NZ611365B2 - Fused aminodihydrothiazine derivatives useful as bace inhibitors - Google Patents

Abstract

Disclosed are fused aminodihydrothiazine derivatives of formula (I): wherein X is hydrogen or fluorine; A is CH or N; Y is methyl, ethyl, monofluoromethyl, difluoromethyl, trifluoromethyl, difluoroethyl, methoxy, ethoxy, methoxymethyl or cyano; and pharmaceutically acceptable salts thereof; which compound has an A? production inhibitory effect or a BACE1 inhibitory effect and is useful as a prophylactic or therapeutic agent for a neurodegenerative disease caused by A? and typified by Alzheimer-type dementia. Examples of compounds of formula (I) are: N-(3-((4aS,5S,7aS)-2-amino-5-(trifluoromethyl)-4a,5,7,7a-tetrahydro-4H-furo[3,4-d][1,3]thiazin-7a-yl)-4-fluorophenyl)-5-cyanopicolinamide; N-(3-((4aS,5S,7aS)-2-amino-5-(trifluoromethyl)-4a,5,7,7a-tetrahydro-4H-furo[3,4-d] [1,3]thiazin-7a-yl)-4-fluorophenyl)-5-(difluoromethyl)pyrazine-2-carboxamide and N-(3-((4aS,5S,7aS)-2-amino-5-(trifluoromethyl)-4a,5,7,7a-tetrahydro-4H-furo[3,4-d][1,3]thiazin-7a-yl)-4-fluorophenyl)-5-methoxypyrazine-2-carboxamide. mpound has an A? production inhibitory effect or a BACE1 inhibitory effect and is useful as a prophylactic or therapeutic agent for a neurodegenerative disease caused by A? and typified by Alzheimer-type dementia. Examples of compounds of formula (I) are: N-(3-((4aS,5S,7aS)-2-amino-5-(trifluoromethyl)-4a,5,7,7a-tetrahydro-4H-furo[3,4-d][1,3]thiazin-7a-yl)-4-fluorophenyl)-5-cyanopicolinamide; N-(3-((4aS,5S,7aS)-2-amino-5-(trifluoromethyl)-4a,5,7,7a-tetrahydro-4H-furo[3,4-d] [1,3]thiazin-7a-yl)-4-fluorophenyl)-5-(difluoromethyl)pyrazine-2-carboxamide and N-(3-((4aS,5S,7aS)-2-amino-5-(trifluoromethyl)-4a,5,7,7a-tetrahydro-4H-furo[3,4-d][1,3]thiazin-7a-yl)-4-fluorophenyl)-5-methoxypyrazine-2-carboxamide.

Description

FUSED AMINODIHYDROTHIAZINE DERIVATIVES USEFUL AS BACE INHIBITORS The present ion relates to a fused aminodihydrothiazine tive and pharmaceutical use thereof. More particularly, the present invention relates to a fused aminodihydrothiazine tive which has an amyloid-B nafter referred to as AB) protein tion inhibitory effect or a beta-site arnyloid-B sor protein cleavage enzyme 1 (hereinafter referred to as BACEl or beta-secretase) inhibitory effect and is effective for treating a neurodegenerative disease caused by AB protein, in particular, Alzheimer-type dementia, Down’s syndrome or the like, and to a pharmaceutical composition comprising the fused aminodihydrothiazine derivative as an active ient.

Alzheimer's disease is a disease characterized by degeneration and loss of neurons as well as formation of senile plaques and neurofibrillary tangles. Currently, only the symptoms of mer’s disease are treated using a symptom‘improving agent typified by an acetylcholinesterase tor, and a fundamental remedy to inhibit progression of the disease has not yet been developed. It is ary to develop a method for controlling the cause of the onset of pathology in order to create a fundamental remedy for mer's disease.

It is assumed that AB-proteins as breakdown products of amyloid precursor proteins (hereinafter referred to as APP) are highly involved in degeneration and loss of neurons and onset of symptoms of dementia. AB—proteins have, as main components, AB4O consisting of 40 amino acids and AB42 with two amino acids added at the C— terminal. The AB4O and AB42 are known to be highly prone to aggregation and to be the main components of senile plaques. Further, it is known that the AB40 and AB42 are increased by mutations in APP and presenilin genes which is observed in familial Alzheimer's disease. Accordingly, a compound that reduces production of AB40 and AB42 is expected to be a disease progression inhibitor or prophylactic agent for Alzheimer's disease.

AB is produced by the ge of APP by beta-secretase (BACEl) and subsequently by secretase. For this reason, ts have been made to create gamma—secretase and beta—secretase inhibitors in order to inhibit AB production.

Published International patent application WOZOl 1/005738 (Eli Lilly and Company) describes compounds of formula (A) and their use as BACE inhibitors: NH2 (A) (R3)p where RI n and p are defined therein.

, R2, R3, X, m, Fused ihydrothiazine compounds of formula (B) have already been disclosed in published International patent application W02009/O91016 (Eisai R&D Management Co., Ltd): R5 R5 R1 N N z \Y ‘R2 e Y ,s R4 R3 wherein ring A represents a €6-14ary1 group or the like; L represents —NReCO— in R8 represents a hydrogen atom or the like] or the like; ring B represents a €6-14aryl group or the like; X represents a C1-3alkylene group or the like; Y represents a single bond or the like; Z ents a C1_3alkylene group or the like; R1 and R2 independently represent a hydrogen atom or the like; and R3, R4, R5 and R6 independently represent a hydrogen atom, a halogen atom or the like. r fused aminodihydrothiazine compounds of formula (C) have been sed in published International patent application WOZOlO/O38686 (Eisai R&D Management Co._. Ltd): R5 R6 R1 % Y” e R4 R3 wherein ring A represents a C6_l4aryl group or the like; L represents ~NReCO— [wherein R6 represents a hydrogen atom or the like] or the like; the ring B represents a C(y14aryl group or the like; X represents a C1_3alkylene group or the like; Y represents a single bond or the like; Z represents an oxygen atom or the like; R1 and R2 each independently represents a hydrogen atom or the like; and R3, R4, R5 and R6 each ndently represents a hydrogen atom, a halogen atom or the like.

The present invention represents a selection from the genus of compounds sed in W02009/091016.

An object of the present invention is to provide further compounds that have an AB production inhibitory effect or a BACEl inhibitory effect and are useful as prophylactic or therapeutic agents for a neurodegenerative disease caused by AB and typified by Alzheimer-type ia, which compounds are fused ihydrothiazine derivatives. The above object is to be read disjunctively with the object of the invention to at least provide the public with a useful alternative.

Thus, the t invention provides the compound of formula (I): /A Y got iN X NH F (1) F30 wherein X is hydrogen or fluorine; A is CH or N; Y is methyl, ethyl, monofluoromethyl, difluoromethyl, trifluoromethyl, difluoroethyl, methoxy, ethoxy, methoxymethyl or —C EN; and pharmaceutically acceptable salts thereof.

In one embodiment of the present invention, X is hydrogen.

In another embodiment of the present invention, A is N.

In another embodiment of the present ion, Y is methyl, oromethyl, difluoromethyl, trifluoromethyl or methoxy.

