe inhibition of a functional response. One such screening procedure involves the use of melanophores which are transfected to express the orexin-1 or orexin-2 receptor, as described in WO 92/01810.

Another screening procedure involves introducing RNA encoding the orexin-1 or orexin-2 receptor into Xenopus oocytes to transiently express the receptor. The receptor oocytes are then contacted with a receptor ligand and a test compound, followed by detection of inhibition of a signal in the case of screening for compounds which are thought to inhibit activation of the receptor by the ligand.

Another method involves screening for compounds which inhibit activation of the receptor by determining inhibition of binding of a labelled orexin-1 or orexin-2 receptor ligand to cells which have the orexin-1 or orexin-2 receptor (as appropriate) on their surface. This method involves transfecting a eukaryotic cell with DNA encoding the orexin-1 or orexin-2 receptor such that the cell expresses the receptor on its surface and contacting the cell or cell membrane preparation with a compound in the presence of a labelled form of an orexin-1 or orexin-2 receptor ligand. The ligand may contain a radioactive label. The amount of labelled ligand bound to the receptors is measured, e.g. by measuring

radioactivity.

Yet another screening technique involves the use of FLIPR equipment for high throughput screening of test compounds that inhibit mobilisation of intracellular calcium ions, or other ions, by affecting the interaction of an orexin-1 or orexin-2 receptor ligand with the orexin-1 or orexin-2 receptor as appropriate.

Throughout the specification and claims which follow, unless the context requires otherwise, the word 'comprise', and variations such as 'comprises' and 'comprising' will be understood to imply the inclusion of a stated integer or step or group of integers or steps but not to the exclusion of any other integer or step or group of integers or steps.

All publications, including but not limited to patents and patent applications, cited in this specification are herein incorporated by reference as if each individual publication were specifically and individually indicated to be incorporated by reference herein as though fully set forth.

The following Examples illustrate the preparation of certain compounds of formula (I) or salts thereof. The Descriptions 1 to 52 illustrate the preparation of intermediates used to make compounds of formula (I) or salts thereof.

In the procedures that follow, after each starting material, reference to a description is typically provided. This is provided merely for assistance to the skilled chemist. The starting material may not necessarily have been prepared from the Description referred to.

The yields were calculated assuming that products were 100 % pure if not stated otherwise.

The compounds described in the Examples described hereinafter have all been prepared as a first step from stereochemically pure starting materials. The stereochemistry of the compounds of the Descriptions and Examples have been assigned on the assumption that the absolute configuration of these centres are retained. The relative stereochemistry of the compounds of the Descriptions and Examples have been assigned on the assumption that the relative stereochemistry is maintained as determined by using Rotating frame 2D ROESY experiments in the chiral intermediates. In some Examples the relative

stereochemistry has been confirmed on the final compounds as well.

Compounds are named using ACD/Name PRO 6.02 chemical naming software

(Advanced Chemistry Development Inc., Toronto, Ontario, M5H2L3, Canada).

Proton Magnetic Resonance ( MR) spectra were recorded either on Varian instrument at 400, 500 or 600 MHz, or on a Bruker instrument at 400 MHz. Chemical shifts are reported in ppm (δ) using the residual solvent line as internal standard. Splitting patterns are designed as s, singlet; d, doublet; t, triplet; q, quartet; m, multiplet; b, broad. The NMR spectra were recorded at a temperature ranging from 25 to 90 °C. When more than one conform er was detected the chemical shifts for the most abundant one is usually reported. Unless otherwise specified, HPLC analyses indicated by HPLC (walk-up): rt (retention time) = x min, were performed on a Agilent 1100 series instrument using a Luna 3u CI 8(2) 100A column (50 x 2.0 mm, 3 μπι particle size) [Mobile phase and Gradient: 100% (water + 0.05% TFA) to 95% (acetonitrile + 0.05% TFA) in 8 min. Column T = 40 °C. Flow rate = 1 mL/min. UV detection wavelength = 220 nm]. Other HPLC analyses, indicated by HPLC (walk-up, 3 min method), were performed using an Agilent Zorbax SB- C18 column (50 x 3.0 mm, 1.8 μπι particle size) [Mobile phase and Gradient: (Solvent A: water + 0.05% TFA) (Solvent B: acetonitrile + 0.05% TFA) Gradient: time 0 min 0% B. From 0 to 95% B in 2.5 min. 95% B for 0.2 min. From 95 to 100% B in 0.2 min. 100% B for 0.4 min. From 100% to 0% B in 0. lmin. Flow rate = 1.5 mL/min. UV detection wavelength = 220 nm]

In the analytical characterization of the described compounds "MS" refers to Mass Spectra taken by Direct infusion Mass or to Mass Spectra associated with peaks taken by UPLC/MS or HPLC/MS analysis, where the Mass Spectrometer used is as mentioned below.

Direct infusion Mass spectra (MS) were run on a Agilent MSD 1100 Mass

Spectrometer, operating in ES (+) and ES (-) ionization mode [ES (+): Mass range: 100- 1000 amu. Infusion solvent: water + 0.1% HC0₂H / CH₃CN 50/50. ES (-): Mass range: 100-1000 amu. Infusion solvent: water + 0.05% H₄OH / CH₃CN 50/50]

MS spectra associated with the peaks were taken on HPLC instrument Perkin Elmer 200 series coupled to an Applied Biosystems API150EX Mass Spectrometer.

UV and MS spectra associated with the peaks were taken on HPLC instrument Agilent 1100 Series coupled to an Agilent LC/MSD 1100 Mass Spectrometer operating in positive or negative electrospray ionization mode and in both acidic and basic gradient conditions

[Acidic gradient LC/MS - ES (+ or -): analyses performed on a Supelcosil ABZ + Plus column (33 x 4.6 mm, 3 μιη). Mobile phase: A - water + 0.1% HC0₂H / B - CH₃CN.

Gradient (standard method): t=0 min 0% (B), from 0% (B) to 95% (B) in 5 min lasting for

1.5 min, from 95% (B) to 0%(B) in 0.1 min, stop time 8.5 min. Column T = room temperature. Flow rate = 1 mL/min. Gradient (fast method): t=0 min 0% (B), from 0% (B) to 95% (B) in 3 min lasting for 1 min, from 95% (B) to 0% (B) in 0.1 min, stop time 4.5 min. Column T = room temperature. Flow rate = 2 mL/min.

Basic gradient LC/MS - ES (+ or -): analyses performed on a XTerra MS CI 8 column (30 x

4.6 mm, 2.5 μιη). Mobile phase: A - 5 mM aq. H₄HC0₃ + ammonia (pH 10) / B - CH₃CN. Gradient: t = 0 min 0% (B), from 0% (B) to 50% (B) in 0.4 min, from 50% (B) to 95% (B) in 3.6 min lasting for 1 min, from 95% (B) to 0% (B) in 0.1 min, stop time 5.8 min. column temperature = room temperature. Flow rate = 1.5 mL/min].

Mass range ES (+ or -): 100-1000 amu. UV detection range: 220-350 nm. The usage of this methodology is indicated by "LC-MS" in the analytic characterization of the described compounds.

Total ion current (TIC) and DAD UV chromatographic traces together with MS and

UV spectra associated with the peaks were taken on a UPLC/MS Acquity™ system equipped with 2996 PDA detector and coupled to a Waters Micromass ZQ™ Mass Spectrometer operating in positive or negative electrospray ionisation mode [LC/MS - ES (+ or -): analyses performed using an AcquityTM UPLC BEH C18 column (50 x 21 mm, 1.7 μιη particle size), column temperature 40 °C]. Mobile phase: A-water + 0.1% HCOOH / B - CH₃CN + 0.075% HCOOH, Flow rate: 1.0 mL/min, Gradient: t=0 min 3% B, t=0.05 min 6% B, t= 0.57 min 70% B, t=l .4 min 99% B, t=l .45 min 3% B). The usage of this methodology is indicated by "UPLC" in the analytic characterization of the described compounds.

[LC/MS - ES (+ or -): analyses performed using an Acquity™ UPLC BEH CI 8 column (50 x 2.1 mm, 1.7 μιη particle size) column temperature 40 °C]. Mobile phase: A - water + 0.1% HC0₂H / B - CH₃CN + 0.06% or 0.1% HC0₂H Gradient: t = 0 min 3% B, t =1.5 min 100% B, t = 1.9 min 100% B, t = 2 min 3% B stop time 2 min. Column T = 40 °C. Flow rate = 1.0 mL/min. Mass range: ES (+): 100-1000 amu or ES(+): 50-800 amu. ES (-): 100- 800 amu. UV detection range: 210-350 nm. The usage of this methodology is indicated by "UPLC (Acid IPQC)" in the analytic characterization of the described compounds.

[LC/MS - ES (+ or -): analyses performed using an Acquity™ UPLC BEH C 18 column (50 x 2.1 mm, 1.7 μιη particle size) column temperature 40 °C]. Mobile phase: A - water + 0.1% HC0₂H / B - CH₃CN + 0.06% or 0.1% HC0₂H. Gradient: t = 0 min 3% B, t = 0.05 min 6% B, t = 0.57 min 70% B, t = 1.06 min 99% B lasting for 0.389 min, t = 1.45 min 3% B, stop time 1.5 min. Column T = 40 °C. Flow rate = 1.0 mL/min. Mass range: ES (+): 100- 1000 amu or ES(+): 50-800 amu, ES (-): 100-800 amu. UV detection range: 210-350 nm. The usage of this methodology is indicated by "UPLC (Acid QC POS 50-800 or

QC POS 70 900 or GEN QC or FF AL QC)" in the analytic characterization of the described compounds.

[LC/MS - ES (+ or -): analyses performed using an Acquity™ UPLC BEH CI 8 column (50 x 2.1 mm, 1.7 μιη particle size) column temperature 40 °C]. Mobile phase: A - water + 0.1% HC0₂H / B - CH₃CN + 0.06% or 0.1% HC0₂H. Gradient: t = 0 min 3% B, t = 1.06 min 99 % B, t = 1.45 min 99 % B, t = 1.46 min 3 % B, stop time 1.5 min. Column T = 40 °C. Flow rate = 1.0 mL/min. Mass range: ES (+): 100-1000 amu. ES (-): 100-800 amu. UV detection range: 210-350 nm. The usage of this methodology is indicated by "UPLC (Acid GEN QC SS)" in the analytic characterization of the described compounds.

Total ion current (TIC) and DAD UV chromatographic traces together with MS and UV spectra associated with the peaks were taken on a UPLC/MS Acquity™ system equipped with PDA detector and coupled to a Waters SQD mass spectrometer operating in positive and negative alternate electrospray ionisation mode [LC/MS - ES(+ or -): analyses performed using an Acquity™ UPLC BEH C 18 column (50 x 2.1 mm, 1.7 μιη particle size) column temperature 40 °C]. Mobile phase: A - 10 mM aqueous solution of NH₄HC0₃ (adjusted to pH 10 with ammonia) / B - CH₃CN. Gradient: t = 0 min 3% B, t = 1.06 min 99% B lasting for 0.39 min, t = 1.46 min 3% B, stop time 1.5 min. Column T = 40 °C. Flow rate = 1.0 mL/min. Mass range: ES (+): 100-1000 amu or ES (+): 50-800 amu. ES (-): 100- 1000 amu. UV detection range: 220-350 nm. The usage of this methodology is indicated by "UPLC (Basic GEN QC or QC POS 50-800)" in the analytic characterization of the described compounds. For reactions involving microwave irradiation, a Personal Chemistry EmrysTM Optimizer was used.

In a number of preparations, purification was performed using Biotage manual flash chromatography (Flash+), Biotage automatic flash chromatography (Horizon, SP1 and SP4), Companion CombiFlash (ISCO) automatic flash chromatography, Flash Master Personal or Vac Master systems.

Flash chromatography was carried out on silica gel 230-400 mesh (supplied by Merck AG Darmstadt, Germany), Varian Mega Be-Si pre-packed cartridges, pre-packed Biotage silica cartridges (e.g. Biotage SNAP cartridge), KP-NH prepacked flash cartridges or ISCO RediSep Silica cartridges.

SPE-SCX cartridges are ion exchange solid phase extraction columns supplied by Varian. The eluent used with SPE-SCX cartridges is DCM and MeOH or ACN or MeOH followed by 2 N ammonia solution in MeOH. The collected fractions are those eluted with the ammonia solution in MeOH.

SPE-Si cartridges are silica solid phase extraction columns supplied by Varian.

ENV+ cartridges are packed with ENV+ a hyper cross-linked hydroxylated polystyrene-divinylbenzene copolymer.

The following table lists the used abbreviations:

ACN Acetonitrile

AcOH Acetic acid

bs or br.s. broad signal

Boc t-Butoxycarbonyl

Burgess reagent Methyl N-(triethylammoniumsulphonyl)carbamate

CBr₄ Carbon tetrabromide

CV Column volumes

Cy Cyclohexanes

DCE Dichloroethane

DCM Dichloromethane

Dess-Martin 1, 1, l-Tris(acetoxy)-l, l-dihydro-l,2-benziodoxol-3-(lH)-one periodinane

DIAD Diisopropyl azodicarboxylate

DIPEA N,N-Diisopropyl-N-ethylamine

DMF Dimethylformamide

DMSO Dimethylsulfoxide

EtOAc Ethyl acetate

EtOH Ethanol

LiOH Lithium hydroxide

MeOH Methanol min Minutes

MTBE Methyl tertiary butyl ether

MP N-Methyl-2-pyrrolidone

Ph Phenyl

pH=3 buffer Citric acid/NaOH/HCl in water solution available from Merck solution KGaA

Grubbs 1^st Benzylidene-bis(tricyclohexylphosphine)dichlororuthenium

generation

(Grubbs I)

rt retention time

T temperature

TBAF Tetrabutyl ammonium fluoride

TBDMS tert-Butyl dimethylsilyl

TBDPS tert-Butyl diphenylsilyl

TBTU O-(benzotriazol-l-yl)-N,N;N'N'-tetramethyl

tetrafluorob orate

t-Bu tert-Butyl

TEA Tri ethyl amine

TEMPO 2,2,6,6-Tetramethylpiperidine- 1 -oxyl

TFA Trifluoroacetic acid

THF Tetrahydrofuran

TMS Trimethylsilyl

Ts ^-Toluensulfonyl

Descriptions

Description 1: 1,1-dimethylethyl (lR,6R)-2-oxo-3-azab icy clo [4.1.0] heptane-4- carboxylate (Dl)

Sodium iodide (391 g, 2.6 mol) was partially dissolved in ACN (1.7 1) after stirring at 20°C for 10 minutes under nitrogen atmosphere. TMS-Cl (0.323 1, 2.5 mol) was added over 10 minutes and the resulting yellow slurry was stirred at 20°C for 1 hour. A solution of (lR,55)-3-oxabicyclo[3.1.0]hexan-2-one (Minakem supplier, 170 g, 1.73 mol) in ACN (340 ml) was added over 5 minutes at 20°C. The suspension was heated to 50°C (internal temperature), then kept for 3 hours and 45 minutes at 50°C. The mixture was diluted with MeOH (1.7 1) at 20°C and concentrated to 5 volumes (850 ml) under reduced pressure. MeOH (1.7 1) was then added followed by TMS-Cl (0.102 1, 0.8 mol). The resulting mixture was stirred at 20°C for 15 hours 30 minutes. The mixture was concentrated under vacuum to 5 volumes (0.85 1), then 2-MeTHF was added (1.7 1) and the solution was concentrated to 5 volumes (0.85 1). 2-MeTHF was added (1.7 1). The dark red solution was washed with aqueous Na₂S0₃ 20% w/w (0.68 1) at 20°C (solution became colourless-light yellow). The biphasic system was separated and the organic layer was washed with water (0.68 1), then concentrated under vacuum to 4 volumes (0.68 1). 2-MeTHF (1.7 1) was added and the solution of methyl (lR,2S)-2-(iodomethyl)cyclopropanecarboxylate was concentrated to 5 volumes (0.85 1), diluted with 2-Me-THF (0.51 1).