One favoured group of compounds of the t invention is the compound of formula (Ia) and pharmaceutically acceptable salts thereof: N Y on/ rN F (la) N NH2 0 Y where Y is as hereinbefore defined. Preferably, Y is methyl, monofluoromethyl, difluoromethyl, trifluoromethyl, oethyl, methoxy, ethoxy or methoxymethyl.

In one embodiment the present invention provides a compound of a (Ia) wherein Y is methoxy or monofluoromethyl.

Another favoured group of compounds of the present invention is the compound of formula (Ib) and pharmaceutically acceptable salts thereof: N Y 0%;1//|N F NH F 0b) N NH2 0 Y where Y is as before defined. Preferably, Y is methyl, monofluoromethyl, difluoromethyl or methoxy. 1 O A further favoured group of compounds of the present invention is the compound of formula (10) and pharmaceutically able salts f: O \ F ('0) N NH2 0 Y where Y is as hereinbefore defined. Preferably, Y is methyl, ethyl, trifluoromethyl, methoxy or -CEN.

Preferred compounds of the present ion are: N-(3—((4aS,SS,7aS)—2—amino-5~(trifluoromethyl)—4a,5,7,7a—tetrahydro—4H—furo[3,4- d][1,3]thiazin-7a-y1)—4—fluorophenyl)—5-methoxypyrazine-2—carboxamide: WO 98213 O \N N—(3—((4aS,SS,7aS)—2-amino(trifluor0methyl)—4a,5,7,7a—tetrahydro-4H-furo[3,4— d] [1,3]thiazin—7a—yl)—4—flu0r0pheny1)—5~cyanopicolinamide: O \ N NH2 0 1/ F PH F ; N-(3-((4aS,SS,7aS)-2—amino(trifluoromethy1)—4a,5 ,7,7a-tetrahydro-4H—furo[3 ,4- d][1,3]thiazin-7a-y1)-4—flu0rophenyl)~5—(difluoromethy1)pyrazine—Z—carboxamide: N;\[H\F O \N N—(3~((4aS,5S,7aS)—2-amino—5~(trifluoromethy1)—4a,5 ,7,7a-tetrahydro-4H—furo[3 ,4— d][1 ,3] thiazin-7a-y1)—4—fluoropheny1)—5 - (trifluoromethy1)picolinamide: N-(3—((4aS,SS,7aS)~2—amino-5—(trifluoromethyl)—4a,5,7,7a—tetrahydro—4H—furo[3,4- d] [1 ,3] thiazin-7a~y1)—4—fluorophenyl)~5~methy1pyrazine—Z—carboxanfide: N-(3-((4aS,SS,7aS)amino-S-(trifluoromethyl)—4a,5,7,7a-tetrahydro—4H—furo[3,4— d][1,3]thiazin-7a—y1)-4—fluoropheny1)—5-methylpicolinmnide: o \1 N NH2 o I FFFH N—(3-((4aS,SS,7aS)amino~5—(trifluor0methyl)~4a,5,7 trahydro-4H-furo[3,4— d] [1 ,3] thiazin—7a-y1)fluorophenyl)—5—ethy1picolinamide: O \ 2012/050833 N-(3—((4aS,SS,7aS)~2—amin0—5—(trifluoromethyl)—4a,5,7,7a—tetrahydro-4H—furo[3,4— d] [ 1 ,3] thiazin-7a—y1)-4—fluoropheny1)(fluoromethyl)pyrazine-Z-carboxamide: N ’ F OVVN| N-(3—((4aS,SS,7aS)-2~amino(trifluoromethyl)—4a,5,7,7a-tetrahydro—4H-furo[3,4— d][1,3]thiazin-7a—y1)-4—fluorophenyl)~5~meth0xypicolinamidez N/ O\ O \I N NH2 O \Y 1 O N-(3—((4aS ,SS ,7as)~2~amino~5-(trifluoromethyl)—4a,5 ,7 ,7a—tetrahydro—4H—furo[3 ,4- d] [ 1 ,3] thiazin—7a—yl)—4-flu0ropheny1)-5—ethoxypyrazine—Z—carboxamide: NWow N-(3-(:(4aS,SS,7aS)—2—amino-S-(trifluoromethyl)—4a,5 ,7,7a—tetrahydro—4H—furo[3,4- l 5 d]L1,3]thiazin—7a—yl)-4—fluorophcnyl)—5—(1,1-difluorocthyl)pyrazinc-2—carboxamidc: PCT/EPZO 12/050833 (4aS,SS,7aS)—2—amin0(trifluoromethyl)—4a,5,7,7a—tetrahydro-4H—furoI3,4— d][1,3]thiazin—7a—y1)—4—flu0r0pheny1)-5—(trifluoromethyl)pyrazine-Z—carboxamide: F F Néféf: O \ N N NH2 O ‘Y F F ; N-(3-((4aS,SS,7aS)~2—amino(trif1uoromethy1)—4a,5,7,7a-tetrahydro-4H-furo[3,4- d][l ,3]thiazin—7a—y1)—4—fluoropheny1)—5—(methoxymethy1)pyrazine-Z—carboxamide: N NH2 O ‘Y N—{3—[(4aS,SS,7aS)—2—amino—5-(trifluoromethyl)-4a:5—dihydro—4H—furo[3,4 d][1,3]thiazin-7a(7H)—y1]—4—flu0r0pheny1}—5—[(2H3)methyloxy]pyrazine—Z-carboxamide: o 2H N,“.r v: ow\N H N-(3{(421858,7218)—2—amino—5-(trifluoromethyl)-4a,5,7,7a—tetrahydr0—4H—fur0[3 ,4— d] [1 ,3]thiazin—7a-yl)-4,5 -difluoropheny1)-5 oromethyl)pyrazine-Z-carboxamide: 0 \N N—(3—((4aS,SS,7aS)—2-amino~5~(trifluoromethyl)~4a,5 ,7,7a-tetrahydro—4H—furo[3 ,4- d] [1,3]thiazin-7a—y1)—4,5—difluor0pheny1)~5-methoxypyrazine-Z-carboxamide: NwOMe ow\N l 0 N—(3{(421358_,7aS)amino-5—(trif1u0r0methy1)-4a,5,7,7a-tetrahydro-4H—fur0[3 ,4— d] [1 ,3]thiazinv7a-y1)—4,5-difluor0pheny1)-5—methylpyrazine-Z—carboxamide: N¢\lrMe OYK/N N-(3-((4aS,SS,7aS)—2—amino-S-(trifluoromethyl)—4a,5,7,7a-tetrahydro—4H—furo[3,4- l 5 d][1,3]thiazin-7a—y1)-4,5-dif1uor0pheny1)—5-(fluoromethyl)—pyrazine-2—carboxamide: 2012/050833 N“I: o§)\/N and pharmaceutically acceptable salts thereof.

In one embodiment, the present invention provides a compound which is N-(3- ((4aS,SS,7aS)—2—amino-5—(trifluoromethyl)-4a,5,7,7a—tetrahydro-4H—furo[3,4— d][l ,3]thiazin—7a—yl)—4—fl uoropheuyl)—5~methoxypyrazine-Z-carhoxamide, or a pharmaceutically able salt thereof.

In another embodiment, the present invention provides a compound which is N— (3—((4aS,SS,7aS)amino-5—(tritluoromethyl)-4a,5,7=7a—tetrahydro—4H-furo[3,4— d][1 ,3]thiazin-7a—yl)-4—fluorophenyl)—5—(fluoromethyl)pyrazine—2—earboxarnide, or a pharmaceutically acceptable salt thereof. c compounds within the scope of this invention include those named in the Examples below and their pharmaceutically acceptable salts.

As used herein, the term "difluoroethyl" refers to an alkyl group having two carbon atoms and substituted with two fluorine atoms. Examples of the group are CH3—CF2-, CHgF-CHF- and CHFQ-CH3-. In the present invention, the group is preferably CH3—CF2—.

The compound of a (I) is not limited to a specific isomer and includes all possible isomers (such as a keto-enol isomer, an imine—enamine isomer and a rotamer) and mixtures thereof. For e, the compound of formula (1) includes the following tautomers: A Y \j/ A Y YEN/ l 0%/ 0 1/ X NH X NH F F N NH H NH O O Y W3 8 F30 H The compounds of the t invention contain three chiral centers located on the ydrofuro—thiazinyl ring within formula (I). The stereochemical configuration at each of these chiral centers is preferably S, i.e. they are (4aS,SS,7aS) stereoisomers. For the avoidance of doubt the (4aS.SS.7aS) stereoisomers of the t invention may be present as a mixture with one or more of the other possible stereoisomers, for example in a racemic mixture.

In one ment, the present ion provides a compound of formula (I) which is stereochemically pure at the (4aS,SS,7aS) chiral centers. In the context of the present specification, the term chemically pure denotes a compound which has 80 % or greater by weight of the (4aS,SS,7aS) stereoisomer and 20% or less by weight of other stereoisomers. In a further embodiment, the compound of formula (I) has 90 % or greater by weight of the S.7aS) stereoisomer and 10% or less by weight of other stereoisomers. In a yet r embodiment, the nd of formula (I) has 95 % or greater by weight of the S,7aS) stereoisomer and 5% or less by weight of other stereoisomers. In a still further embodiment, the compound of formula (I) has 97 % or greater by weight of the (4aS,SS,7aS) stereoisomer and 3% or less by weight of other stereoisomers.

In the present specification, although crystal polymorphs of the compound may be present, the compound is similarly not limited thereto and may be present as a single crystal form or a mixture of single crystal forms. The compound may be an anhydride or a hydrate. Any of these forms is included in the claims of the present cation.

The present invention also includes isotopically—labelled compounds, which are identical to the compounds of formula (I), except 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 usually found in nature. Examples of isotopes that can be incorporated into compounds of the invention include isotopes of hydrogen, carbon, nitrogen, oxygen, fluorine, phosphorous, chlorine, technetium and iodine, such as 2H, 3H, 11C, 14C, ”N. 150.. 181:, 3-1), 99mTC, 123I and 1311.

Compounds of the present invention and pharmaceutically acceptable derivatives (e. g. 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— 3O labelled compounds of the t invention, for example those into which radioactive isotopes such as H and/or 14C are incorporated, are useful in drug and/or substrate tissue distribution assays. H and 14C are considered useful due to their ease of preparation and detectability. 11C, 15Oand 18F es are considered useful in PET ron emission tomography), and 99mTc, 1231 and 1311 isotopes are considered useful in SPECT (single photon enrission computerized tomography), all useful in brain imaging. Substitution with heavier isotopes such as 2H can afford certain therapeutic advantages resulting from greater metabolic ity, for example increased in viva half—life or reduced dosage ements and, hence, are considered useful in some stances. Isotopically labelled compounds of formula (I) of this invention can generally be prepared by ng out the procedures disclosed in the s and/or in the Examples below, by substituting a readily available ically labelled reagent for a non-isotopically labelled reagent.

The fused aminodihydrothiazine derivative of the formula (I) according to the present invention may be a pharmaceutically acceptable salt. Pharmaceutically acceptable salts include those described by Berge, Bighley and Monkhouse, J. Pharm.

Sci, 1977, 766, 1—19. Specific examples of the pharmaceutically acceptable salt include inorganic acid salts (such as sulfates, es, orates, phosphates, carbonates, bicarbonates. hydrofluorides, hydrochlorides, hydrobromides and hydroiodides), c carboxylates (such as es, oxalates, maleates, tartrates, fumarates, citrates, malonates and lactates), organic sulfonates (such as methanesulfonates, trifluoromethanesulfonates, ethanesulfonates, benzenesulfonates, toluenesulfonates and camphorsulfonates), amino acid salts (such as aspartates and glutamates), nary amine salts, alkali metal salts (such as sodium salts and potassium salts) and alkali earth metal salts (such as magnesium salts and calcium salts).

The compound of the formula (1) according to the present invention can be converted to a pharmaceutically acceptable salt by a conventional method where necessary. The salt can be prepared by a method in which methods lly used in the field of organic tic chemistry and the like are riately combined.

Specific examples of the method include neutralization titration of a free solution of the compound of the present invention with an acid solution.

The fused aminodihydrothiazine derivative of the formula (I) or pharmaceutically acceptable salt according to the present invention may be a solvate thereof. Examples of a solvate include a hydrate.

The compound of the formula (1) according to the present invention can be converted to a solvate by subjecting the compound to a solvate g reaction known per 56 where ary.

The present invention further es a compound of formula (I) or a 3O pharmaceutically acceptable salt thereof for use in therapy.

The fused aminodihydrothiazine derivative or pharmaceutically acceptable salt thereof or solvate thereof according to the present ion has an excellent AB production inhibitory effect or BACEl inhibitory effect and is useful as a prophylactic or therapeutic agent for a neurodegenerative disease caused by AB and typified by Alzheimer-type dementia. The compounds of the invention reduce both AB4O and AB42.

Furthermore, the compounds of the present invention may have a BACE 2 inhibitory effect.

WO 98213 Thus, in another aspect, the present invention provides a compound of formula (I) as defined above, or a pharmaceutically acceptable salt thereof, for inhibiting production of amyloid-B protein.

In a further aspect, the present invention provides a nd of formula (I) as d above, or a pharmaceutically acceptable salt thereof, for inhibiting ite amyloid—B precursor protein cleaving enzyme 1 (BACE 1).

In a further , the present invention provides a compound of formula (I) as d above, or a pharmaceutically acceptable salt thereof, for ng a neurodegenerative disease. Examples of neurodegenerative diseases include Alzheimer—type dementia (AD), Down’s syndrome, cerebrov ascular amyloid angiopathy (CAA), mild cognitive impairment (MCI), memory loss, presenile dementia, senile dementia, hereditary cerebral hemorrhage with amyloidosis, and other degenerative dementias such as dementias of mixed ar and degenerative origin, dementia associated with supranuclear palsy, ia associated with cortical basal degeneration, dementia associated with Parkinson's Disease (PD), and dementia associated with diffuse Lewy Body type of AD. In one embodiment, the neurodegenerative disease is Alzheimer—type dementia (AD).