N-(Diphenylmethylene)glycine t-butylester (503.2g, 1.7mol, 1.2 eq) was suspended in dry Me-THF (1.7L) at 20°C under nitrogen. The mixture was cooled to 0°C and KOtBu (195.5g, 1.74mol, 1 eq) was added in three portions. The slurry was become a yellow- orange solution and was stirred at 0°C for 30 minutes. The previous solution of methyl (lR,2S)-2-(iodomethyl)cyclopropanecarboxylate in 2-MeTHF was slowly added over 25 minutes, keeping the temperature lower than 5°C during the addition. The mixture was stirred at 0°C for 2.5 hours. The mixture was quenched with buffer pH=7

(KH₂P0₄/Na₂HP0₄) (340ml) at 0°C. The biphasic system was warmed at 20°C. The water phase was discharged. To the organic phase at 0°C was added citric acid 30% w/w (1.36L) keeping the temperature 0-5°C and the biphasic system was stirred for 16 hours 20 minutes at 20°C. Cyclohexane (3.4L) was added and the phases were separated. The water phase was washed with cyclohexane (3.4L). Ethyl acetate (3.4L) was added to the water phase, and then the system was basified to pH=8.5 with aqueous saturated K₂C0₃ (0.85L) then diluted with water (0.425L). The biphasic system was separated. The aqueous layer was back extracted with ethyl acetate (3.4L). The combined organic phases were washed with water (0.51L), concentrated to 10 volumes (1.7L). Toluene (3.4L) was added and the solution was concentrated to 10 volumes (1.7L), diluted again with toluene (0.85L). To this solution was added HC1 37% (0.85ml, catalytic amount). The solution was heated to 105°C for 20 hours. The solution was cooled at 40°C, reduced to 4 volumes (0.68L) under reduced pressure and heptane (1.19L) was added over lhour. The mixture was stirred at 40°C for 30 minutes and then cooled at 15°C over 1 hour: a solid was precipitated. The slurry was stirred at 15° for approximately 16 hours and then filtered. The solid was washed with heptane (2x0.425L), dried in a vacuum oven at 40°C for 20 hours and 30 minutes. 1,1 -dimethyl ethyl (lR,6R)-2-oxo-3-azabicyclo[4.1.0]heptane-4-carboxylate (syn/anti mixture, 194g) Dl was obtained as white solid.

1H MR (600 MHz, DMSO-d6) δ ppm 6.86 - 7.39 (1 H, 2 m), 3.81 (1 H, 2 dd), 2.20 - 2.33 (1 H, 2 m), 1.74 - 2.11 (1 H, 2 m), 1.42 (9 H, s), 1.4 - 1.6 (1 H, m), 0.90 - 1.12 (1 H, 2 m), 0.69 - 0.88 (1 H, 2 m)

Description 2: l,l-dimethylethyl(lR,4S,6R)-4-(hydroxymethyl)-3- azabicyclo [4.1.0] heptane-3-carboxylate (D2)

The 1,1 -dimethyl ethyl (lR,6R)-2-oxo-3-azabicyclo[4.1.0]heptane-4-carboxylate Dl (150g, 1 eq) was dissolved in toluene (0.450L) and MeOH (1.05L) stirred for 5 minutes at 20°C. The temperature was cooled to 15°C and KOH (60g, 1.06 mol, 1.5eq) was added in two portions. The solution was stirred at 20°C for 3hours. The solution was cooled to 10°C, TMSCl (0.36L, 2.84 mol, 4eq) was added keeping the temperature around 10-15°C over 40 minutes. White solid was precipitated (KC1). The slurry was stirred at room temperature overnight. The pH of the organic phase was measured and found to be 1. NaHC0₃ solid (240g) was added in four portions to reach pH=5.5. The volume was reduced to 4 volumes (0.6L). THF (1.5L) was added and the volume is reduced to 4 volumes (0.6L) by distillation under reduced pressure. The solid was filtered (note: 60ml of the slurry was collected prior to filtration, so 10% of input was removed) and washed with THF (3x0.3L). The filtrate appeared cloudy. The solution was reduced to 2.2 volumes (0.337L) by distillation under reduced pressure and BF₃.THF (422.55mL, 3.83 mol, 6eq considering the 10% removed) was added under stirring whilst maintaining an internal temperature of 25°C. The resulting solution was added slowly to a solution of L1BH₄ (4M in THF) (0.648L, 2.59 mol, 4eq) diluted with THF (0.405L) keeping the temperature at 25-30°C (the line was washed with THF (0.337L)). The mixture was stirred at 30°C overnight (17 hours). The mixture was quenched slowly with MeOH (0.54L) at 25-30°C. The solution was stirred at 50°C for approximately 1 hour. After this time, the solution was reduced to 5.5 volumes (742.5 ml) by distillation under reduced pressure. HC1 3M (0.540L) was then added at 10-15°C. The mixture was stirred at 20°C for 1 hour and toluene (0.54L) was added. The phases were separated. The aqueous phase was washed with toluene (3x0.54L).The aqueous layer was basified with 6M NaOH (405mL) until pH=9. To the basic aqueous solution at 25°C were successively added THF (67.5 mL) and a solution of ditert-butyl dicarbonate in THF

(50%wt/vol, d=0.92, 0.25L, 0.626 mol, 0.93eq) The pH was adjusted to pH=8.5 by addition of 6M NaOH (0.135L). The resulting slurry was stirred for 30 minutes at 25°C and the pH was adjusted to pH=9 by addition of 6M NaOH (0.135L). The slurry was then stirred for 3hours and then filtered. The inorganic salts were washed with MTBE (2x0.27L). The filtrate was diluted with MTBE (1.08L). The biphasic system was separated. The organic phase was washed with NaCl 20%w/w (0.54L) and then concentrated under reduced pressure to 2.5 volumes (337.5 mL). Heptane (1.35L) was added and the solution was reduced to 5 volumes (0.675L), diluted with heptane (0.675L) and concentrated to 5 volumes (0.675L) by distillation under reduced pressure. Seed (135mg) of the title compound was added at 40°C and the slurry was cooled at 20°C in 1 hour. The slurry was stirred for at least 4 hours and filtered. The solid was washed with cold heptane (0.27L) and dried in a vacuum oven at 40°C for 14 hours and 30 minutes. l,l-dimethylethyl(lR,4S,6R)- 4-(hydroxymethyl)-3-azabicyclo[4.1.0]heptane-3-carboxylate (98g) D2 was obtained as white solid.

1H MR (600 MHz, DMSO-d6) δ ppm 4.67 (1 H, br. s.), 3.6 - 3.9 (2 H, m), 3.2 - 3.5 (3 H, m), 1.89 (1 H, m), 1.54 (1 H, m), 1.37 (9 H, br. s.), 0.90 (2 H, m), 0.58 (1 H, m), -0.09 (1 H, q)

Description 3 : l,l-dimethylethyl(lR,4S,6R)-4-[(l,3-dioxo-l,3-dihydro-2H-isoindol-2- yl)methyl]-3-azabicyclo[4.1.0]heptane-3-carboxylate (D3)

l,l-dimethylethyl(lR,4S,6R)-4-(hydroxymethyl)-3-azabicyclo[4.1.0]heptane-3-carboxylate D2 (1.43 g) was dissolved in THF (50 ml) then lH-isoindole-l,3(2H)-dione (1.111 g, 7.55 mmol) and triphenylphosphine (2.475 g, 9.44 mmol) were added. This solution was warmed up to 50 °C and then DIAD (1.835 ml, 9.44 mmol) was added dropwise. The reaction was stirred at 50 °C for 30 minutes, then it was cooled to room temperature and all volatiles were removed under vacuum. The residue was purified by Silica Gel Chromatography (Biotage SP-column size 100 g SNAP) eluting with Cy:EtOAc=8:2 to 5:5 as eluent. It was recovered the title compound D3 (1.85 g). UPLC: (Acid Final QC): rt= 0.81, peak observed: 357 (M+1). C₂₀H₂₄N₂O₄ requires 356. 1H NMR (400 MHz, DMSO-d6) δ ppm 8.04-7.69 (m, 4H) 4.35-4.12 (m, 1H) 4.00-3.83 (m, 1H) 3.77-3.40 (m, 3H) 2.07-1.81 (m, 1H) 1.77-1.55 (m, 1H) 1.13-0.94 (m, 9H) 0.75-0.59 (m, 1H) 0.07- -0.19 (m, 1H).

Description 4 : l,l-dimethylethyl(lR,4S,6R)-4-(aminomethyl)-3- azabicyclo [4.1.0] heptane-3-carboxylate (D4)

l,l-dimethylethyl(lR,4S,6R)-4-[(l,3-dioxo-l,3-dihydro-2H-isoindol-2-yl)methyl]-3- azabicyclo[4.1.0]heptane-3-carboxylate D3 (1.85 g) was dissolved in EtOH (20 ml) then hydrazine (2.036 ml, 51.9 mmol) was carefully added and the reaction stirred at room temperature overnight. All volatiles were removed under vacuum and the solid residue was triturated with Et₂0. These organic phases were collected together and concentrated to dryness to give the title compound D4 as pale yellow oil (1.1 g). UPLC: (Acid Final QC): rt= 0.45, peak observed: 227 (M+1). C₁₂H₂₂N₂O₂ requires 226. 1H NMR (400 MHz, CDCI₃) δ ppm 4.09-3.64 (m, 2H) 3.41-3.18 (m, 1H) 2.99-2.83 (m, 1H) 2.77-2.59 (m, 1H) 1.90- 1.71 (m, 2H) 1.67-1.48 (m, 2H) 1.47 (s, 1H) 1.02-0.84 (m, 2H) 0.74-0.55 (m, 1H) 0.18- - 0.18 (m, 1H). Description 5: l-(l,l-dimethylethyl) 2-methyl (2S)-3,6-dihydro-l,2(2H>

pyridinedicarboxylate (D5)

To a solution of (2S)-l-{ [(l, l-dimethylethyl)oxy]carbonyl}-l,2,3,6-tetrahydro-2- pyridinecarboxylic acid (1.50 g, 6.60 mmol) in DMF (6 ml), DIPEA (6.92 ml, 39.60 mmol) and TBTU (2.97 g, 9.24 mmol) were added and the mixture stirred at room temperature for 45 min. MeOH (1.42 ml, 35.10 mmol) was added and the resulting reaction mixture stirred for 2 hours. The mixture was diluted with DCM and washed with a saturated NaHC0₃ aqueous solution. The organic layer was separated, dried (Na₂S0₄), filtered through a phase separator tube and concentrated under reduced pressure. The crude material was purified by flash chromatography on silica gel (Flash Master 70 g, Cy/EtOAc 90/10). Collected fractions gave the title compound D5 (1.10 g). MS: (ES/+) m/z: 242 (M+1), 186 [M+l- C(Me)₃)] and 142 (M+l-Boc). Ci₂Hi₉N0₄ requires 241. 1H- MR (400 MHz, CDC1₃) 6(ppm): 5.60 - 5.82 (m, 2 H), 4.84 - 5.15 (m, 1 H), 4.01 - 4.19 (m, 1 H), 3.75 - 3.89 (m, 1 H), 3.69 - 3.76 (m, 3 H), 2.44 - 2.72 (m, 2 H), 1.45 - 1.55 (m, 9 H).

Description 6: 1,1-dimethylethyl (2S)-2-(hydroxymethyl)-3,6-dihydro-l(2H)- pyridinecarboxylate (D6)

A solution of 1 -(1, 1-dimethylethyl) 2-methyl (2S 3,6-dihydro-l,2(2H)- pyridinedicarboxylate D5 (1.10 g) in THF (25 ml) was cooled down to 0 °C and lithium borohydride (2.3 M solution in THF, 4.96 ml, 1 1.40 mmol) was added dropwise. The resulting reaction mixture was stirred at room temperature overnight. Further lithium borohydride (9.92 ml, 22.80 ml) was added; the mixture was stirred for 6 hours and then quenched with brine and extracted with EtOAc. The organic phase was separated, dried (Na₂S0 ), filtered through a phase separator tube and concentrated under reduced pressure to afford the title compound D6 (0.98 g). The material was used in the next step without any further purification. MS: (ES/+) m/z: 214 (M+1), 158 [M+1-C(CH₃)₃)] and 114 (M+l-Boc). CiiHi₉N0₃ requires 213. 1H- MR (400 MHz, CDC1₃) 6(ppm): 5.61 - 5.82 (m, 2 H), 4.35 - 4.64 (m, 1 H), 3.98 - 4.30 (m, 1 H), 3.48 - 3.73 (m, 3 H), 2.35 - 2.48 (m, 1 H), 1.96 - 2.15 (m, 1 H), 1.50 (m, 9 H).

Description 7: 1,1-dimethylethyl (2S)-2-({[(l,l- dimethylethyl)(diphenyl)silyl]oxy}methyl)-3,6-dihydro-l(2H)-pyridinecarboxylate

To a solution of 1,1-dimethylethyl (2,S)-2-(hydroxymethyl)-3,6-dihydro-l(2H)- pyridinecarboxylate D6 (0.98 g of the crude material obtained in the Description 2) in DMF (5 ml), imidazole (1.56 g, 22.97 mmol) and chloro(l,l-dimethylethyl)diphenylsilane (1.52 g, 5.52 mmol) were added and the reaction mixture was left under stirring at room temperature for 3 hours. The mixture was diluted with brine and extracted with EtOAc. The organic phase was separated, dried (Na₂S0₄), filtered through a phase separator tube and concentrated under reduced pressure. The residue was purified by flash chromatography on silica gel (Flash Master 70 g, Cy/EtOAc 90/10) to afford the title compound D7 (1.81 g). MS: (ES/+) m/z: 452 (M+l) and 474 (M+Na). C₂₇H37N0₃Si requires 451. 1H- MR (400 MHz, CDC1₃) 6(ppm): 7.57 - 7.78 (m, 4 H), 7.32 - 7.51 (m, 6 H), 5.44 - 5.75 (m, 2 H), 4.37 - 4.80 (m, 1 H), 4.02 - 4.31 (m, 1 H), 3.53 - 3.72 (m, 2 H), 3.28 - 3.51 (m, 1 H), 1.99 - 2.44 (m, 2 H), 1.48 (s, 9 H), 1.07 (s, 9 H).

Description 8: (2S)-2-({[(l,l-dimethylethyl)(diphenyl)silyl]oxy}methyl)-l,2,3,6- tetrahydropyridine (D8):

To a solution of 1,1-dimethylethyl (25)-2-({[(l,l- dimethylethyl)(diphenyl)silyl]oxy}methyl)-3,6-dihydro-l(2H)-pyridinecarboxylate D7 (1.81 g) in DCM (40 ml), TFA (20 ml) was added and the reaction mixture stirred at room temperature for 1 hour. Volatiles were removed under reduced pressure and the residue was eluted through a SCX column. Collected fractions gave the title compound D8 (1.35 g). MS: (ES/+) m/z: 352 (M+l). C₂₂H₂₉NOSi requires 351. 1H- MR (300 MHz, CDC1₃) 6(ppm): 7.57 - 7.78 (m, 4 H), 7.32 - 7.51 (m, 6 H), 5.71 - 5.76 (m, 2 H), 3.54 - 3.72 (m, 2 H), 3.34 - 3.53 (m, 2 H), 2.89 - 3.02 (m, 1 H), 1.83 - 1.92 (m, 2 H), 1.07 (s, 9 H).

Description 9A and 9B: (2S)-2-({[(l,l-dimethylethyl)(diphenyl)silyl]oxy}methyl)-l-[(4- methylphenyl)sulfonyl]-l,2,3,6-tetrahydropyridine (D9A D9B):

A) To a solution of (25)-2-({[(l,l-dimethylethyl)(diphenyl)silyl]oxy}methyl)-l,2,3,6- tetrahydropyridine D8 (1.35 g) in DCM (25.60 ml), TEA (1.07 ml, 7.68 mmol) and 4- methylbenzenesulfonyl chloride (0.80 g, 4.22 mmol) were added and the resulting reaction mixture was stirred at room temperature overnight. The mixture was washed with a saturated aqueous H₄C1 solution. The organic layer was separated, dried ( a₂S04), filtered through a phase separator tube and concentrated under reduced pressure. The residue was purified by flash chromatography on silica gel (Biotage SP 40 M, from Cy 100 to Cy/EtOAc 90/10) to afford the title compound D9A (1.90 g). MS: (ES/+) m/z: 506 (M+l) and 528 (M+Na). C₂₉H₃₅N0₃SSi requires 505. 1H- MR (300 MHz, CDC1₃) 6(ppm): 7.29 - 7.76 (m, 12 H), 7.15 (d, 2 H), 5.45 - 5.67 (m, 2 H), 4.42 - 4.37 (m, 1 H), 3.92 - 4.11 (m, 1 H), 3.51 - 3.61 (m, 2 H), 3.35 - 3.50 (m, 1 H), 2.37 (s, 3 H), 2.04 - 2.33 (m, 2 H), 1.03 (s, 9 H).