In another aspect, the invention provides the use of a compound of formula (I) as defined above, or a ceutically able salt thereof, for the manufacture of a medicament for the treatment or tion of a neurodegenerative disease, such as Alzheimer—type ia (AD), Down’s syndrome, cerebrovascular d angiopathy (CAA), mild cognitive impairment (MCI), memory loss, presenile dementia, senile dementia, hereditary cerebral hemorrhage with amyloidosis, and other degenerative dementias such as dementias of mixed vascular and degenerative origin, dementia associated with supranuclear palsy, dementia associated with cortical basal degeneration, dementia associated with Parkinson's Disease (PD), and dementia associated with e Lewy Body type of AD. In one ment, the neurodegenerative disease is Alzheimer~type dementia (AD).

In another aspect, the invention provides a method of inhibiting production of amyloid—B protein and/or of ng or preventing a neurodegenerative disease, such as Alzheimer-type dementia (AD), Down’s syndrome, cerebrovascular amyloid athy (CAA), mild cognitive impairment (MCI), memory loss, presenile dementia, senile dementia, hereditary cerebral hemorrhage with amyloidosis, and other degenerative dementias such as ias of mixed vascular and degenerative origin, dementia associated with supranuclear palsy, dementia associated with cortical basal degeneration, dementia associated with Parkinson's Disease (PD), and dementia associated with diffuse Lewy Body type of AD to a human , involving administering subject in need thereof a therapeutically or prophylactically effective amount of a 20121’050833 compound of formula (I) or a pharmaceutically acceptable salt thereof. Examples of neurodegenerative diseases include those listed above. In one ment, the neurodegenerative disease is Alzheimer-type dementia (AD). “Effective amount” means an amount sufficient to cause a benefit to the subject or at least to cause a change in the subject’s condition. onal conditions which may be treated by the compounds of the present invention include type 2 diabetes, Creutzfield-Jakob Disease (CJD), peripheral nerve injury, peripheral neuropathy, progressive supra-nuclear palsy, stroke, amyotrophic lateral sclerosis (ALS), autoimmune diseases, inflammation, arterial thrombosis, anxiety disorders, psychotic disorders, epilepsy, seizures, convulsions, stress disorders, ar amyloidosis, pain, Gerstmann—Straeussler—Scheinker syndrome, scrapie, encephalopathy, spino cerebellar ataxia, Wilson‘s e, Graves Disease, gton's Disease, Whipple's Disease, Kostmann Disease, glaucoma, hereditary cerebral hemorrhage with amyloidosis, cerebral hemorrhage with amyloidosis, vascular amyloidosis, brain inflammation, e X syndrome, stroke, Tourette's syndrome, inclusion body myositis, stress ers, depression, bipolar disorder and obsessive compulsive disorder.

In one aspect the present invention further provides a compound of formula (I) as defined above, or a pharmaceutically acceptable salt thereof, for treating type 2 diabetes.

In a further aspect the present invention further provides the use of a compound of formula (I) as defined above, or a pharmaceutically acceptable salt thereof, for the manufacture of a medicament for the treatment or prevention of type 2 diabetes.

In a yet furher aspect the present invention further provides a method of inhibiting production of amyloid—B protein and/or of treating or ting type 2 diabetes involving administering to a human subject in need thereof a eutically or prophylactically effective amount of a compound of a (I) or a pharmaceutically acceptable salt thereof.

A r aspect of the invention provides a pharmaceutical composition comprising a compound of formula (I) as d above, or a pharmaceutically acceptable salt thereof, as active ingredient in association with a pharmaceutically acceptable carrier. The composition may be in any suitable form, depending on the intended method of administration. It may for example be in the form of a tablet, capsule or liquid for oral stration, or of a solution or suspension for administration parenterally.

The fused aminodihydrothiazine derivative or pharrnaceutically acceptable salt thereof according to the present invention can be formulated by a conventional method.

Preferable examples of the dosage form include s, coated tablets such as film s and sugar-coated s, fine granules, es, s, capsules, syrups, s, inhalants, suppositories, injections, ointments, eye drops, nasal drops, ear drops, asms and lotions.

These solid preparations such as tablets, capsules, granules and s can contain generally 0.01 to 100 wt%, and preferably 0.1 to 100 wt% of the fused aminodihydrothiazine derivative or pharmaceutically able salt thereof according to the present invention as an active ingredient.

The active ingredient is formulated by blending ingredients generally used as materials for a pharmaceutical preparation and adding an excipient, a disintegrant, a binder, a lubricant, a colorant and a corrective typically used, and adding a stabilizer, an emulsifier, an absorbefacient, a surfactant, a pH adjuster, a preservative and an antioxidant where necessary, for example, using a conventional method. es of such ingredients e animal and vegetable oils such as soybean oil, beef tallow and synthetic glyceride; hydrocarbons such as liquid paraffin, squalane and solid paraffin; ester oils such as odecyl myristate and isopropyl myristate; higher alcohols such as earyl alcohol and behenyl alcohol; a ne resin; silicone oil; surfactants such as polyoxyethylene fatty acid ester, sorbitan fatty acid ester, ol fatty acid ester, polyoxyethylene an fatty acid ester, polyoxyethylene hydrogenated castor oil and a polyoxyethylene—polyoxypropylene block copolymer; water—soluble polymers such as hydroxyethylcellulose, polyacrylic acid, a carboxyvinyl polymer, polyethylene glycol, polyvinylpyrrolidone and methylcellulo se; lower alcohols such as ethanol and isopropanol; polyhydric alcohols such as glycerol, ene glycol, dipropylene glycol and ol; sugars such as glucose and sucrose; inorganic powders such as silicic anhydride, magnesium aluminum silicate and aluminum silicate; and purified water.

Examples of the excipient used include lactose, corn starch, saccharose, glucose, mannitol, sorbitol, crystalline cellulose and silicon dioxide. es of the binder used include nyl alcohol, polyvinyl ether, methylcellulose, ethylcellulose, gum arabic, tragacanth, n, shellac, hydroxypropylmethylcellulose, hydroxypropylcellulose, polyvinylpyrrolidone, a polypropylene glycol-polyoxyethylene block copolymer and nieglumine. Examples of the disintegrant used include starch, agar, gelatin , crystalline cellulose, calcium carbonate, sodium bicarbonate, m citrate, dextrin, pectin and carboxymethylcellulose calcium. Examples of the lubricant used include magnesium stearate, talc, polyethylene , silica and hydrogenated vegetable oil. Examples of the colorant used include those permitted to be added to pharmaceuticals. Examples of the corrective used include cocoa powder, menthol, empasm, mentha oil, borneol and cinnamon powder. Obviously, the ients are not limited to the above additive ingredients.

For example, an oral preparation is prepared by adding the fused aminodihydrothiazine derivative or pharmaceutically acceptable salt thereof according -15_ PCT/EP20121050833 to the present invention as an active ingredient, an ent and, where ary, a binder, a disintegrant, a lubricant, a colorant, a corrective and the like, and then forming the mixture into powder, fine granules, granules, s, coated tablets, capsules or the like by a conventional method. Obviously, tablets or granules may be appropriately coated, for example, sugar coated, where necessary.

For example, a syrup or an injection preparation is prepared by adding a pH adjuster, a solubilizer, an isotonizing agent and the like, and a solubilizing agent, a stabilizer and the like where necessary by a conventional method. The injection may be a previously prepared solution, or may be powder itself or powder containing a le additive. which is dissolved before use. The injection can contain usually 0.01 to 100 wt%, and preferably 0.1 to 100 wt% of the active ingredient. Further, a liquid preparation for oral stration such as a suspension or a syrup can contain usually 0.01 to 100 wt%, and preferably 0.1 to 100 wt% of the active ingredient.

For example, an al preparation can be prepared by any tional method without specific limitations. As a base material, any of s materials usually used for a pharmaceutical, a quasi drug, a cosmetic or the like can be used.

Examples of the base material e materials such as animal and vegetable oils, mineral oils, ester oils. waxes, higher alcohols, fatty acids, silicone oils, surfactants, phospholipids, alcohols, polyhydric alcohols, water-soluble polymers, clay minerals and purified water. A pH adjuster, an antioxidant, a chelator, a preservative and fungicide, a colorant, a flavor or the like can be added where necessary. r, ingredients such as an ingredient having a entiation inducing effect, a blood flow enhancer, a bactericide, an antiphlogistic, a cell activator, Vitamin, amino acid, a humectant and a keratolytic agent can be blended where necessary.

The dose of the fused aminodihydrothiazine derivative or pharmaceutically acceptable salt thereof according to the present invention varies according to the degree of symptoms, age, sex, body weight, mode of administration, type of salt and specific type of disease, for example. Typically, the active ingredient is orally administered to an adult at about 30 pg to 10 g, ably 100 pg to 5 g, and more preferably 100 pg to l g per day, or is administered to an adult by injection at about 30 pg to 1 g, preferably 100 pg to 500 mg, and more preferably 100 pg to 300 mg per day, in one or several doses, respectively. nds of the formula (I) may be used in ation with other therapeutic agents, for example medicaments claimed to be useful as either disease modifying or symptomatic treatments of a neurodegenerative disease such as Alzheimer’s disease. Thus, in a further aspect, the present invention provides a pharmaceutical product comprising, in combination, a first active ingredient which is a compound of formula (I) or a pharmaceutically able salt thereof and at least one ~16- 20121050833 r active ingredient useful in treating a neurodegenerative disease. In one embodiment of the invention, the neurodegenerative disease is Alzheimer—type dementia (AD). Suitable examples of such further active ingredients may be symptomatic agents, for example those known to modify ergic transmission such as M1 and M3 muscarinic or agonists or allosteric modulators, M2 muscaiinic antagonists, M4 ts or positive allosteric modulators (PAMs), acetylcholinesterase inhibitors (such as ydroaminoacridine, donepezil hydrochloride and rivastigrnine), nicotinic receptor agonists or allosteric modulators (such as 0:7 agonists or allosteric modulators or (1432 agonists or allosteric modulators), PPAR agonists (such as PPARy ts), 5—HT4 receptor agonists or partial agonists, histamine H3 antagonists, 5—HT6 receptor antagonists or SHTIA receptor ligands and NMDA receptor antagonists or modulators, 5-HT3A antagonists, 5—HT7 nists, Dl agonists or PAMs, D4 agonists or PAMs, D5 agonists or PAMs, GABA—A (15 e agonists or negative allosteric modulators (NAMs), GABA-A (12/3 agonists or PAMs, mGluRZ modulators (PAMs or NAMs), mGluR3 PAMs, mGluRS PAMs, PDE 1 inhibitors, PDE 2 inhibitors, PDE 4 inhibitors, PDE 5 inhibitors, PDE 9 inhibitors, PDE 10 tors, GlyTl tors, DAAO inhibitors, ASCl inhibitors, AMPA modulators, SIRTl activators or inhibitors, AT4 antagonists. GalRi nists, GalR3 ligands, adenosine A1 antagonists, adenosine A2a antagonists, (12A antagonists or agonists, selective and unselective norepinephrine reuptake inhibitors (SNRIS), or potential disease modifying agents such as gamma secretase inhibitors or modulators, alpha secretase activators or tors, amyloid aggregation inhibitors, amyloid antibodies, tau aggregation inhibitors or tau orylation/kinase inhibitors, tau dephosphorylation / atase activators, mitogen—activated protein kinase kinase 4 (MKK4/MEK4/MAP2K4) inhibitors, c~Jun N—terminal kinase (JNK) inhibitors, casein kinase inhibitors, MKZ (mitogen activated protein —activated protein kinase 2) inhibitors, MARK (microtubule affinity regulating kinase) inhibitors, CDKS (cyclin dependent kinase 5) inhibitors, GSK—3 (glycogen synthase kinase~3) inhibitors and tau-tubulin kinase—l (TTBKl) inhibitors.

Further examples of such other therapeutic agents may be calcium channel blockers, 3O A (3-hydroxy—3—rnethyl~glutaryl-COA) reductase inhibitors (statins) and lipid lowering , NGF (nerve growth factor) mimics, antioxidants, GPR3 ligands, plasmin activators, neprilysin (NEP) activators, IDE (insulin degrading enzyme) activators, melatonin MTl and/or MT2 agonists, TLX/NRZEl (tailless X receptor) ligands, GluRl ligands, RAGE (receptor for advanced ion end—products) antagonists, EGFR (epidermal growth factor receptor) inhibitors, FPRL-l (formyl peptide-like receptor-1) ligands, GABA antagonists, and MICAL (molecule interacting with casL) inhibitors, eg. oxoreductase inhibitors, CB1 antagonists/inverse agonists, non—steroidal nflammatory drugs (NSAIDs), anti-inflammatory agents (for -17_ example agents that could be used to treat neuroinflarnmation either by enhancing or reducing neuroinflammation). amyloid sor protein (APP) ligands, anti-amyloid vaccines and / or antibodies, agents that promote or enhance amyloid efflux and / or clearance, histone deacetylase (HDAC) inhbitors, EP2 antagonists, ll—beta HSDl (hydroxysteroid dehydrogenase) inhibitors, liver X receptor (LXR) agonists or PAMs, lipoprotein receptor-related protein (LRP) mimics and / or ligands and/or ers and/or inhibitors, butyryl cholinesterase inhibitors, kynurinic acid antagonists and / or inhibitors of kynurenine aminotransferease (KAT), orphanin FQ / nociceptin (NOP) / opioid—like receptor 1 (ORLl) antagonists, excitatory amino acid transporter (EAAT) ligands (activators or inhibitors), and plasminogen activator inhibitor-1 ) inhibitors, niacin and /or GPR109 agonists or PAMs in combination with cholesterol ng agents and / or HMGCoA reductase inhibitors (statins), dimebolin or similar agents, antihistamines, metal binding / ing , antibiotics, growth hormone secretagogues, cholesterol lowering agents, vitamin E, cholesterol absorption tors, cholesterol efflux promoters and / or tors, and insulin upregulating agents.

In one embodiment, the present invention provides a ceutical product sing, in combination, a first active ingredient which is a compound of formula (1) or a pharmaceutically acceptable salt thereof and at least one further active ingredient selected from:- 0 cholinesterase inhibitors, e.g. donepezil, galantamine, rivastigamine, tetrahydroaminoacricline and phartnaceutically acceptable salts thereof, 0 5—HT6 antagonists, e.g. 457 and pharmaceutically acceptable salts thereof, 0 HMGCOA reductase inhibitors e.g. lovastatin, statin, atorvastatin, simvastatin, fluvastatin, pitavastatin, pravastatin and pharmaceutically able salts thereof.