B) An alternative method to make D5 is as follows: N-[(1S)-1-({[(1,1- dimethyl ethyl)(diphenyl)silyl]oxy }methyl)-3-buten- 1 -yl]-4-m ethyl -N-2-propen- 1 - ylbenzenesulfonamide D13 (7.46 g) was dissolved in DCM (50 ml) then Grubbs I (1.170 g, 1.398 mmol) was added and the mixture was stirred at room temperature overnight. All volatiles were removed under vacuum and the resulting crude product was purified by silica gel chromatography (Biotage SP - column size 340 g SNAP, Cy to Cy/EtOAc 80/20) to afford the title compound D9B (7.4 g). MS: (ES/+) m/z: 506 (M+l) and 528 (M+Na).

C₂₉H₃₅N0₃SSi requires 505. 1H-NMR (400 MHz, CDC1₃) 6(ppm): 7.67-7.58 (m, 5 H), 7.47-7.35 (m, 5 H), 7.21-7.16 (m 2 H), 5.5-4.8 (m, 2 H), 4.42 - 4.37 (m, 1 H), 4.11-3.92 (m, 1 H), 3.62-3.50 (m, 2 H), 3.50 - 3.35 (m, 1 H), 2.40 (s, 3 H), 2.33-2.11 (m, 2 H), 2.00-1.08 (m, 2 H), 1.05 (s, 9 H).

Description 10: (lR,4S,6R)-4-({[(l,l-dimethylethyl)(diphenyl)silyl]oxy}methyl)-3-[(4- methylphenyl)sulfonyl]-3-azabicyclo[4.1.0]heptane (D10):

A solution of diethylzinc (1 M solution in hexanes, 21.35 ml, 21.35 mmol) in DCM (10 ml) was cooled down to 0 °C and TFA (1.64 ml, 21.35 mmol) was added dropwise. After 20 minutes stirring, diiodomethane (1.73 mol, 21.35 mmol) was added and the mixture left stirring for a further 20 minutes. A solution of (25)-2-({[(l,l- dimethylethyl)(diphenyl)silyl]oxy}methyl)-l-[(4-methylphenyl)sulfonyl]-l,2,3,6- tetrahydropyridine D9A (1.35 g) in DCM (5 ml) was then added, the resulting reaction mixture was allowed to warm up to room temperature and stirred for 6 hours. A solution of diethylzinc (8 eq), TFA (8 eq) and diiodomethane (8 eq) in DCM was prepared and added to the previous mixture at 0 °C. The resulting reaction mixture was left under stirring at room temperature overnight and washed with a saturated aqueous NH₄C1 solution. The aqueous layer was back-extracted with EtOAc. The collected organic layers were dried (Na₂S0₄), filtered through a phase separator tube and concentrated under reduced pressure. The residue was purified by flash chromatography on silica gel (Biotage SP 40 M, from Cy 100 to Cy/EtOAc 90/10) to afford the title compound D6 (0.83 g). MS: (ES/+) m/z: 520 (M+l) and 542 (M+Na). C₃₀H₃₇NO₃SSi requires 519. 1H- MR (300 MHz, CDC1₃) 6(ppm): 7.50 - 7.75 (m, 6 H), 7.28 - 7.49 (m, 6 H), 7.15 (d, 2 H), 3.78 - 3.90 (m, 1 H), 3.52 - 3.70 (m, 2 H), 3.20 - 3.41 (m, 2 H), 2.37 (s, 3 H), 2.17 - 2.29 (m, 1 H), 1.31 - 1.41 (m, 1 H), 1.03 (s, 9 H), 0.56 - 0.93 (m, 3 H), -0.01 (q, 1 H).

Description 11 N-[(lS)-l-(hydroxymethyl)-3-buten-l-yl]-4-methylbenzenesulfonamide

A solution of (25)-2-amino-4-pentenoic acid (5 g, 43.4 mmol) in THF (200 ml) was cooled down to 0 °C and LiAlH₄ (1 M solution in THF, 54.3 ml, 54.3 mmol) was added dropwise. The resulting reaction mixture was allowed to warm-up to room temperature and stirred overnight. The mixture was then cooled down to 0 °C and quenched with a 2 M aqueous NaOH solution. The solid was filtered off and extracted with boiling THF for 1 hour. The combined ethereal extracts were concentrated under reduced pressure and the remaining aqueous mixture extracted with DCM. The combined organic phases were washed with brine, dried (Na₂S0₄) and evaporated under reduced pressure to afford the crude intermediate (25)-2-amino-4-penten-l-ol (3.82 g) that was used in the next step without any further purification.

A solution of sodium carbonate (6.40 g, 60.4 mmol) in water (35 ml) was left under stirring for 20 minutes at room temperature. (25)-2-amino-4-penten-l-ol (3.82 g) was added, followed by EtOAc (80 ml). After 30 minutes stirring, a solution of ^-toluenesulfonyl chloride (5.59 g, 29.3 mmol) in EtOAc (10 ml) and THF (10 ml) was added over 30 minutes. The reaction mixture was stirred at room temperature for 5 hours. Water (30 ml) and EtOAc (100 ml) were then added. The organic phase was separated and the aqueous one extracted with EtOAc (2 x 50 ml). The combined organic layers were dried (Na₂S0₄), filtered and concentrated under reduced pressure. The residue was purified by flash chromatography on silica gel (Biotage SP 340 g SNAP, from Cy/EtOAc 70/30 to EtOAc 100) to afford the title compound Dll (4.23 g). MS: (ES/+) m/z: 256 (M+l). Ci₂Hi₇N0₃S requires 255. 1H-NMR (400 MHz, DMSO-i¾) 6(ppm): 7.68 (d, 2 H), 7.48 (d, 1 H), 7.37 (d, 2 H), 5.48 - 5.63 (m, 1 H), 4.82 - 4.98 (m, 2 H), 4.66 (t, 1 H), 3.18 - 3.27 (m, 1 H), 3.00 - 3.17 (m, 2 H), 2.39 (s, 3 H), 2.17 - 2.27 (m, 1 H), 1.91 - 2.03 (m, 1 H).

Description 12: N-[(lS)-l-({[(l,l-dimethylethyl)(diphenyl)silyl]oxy}methyl)-3-buten-l- yl]-4-methylbenzenesulfonamide (D12):

To a solution ofN-[(l,S)-l-(hydroxymethyl)-3-buten-l-yl]-4-methylbenzenesulfonamide Dll (4.23 g) in DMF (35 ml), imidazole (2.98 g, 43.7 mmol) and TBDPSC1 (7.49 ml, 29.2 mmol) were added and the resulting reaction mixture was left under stirring overnight at room temperature. The mixture was diluted with H₂0 (300 ml) and extracted with EtOAc (5 x 50 ml). The combined organic phases were dried (Na₂S0₄), filtered and concentrated under reduced pressure to give a yellow oil. The residue was purified by flash

chromatography on silica gel (Biotage SP 340 g SNAP, from Cy 100 to Cy/EtOAc 90/10) to afford the title compound D12 (8.07 g) as a crude material which was used in the next step without any further purification. MS: (ES/+) m/z: 494 (M+l) and 516 (M+Na).

C₂₈H₃₅N0₃SSi requires 493. 1H-NMR (400 MHz, CDC1₃) 6(ppm): 7.69 (d, 2 H), 7.35 - 7.77 (m, 10 H), 7.24 (d, 2 H), 5.47 - 5.63 (m, 1 H), 5.01 (bs, 1 H), 4.96 - 5.00 (m, 1 H), 4.77 (bd, 1 H), 3.57 (dd, 1 H), 3.44 (dd, 1 H), 3.25 - 3.37 (m, 1 H), 2.43 (s, 3 H), 2.30 - 2.37 (m, 2 H), 1.05 (s, 9 H).

Description 13: N-[(lS)-l-({[(l,l-dimethylethyl)(diphenyl)silyl]oxy}methyl)-3-buten-l- yl]-4-methyl-N-2-propen-l-ylbenzenesulfonamide (D9):

To a solution of N-[(l,S)-l-({[(l,l-dimethylethyl)(diphenyl)silyl]oxy}methyl)-3-buten-l- yl]-4-methylbenzenesulfonamide D12 (8.07 g of the crude material obtained in the

Description 8) in DMF (30 ml), cesium carbonate (7.46 g, 22.9 mmol) and 3-bromo-l- propene (1.38 g, 11.4 mmol) were added and the mixture was stirred at room temperature overnight. The mixture was diluted with H₂0 (300 ml) and extracted with Et₂0 (5 x 50 ml). The combined organic phases were dried (Na₂S0₄), filtered and concentrated under reduced pressure. The residue was purified by flash chromatography on silica gel (Biotage SP 340 g SNAP, from Cy 100 to Cy/EtOAc 90/10) to afford the title compound D13 (7.46 g). MS: (ES/+) m/z: 534 (M+1) and 556 (M+Na). C₃₁H₃₉NO₃SS1 requires 533. 1H-NMR (400 MHz, CDCI₃) 6(ppm): 7.35 - 7.79 (m, 12 H), 7.20 (d, 2 H), 5.72 - 5.86 (m, 1 H), 5.47 - 5.62 (m, 1 H), 4.88 - 5.16 (m, 4 H), 3.90 - 4.05 (m, 2 H), 3.77 - 3.88 (m, 1 H), 3.59 - 3.71 (m, 2 H), 2.40 (s, 3 H), 2.38 - 2.51 (m, 1 H), 2.22 - 2.33 (m, 1 H), 1.04 (s, 9 H). Description 14: (lR,4S,6R)-4-({[(l,l-dimethylethyl)(diphenyl)silyl]oxy}methyl)-3- azabicyclo[4.1.0]heptane (D14)

To a solution of (lR,4S,6R)-4-({[(l,l-dimethylethyl)(diphenyl)silyl]oxy}methyl)-3-[(4- methylphenyl)sulfonyl]-3-azabicyclo[4.1.0]heptane D10 (3.6 g) in MeOH (500 ml), under a nitrogen atmosphere, magnesium (9.76 g, 402 mmol) (turnings, previously flame dried) and NH₄CI (10.37 g, 194 mmol) were subsequently added and the reaction mixture was vigorously stirred at 23 °C. After 2 hours further Mg (5 g) was added and the reaction mixture was stirred for other 2.5 hours. About 25 % of starting material was present and DCM (300 ml) and aqueous NH₄CI (saturated solution 200 ml) were added. The organic layer was separated and washed with brine (80 ml), filtered through a hydrophobic filter and evaporated under reduced pressure to give a colorless oil which was charged on a SCX (20 g) to afford the title compound D14 (1.81 g). UPLC (Acid IPQC): rtl = 1.00 minutes, peaks observed: 365 (M+1). C₂₃H₃iNOSi requires 364. 1H NMR (400 MHz, DMSO-^6) δ ppm 0.11 - 0.19 (m, 1 H) 0.50 - 0.60 (m, 1 H) 0.86 - 1.07 (m, 11 H) 1.40 - 1.56 (m, 2 H) 1.63 - 1.75 (m, 1 H) 2.23 - 2.37 (m,l H) 2.55 - 2.65 (m, 1 H) 3.43 - 3.51 (m, 2 H) 7.36 - 7.51 (m, 6 H) 7.55 - 7.67 (m, 4 H). Description 15: (lR,4S,6R)-4-({[(l,l-dimethylethyl)(diphenyl)silyl]oxy}methyl)-3-{[6- methyl-3-(2-pyrimidinyl)-2-pyridinyl] carbonyl}-3-azabicyclo [4.1.0] heptane (Dl 5)

To a solution of (lR,4S,6R)-4-({[(l,l-dimethylethyl)(diphenyl)silyl]oxy}methyl)-3- azabicyclo[4.1.0]heptane D14 (1.5 g) in dry DCM (30 ml) at room temperature under N₂ flux , 6-methyl-3-(2-pyrimidinyl)-2-pyridinecarboxylic acid D33 (2.94 ) and DIPEA (2.150 ml, 12.31 mmol) were added, followed by TBTU (1.534 g, 4.78 mmol), and the yellow suspension was left stirring at room temperature for 1.5 hour. Mixture was diluted with DCM and washed twice with NaHC0₃ saturated solution; aqueous phase was back- extracted with DCM and the collected organic phases were washed with water and brine. Organic layers were collected, dried over Na₂S0₄, filtered and evaporated; resulting dark green oil was purified by flash chromatography on KP- H column (SNAP 110 g eluting with Cy/EtOAc 1 : 1) affording the title compound D15 (1.79 g) as light yellow oil. 1H NMR (400 MHz, CDC1₃) δ ppm 0.46 - 0.54 (m, 1 H) 0.56 - 0.65 (m, 1 H) 0.73 - 0.98 (m, 2 H) 1.11 (s, 9 H) 1.79 - 1.87 (m, 1 H) 2.42 - 2.50 (m, 1 H) 2.60 (s, 3 H) 3.21 - 4.30 (m, 4 H) 4.66 - 4.78 (m, 1 H) 7.01 (t, 1 H) 7.19 - 7.80 (m, 11 H) 8.42 (d, 2 H) 8.49 (d, 1 H)

Description 16: ((lR,4S,6R)-3-{[6-methyl-3-(2-pyrimidinyl)-2-pyridinyl]carbonyl}-3- azabicyclo [4.1.0] hept-4-yl)methanol (Dl 6).

To a stirring solution of (lR,4S,6R)-4-({[(l,l-dimethylethyl)(diphenyl)silyl]oxy}methyl)-3- { [6-methyl-3-(2-pyrimidinyl)-2-pyridinyl]carbonyl}-3-azabicyclo[4.1.OJheptane D15 (1.79 g) in dry THF (25 ml) at room temperature under nitrogen flux, TBAF (3.50 ml, 3.50 mmol) was added slowly and the mixture was stirred at room temperature for 2 hours.

Mixture was diluted with EtOAc and washed with NH₄C1 saturated solution and brine; organic phases collected were dried (Na₂S0₄), filtered and evaporated, and the resulting crude purified by flash chromatography (on KP-Sil SNAP 100 g column eluting with DCM/MeOH 95:5), affording the title compound D16 (600 mg) as white foam. A second batch of the title compound D16 (295 mg, 0.909 mmol, 28.6 % yield) was obtained as slightly impure product. UPLC (Acid GEN QC SS): rtl = 0.58 minutes, peaks observed: 325 (M+l). Ci₈H₂₀N₄O₂ requires 324. 1H NMR (500 MHz, DMSO-d₆) δ ppm 8.80 - 8.94 (m, 2 H), 8.43 (d, 1 H), 7.39 - 7.52 (m, 2 H), 4.74 (t, 1 H), 4.22 - 4.30 (m, 1 H), 3.50 - 3.71 (m, 2 H), 3.45 (dd, 1 H), 3.18 - 3.23 (m, 1 H), 2.53 - 2.56 (m, 3 H), 2.15 - 2.24 (m, 1 H), 1.58 - 1.67 (m, 1 H), 0.83 - 1.10 (m, 2 H), 0.52 - 0.61 (m, 1 H), 0.41 - 0.47 (m, 1 H). Description 17: 2-[((lR,4S,6R)-3-{[6-methyl-3-(2-pyrimidinyl)-2-pyridinyl]carbonyl}- 3-azabicyclo[4.1.0]hept-4-yl)methyl]-lH-isoindole-l,3(2H)-dione (D17)

To a solution of ((lR,4S,6R)-3-{[6-methyl-3-(2-pyrimidinyl)-2-pyridinyl]carbonyl}-3- azabicyclo[4.1.0]hept-4-yl)methanol D16 (270 mg) in dry THF (7 ml), phthalimide (147 mg, 0.999 mmol) and triphenylphosphine (327 mg, 1.249 mmol) were added. Mixture was brought to 50 °C, then DIAD (0.243 ml, 1.249 mmol) was added dropwise and the solution was stirred at the same temperature for 1 hour. After cooling to room temperature 0.1 ml of water were added and volatiles were evaporated under reduced pressure; resulting crude was purified by flash chromatography on (KP-Sil column SNAP 25 g eluting with EtOAc 100%), affording the title compound D17 (297 mg) as white solid. UPLC (Acid

GEN_QC_SS): rtl = 0.75 minutes and rt2 = 0.82 minutes (rotamers present), peaks observed: 454 (M+l). C₂6H₂3N₅03 requires 453. 1H NMR (400 MHz, CDC1₃) δ ppm 0.37 - 0.43 (m, 1 H) 0.81 - 0.91 (m, 1 H) 0.98 - 1.31 (m, 2 H) 1.69 - 2.16 (m, 2 H) 2.43 (s, 3 H) 3.63 - 3.99 (m, 3 H) 4.23 - 4.37 (m, 1 H) 4.60 (d, 1 H) 7.03 (t, 1 H) 7.25 (d, 1 H) 7.61 - 7.92 (m, 4 H) 8.51 (d, 1 H) 8.56 (d, 2 H).