The individual components of such combinations may be administered either sequentially or simultaneously in separate or combined pharmaceutical formulations.

Consequently, the pharmaceutical product may, for example be a pharmaceutical composition comprising the first and further active ients in admixture. 3O Alternatively, the pharmaceutical product may for example comprise the first and further active ingredients in separate ceutical preparations suitable for simultaneous, sequential or separate administration to a patient in need thereof The combinations referred to above may conveniently be presented for use in the form of a pharmaceutical formulation and thus pharmaceutical formulations comprising a ation as defined above together with a pharmaceutically acceptable carrier or excipient comprise a further aspect of the invention.

When a compound of formula (I) or a pharmaceutically able salt thereof is used in combination with a second therapeutic agent active, the dose of each compound —18- may differ from that when the compound is used alone. Appropriate doses will be readily appreciated by those skilled in the art.

Thus, an additional aspect of the invention provides a method of preparation of a pharmaceutical composition, involving admixing at least one nd of formula (I) as defined above, or a pharmaceutically acceptable salt therof, with one or more ceutically acceptable adjuvants, diluents or carriers and/or with one or more other therapeutically or prophylactically active agents.

In one embodiment, the present invention es a ceutical composition comprising a compound of formula (I) or a pharmaceutically acceptable salt thereof, one or more other agents for the treatment of Alzheimer’s disease, such as an M1 and M3 muscarinic receptor agonist or eric modulator, an M2 muscarinic nist, an acetylcholinesterase tor, a nicotinic receptor agonist or allosteric tor, a PPAR agonist, a 5~HT4 receptor agonist or partial agonist, a histamine H3 antagonist, a 5—HT6 receptor antagonist, a SHTIA receptor ligand, a NMDA receptor antagonist or modulator, a 5-HT2A antagonist, a 5—HT; antagonist, a D1 agonist or positive allosteric modulator (PAM), a D4 agonist or PAM, a GABA—A a5 inverse agonist or negative allosteric modulator (NAM), a GABA—A (12/3 agonist or PAM, a mGluR2 modulator (PAM or NAM), a mGluR3 PAM, a mGluRS PAM, a PDE 1 inhibitor, a PDE 2 inhibitor, a PDE 4 inhibitor, a PDE 5 inhibitor, a PDE 9 inhibitor, a PDE 10 inhibitor, a GlyTl inhibitor, a DAAO inhibitor, a ASCl inhibitor, a AMPA tor, a SIRTl activator or inhibitor, a AT4 antagonist, a GalRl antagonist, a GalR3 , an adenosine A1 antagonist, an adenosine A221 antagonist, an a2A antagonist or agonist, a selective or unselective norepinephrine reuptake inhibitor (SNRI), a gamma secretase inhibitor or modulator, an alpha secretase activator or tor, an amyloid aggregation inhibitor, an d antibody, a tau aggregation inhibitor, a tau orylation inhibitor, a MK2 (mitogen activated protein ldnase— activated protein kinase 2) inhibitor, a MARK (microtubule affinity regulating kinase) inhibitor, a CDK5 (cyclin dependent kinase 5) inhibitor, 21 GSK—3 (glycogen se -3) inhibitor, a calcium channel blocker, a HMG—CoA (3—hydroxymethyl~ 3O glutaryl—COA) reductase inhibitor (statin) and a lipid lowering agent, a NGF (nerve growth factor) mimic, an antioxidant, a GPR3 ligand, 21 plasmin tor, a neprilysin (NEP) activator, an lDE (insulin degrading enzyme) activator, a melatonin MTl and/or MT2 agonist, a TLX (tailless X receptor) ligand, a GluRl ligand, a RAGE (receptor for advanced glycation end-products) antagonist, an EGFR (epidermal growth factor receptor) inhibitor, a FPRL—l (formyl peptide-like receptor—l) ligand, a GABA antagonist or a MICAL ule interacting with casL) tor such as an oxoreductase inhibitor, in association with a ceuticalyy acceptable r. In a further embodiment the present invention provides a combination comprising a compound of formula (I) or a pharmaceutically acceptable salt thereof, er with a further therapeutic agent as described herein above for sequential or simultaneous administration in separate or combined pharmaceutical formulations.

In a further aspect, the invention provides a method of inhibiting production of amyloid—[3 protein and/or of ng or ting a neurodegenerative disease, such as Alzheimer—type dementia (AD), Down’s syndrome, cerebrovascular amyloid angiopathy (CAA), mild cognitive impairment (MCI), memory loss, presenile dementia, senile dementia, hereditary cerebral hemorrhage with amyloidosis, and other degenerative dementias such as dementias of mixed vascular and degenerative origin, dementia associated with supranuclear palsy, dementia associated with cortical basal degeneration, dementia associated with Parkinson's Disease (PD), and dementia associated with diffuse Lewy Body type of AD, the method involving administering to a human subject suffering from the condition a therapeutically or prophylactically ive amount of the pharmaceutical composition described above or of a compound of formula (I) as defined above, or a pharmaceutically acceptable salt thereof. tive amount” means an amount sufficient to cause a benefit to the subject or at least to cause a change in the subject’s ion.

Alzheimer’s Disease (AD) is characterized pathologically by the presence of neurofibrillary tangles (NFTs) and plaques, consisting of amyloid (AB) es of varying length, for e 42 amino acids (A642) and 40 amino acids . In on to these pathological markers, brain y is also evident. The build up of plaques is believed to be due to the ation of AB peptides. AB peptides are formed in the brain by the sequential cleavage of amyloid precursor protein (APP) by {3- secretase l) and 'y—secretase. Therefore potential AD drugs aimed at inhibiting amyloid formation by inhibiting BACE—l or ’y—secretase, must be able to achieve adequate exposure in the brain, in order to exert an effect on AD.

Although BACE~l represents an attractive target to halt or slow the production of amyloid peptides, various groups have found it challenging to identify BACE-l inhibitors that can penetrate the central nervous system (CNS) and thus inhibit the 3O enzyme at the site of action.

The brain is protected by several barriers including the blood brain barrier (BBB) and transporters (Hitchcock and Pennington, J Med Chem 2006, 29, 7559; Ueno, Curr.

Med. Chem. 2007, I4, ll99; Gloor et al., Brain Res. Rev. 2001, 36, 258). Several efflux transporters have been terised which prevent nds entering the brain.

One of the best characterised and most prominent in preventing the CNS penetration of xenobiotics is P~g1ycoprotein (ng) ara and Sugiyama, Drug Discovery Today, 2001, 6, 150', Mahar Doan et al., J. Pharm. Expt. Ther. 2002, 303, 1029; Lin, Drugs of Today 2004, 40, 5; Lin & Yamazaki, Clin Pharmacokinet. 2003, 42, 59; Schinkel, Adv.

PCT/EP20121050833 Drug Deliv. Rev. 1999, 36, 179). It has been shown that ng efflux is important for BACE—l inhibitors (Hussain et 211., J. Neurochem. 2007, 100, 802). Thus. ming ng efflux is important.

Those skilled in the art will iate that there are l ways to measure or predict CNS penetration in vitro or in. vivo. The potential for CNS penetration can be assessed in vitro by determining r a compound can be ted to ng efflux, i.e. by conducting an in vitro ng assay. Those skilled in the art will appreciate that a number of cell lines can be used and that these cell lines may or may not affect the results of the assay. One such assay is described below (Cyprotex UK).

The following MDR-J MDCK assay was used to assess ng efi‘lax. The assay was conducted at Cyprotex Discovery Ltd. 15 Beech Lane, Macclesfield, Cheshire, UK, SKIO ZDR MDRl-MDCK Permeability (Bi-directional; pH 7.4/pH 7.4) Protocol Summary MDCK cells are an lial cell line of canine kidney origin. These cells can be transfected to stably express active P—glycoprotein (MDRl-MDCK) and are ideal for studying drug efflux. Test nd was added to either the apical or basolateral side of a confluent monolayer of MDRl—MDCK cells and permeability was measured by monitoring the appearance of the test compound on the opposite side of the membrane using LC—MS/MS. From this an apparent permeability (Papp) coefficient and efflux ratio was measured/calculated.

Objective To measure the permeability of test compound in the apical to basolateral (AB) and basolateral to apical (B—A) direction across MDRl-MDCK cells. A ratio of B—A and A-B permeabilities was calculated (efflux ratio) to show whether the compound undergoes P-glycoprotein efflux. nds were provided as a 200 uL solution of 10 mM test compound in DMSO.

Experimental Procedure MDRl—MDCK cells obtained from the NIH (Rockville, MD, USA) were used.

Following culture to confluency, the monolayers were prepared by g both basolateral and apical surfaces twice with pH 7.4 buffer at 37°C. Cells were then ted with pH 7.4 buffer in both apical and basolateral compartments for 40 min to stabilise physiological parameters.

PCT/EP20121’050833 Buffer at pH 7.4 was then removed from the apical compartment and replaced with test compound dosing solutions. The solutions were prepared by diluting 10 mM test compound in DMSO with buffer to give a final test compound concentration of 10 uM (final DMSO tration adjusted to 1%). The fluorescent integrity marker Lucifer yellow was also included in the dosing solution. The apical compartment. inserts were then placed into ‘companion’ plates containing fresh buffer at pH 7.4.

Analytical standards were made from dosing ons.

For basolateral to apical (B—A) experiments the experiment was initiated by replacing buffer in the s then g them in companion plates containing dosing solutions. Incubations were carried out in an atmosphere of 5% CO; with a relative humidity of 95% at 37°C for 60 minutes.

After the incubation period, the companion plate was d and apical and basolateral samples diluted for analysis by LC-MS/MS. Test compound permeability was ed in duplicate. On each plate compounds of known permeability characteristics were run as controls.

Test and control compounds were quantified by LC—MS/MS cassette analysis using a 5-point calibration with appropriate dilution of the samples. Cyprotex generic analytical conditions were used. The starting tration (CO) was determined from the dosing solution and the experimental recovery calculated from C0 and both apical and teral compartment concentrations.

The integrity of the monolayers throughout the experiment was checked by monitoring r yellow permeation using fluorimetric analysis. Lucifer yellow permeation is low if monolayers have not been damaged. If a r yellow Papp value was above QC limits in one individual test compound well, then an n=l result was reported. If Lucifer yellow Papp values were above QC limits in both replicate wells for a test compound, the compound was re—tested. If on repeat. high Lucifer yellow permeation was observed in both wells then toxicity or inherent fluorescence of the test compound was assumed. No further experiments were performed in this ce.

Data Analysis The permeability cient for each compound (Paw) was calculated from the ing equation: Papp = (dQ + dt) + (C0 x A) Where dQ/dt is the rate of permeation of the drug across the cells, C0 is the donor compartment concentration at time zero and A is the area of the cell monolayer. Co was obtained from analysis of the dosing solution at the start of the experiment.

In addition, an efflux ratio (ER) was calculated from mean A-B and B-A data.

This is derived from: PCT/EP20121050833 ER = ((Papp (B “ A» + ((Papp (A _ B» Two control compounds were screened ide the test compounds, propranolol (highly permeable) and prazosin (a substrate for P-glycoprotein).

Suipi'isingly, compounds from the present invention wene found to display a lower ng efflux ratio than compounds exemplified in W02009/091016 indicating that they have the potential to show higher CNS penetration. Data for ed examples are shown in Table 1 below. 1 0 Table I: MDR-I MDCK P I assa data Exam-1e Pu ER Comparative Example 1 26.2 Comarative Exam-1e 2 16.6 Com. arative Exam 1e 3 24.0 Com - arative Exam le 4 20.7 Com - arative Exam n 16 5 Com-arative Examle 6 53.439??? mqqooq 18 t—‘Qf‘f‘t‘pt‘t‘FLFOQOOONQOOOUI Note: Example 14 is intentionally not included.

Comparative Examples 1 to 6 are covered by hed International patent application W02009/091016; Comparative Examples 1 to 4 are specifically described in WO2009/091016 as Examples 32, 35, 54 and 73 respectively.

Comparative Examples 5 and 6 are N—(3—((4aS,SS,7aS)—2—amino—5—(fluoromethyl)—4a,5,7,7a-tetrahydro—4H-furo[3,4- d][l ,3]thiazin—7a—y1)-4—flu0ropheny1)—5-ethoxypicolinarnide, and N—(3-((4aS,5R,7aS)—2—aminomethyl-4a,5,7,7a—tetrahydr0-4H—furo[3,4-d][1,3]thiazin— 7a—yl)fluorophenyl)—5«ethoxypicolinamide respectively.

The data demonstrate that compounds of the present invention and specific examples 1 - 13 and 15 ~ 18 have lower ng efflux and therefore potentially higher CNS penetration than representative examples from W02009/091016 using the aforementioned recognized method of assessing CNS penetration. For example, Comparative Examples 1 to 6 have higher ng efflux ratios than compounds of the present invention. Furthermore, Comparative Examples 5 and 6 have higher ng efflux ratios than a close analogue, Example 10 of the present invention, which clearly demonstrates the beneficial effect the trifluoromethyl group on the tetrahydrofuran ring exerts on ng efflux, i.e. the trifluoromethyl group reduces ng efflux.

Those skilled in the art will iate that the in vitro ng assay bed above is a predictive assay of in vivo CNS penetration. It is thus also highly desirable if the decreased ng-mediated efflux translates to the in. viva situation. Those skilled in the art will appreciate there are many ways to assess the CNS penetration of compounds in viva. For example, one can quantify compound concentrations in blood or plasma and brain and calculate a blood (BrzBl) or brainzplasma (BrzPl) ratio.

This method has been used historically and has been widely accepted as a method of determining CNS penetration (Summerfield et al., J col. Expt. Ther. 2007, 322, 205). Those skilled in the alt will appreciate that this type of assay could be conducted at steady state, a single time point, multiple time points or could be done by quoting Area Under the Curve (AUC) ratios. All methods are equally valid but each may have 3O certain caveats that will be appreciated by those skilled in the art. Recent literature has been published to suggest that it is important to er the free concentrations in vivo and that when no efflux occurs from the brain the free plasma concentration should be the same or equivalent to the free brain concentration (Kalvass and , Biopharmaceutics & Drug Disposition 2002, 23, 327; Mauer et a1, Drug Metab.