Description 18: [((lR,4S,6R)-3-{[6-methyl-3-(2-pyrimidinyl)-2-pyridinyl]carbonyl}-3- azabicyclo [4.1.0] hept-4-yl)methyl] amine (D 18)

2-[((lR,4S,6R)-3-{[6-methyl-3-(2-pyrimidinyl)-2-pyridinyl]carbonyl}-3- azabicyclo[4.1.0]hept-4-yl)methyl]-lH-isoindole-l,3(2H)-dione D17 (297 mg) was dissolved in EtOH (7 ml), then hydrazine (0.206 ml, 6.55 mmol) was added and the mixture was stirred at room temperature overnight. The morning after a white solid was precipitated and TLC (DCM/MeOH 9: 1) and UPLC showed reaction was complete. Solvent was removed at reduced pressure, the residue redissolved in MeOH and charged on a SCX cartridge (5 g) and the cartridge was eluted. Fractions containing desired product were evaporated and the residue purified by flash chromatography (on KP- H column SNAP 11 g eluting with EtOAc 100%) affording the title compound D18 (150 mg) as white foam. 1H NMR (500 MHz, DMSO-d₆) δ ppm 8.82 - 8.89 (m, 2 H), 8.39 (d, 1 H), 7.45 - 7.52 (m, 1 H), 7.40 - 7.46 (m, 1 H), 4.19 - 4.28 (m, 1 H), 3.45 (d, 1 H), 3.24 (d, 1 H), 2.85 - 2.93 (m, 1 H), 2.69 - 2.76 (m, 1 H), 2.53 (s, 3 H), 2.00 - 2.13 (m, 1 H), 1.65 - 1.75 (m, 1 H), 1.45 - 1.68 (m, 2 H), 0.90 - 1.00 (m, 1 H), 0.81 - 0.91 (m, 1 H), 0.47 - 0.60 (m, 2 H).

Description 19: (2S)-2-amino-4-penten-l-ol (D19)

In a 20 L reactor, to a suspension of (2S)-2-amino-4-pentenoic acid (available from Sigma - Aldrich #285013) (200 g, 1319 mmol) in THF dry (3200 ml) stirred under nitrogen at 0 °C was added a solution of LiAlFL (1600 ml, 1600 mmol) 1 M in THF dropwise in 1.5 hours (maintaining internal temperature between 0 °C and 5 °C). The reaction mixture was stirred at 25 °C for 2 hours (white suspension). The check by TLC (DCM/MeOH 1/1, AcOH 0.5% ninihydrine) showed reaction to be completed. The reaction mixture was cooled to 0 °C and was quenched by adding in sequence: 60.7 ml of water (1 ml H₂0 x 1 g of LiAlH₄) + 60.7 ml of NaOH 1 N (1 ml NaOH 1M x 1 g of LiAlH₄) + 182 ml of water (3 ml H₂0 x 1 g of LiAlFU). The suspension was stirred at room temperature for 1 hour then the precipitate was filtered over sodium sulphate (gooch n3) and washed with Et₂0 (6 L) and DCM (4 L). The solvent was evaporated (temperature bath 30 °C) of the crude title compound D19 (110 g) as pale-orange oil. MS: (ES/+) m/z: 102 (M+l). C₅HnNO requires 101. 1H NMR (400 MHz, DMSO-d₆) δ ppm 1.39 (br. s, 2 H) 1.81 - 1.96 (m, 1 H) 2.06 - 2.19 (m, 1 H) 2.59 - 2.73 (m, 1 H) 3.14 (dd, 1 H) 3.26 (dd, 1 H) 4.48 (br. s, 1 H) 4.91 - 5.09 (m, 2 H) 5.71 - 5.92 (m, 1 H)

Description 20: 1,1-dimethylethyl (2-{[(lS)-l-(hydroxymethyl)-3-buten-l-yl]amino}-2- oxoethyl)carbamate (non-preferred name) (D20)

In a 5 L reactor, to a solution of (2S)-2-amino-4-penten-l-ol D19 (110 g of the crude title compound prepared in the description D19) in THF (660 ml) and MeOH (440 ml) stirred at 0 °C (+5 °C internal) was added triethylamine (182 ml, 1305 mmol) and 2,5-dioxo-l- pyrrolidinyl N-{[(l,l-dimethylethyl)oxy]carbonyl}glycinate (available from Sigma -Aldrich #15423) (237 g, 870 mmol) portionwise over 15 min. The reaction mixture was stirred at 2 °C (internal temperature) for 3 hours. TLC check (TLC-NH₂, DCM/MeOH 95/5, potassium permanganate) showed residual starting material. Further 2,5-dioxo-l -pyrrolidinyl N-{[(1,1- dimethylethyl)oxy]carbonyl}glycinate (60 g, 220 mmol) was added and the mixture stirred at 2 °C for 1 hour. TLC check (TLC- H₂, DCM/MeOH 95/5, potassium permanganate) showed residual starting material. Further 2, 5-dioxo-l-pyrrolidinyl N-{[(1,1- dimethylethyl)oxy]carbonyl}glycinate (40 g, 146 mmol) was added and the mixture stirred at 2 °C for 1 hours. TLC check (TLC- H₂, DCM/MeOH 95/5, potassium permanganate) showed residual starting material but the work-up was carried out. The reaction mixture was poured into aqueous saturated solution of H₄CI (3400 ml) and EtOAc (1375 ml), then the phases were separated and the aqueous layer was back-extracted with EtOAc (1375 ml). The combined organic layers were washed with NaHC0₃ aqueous saturated solution (1031 ml) dried (Na₂S0₄) and evaporated to give crude material (268 g, deep brown). This residue was triturated with Et₂0 (687 ml) for 1 hour at 25 °C. The solid was filtered (gooch n3), washed with Et₂0 (200 ml) and dried under vacuum to give the title compound D20 (87 g) as pale brown solid. Mother liquors (deep brown) were evaporated and the residue chromatographed (Biotage 75 L, silica column, eluting with DCM/MeOH 98/2, 95/5) to give (34 g) of residual brown product that was triturated with Et₂0 (200 ml). The solid was filtered, washed with Et₂0 and dried under vacuum to give a further batch of the title compound D20 (26 g) as pale brown solid. MS: (ES/+) m/z: 259 (M+l). Ci₂H₂₂N₂0₄ requires 258. 1H MR (400 MHz, CDC1₃) δ ppm 1.47 (s, 9 H) 2.22 - 2.43 (m, 2 H) 2.68 - 2.83 (m, 1 H) 3.50 - 3.86 (m, 4 H) 3.94 - 4.09 (m, 1 H) 5.00 - 5.25 (m, 3 H) 5.64 - 5.89 (m, 1 H) 6.17 - 6.44 (m, 1 H)

Description 21: 1,1-dimethylethyl {2-[(4S)-2,2-dimethyl-4-(2-propen-l-yl)-l,3- oxazolidin-3-yl]-2-oxoethyl} carbamate (non-preferred name) (D21)

To a suspension of 1,1-dimethylethyl (2-{[(l S)-l-(hydroxymethyl)-3-buten-l-yl]amino}-2- oxoethyl)carbamate D20 (37 g) in toluene (370 ml) stirred at 25 °C were added 2,2- bis(methyloxy)propane (370 ml, 3020 mmol) and p-toluenesulfonic acid monohydrate (3.7 g, 19.45 mmol). The reaction mixture was stirred at reflux (85 °C internal, oil bath 105 °C) for 1.5 hour (clear solution). The check by TLC (DCM/MeOH 95/5) showed the reaction to be completed. The solvent was evaporated to obtain a brown oil that was chromatographed (Biotage 75 L, silica, eluting with Cy/EtOAc 8/2, 7/3) to give the title compound D21 (30 g) as yellow oil. UPLC (Acid GEN QC): rt = 0.69 minutes, peak observed: 299 (M+l).

Ci₅H₂₆N₂0₄ requires 298. 1H MR (400 MHz, CDC1₃) δ ppm 1.46 (s, 9 H) 1.54 (s, 3 H) 1.67 (s, 3 H) 2.30 - 2.49 (m, 2 H) 3.71 - 4.05 (m, 5 H) 5.04 - 5.22 (m, 2 H) 5.37 - 5.52 (m, 1 H) 5.65 - 5.81 (m, 1 H)

Description 22: trifluoromethanesulfonic acid - {2-[(4S)-2,2-dimethyl-4-(2-propen-l- yl)-l,3-oxazolidin-3-yl]-2-oxoethyl}amine (1:1) (D22)

To a solution of 1,1 -dimethyl ethyl {2-[(4S)-2,2-dimethyl-4-(2-propen-l-yl)-l,3-oxazolidin- 3-yl]-2-oxoethyl}carbamate D21 (28.67 g) in DCM (300 ml) 2,6-dimethylpyridine (27.9 ml, 240 mmol) was added followed by trimethylsilyl trifluoromethanesulfonate (34.7 ml, 192 mmol) the mixture was stirred at room temperature for 30 min. The reaction was quenched with 2 ml of water and the solvent was removed under reduced pressure, the residue was charged on a Biotage 75 L column eluting with [DCM/MeOH 100:0 then 98:2 then 96:4 then 85: 15]. Evaporation of the solvent gave: the title compound D22 (21 g). UPLC (Acid FINAL QC): rt = 0.36 minutes, peak observed: 199 (M+l- CHF₃0₃S)

CioHi₈N₂0₂-CHF₃0₃S requires 348. 1H NMR (400 MHz, DMSO-d₆) δ ppm 1.40 (s, 3 H), 1.50 (s, 3 H), 2.17-2.43 1.40 (m, 2 H), 3.68-3.98 (m, 4 H), 3.99-4.09 (m, 1 H), 4.83-5.40 (m, 2 H), 5.58-5.97 (m, 1 H), 7.63-8.36 (br.s., 2 H)

Description 23: (4S)-3-(diazoacetyl)-2,2-dimethyl-4-(2-propen-l-yl)-l,3-oxazolidine

{2-[(4S)-2,2-dimethyl-4-(2-propen-l-yl)-l,3-oxazolidin-3-yl]-2-oxoethyl}amine

trifluoromethansulfonate D22 (67.0 g) was dissolved in DCM (670 ml) and pH = 5 Buffer solution (670 ml) and cooled to 2 °C (internal). Sodium nitrite (26.5 g, 385 mmol) dissolved in water (134 ml) was added dropwise to the reaction mixture stirred at 2 °C over 30 min. The reaction mixture was stirred at 3 °C for 2.5 hours. Phases were separated. Water phase was back-extracted with DCM (1 x 670 ml, 1 x 335 ml). The combined organic layers, dried (Na₂S0₄), were evaporated (bath temperature 30 °C) to give 43 g of crude product. This crude was purified over silica pad [(230-400 Mesh) eluting with Cy/EtOAc 8/2, 7/3, 6/4] to give the title compound D23 (36.58 g) as pale yellow oil. UPLC (Acid GEN QC): rt = 0.59 minutes, peak observed: 210 (M+l) Ci₀Hi₅N₃O₂ requires 209. 1H NMR (400 MHz, CDC1₃) δ ppm 1.58 (s, 3 H) 1.69 (s, 3 H) 2.25 - 2.50 (m, 2 H) 3.43 - 3.70 (m, 1 H) 3.82 - 4.01 (m, 2 H) 4.84 (s, 1 H) 5.09 - 5.24 (m, 2 H) 5.63 - 5.84 (m, 1 H)

Description 24: (5aS,6aS,7aS)-3,3-dimethylhexahydro-5H- cyclopropa[</|[l,3]oxazolo[3,4- ]pyridin-5-one and (5aR,6aR,7aS)-3,3- dimethylhexahydro-5H-cyclopropa [d\ [1 ,3] oxazolo [3,4- ]pyridin-5-one

syn D24B anti)

A syn B anti

(4S)-3-(diazoacetyl)-2,2-dimethyl-4-(2-propen-l-yl)-l,3-oxazolidine D23 (36.5 g) dissolved in DCM (365 ml) was added dropwise at 25 °C to a suspension of rhodium(II) acetate dimer (3.85 g, 8.72 mmol) in DCM (183 ml) over 2.5 hours. The resulting mixture was stirred at 25 °C for 30 minutes. From TLC (Cy/ EtOAc 1/1): no more starting material. The mixture was filtered (gooch n 3), concentrated and chromatographed twice (over silica 230-400 Mesh, eluting with Cy/ EtOAc 7/3, 6/4) to give three fractions that after trituration with n- heptane (40 ml, for each fraction) gave the following three batches:

D24B D24A 95:3 (10.3 g, anti as major isomer anti/syn 95/3) HPLC (walk up): rtl = 3.09 rt2 = 3.14 minutes;

D24A D24B 31:68 (4.47 g, anti/syn roughly 31/68) HPLC (walk up): rtl = 3.05 rt2 = 3.11 minutes;

D24A D24B (10.5 g, D 24A syn as major isomer). HPLC (walk up): rtl = 3.08 rt2 = 3.16 minutes. 1H MR (500 MHz, CDC1₃) δ ppm 3.87 - 4.02 (m, 2 H), 3.32 (t, 1 H), 2.29 - 2.38 (m, 1 H), 1.57 (s, 3 H), 1.45 - 1.51 (m, 1 H), 1.43 (s, 3 H), 1.36 - 1.42 (m, 1 H), 1.12 - 1.20 (m, 1 H), 1.06 - 1.12 (m, 0 H), 0.45 - 0.54 (m, 1 H)

662 mg of this third batch of D24A D24B were taken and purified by flash chromatography (Snap-50 g silica gel column, EtOAc /Cy from 100 % Cy to 30:70). From this purification it was obtained a batch of almost pure cis isomer (the title compound D24A) (298 mg) as white solid, and a 347g batch of a mixture of cis/ trans isomers (75/25) as a colourless oil. UPLC (Basic GEN QC): rt = 0.48 minutes, peak observed: 182 (M+l). Ci₀Hi₅NO₂ requires 181. 1H MR (400 MHz, CDC1₃) δ ppm 3.98 - 4.10 (m, 2 H) 3.36 - 3.45 (m, 1 H) 2.37 - 2.47 (m, 1 H) 1.66 (s, 3 H) 1.53 - 1.61 (m, 1 H) 1.52 (s, 3 H) 1.42 - 1.50 (m, 1 H) 1.14 - 1.29 (m, 2 H) 0.59 (m, 1 H)

Description 25: (lS,4S,6S)-4-(hydroxymethyl)-3-azabicyclo[4.1.0]heptan-2-one (D25)

(5aS,6aS,7aS)-3,3-dimethylhexahydro-5H-cyclopropa[d][l,3]oxazolo[3,4-a]pyridin-5-one D24 (3.56 g) was dissolved in HC1 (25 ml, 150 mmol)(6 M in water) into a 250 ml-round bottomed flask and the mixture was stirred at 40 °C: after 4 hours the reaction was complete. The solvent was evaporated at reduced pressure using a rotavapor (bath temperature: 40 °C). The oily residue was stripped with toluene and the residue dried under high vacuum for 3 hours, obtaining the title compound D25 as white solid (2.843 g). UPLC (Acid IPQC): rt = 0.31 minutes, peak observed: 142 (M+l). C₇HnN0₂ requires 141. 1H NMR (500 MHz, DMSO-i¾) d ppm 0.56 - 0.68 (m, 1 H) 0.93 - 1.05 (m, 1 H) 1.30 - 1.39 (m, 1 H) 1.39 - 1.48 (m, 1 H) 1.57 - 1.67 (m, 1 H) 1.98 - 2.09 (m, 1 H) 3.17 - 3.29 (m, 2 H) 3.31 - 3.40 (m, 1 H) 6.89 - 7.13 (m, 1 H) Description 26: 1,1-dimethylethyl (lS,4S,6S)-4-(hydroxymethyl)-3- azabicyclo[4.1.0]heptane-3-carboxylate (D26)

(lS,4S,6S)-4-(hydroxymethyl)-3-azabicyclo[4.1.0]heptan-2-one D25 (3.839 g) was suspended in THF (40 ml) then BH₃.THF (1 M THF solution, 136 ml, 136 mmol) was added slowly (over 5 minutes) and the resulting mixture stirred at reflux for 2 hours. The mixture was cooled to room temperature and then to 0 °C using an ice/water bath. MeOH (25 ml) was slowly added and, when the gas evolution stopped, HCl (3 M water solution, 140 ml, 420 mmol) was slowly added and the resulting mixture was stirred again at 85 °C for 1 hour. The mixture was cooled again to room temperature.

A second reaction mixture was prepared: (lS,4S,6S)-4-(hydroxymethyl)-3- azabicyclo[4.1.0]heptan-2-one D25 (100 mg) was suspended in THF (0.5 mL), then BH₃.THF (3.6 mL, 3.60 mmol) was added slowly (over 1 minute) and the resulting mixture stirred at reflux for 2 hours. This second mixture was chilled to room temperature, then HCl (3.6 mL, 10.80 mmol) was slowly added and the resulting mixture was stirred again at 75°C for 1 hour. This mixture was chilled again to room temperature and then it was added to the first mixture to form a single mixture.

NaOH (3 M water solution, 140 ml, 420 mmol) was slowly added to the acidic mixture, then additional NaOH (50 ml, 150 mmol) was added in order to get a pH value of about 10. Boc₂0 (7.13 ml, 30.7 mmol) was added dissolved in THF (30 ml) and the resulting biphasic mixture was stirred vigorously at room temperature overnight. New Boc₂0 (4.57 ml, 19.70 mmol) was added dissolved in THF (20 ml) and the mixture stirred vigorously at room temperature for 1.5 hours. EtOAc (100 ml) was added to the mixture and the phases were separated. The water phase was extracted with EtOAc (3 x 100 ml) and all the organic fractions were mixed together. The so obtained organic solution was washed with brine (3 x 150 ml), dried over Na₂S0₄ and evaporated at reduced pressure, obtaining the crude target material as pale yellow oil (14 g). This material was purified by Biotage (Snap-340 g silica gel column, from pure Cy to EtOAc /Cy 70:30). It was obtained the title compound D26 (5.695 g) as colourless oil. MS: (ES/+) m/z: 228 (M+l) 128 (M+l-Boc). Ci₂H₂iN0₃ requires 227. 1H NMR (500 MHz, DMSO-d₆) δ ppm 4.43 (t, 1 H), 4.00 - 4.20 (m, 1 H), 3.34 - 3.45 (m, 1 H), 3.25 - 3.31 (m, 2 H), 2.10 - 2.23 (m, 1 H), 2.00 - 2.10 (m, 1 H), 1.30 (s, 9 H), 0.86 - 0.99 (m, 2 H), 0.66 - 0.77 (m, 1 H), 0.51 - 0.61 (m, 1 H), -0.04 - 0.05 (m, 1 H)

Description 27: 1,1-dimethylethyl (lS,4S,6S)-4-[(l,3-dioxo-l,3-dihydro-2H-isoindol-2- yl)methyl]-3-azabic clo[4.1.0]heptane-3-carboxylate (D27)

1,1-dimethylethyl (lS,4S,6S)-4-(hydroxymethyl)-3-azabicyclo[4.1.0]heptane-3-carboxylate D26 (310 mg), triphenylphosphine (1073 mg, 4.09 mmol) and phthalimide (502 mg, 3.41 mmol) were collected together and dissolved in THF (7 ml). The mixture was brought to 50 °C and then DIAD (0.796 ml, 4.09 mmol) was added dropwise. The mixture was stirred for 1 hour at 50 °C then 1 ml of water was added. Volatiles were removed in vacuum and the resulting crude was purified by silica gel chromatography (column size 10 g) using

Cy/EtOAc = 8:2 to Cy/EtOAc = 5:5 to EtOAc 100%. It was recovered the title compound D27 (488 mg). UPLC: (Acid Final_QC): rt = 0.80 minutes, peaks observed: 357 (M+1). C20H24N2O4 requires 356.

Description 28: 1,1-dimethylethyl (lS,4S,6S)-4-(aminomethyl)-3- azabicyclo [4.1.0] heptane-3-carboxylate (D28)

1,1-dimethylethyl (lS,4S,6S)-4-[(l,3-dioxo-l,3-dihydro-2H-isoindol-2-yl)methyl]-3- azabicyclo[4.1.0]heptane-3-carboxylate D27 (1.8 g) was dissolved in EtOH (8 ml) then hydrazine (2.476 ml, 50.5 mmol) was carefully added and the reaction stirred at room temperature for 2 hours. All volatiles were removed under vacuum and the solid residue was purified by SCX chromatography (column size 20 g using MeOH 100% to MeOH/NH3 2M). The product recovered showed a low purity so it was purified again by silica - H chromatography (Biotage SP - column size 28 g using EtOAc 100 % as eluent). It was recovered the title compound D28 (793 mg). UPLC: (Acid Final QC): rt = 0.46 minutes, peaks observed: 227 (M+1). C12H22N2O2 requires 226. 1H MR (400 MHz, CDC1₃) δ ppm 4.41 (br. s., 1H) 3.66 (br. s., 1H) 2.90-2.65 (m, 2H) 2.42-2.17 (m, 2H) 1.60-1.40 (m, 11 H) 1.21-1.08 (m, 1H) 1.06-0.84 (m, 2H) 0.81-0.68 (m, 1H) 0.30-0.04 (m, 1H). Description 29: N-[(lS,4S,6S)-3-azabicyclo[4.1.0]hept-4-ylmethyl]-5-(trifluoromethyl)-

1,1 -dimethyl ethyl (lS,4S,6S)-4-(aminomethyl)-3-azabicyclo[4.1.0]heptane-3-carboxylate D28 (50 mg), 2-chloro-5-(trifluoromethyl)-l,3,4-thiadiazole (41.7 mg, 0.221 mmol) and DIPEA (0.077 ml, 0.442 mmol) were reacted in DMSO (3 ml) at 80 °C for 2 hours, then solvent was removed under vacuum, and the crude redissolved in DCM (5 ml) and reacted with TFA (2 ml) at room temperature for 2 hours. Solvent was evaporated and the crude purified over an SCX (1 g, washing with MeOH and eluting with 2M ammonia in MeOH). The yellow oil obtained was then purified with Biotage SPl, over an 1 lg KP-NH SNAP column, eluting with a gradient of cyclohexane and IsPrOH. The title compound D29 was eluted with ca 15% iPrOH and recovered as light yellow solid (46 mg).

UPLC (GEN QC SS) rt = 0.45 minutes, peak observed: 279 (M+l) C₁₀H₁₃F₃N₄S requires 278.

1H NMR (400 MHz, CHLOROFORM-^ δ ppm 0.17 (q, 1 H) 0.71 (td, 1 H) 0.81 - 0.94 (m, 1 H) 1.00 - 1.14 (m, 1 H) 1.16 - 1.33 (m, 2 H) 2.05 (ddd, 1 H) 2.68 (m, 1 H) 2.97 (dd, 1 H) 3.09 - 3.31 (m, 2 H) 3.46 (dd, 1 H)

Description 30: N-[(lR,4S,6R)-3-azabicyclo[4.1.0]hept-4-ylmethyl]-5-(trifluoromethyl)- e

1,1 -dimethyl ethyl (lR,4S,6R)-4-(aminomethyl)-3-azabicyclo[4.1.0]heptane-3-carboxylate D4 (0.2 g), 2-chloro-5-(trifluoromethyl)-l,3,4-thiadiazole (0.167 g, 0.884 mmol) and DIPEA (0.309 ml, 1.767 mmol) were collected in DMSO (3 ml) and shaken at 80 °C for 2 hours. Solvent was removed under vacuum, and the resulting crude was dissolved in DCM (5 ml) and reacted at room temperature with TFA (2 ml). Solvent was removed, and the crude purified with 5 g SCX, washing with MeOH and eluting with 2M ammonia in MeOH. The resulting crude was purified with Biotage SPl (over a 25g KP-NH column, eluting with a gradient of cyclohexane and 2-propanol). The title compound D30 was eluted with 15% 2- propanol and recovered as yellow oil (0.2 g) UPLC (GEN QC SS): rt = 0.46 minutes, peak observed: 279 (M+l). C₁₀H₁₃F₃N₄S requires 278. 1H NMR (400 MHz, CHLOROFORM-<f) d ppm 0.22 (q, 1 H) 0.73 (td, 1 H) 0.98 - 1.14 (m, 2 H) 1.53 (ddd, 1 H) 1.91 (dd, 1 H) 2.48 - 2.58 (m, 1 H) 2.74 (dd, 1 H) 3.10 (dd, 1 H) 3.42 - 3.50 (m, 2 H) Description 31: 3-(5,5-Dimethyl-l,3,2-dioxaborinan-2-yl)-6-methyl-2- pyridinecarbonitrile (D31)

2,2,6,6-tetramethylpiperidine (3.49 ml, 20.52 mmol) was dissolved in dry THF (25ml) under argon and stirred at -30 °C; BuLi (13.33 ml, 21.33 mmol) 1.6 M in hexane was added over 5 min (the temperature never exceeded -25 °C). The yellow solution was stirred at -30 °C for 20 min, then chilled at -78 °C and tris(l-methylethyl) borate (4.38 ml, 18.96 mmol) was added over 5 min (the temperature never exceeded -73 °C).

After 10 min at -78 °C, 6-methyl-2-pyridinecarbonitrile (2.0 g, 16.93 mmol) dissolved in dry THF (14 ml) was added dropwise (over 20 min) maintaining internal temperature below -73 °C and the mixture became dark-brown. The mixture was stirred at -73 °C for 2 hours. The mixture was quenched with AcOH (2.374 ml, 41.5 mmol) dropwise at -73 °C (the temperature never exceeded -60 °C and the mixture became brilliant orange). The cooling bath was removed and the mixture left to reach the room temperature: during this period the mixture became thick and new THF (8 ml) had to be added in order to have a better stirring. The mixture was stirred 10 min at room temperature then 2,2-dimethyl- 1,3 -propanediol (2.409 g, 23.13 mmol) was added in one portion and the mixture stirred at room temperature overnight. The solvent was evaporated and the orange residue taken-up with DCM (100 ml) and 10 % water solution of KH₂PO₄ (100 ml). The phases were separated and the water phase was back-extracted with DCM (50 ml). The combined organic phases were washed with 10 % water solution of KH₂P0₄ (50 ml). The DCM was evaporated. The residue was dissolved in Et₂0 (100 ml) and extracted with NaOH 0.05 M (5 x 50 ml, boronic ester in water phase). The aqueous phases were joined together and the pH was adjusted between pH = 4 and pH = 5 with 10 % water solution of KH₂P0₄ (50 ml). The so obtained yellow solution was extracted with EtOAc (3 x 200 mis). All the organics joined together were dried (Na₂S0₄) and evaporated the title compound D31 (2.29 g) of as yellow oil, that solidified on standing. C₁₂H₁₅BN₂O₂ requires 230. 1H MR (400 MHz, CDC1₃) δ ppm 7.97 - 8.15 (m, 1 H), 7.31 - 7.36 (m, 1 H), 3.85 (m, 4 H), 2.52 - 2.73 (s, 3 H), 0.97 - 1.10 (m, 6 H).

Description 32: 6-Methyl-3-(2-pyrimidinyl)-2-pyridinecarbonitrile (D32)

A) Isopropylmagnesium chloride-LiCl (37.9 ml, 36.5 mmol) was added portion wise (in overall 10 min) to a solution of 3-bromo-6-methyl-2-pyridinecarbonitrile (4 g, 20.30 mmol) in THF (150 ml) cooled to -70 °C (internal temperature). The reaction was kept to that temperature for 15 min. Then it was allowed to gently warm up to -40 °C in overall 1 hour. Then, it was cooled to -78 °C and zinc chloride (3.32 g, 24.36 mmol) was added. The resulting mixture was allowed to warm up to room temperature in 1 hour. Pd(Ph₃P)₄ (2.346 g, 2.030 mmol), 2-chloropyrimidine (3 g, 26.2 mmol) were added and the mixture was refluxed (external temperature 100 °C) until complete consumption of starting

chloropyrimidine (3 hours). The reaction mixture was cooled to room temperature and poured into water (200 ml) cooled to 10 °C. It was then extracted with EtOAc (5 x 200 ml). The collected organic phases, containing large amount of colloid material and water, were washed with brine (200 ml). The water phase was filtered over a gouch, and the solid material was washed with further EtOAc (2 x 300ml). The collected organic phases were dried overnight over Na₂S0₄, filtered and concentrated to give the crude material (7 g) which was purified (Biotage Spl over a 240 g Silica Analogix column, with a 25 g pre- column) to give the title compound D32 as yellow solid (1.8 g). UPLC (Acid

GEN_QC_SS): rt = 0.58 minutes, peak observed: 197 (M+l). CnH₈N₄ requires 196. B) An alternative method to make D32 is: 3-(5,5-dimethyl-l,3,2-dioxaborinan-2-yl)-6- methyl-2-pyridinecarbonitrile D31 (50.6 mg) was dissolved 1,4-Dioxane (1 ml) under nitrogen in a vial, then 2-bromopyrimidine (42.0 mg, 0.264 mmol), CsF (67 mg, 0.441 mmol), Pd(Ph₃P)₄ (12 mg, 10.38 μιηοΐ) and Cul (7 mg, 0.037 mmol) were added in sequence. The vial was then capped and stirred at 65 °C, after 1 hour the solvent was removed at reduced pressure and the residue partitioned between EtOAc (10 ml) and

NaHC0₃ (saturated solution, 10 ml). The phases were separated and the water was extracted with EtOAc (2 x 10 ml). The organic fraction were joined together, dried over Na₂S0₄ and evaporated at reduced pressure, obtaining an orange oily residue which was purified (Biotage, Snap 25 g silica gel column, EtOAc/Cy from pure Cy to 50:50 in 10 column volumes) to obtain the title compound D32 as pale yellow solid (27.6 mg).

Description 33: 6-Methyl-3-(2-pyrimidinyl)-2-pyridinecarboxylic acid (D33)

A) 6-methyl-3-(2-pyrimidinyl)-2-pyridinecarbonitrile D32 (0.8 g) was reacted in 6 M aqueous HC1 (40 ml, 240 mmol) at 80 °C for 3 hours, then solvent was removed under vacuum, and the resulting crude was purified (70 g Varian CI 8 column conditioning with MeOH (120 ml), then water (120 ml), loading in water, washing with water (200 ml), product eluted with 100 % MeOH) to give the title compound D33 (0.6 g) as yellow solid. UPLC (Acid GEN_QC_SS): rt = 0.30 minutes, peak observed: 216 (M+l). CnH₉N₃0₂ requires 217. 1H MR (400 MHz, DMSO-i¾) δ ppm 13.07 (bs, 1 H), 8.78 - 9.01 (m, 2 H), 8.39 (m, 1 H), 7.39 - 7.67 (m, 2 H), 2.56 - 2.67 (s, 3 H).

B) An alternative method to make D33 is as follows: 6-methyl-3-(2-pyrimidinyl)-2- pyridinecarbonitrile D32 (0.481 g) was suspended in EtOH (5 ml) and a solution of NaOH (0.490 g, 12.26 mmol) in water (5 ml) was added. The yellow mixture was stirred at 100 °C overnight. The yellow solution was cooled to 25 °C and HC1 6 M (1.0 ml) was added dropwise till pH = 4.5. The solvent was removed to give a yellow powder that was dried at 50 °C/vacuum for 1.5 hours to give the title compound D33 (1.242 g).

Description 34: 2-methylfuro [3,4-b]pyridine-5,7-dione (D34)

In a 100 ml round-bottomed flask 6-methyl-2,3-pyridinedicarboxylic acid (10 g, 55.2 mmol) and acetic anhydride (26 ml, 276 mmol) were added and heated at 100 °C under nitrogen for 5 hours. After this time the volatiles were removed under vacuum to give the title compound D34 (8.2 g) as a slightly brown solid. 1H NMR (400 MHz, DMSO-i¾) δ ppm 8.41 (d, 1 H), 7.82 (d, 1 H), 2.73 (s, 3 H).

Description 35: 6-methyl-2-[(methyloxy)carbonyl]-3-pyridinecarboxylic acid (D35)

2-methylfuro [3,4-b]pyridine-5,7-dione D34 (3 g) was added portionwise over 5 minutes to stirred MeOH (20 ml) at 0 °C. The mixture was stirred at 0 °C for 30 minutes then at room temperature for other 2.5 hours. The solution was evaporated at reduced pressure and the residue recrystallized from toluene (50 ml). The solid was filtered and dried under high vacuum for 30 minutes, obtaining a first batch of the title compound D35 (1.16 g) as pale brown solid. From the toluene solution new solid precipitated: this solid was filtered and dried under high vacuum for 30 minutes, obtaining a second batch of the title compound D35 (352 mg) as pale yellow solid. The toluene solution was then evaporated at reduced pressure and the residue recrystallized again from toluene (25 ml). The solid was filtered and dried under high vacuum for 30 minutes, obtaining a third batch of the title compound D35 (615 mg) as pale yellow solid. UPLC (Basic GEN QC): rt = 0.23 minutes, peak observed: 195 (M+1). C₉H₉N0₄ requires 196. 1H MR (400 MHz, DMSO-i¾) δ ppm 13.61 (br. s., 1 H), 8.09 - 8.31 (m, 1 H), 7.51 (m, 1 H), 3.82 (s, 3 H), 2.55 (s, 3 H). Description 36: methyl 3-({[(l,l-dimethylethyl)oxy]carbonyl}amino)-6-methyl-2- pyridinecarboxylate (D36)

6-methyl-2-[(methyloxy)carbonyl]-3-pyridinecarboxylic acid D35 (1.15 g) was suspended in toluene (40 ml) and DIPEA (1.25 ml, 7.16 mmol) was added, causing the complete dissolution of the solid. This mixture was stirred 10 minutes at room temperature, then diphenyl azidophosphate (1.35 ml, 6.26 mmol) was added in one portion and the mixture was stirred at reflux for 1 hour. The solution was cooled at room temperature and t-BuOH (2.5 ml, 26 mmol) was added in one portion. The mixture was then stirred at 70 °C for 1 hour and then cooled at room temperature, Et₂0 (50 ml) was added and the resulting solution washed with NaHC0₃ saturated solution (3 x 60 ml). The water phases were joined together and back-extracted with Et₂0 (50 ml). The two organic solutions were j oined together, dried over Na₂S0₄ and evaporated at reduced pressure, obtaining the crude target material as pale yellow oil. This material was purified by flash chromatography on silica gel (Biotage, EtOAc/Cy from 10/90 to 70/30; Snap-100 g column). The title compound D36 (1.315 g) was obtained as white solid. UPLC (Basic GEN QC): rt = 0.68 minutes, peak observed: 267 (M+1). Ci₃Hi₈N₂0₄ requires 266. 1H NMR (400 MHz, CDC1₃) δ ppm 10.13 (bs., 1 H), 8.77 (d, 1 H), 7.34 (d, 1 H), 4.03 (s, 3 H), 2.59 (s, 3 H), 1.53 - 1.56 (m, 9 H).

Description 37: methyl 3-amino-6-methyl-2-pyridinecarboxylate (D37)

Methyl 3-({[(l,l-dimethylethyl)oxy]carbonyl}amino)-6-methyl-2-pyridinecarboxylate D36 (1.3 g) was dissolved in DCM (80 ml) and the mixture stirred at 0 °C. A solution of TFA (5 ml, 64.9 mmol) in DCM (10 ml) was dropped into the cold mixture over 3 minutes. The resulting solution was left under stirring at 0 °C for 30 minutes, then the mixture was left still at room temperature overnight. TFA (4 ml, 51.9 mmol) dissolved in DCM (10 ml) was added over 3 minutes and the mixture stirred again at room temperature for 5 hours. The solution was loaded onto an SCX-25 g column and the column was eluted firstly with DCM (100 ml) and then MeOH (20 ml). The material was collected eluting with NH₃ (2M in MeOH, 100 mis) and after evaporation under reduced pressure of the ammonia solution it was obtained the title compound D37 (770 mg) as a white solid. UPLC (Basic GEN QC): rt = 0.44 minutes, peak observed: 167 (M+1). C₈Hi₀N₂O₂ requires 166. 1H NMR (400 MHz, CDC1₃) δ ppm 7.14 (d, 1 H), 7.01 (d, 1 H), 3.99 (s, 3 H), 2.52 (s, 3 H).

Description 38: methyl 3-iodo-6-methyl-2-pyridinecarboxylate (D38)

HC1 6 M solution in water (4.5 ml, 27.0 mmol) was added to methyl 3-amino-6-methyl-2- pyridinecarboxylate D37 (768 mg) and the resulting pale yellow mixture was sequentially diluted with water (4 x 5 ml) and chilled at 0 °C (internal temperature).

A solution of sodium nitrite (480 mg, 6.96 mmol) in water (2 ml) was dropped into the mixture over 1 minute. After this addition the mixture was stirred at 0 °C for 30 minutes, then a solution of KI (1.69 g, 10.18 mmol) in water (2 ml) was added over 1 minute, causing the formation of a dark violet crust (moderate gas evolution). The mixture was left under stirring for 1 hour: during this period the temperature passed from 0 °C to + 5 °C. EtOAc (50 ml) was then added to the stirred mixture, causing the dissolution of the dark solid. Water (50 ml) and EtOAc (50 ml) were added and the whole mixture was poured into a separator funnel. After the separation of the two phases, the water phase was extracted with EtOAc. All the organic phases were joined together and washed with NaHC0₃ saturated solution; the acidic water phase was neutralized by the addition of the previously used NaHC0₃ saturated solution and the resulting mixture extracted with EtOAc (2 x 50 ml). All the organic phases were joined together, dried over Na₂S0₄ and evaporated at reduced pressure, obtaining the crude target material as dark brown/violet oil. This material was purified by silica gel chromatography (Biotage SP4 Snap-100 g column, EtOAc /Cy from 10/90 to 30/70). The title compound D38 was obtained as a pale brown solid (1.1 g). UPLC (Basic GEN QC): rt = 0.68 minutes, peak observed: 278 (M+l). C₈H₈IN0₂ requires 277. 1H MR (400 MHz, CDC1₃) δ ppm 8.12 (d, 1 H), 7.01 (d, 1 H), 4.01 (s, 3 H), 2.58 (s, 3 H).

Description 39: methyl 6-methyl-3-(2H-l,2,3-triazol-2-yl)-2-pyridinecarboxylate (D39)

DMF (1.5 ml) was added to a mixture of methyl 3-iodo-6-methyl-2-pyridinecarboxylate D38 (100 mg), lH-l,2,3-triazole (49.9 mg, 0.722 mmol), (lR,2R)-N,N'-dimethyl-l,2- cyclohexanediamine (10.27 mg, 0.072 mmol), Cul (3.44 mg, 0.018 mmol) and Cs₂C0₃ (235 mg, 0.722 mmol) in a microwave vial. The mixture was degassed via three vacuum/nitrogen cycles then irradiated in a single mode microwave reactor to 120 °C for 20 minutes. The mixture was irradiated in a single mode microwave reactor to 120 °C for a further 40 minutes. The reaction mixture was cooled and filtered washing the solids with EtOAc (20 ml). The solids were dissolved in ph=3 buffer solution (5ml); UPLC check of this aqueous solution showed that it contained a considerable quantity of 6-methyl-3-(2H-l,2,3-triazol-2- yl)-2-pyridinecarboxylic acid. The aqueous phase was extracted repeatedly with DCM; the combined DCM extracts were diluted with MeOH (50ml) and treated with TMS- diazomethane. The volatiles were evaporated to give a yellow residue that was purified by flash chromatography on silica gel (Biotage, SNAP lOg column, 10%-50% EtOAc/Cy) to give the title compound D39 (38 mg) as a white solid. UPLC (Basic QC POS 50-800): rt = 0.57 minutes, peak observed: 219 (M+l). C₁₀H₁₀N₄O₂ requires 218. 1H NMR (400 MHz, CDCI₃) δ ppm 8.20 (d, 1 H), 7.87 (s, 2 H), 7.44 (d, 1 H), 3.94 (s, 3 H), 2.71 (s, 3 H).

Description 40A/40B: 6-methyl-3-(2H-l,2,3-triazol-2-yl)-2-pyridinecarboxylic acid

A) A solution of methyl 6-methyl-3-(2H-l,2,3-triazol-2-yl)-2-pyridinecarboxylate D39 (36 mg) and LiOH (5.93 mg, 0.247 mmol) in THF/water (2: 1, 3 ml) was stirred overnight. The mixture was evaporated under reduced pressure; the residue was taken up in water (2 ml) and neutralised with 1 M HCl water solution and then loaded onto a pre- conditioned CI 8 5 g column (the column was eluted with water and then MeOH). The methanol fractions were evaporated under reduced pressure to give the title compound

D40A (34 mg) as a white solid. UPLC (Basic QC POS 50-800): rt = 0.30 minutes, peak observed: 205 (M+l). C₉H₈N₄0₂ requires 204. 1H NMR (400 MHz, MeOD) δ (ppm) 8.24 (d, 1 H), 7.99 (s, 2 H), 7.61 (d, 1 H), 2.67 (s, 3 H).

B) An alternative method to prepare D40 is as follows: (N10902-73-1)

6-methyl-3-(2H-l,2,3-triazol-2-yl)-2-pyridinecarbonitrile D46 (1 g) in EtOH (10 mL) was treated with 6M sodium hydroxide (5.40 mL, 21.60 mmol) and the resulting pale yellow solution was heated at 100°C (external bath temperature, internal reflux) for 14 hours. UPLC check confirmed reaction was over, thus reaction mixture was cooled down to room temperature and acidified with aqueous HCl 6M to pH 4/5. Resulting slurry was evaporated to dryness and the obtained yellow solid was triturated with EtOH (10ml) and dried at high vacuum at 50°C for 1 hour to get the title compound D40B (2.85 g) as white solid. Mother liquor were evaporated to dryness to get D40B (0.1 g) as pale yellow solid.

UPLC (IPQC): rt = 0.42 minutes peak observed: 205 (M+l) C₉H₈N₄0₂ requires 204. Description D41: methyl 6-methyl-3-(lH-pyrazol-l-yl)-2-pyridinecarboxylate (D41)

DMF (1.5 ml) was added to a mixture of methyl 3-iodo-6-methyl-2-pyridinecarboxylate D38 (200 mg), lH-pyrazole (98 mg, 1.444 mmol), (lR,2R)-N,A imethyl-l,2- cyclohexanedi amine (20.54 mg, 0.144 mmol), bis(copper(I) trifluoromethanesulfonate), benzene complex (18.17 mg, 0.036 mmol) and cesium carbonate (470 mg, 1.444 mmol) in a screw-topped vial. The mixture was degassed via 3 vacuum/nitrogen cycles and heated with shaking to 120 °C for 1 hour. The reaction mixture was evaporated to dryness under reduced pressure. The residue was dissolved in water/MeOH (1 : 1, 3 ml) and acidified to pH=2 by addition of 4 M HC1 solution. The resulting mixture was evaporated to dryness under reduced pressure then the residue was triturated with DCM/MeOH (3 : 1, 20 ml). The mixture was filtered washing with more DCM/MeOH (3: 1, 5 ml). The filtrate was treated with TMS-diazomethane solution (2 M in hexanes, 2 ml, 4 mmol) to re-esterify the acid. The reaction mixture was evaporated under reduced pressure and the residue was purified twice by flash chromatography on silica gel (Biotage Snap 10 g column, EtOAc/Cy from 20/80 to 50/50 and then Biotage KP- H Snap 11 g column, EtOAc/DCM isocratic 1/99) to give the title compound D41 (107 mg) as a colourless gum.

UPLC (Basic QC POS 50-800): rt = 0.51 minutes, peak observed: 218 (M+l). C₁₁H₁₁N₃O₂ requires 217. 1H MR (400 MHz, CDCl₃) δ ppm 7.63 - 7.86 (m, 3 H), 7.39 (m, 1 H), 6.48 (m, 1 H), 3.85 (s, 3 H), 2.68 (s, 3 H). Description D42: 6-methyl-3-(lH-pyrazol-l-yl)-2-pyridinecarboxylic acid (D42)

A solution of methyl 6-methyl-3-(lH-pyrazol-l-yl)-2-pyridinecarboxylate D41 (106 mg) and LiOH (17.53 mg, 0.732 mmol) in THF/water (2: 1, 6 ml) was stirred overnight. The mixture was evaporated under reduced pressure; the residue was taken up in water (2 ml) and the pH was adjusted to pH=2 with 1 M HC1 solution. The mixture was loaded onto a pre-conditioned C18 column (5 g, eluted with water and then MeOH). The methanol fractions were evaporated under reduced pressure to give the title compound D42 (98 mg) as a white solid.

UPLC (Basic QC POS 50-800): rt = 0.30 minutes, peak observed: 160 [(M-C0₂)+1]. CioH₉N₃0₂ requires 203. 1H MR (400 MHz, methanol-^) δ ppm 7.77 - 8.03 (m, 2 H), 7.74 (m, 1 H), 7.58 (m, 1 H), 6.55 (m, 1 H), 2.66 (s, 3 H).

Description 43: 2-(2H-l,2,3-triazol-2-yl)benzoic acid (D43)

2-iodobenzoic acid (1 g, 4.03 mmol), 2H-l,2,3-triazole (0.334 g, 4.84 mmol), copper(I) iodide (0.768 g, 4.03 mmol), cesium carbonate (2.63 g, 8.06 mmol) were reacted in DMF at 120 °C under microwave irradiation for 20 minutes. Then the reaction was monitored confirming the presence of the required product. Large amount of starting material resulted left. The reaction was further irradiated for 20 minutes at 120 °C. Solvent was then removed under vacuum, poured into water (100 ml) and neutralised with 37% HC1. Aqueous phase was extracted with Et₂0 (3x100 ml), and the collected organic phases were concentrated to give 1 g of crude material. This was purified with Biotage SP1 (over a 340g C18 column) eluting with a gradient of ACN and water (modified with 0.5% HCOOH). Fractions were collected and concentrated under vacuum to give the title compound D43 as colourless solid (140 mg). UPLC (GEN QC Basic): rt = 0.29 minutes, peak observed: 188 (M-l) C₉H₇N₃0₂ requires: 189. 1H NMR (400 MHz, CHLOROFORM-<f) δ ppm 6.47 (br. s., 1 H) 7.47 - 7.59 (m, 1 H) 7.66 (t, 1 H) 7.75 - 7.87 (m, 3 H) 7.91 (d, 1 H)

Description 44: 5-methyl-2-(2H-l,2,3-triazol-2-yl)benzoic acid (D44)

To 2-iodo-5-methylbenzoic acid (1.5 g, 5.72 mmol), copper(I) iodide (1.090 g, 5.72 mmol) and (lR,2R)-N,N'-dimethyl-l,2-cyclohexanediamine (0.814 g, 5.72 mmol) was added DMF (10 ml) under nitrogen and then 2H-l,2,3-triazole (0.474 g, 6.87 mmol). The reaction mixture was heated at 120 °C at the microwave Personal Chemistry for 15 minutes. UPLC- MS monitor showed that the reaction was complete. After cooling at room temperature water was added and the aqueous washed with EtOAc. Then the aqueous acidified to pH =1 with aqueous 1 M HC1 and extracted with EtOAc. The phases were separated on a hydrophobic frit, the combined organic solvent was removed to give the crude product (2.4 g).

The crude compound was purified by silica gel chromatography (SNAP KP-Sil lOOg cartridge) eluted with 5 CV of a mobile phase prepared with DCM (959.04 ml), acetic acid (0.96 ml) and MeOH (40 ml). The evaporation of proper fractions gave a yellow oil which was triturated with toluene (4 x 50 ml) and the title compound D44 was obtained as yellow solid (540 mg).

UPLC (Basic GEN QC): rt = 0.35 minutes, peak observed: 204 (M+l) Ci₀H₉N₃O₂ requires: 203.

1H NMR (400 MHz, DMSO-i¾) δ ppm 2.44 (s, 3 H) 7.52 (dd, 1 H) 7.59 (s, 1 H) 7.64 (d, 1 H) 8.06 (s, 2 H).

Description 45: 6-methyl-3-(lH-l,2,3-triazol-l-yl)-2-pyridinecarbonitrile (D45) and description 46: 6-methyl-3-(2H-l,2,3-triazol-2-yl)-2-pyridinecarbonitrile (D46)

D45 D46

A solution of 3-bromo-6-methylpyridine-2-carbonitrile (3 g, 15.23 mmol), lH-l,2,3-triazole (2.103 g, 30.5 mmol), (lR^^-^jV-dimethyl-l^-cyclohexanediamine (0.433 g, 3.05 mmol), cesium carbonate (9.92 g, 30.5 mmol) in dry DMF (30 ml) was degassed with argon bubbling for 5 minutes. Then bis(copper(I)trifluoromethanesulfonate)benzene complex (0.383 g, 0.761 mmol) was finally added and the resulting mixture was further de-gassed with argon bubbling for 30 seconds. Reaction mixture was heated at 120°C for 4 hours. Reaction mixture was cooled down to room temperature and water (50 ml) was added. Mixture was taken up with EtOAc (100 ml). Emulsion formed, separation of phases was difficult, so further water 50 ml and EtOAc (50 ml) were added but separation still remained very difficult. Aqueous layer was back extracted with EtOAc (2x50 ml). Combined organic phases were washed with brine (30ml) and evaporated to dryness to get crude material (3.7 g) as thick brown oil that was purified over Si0₂ Biotage SNAP lOOg (eluting with

Cy/EtOAc 9/1 to 6/4). Evaporation of solvent afforded 3 fractions:

6-methyl-2-pyridinecarbonitrile (0.4 g, 3.39 mmol, 22.24 % yield) as white solid;

UPLC (IPQC): rt = 0.60 minutes peak observed: 119 (M+l);

D45 6-methyl-3-(lH-l,2,3-triazol-l-yl)-2-pyridinecarbonitrile (0.68 g) as white solid UPLC (IPQC): rt = 0.58 minutes peak observed: 186 (M+l) C₉H₇N₅ requires 185.

1H NMR (400 MHz, CHLOROFORM-^ δ ppm 2.74 (s, 3 H) 7.62 (d, 1 H) 7.97 (d, 1 H) 8.22 (d, 1 H) 8.39 (d, 1 H)

D46 6-methyl-3-(2H-l,2,3-triazol-2-yl)-2-pyridinecarbonitrile (1 g) as white solid

UPLC (IPQC): rt = 0.74 minutes, peak observed: 186 (M+l) C₉H₇N₅ requires 185

1H NMR (400 MHz, CHLOROFORM-^ δ ppm 2.71 (s, 3 H) 7.54 (d, 1 H) 7.99 (s, 2 H) 8.34 (d, 1 H)

Description 47: 6-methyl-3-(lH-l,2,3-triazol-l-yl)-2-pyridinecarboxylic acid sodium salt (D47)

6-methyl-3-(lH-l,2,3-triazol-l-yl)-2-pyridinecarbonitrile D45 (650 mg) was reacted with NaOH (2.340 ml, 14.04 mmol) in EtOH (2 ml) at 100 °C for 2 hours. Solvent was removed under vacuum and the crude was dissolved in 15 ml of water and neutralised with 37% HCl to pH 6-7. Water was removed under vacuum to give the title compound D47 as colourless solid (2.2 g). This compound was contaminated by NaCl and purity was estimated considering the theoretical amount of product deriving from a quantitative reaction.

UPLC (Final QC_neg): rt = 0.32 minutes, peak observed: 203 (M-l) C₉H₉N₄0₂ requires 204. C₉H₈N₄0₂-Na+ requires 227. escription 48: l,l₉l-trifluoro-3-(phenylsulfonyl)-2-propanol (D48)

NaH (3.84 g, 96 mmol) was added portionwise to a solution of (methylsulfonyl) benzene (10 g, 64.0 mmol) in THF (200 ml) cooled to 0 °C. The resulting mixture was stirred at that temperature for 30 minutes, then ethyl trifluoroacetate (18.19 g, 128 mmol) was added and the mixture was refluxed for 3 hours. Then the resulting solution was cooled to room temperature and poured into Et₂0 (1 1) and washed with brine (2x150 ml), dried over Na₂S0₄, filtered and concentrated. The resulting yellow foam (20 g) was dissolved in MeOH (200 ml) and cooled to 0°C. NaBH₄ (2.422 g, 64.0 mmol) was added portionwise and the mixture was allowed to warm up to room temperature and stirred overnight at room temperature. Solvent was removed under vacuum, and the crude was poured in brine (300 ml) and extracted with Et₂0 (3x300 ml). The collected organic phases were washed with brine (150 ml), dried over Na₂S0₄, filtered and concentrated. The colourless solid (16 g) was dissolved in Et₂0 (100 ml), then cyclohexane was added (100 ml) and the solution was left at -4 °C overnight. The resulting solid was filtered to give the title compound D48A (7.4 g). The solution was concentrated and then suspended in cyclohexane (100 ml), warmed to 60 °C and quickly filtered. The resulting cloudy mixture was warmed to 40°C and Et₂0 added until a clear solution was obtained. This solution was left standing at room temperature and then at 4 °C to give, after filtration, the title compound D48B (4 g).

UPLC (GEN QC SS) rt = 0.71 minutes, peak observed: 255 (M+l), C₉H₉F₃O₃S requires 254.

1H NMR (400 MHz, CHLOROFORM-^ δ ppm 3.35 - 3.57 (m, 2 H) 3.65 - 3.91 (m, 1 H) 4.55 - 4.75 (m, 1 H) 7.56 - 7.71 (m, 2 H) 7.71 - 7.84 (m, 1 H) 7.94 - 8.09 (m, 2 H)

Description 49: {[(l£)-3,3,3-trifluoro-l-propen-l-yl]sulfonyl}benzene (D49)

p-toluenesulfonyl chloride (8.55 g, 44.8 mmol) was added to a solution of l,l,l-trifluoro-3- (phenylsulfonyl)-2-propanol D48A and D48B (11.4 g) and TEA (12.50 ml, 90 mmol) in DCM (200 ml). The resulting mixture was refluxed for 3 hours, then DBU (6.76 ml, 44.8 mmol) was added and the mixture further stirred under reflux for 2 hours.

The reaction was then poured into brine (200 ml). The aqueous phase was extracted with DCM (3x200) and the collected organic phases (800 ml) were washed with brine (2x200 ml) and 1M aqueous HC1 (3x200 ml), dried overNa₂S0₄, filtered and concentrated. Several attempts to cristallize the product from Cy, Cy/Et₂0, Cy/EtOAc were unsuccessful and the crude was purified with Biotage SP1 (over a 150 g Analogix column, with a gradient of Cy and EtOAc). The title compound D49 was recovered as colourless solid (5.9 g) 1H MR (400 MHz, CHLOROFORM-^ δ ppm 6.84 (dq, 1 H) 6.99 - 7.09 (m, 1 H) 7.60 - 7.67 (m, 2 H) 7.69 - 7.77 (m, 1 H) 7.89 - 8.02 (m, 2 H)

Description 50: 5-(trifluoromethyl)-l,3-thiazol-2-amine (D50)

tert-butyl hydroperoxide (1.422 ml, 7.11 mmol) was added dropwise to a solution of BuLi (4.45 ml, 7.11 mmol) in THF (20 ml) cooled to -78 °C (small exotermic effect). The resulting solution was stirred at that temperature for 1 hour, then a solution of {[(1Ε)-3,3,3- trifluoro-l-propen-l-yl]sulfonyl}benzene D49 (1.4 g) in THF (20 ml) was added; the mixture was stirred at -78°C for 1 hour and then allowed to gently warm up to 0 °C in 1 hour. The reaction was quenched at 0 °C with 10 ml of 10% HCl, then was poured in water and extracted with DCM (3x150 ml). The collected organic phases were washed with water and dried over Na₂S0₄, then filtered and concentrated. The crude was purified with a Biotage SNAP 50 g Silica column, eluting with DCM. A colourless solid was recovered, and this was reacted with thiourea (0.902 g, 11.85 mmol) in DMF (5 ml) at 90 °C overnight. The black mixture was diluted with Et₂0 (500 ml) and washed with water (4x150 ml), then dried over Na₂S0₄, filtered and concentrated. The black crude was purified with Biotage SNAP 100 g column, eluting with a gradient of Cy and EtOAc. The title compound D50 was eluted with ca 50% EtOAc and recovered as dark wax (200 mg).

UPLC (GEN QC SS) rt = 0.57 minutes, peak observed: 169 (M+l), C₄H₃F₃N₂S requires 168.

1H NMR (400 MHz, CHLOROFORM-^ δ ppm 5.55 (br. s., 2 H) 7.41 (d, 1 H) Description 51: 2-chloro-5-(trifluoromethyl)-l,3-thiazole (D51)

tert-butyl nitrite (140 mg, 1.356 mmol) was added dropwise at room temperature to a suspension of 5-(trifluoromethyl)-l,3-thiazol-2-amine D50 (190 mg) and copper(II) chloride (182 mg, 1.356 mmol) in ACN (15 ml). The resulting mixture was stirred for 30 minutes at room temperature, then was taken up with Et₂0 (200 ml) and washed with water (3x100 ml). The organic phase was dried over Na₂S0₄, then filtered and distillated in a Claisen apparatus to remove the volatiles. The title compound D51 (200 mg) was used in the next step without any further purification.

1H NMR (400 MHz, CHLOROFORM-^ δ ppm 7.92 (d, 1 H

4-(trifluoromethyl)-l,3-thiazol-2-amine (0.25 g, 1.487 mmol), copper(II) chloride (0.240 g, 1.784 mmol), tert-butyl nitrite (0.230 g, 2.230 mmol) were reacted in ACN (20 ml) at room temperature for 3 hours. The reaction mixture was poured into 100 ml 10% aqueous HC1 and extracted with Et₂0 (3x100 ml). The collected organic phases were washed with water, dried over Na₂S0₄, filtered and concentrated at rotavapor at max 650 mBar/35 °C and then in a Claisen apparatus to remove ACN to give the title compound D52 as yellow oil (170 mg).

1H NMR (400 MHz, CHLOROFORM-^ δ ppm 7.72 (s, 1 H) EXAMPLES

Example 1 : N-[((lR,4S,6R)-3-{[6-methyl-3-(2H-l,2,3-triazol-2-yl)-2- pyridinyl]carbonyl}-3-azabicyclo[4.1.0]hept-4-yl)methyl]-5-(trifluoromethyl)-l,3,4- thiadiazol-2-amine (El)

6-methyl-3-(2H-l,2,3-triazol-2-yl)-2-pyridinecarboxylic acid D40 (29.3 mg) was reacted with TBTU (55.4 mg, 0.172 mmol) and DIPEA (0.025 ml, 0.144 mmol) in DCM (3 ml) for 20 minutes, then N-[(lR,4S,6R)-3-azabicyclo[4.1.0]hept-4-ylmethyl]-5-(trifluoromethyl)- l,3,4-thiadiazol-2-amine D30 (40 mg) was added and the resulting solution was shaken at room temperature for 2 hours. The solvent was removed and the crude was purified with a column stacking of two SNAP 10 g C18, eluting with ACN and water (modified with 0.1% of HCOOH). Fractions containing the required product were neutralised over an SCX column (2 g, washing with MeOH and eluting with 2M ammonia in MeOH) to give the title compound El (55 mg)

UPLC (Final QC) rt = 0.77 minutes, peak observed: 465 (M+l). Ci₉Hi₉F3N₈0₆ requires 464

1H NMR (500 MHz, DMSO-i¾) δ ppm 0.16 - 0.24 (m, 1 H) 0.67 - 0.75 (m, 1 H) 0.91 - 1.01 (m, 1 H) 1.05 - 1.12 (m, 1 H) 1.64 - 1.80 (m, 2 H) 2.36 (s, 3 H) 3.26 - 3.81 (m, 4 H) 4.38 (d, 1 H) 7.44 (d, 1 H) 8.00 - 8.30 (m, 3 H) 8.49 - 8.77 (m, 1 H)

Example 2: N-[((lR,4S,6R)-3-{[6-methyl-3-(lH-pyrazol-l-yl)-2-pyridinyl]carbonyl}-3- azabicyclo[4.1.0]hept-4-yl)methyl]-5-(trifluoromethyl)-l,3,4-thiadiazol-2-amine (E2)

6-methyl-3-(lH-pyrazol-l-yl)-2-pyridinecarboxylic acid D42 (29.2 mg) was reacted with TBTU (55.4 mg, 0.172 mmol) and DIPEA (0.025 ml, 0.144 mmol) in DCM (3 mL) at room temperature for 30 minutes.

N-[(lR,4S,6R)-3-azabicyclo[4.1.0]hept-4-ylmethyl]-5-(trifluoromethyl)-l,3,4-thiadiazol-2- amine D30 (40 mg) was added and the resulting mixture was shaken for 2 hours at room temperature. The reaction was concentrated under vacuum and the resulting crude was purified with a column stacking of two SNAP 10 g CI 8, eluting with ACN and water (modified with 0.1% of HCOOH). Fractions containing the required product were

neutralised over an SCX column (2 g, washing with MeOH and eluting with 2M ammonia in MeOH) to give the title compound E2 (56 mg).

UPLC (Final QC): rt = 0.78 minutes, peak observed: 464 (M+l). C₂oH2oF₃N₇0₆ requires 463.

1H NMR (500 MHz, DMSO-i¾) δ ppm -0.09 (br. s., 1 H) 0.63 (d, 1 H) 0.71 - 0.94 (m, 1 H) 0.94 - 1.17 (m, 1 H) 1.54 - 1.76 (m, 1 H) 2.31 (br. s., 2 H) 2.36 (br. s., 1 H) 2.63 (br. s., 2 H) 3.30 (s, 1 H) 3.51 (d, 1 H) 3.55 - 3.78 (m, 1 H) 4.38 (d, 1 H) 6.55 (s, 1 H) 7.38 (d, 1 H) 7.77 (s, 1 H) 7.91 (d, 1 H) 8.04 - 8.11 (m, 1 H) 8.67 (br. s., 1 H)

The following compounds were made using a similar method to that described above for Example 2.

Amide coupling Characterising data

Reactants

D30 and D44 N-[((lR,4S,6R)-3-{[5-methyl-2-(2H-l,2,3-triazol-2- yl)phenyl] carbonyl}-3-azabicyclo [4.1.0] hept-4- yl)methyl]-5-(trifluoromethyl)-l,3,4-thiadiazol-2- amine

1H NMR (500 MHz, DMSO-i¾) δ ppm 0.53 - 0.71 (n 1 H) 0.78 - 0.95 (m, 1 H) 0.95 - 1.10 (m, 1 H) 1.58 -

1.78 (m, 1 H) 2.00 - 2.14 (m, 2 H) 2.33 - 2.51 (m, 2 H

2.54 - 2.72 (m, 2 H) 3.19 - 3.33 (m, 2 H) 3.50 - 3.73 (m, 2 H) 4.39 - 4.52 (m, 1 H) 7.20 - 7.35 (m, 1 H) 8.0 - 8.14 (m, 2 H) 8.76 (br. s., 1 H)

Example 6: N-[((lR,4S,6R)-3-{[6-methyl-3-(2-pyrimidinyl)-2-pyridinyl]carbonyl}-3- azabicyclo[4.1.0]hept-4-yl)methyl]-5-(trifluoromethyl)-l,3-thiazol-2-amine (E6)

[((lR,4S,6R)-3-{[6-methyl-3-(2-pyrimidinyl)-2-pyridinyl]carbonyl}-3- azabicyclo[4.1.0]hept-4-yl)methyl]amine D18 (50 mg), 2-chloro-5-(trifluoromethyl)-l,3- thiazole D51 (29.0 mg) and DIPEA (0.081 ml, 0.464 mmol) were reacted in DMSO (4 ml) at 80 °C for 3 hours. Solvent was removed under vacuum, and the crude purified with 12 g CI 8 SNAP column, eluting with a gradient of ACN and water, modified with 0.5%

HCOOH. The compound was eluted with ca 80% ACN and the fractions were collected and then neutralised over a 0.5 g SCX cartridge, washing with MeOH and eluting with 2M ammonia in MeOH. The title compound E6 was recovered as yellow oil (21 mg)

UPLC (IPQC) rtl= 1.11 minutes, rt2= 1.14 minutes (rotamers present) peaks observed: 475, C₂2H₂iF₃N₆OS requires: 474.

1H NMR (400 MHz, DMSO-i¾) δ ppm 0.22 - 0.34 (m, 1 H) 0.72 - 0.83 (m, 1 H) 0.99 - 1.06 (m, 1 H) 1.07 - 1.16 (m, 1 H) 1.60 - 1.75 (m, 1 H) 1.75 - 1.86 (m, 1 H) 2.47 - 2.53 (m, 3 H) 3.14 - 3.77 (m, 4 H) 4.33 (d, 1 H) 7.41 - 7.53 (m, 3 H) 8.41 (d, 1 H) 8.44 - 8.52 (m, 1 H) 8.91 (d, 2 H)

Example 7: N-[((lR,4S,6R)-3-{[6-methyl-3-(2-pyrimidinyl)-2-pyridinyl]carbonyl}-3- azabicyclo[4.1.0]hept-4-yl)methyl]-5-(trifluoromethyl)-l,3,4-thiadiazol-2-amine (E7)

[((lR,4S,6R)-3-{[6-methyl-3-(2-pyrimidinyl)-2-pyridinyl]carbonyl}-3- azabicyclo[4.1.0]hept-4-yl)methyl]amine D18 (20 mg), 2-chloro-5-(trifluoromethyl)-l,3,4- thiadiazole (11.66 mg, 0.062 mmol) and DIPEA (0.022 ml, 0.124 mmol) were collected in DMSO (2 ml) and shaken at 80 °C for 2 hours. Solvent was removed under vacuum, and the resulting crude was purified with Biotage SP1 (over a column stacking of 2x12 g CI 8 SNAP columns, eluting with a gradient of ACN and water, modified with 0.5% HCOOH). Fractions containing the required product were collected and neutralised over a 1 g SCX column, washing with MeOH and eluting with 2M ammonia in MeOH. The resulting compound (25 mg) was further purified with Biotage Spl (over a column stacking of Analogix 2x4 g Silica columns, using DCM/MeOH as eluent) affording the title compound E7 (21 mg).

1H NMR (400 MHz, DMSO-i¾) δ ppm 0.26 - 0.33 (m, 1 H) 0.73 - 0.82 (m, 1 H) 0.91 - 1.19 (m, 2 H) 1.61 - 1.85 (m, 2 H) 2.43 (s, 3 H) 3.35 - 3.42 (m, 1 H) 3.49 - 3.86 (m, 3 H) 4.31 - 4.42 (m, 1 H) 7.35 - 7.54 (m, 2 H) 8.37 (d, 1 H) 8.53 - 8.79 (m, 1 H) 8.81 - 8.97 (m, 2 H)

Example 8: N-[((lR,4S,6R)-3-{[6-methyl-3-(2-pyrimidinyl)-2-pyridinyl]carbonyl}-3- azabicyclo[4.1.0]hept-4-yl)methyl]-4-(trifluoromethyl)-l,3-thiazol-2-amine (E8)

[((lR,4S,6R)-3-{[6-methyl-3-(2-pyrimidinyl)-2-pyridinyl]carbonyl}-3- azabicyclo[4.1.0]hept-4-yl)methyl]amine D18 (20 mg), 2-chloro-4-(trifluoromethyl)-l,3- thiazole D52 (11.60 mg) and DIPEA (0.022 ml, 0.124 mmol) were collected in DMSO (2 ml) and shaken at 100 °C overnight. Then solvent was removed under vacuum, and the crude was purified with Biotage SPl, over a column stacking of 2 SNAP CI 8 12 g, eluting with a gradient of ACN and water (modified with 0.5% of HCOOH). Fraction containing the required product were collected and neutralised on a 0.5 g SCX (washing with MeOH and eluting with 2M ammonia in MeOH). Resulting compound was further purified with Biotage Spl (over a 25 g Silica Analogix column, eluting with a gradient of DCM and MeOH), affording the title compound E8 (4 mg).

1H NMR (500 MHz, DMSO-i¾) δ ppm 0.24 - 0.33 (m, 1 H) 0.74 - 0.81 (m, 1 H) 0.92 - 1.18 (m, 2 H) 1.63 - 1.86 (m, 2 H) 2.47 - 2.52 (m, 3 H) 3.15 - 3.45 (m, 2 H) 3.53 - 3.74 (m, 2 H) 4.31 - 4.38 (m, 1 H) 7.31 (s, 1 H) 7.39 - 7.51 (m, 2 H) 8.08 - 8.17 (m, 1 H) 8.38 (d, 1 H) 8.89 (d, 2 H)

Example 9: N-[((lS,4S,6S)-3-{[6-methyl-3-(2-pyrimidinyl)-2-pyridinyl]carbonyl}-3- azabicyclo[4.1.0]hept-4-yl)methyl]-5-(trifluoromethyl)-l,3,4-thiadiazol-2-amine (E9)

6-methyl-3-(2-pyrimidinyl)-2-pyridinecarboxylic acid D33 (121 mg) was reacted with

TBTU (54.2 mg, 0.169 mmol) and DIPEA (0.029 ml, 0.169 mmol) in DCM (3 ml) at room temperature for 30 minutes, then N-[(l S,4S,6S)-3-azabicyclo[4.1.0]hept-4-ylmethyl]-5- (trifluoromethyl)-l,3,4-thiadiazol-2-amine D29 (47 mg) was added and the mixture was shaken at room temperature for 2 hours. Solvent was then removed and the resulting crude was purified with a column stacking of two SNAP 10 g CI 8, eluting with ACN and water (modified with 0.1% of HCOOH). Fractions containing the required product were neutralised over an SCX column (2 g, washing with MeOH and eluting with 2M ammonia in MeOH) to give the title compound E9 (46 mg).

UPLC (Final QC) rtl = 0.70 minutes, rt2 = 0.71 minutes (rotamers observed), peaks observed: 476 (M+l) C₂₁H₂₀F₃N₇OS requires 475. 1H MR (500 MHz, DMSO-i¾) δ ppm 0.15 - 0.23 (m, 1 H) 0.61 - 0.72 (m, 1 H) 0.90 - 0.99 (m, 1 H) 0.99 - 1.08 (m, 1 H) 1.10 - 1.23 (m, 1 H) 2.39 - 2.48 (m, 1 H) 2.56 (s, 3 H) 2.59 - 2.72 (m, 1 H) 3.42 - 3.54 (m, 1 H) 3.54 - 3.64 (m, 1 H) 3.81 (d, 1 H) 4.08 - 4.25 (m, 1 H) 7.40 - 7.51 (m, 2 H) 8.50 (d, 1 H) 8.61 (br. s., 1 H) 8.78 - 8.89 (m, 2 H)

Example 10: Determination of antagonist affinity at human Orexin-1 and 2 receptors using FLIPR

Cell Culture

Adherent Chinese Hamster Ovary (CHO) cells, stably expressing the recombinant human Orexin-1 or human Orexin-2 receptors or Rat Basophilic Leukaemia Cells (RBL) stably expressing recombinant rat Orexin-1 or rat Orexin-2 receptors were maintained in culture in Alpha Minimum Essential Medium (Gibco/Invitrogen, cat. no.; 22571-020), supplemented with 10% decomplemented foetal bovine serum (Life Technologies, cat. no. 10106-078) and 400 μg/mL Geneticin G418 (Calbiochem, cat. no.345810). Cells were grown as monolayers under 95%:5% air:C0₂ at 37 °C.

The sequences of the human orexin 1, human orexin 2, rat orexin 1 and rat orexin 2 receptors used to test the compounds of examples 1 to 9 were as published in Sakurai, T. et al (1998) Cell, 92 pp 573 to 585.

Measurement of [C^J, using the FLIPR™

Cells were seeded into black clear-bottom 384-well plates (density of 20,000 cells per well) in culture medium as described above and maintained overnight (95%:5% air:C0₂ at 37°C). On the day of the experiment, culture medium were discarded and the cells washed three times with standard buffer (NaCl, 145 mM; KC1, 5 mM; HEPES, 20 mM; Glucose, 5.5 mM; MgCl₂, 1 mM; CaCl₂, 2 mM) added with Probenecid 2.5 mM. The plates were then incubated at 37 °C for 60 minutes in the dark with 2 μΜ FLUO-4AM dye to allow cell uptake of the FLUO-4AM, which is subsequently converted by intracellular esterases to FLUO-4, which is unable to leave the cells. After incubation, cells were washed three times with standard buffer to remove extracellular dye and 30 μΕ of buffer were left in each well after washing.

Compounds of the invention were tested in a final assay concentration range from 1.66xlO^"5M to 1.58xl0^"uM. Compounds of the invention were dissolved in

dimethylsulfoxide (DMSO) at a stock concentration of 10 mM. These stock solutions were serially diluted with DMSO and 1 μΐ, of each dilution was transferred to a 384 well compound plate. Immediately before introducing compound to the cells, buffer solution (50 μΐ/well) was added to this plate. To allow agonist stimulation of the cells, a stock plate containing a solution of human orexin A (hOrexin A) was diluted with buffer to final concentration just before use. This final concentration of hOrexin A was equivalent to the calculated EC80 for hOrexinA agonist potency in this test system. This value was obtained by testing hOrexinA in concentration response curve (at least 16 replicates) the same day of the experiment. The loaded cells were then incubated for lOmin at 37°C with test compound. The plates were then placed into a FLIPR™ (Molecular Devices, UK) to monitor cell fluorescence (Xg_X = 488nm, λ_ΕΜ = 540nm) (Sullivan E, Tucker EM, Dale IL. Measurement of [Ca²⁺]i using the fluometnc imaging plate reader (FLIPR). In: Lambert DG (ed.), Calcium Signaling Protocols. New Jersey: Humana Press, 1999, 125-136). A baseline fluorescence reading was taken over a 5 to 10 second period, and then 10 μΕ of EC80 hOrexinA solution was added. The fluorescence was then read over a 4-5 minute period.

Data Analysis

Functional responses using FLIPR were measured as peak fluorescence intensity minus basal fluorescence and expressed as a percentage of a non-inhibited Orexin-A- induced response on the same plate. Iterative curve-fitting and parameter estimations were carried out using a four parameter logistic model and Microsoft Excel (Bowen WP, Jerman JC. Nonlinear regression using spreadsheets. Trends Pharmacol. Sci. 1995; 16: 413-417). Antagonist affinity values (IC₅₀) were converted to functional pIQ values using a modified Cheng-Prusoff correction (Cheng YC, Prusoff WH. Relationship between the inhibition constant (IQ) and the concentration of inhibitor which causes 50 percent inhibition (IC₅₀) of an enzymatic reaction. Biochem. Pharmacol. 1973, 22: 3099-3108).

Where [agonist] is the agonist concentration, EC₅₀ is the concentration of agonist giving 50% activity derived from the agonist dose response curve and n=slope of the dose response curve. When n=l the equation collapses to the more familiar Cheng-Prusoff equation.

Compounds of examples 1 to 9 were tested according to the method of example 7.

All compounds gave fpKi values from 7.1 to 9.5 at the human cloned orexin-1 receptor and from 6.7 to 8.9 at the human cloned orexin-2 receptor with the exception that the compound of example 4, N-[((lR,4S,6R)-3-{[6-methyl-3-(lH-l,2,3-triazol-l-yl)-2- pyridinyl]carbonyl}-3-azabicyclo[4.1.0]hept-4-yl)methyl]-5-(trifluoromethyl)-l,3,4- thiadiazol-2-amine, has an fpKi value of less than 4.8 at the orexin 2 receptor.

Claims

1. A compound of formula (I)

(I)

wherein:

Y is C or N;

m is 0 or 1; and

n is 0 or 1;

or a pharmaceutically acceptable salt thereof.

2. A compound according to claim 1 where the stereogenic centres of the compounds of formula (I) are in a cis (IS, 4S,6S)- configuration, or a pharmaceutically acceptable salt thereof.

3. A compound according to claim 1 where the stereogenic centres of the compounds of formula (I) are in a trans (\R,4S,6R)- configuration, or a pharmaceutically acceptable salt thereof.

4. A compound according to any one of claims 1 to 3 where Het is thiazolyl, or a pharmaceutically acceptable salt thereof.

5. A compound according to any one of claims 1 to 3 where Het is thiadiazolyl, or a pharmaceutically acceptable salt thereof.

6. A compound according to any one of claims 1 to 5 where Het is substituted with trifluoromethyl or a pharmaceutically acceptable salt thereof.

7. A compound according to any one of claims 1 to 6 where Ri is triazolyl, or a pharmaceutically acceptable salt thereof.

8. A compound according to any one of claims 1 to 6 where Ri is pyrazolyl, or a pharmaceutically acceptable salt thereof.

9. A compound according to any one of claims 1 to 6 where Ri is pyrimidinyl, or a pharmaceutically acceptable salt thereof.

10. A compound of formula (I) selected from the group consisting of:

^-[((lR^^e^-S-ICe-methyl-S-C H-l^^-triazol- -y - -pyridiny^carbonyll-S- azabicyclo[4.1.0]hept-4-yl)methyl]-5-(trifluoromethyl)-l,3,4-thiadiazol-2-amine;

N-[((lR,4S,6R)-3-{[6-methyl-3-(lH-pyrazol-l-yl)-2-pyridinyl]carbonyl}-3- azabicyclo[4.1.0]hept-4-yl)methyl]-5-(trifluoromethyl)-l,3,4-thiadiazol-2-amine;

^-[((lR^^e^-S-ICS-methyl- -C H-l^^-triazol- -y pheny^carbonyll-S- azabicyclo[4.1.0]hept-4-yl)methyl]-5-(trifluoromethyl)-l,3,4-thiadiazol-2-amine;

N-[((lR,4^,6R)-3-{[2-(2H-l,2,3-triazol-2-yl)phenyl]carbonyl}-3-azabicyclo[4.1.0]hept-4- yl)methyl]-5-(trifluorom ethyl)- 1 ,3 ,4-thiadiazol-2-amine;

N-[((lR,4^,6R)-3-{[6-methyl-3-(2-pyrimidinyl)-2-pyridinyl]carbonyl}-3- azabicyclo[4.1.0]hept-4-yl)methyl]-5-(trifluoromethyl)-l ,3-thiazol-2-amine;

N-[((lR,4^,6R)-3-{[6-methyl-3-(2-pyrimidinyl)-2-pyridinyl]carbonyl}-3- azabicyclo[4.1.0]hept-4-yl)methyl]-5-(trifluoromethyl)-l ,3,4-thiadiazol-2-amine;

N-[((l^,4^,65)-3-{[6-methyl-3-(2-pyrimidinyl)-2-pyridinyl]carbonyl}-3- azabicyclo[4.1.0]hept-4-yl)methyl]-5-(trifluoromethyl)-l,3,4-thiadiazol-2-amine, or a pharmaceutically acceptable salt thereof.

11. The compound as defined in any one of claims 1 to 10, or a pharmaceutically acceptable salt thereof, for use in therapy.

12. The compound as defined in any one of claims 1 to 10, or a pharmaceutically acceptable salt thereof, for use in the treatment of a disease or disorder where an antagonist of a human orexin receptor is required.

13. The compound according to claim 12, or a pharmaceutically acceptable salt thereof, wherein the disease or disorder is a sleep disorder, a depression or mood disorder, an anxiety disorder, a substance-related disorder or a feeding disorder.

14. The compound according to claim 13, or a pharmaceutically acceptable salt thereof, wherein the disease or disorder is a sleep disorder.

15. A pharmaceutical composition comprising a) the compound as defined in any one of claims 1 to 10, or a pharmaceutically acceptable salt thereof, and b) one or more

pharmaceutically acceptable carriers.