Disposition 2005, 33, 175; r Expert Opin. Drug Discov. 2007, 2, 51). Thus, a compound that can freely penetrate the CNS and is not ted to active efflux, for example by ng or another orter, should demonstrate a free free plasma (Brfr: Pifr) or an unbound brainzunbound plasma (Bru : Plu) of approximately 1:1.

WO 98213 Those skilled in the art will appreciate that the free or unbound concentrations can be calculated by multiplying the total brain or total plasma concentration by the fraction unbound in brain tissue or plasma, which can be measured by the assay described below.

Those skilled in the art will appreciate that the fraction unbound may change with experimental factors, for example concentration, or ature, etc. Those skilled in the art will be able to assess this and select the most appropriate set of conditions.

Those skilled in the art will also appreciate that as long as the ions are the same for each compound screened then the assay will give consistent data for the range of compounds tested thus minimising any discrepancies. It has also been proposed that drug concentrations in cerebrospinal fluid (CSF) are equivalent to free brain concentrations for compounds which are not actively effluxed from the brain (He et al., Xenobiotica 2009, 39, 687). Thus another method of determining CNS penetration would be to assess the CSF2free plasma (CSF : Pifr) or CSF :unbound plasma (CSF : Plu). If the free drug in plasma is able to permeate into the CNS and is not actively influxed or effluxed then the CSFZPIfi» or CSF:P1u should be approximately 1:1. Those skilled in the art will appreciate the issues associated with ining CSF drug concentrations and extracting CSF, for e CSF can be contaminated by blood depending on the method of awal. also the CSF concentrations may be of lower accuracy, depending on the dose used.

Thus it has been shown that a BACE inhibitor from GlaxoSmithKline (GSK188909), BACE-l leo SnM, which has low CNS exposure was ineffective at lowering A1340 production in the brains of TASTPM mice (which overexpress both [MS961 human APPsweKj95N and PS—1M146V) upon acute administration (Hussain et al., J.

Neurochem. 2007, 100, 802 — 809). Following an oral dose of 250mg/kg the brain tration of GSKl88909 in TASTPM mice was 0.62uM. When a ng inhibitor (GF120918) was dosed 5 hours before the oral administration of GS Kl 88909, the brain concentration of GSK188909 was found to be 5.43uM following an oral dose of 250mg/kg, i.e. the inistration of a ng inhibitor caused an almost 9-fold increase in CNS penetration, showing ng efflux is an important mechanism in preventing BACE inhibitors from penetrating the CNS. Furthermore, in the absence of a ng inhibitor, 21 250mg/kg oral dose of GSK188909 did not have any effect on brain A840 levels in TASTPM mice, whereas when a ng inhibitor was inistered (5 hours prior to the stration of 909) a 68% reduction in brain A840 levels relative to vehicle treated mice was observed. r paper has reported a similar effect with three BACE-l inhibitors from Bristol-Myers Squibb (Meredith et al., J. Pharm. Expt. Ther. 2008, 326, 502-513). The three reported compounds were found to be ng substrates in vitro. When dosed to mice, the three compounds showed low CNS penetration and did not lower amyloid 2012/050833 levels in the brain but were able to lower plasma amyloid levels. When the same three compounds were administered to ng knockout (KO) mice, the level of CNS penetration increased and the nds were able to lower amyloid levels in the brain.

Researchers at Schering-Plough have also published papers oh et al._.

Bioorg. Med. Chem. Lett. 2008, 18, 418) to show that BACE-l inhibitors from their series (e.g. example 11 from the aforementioned reference), are subject to ng efflux, as a result of which the compound was found to display a low BrzPl (<01) in the rat.

The literature cited above emphasises the difficulties in identifying BACE—l inhibitors which are not subjected to ng efflux. Such inhibitors would be highly desirable and many research groups have attempted to discover such compounds without success. Thus BACE-l tors which are not ng substrates and can therefore readily penetrate the CNS and lower amyloid in the brain would be ble.

More recently, researchers at. Wyeth have reported extensive work to overcome ng efflux in a series of cyclic acylguanidine BACE~1 inhibitors (Malamas et al, Bioorg.

Med. Chem. Lett. 2010, 20, 6597). Compounds were discovered that were weak ng substrates and with BrzPl approaching 1: 1. However, two lead examples with d ng efflux (84 and 89 from the aforementioned reference), did not lower A040 in the brain of Tg2576 mice 8 hours after a 30 mg/kg oral dose. The lack of efficacy was attributed to the fact the compounds showed high brain tissue binding. Thus, it is important to discover BACE-l tors that are not ng substrates but still have a reasonable unbound fraction in brain tissue and are able to lower amyloid in the brain.

It has also been shown that BACE inhibitors that are not ng substrates in vitro can penetrate the CNS (e.g. TC-l from Merck), and can lower A040 levels in the brain of C mice and monkeys (Sankaranarayanan et al., J. Pharmacol. Expt. Ther. 2009, 328, 0). Thus, in vitro ng assays showed TC-l not to be a ng substrate and when TC-l was dosed to APP~YAC mice (lOOmg/kg i.p.) it was able to modestly penetrate the CNS as shown by the brain concentrations and the brainzplasma ratio and this ability ed in moderate lowering of brain amyloid.

Time Plasma conc. Brain conc. Br:Pl Reductionin brain (HM) (HM) A40 (%) _——m— Brain and plasma concentration of T01 following lOOmg/kg i.p. dose and corresponding effects on brain Ab40 levels, in APP—YAC mice.

In separate experiments it was shown that TC-l could penetrate the CSF of monkeys when co—administered with a CYP3A4 tor (ritonavir). In these —26~ PCTIEP2012/050833 experiments the e plasma concentration of TC-l was found to be 2.7uM, whilst the CSF concentration was found to be 0.025uM. However. as TC—l is ~99% bound to plasma proteins the free plasma concentration was calculated to be ~0.027nM. It was found that CSF A040 levels showed a 42% decrease relative to a vehicle treated l group. Thus, a BACE inhibitor that can freely penetrate the CNS would be expected to be able to lower amyloid levels in the CNS. It would be beneficial not to have to be co—dosed with a CYP3A4 inhibitor.

The compounds of the present invention have been shown to lower AB production in cellular assays which correlates with their ability to lower AB production in animals. Thus, the compounds of the present invention will have utility in lowering AB production in humans and thus will be useful in the treatment of neurodegenerative diseases such as Alzheimer’s disease.

Rat in viva CNS penetration Male Sprague Dawley rats were acquired from Charles River UK Ltd. (Margate, UK) and housed according to UK Home Office guidelines. Drugs were made up to the appropriate concentrations in 0.5% methyl ose. Animals were dosed orally (2mL/kg) by gavage at the doses outlined in Tables 2 to 4 below.

At the time points post-dosing, specified in Tables 2 to 4 below the animals were administered an i.p. injection of sodium pentobarbitone (approximately 330mg/kg for terminal anaesthesia).

Using a tine. the animals were decapitated and trunk blood ted into 15ml Falcon tubes containing 100 IU heparin. Blood was ed followed by fugation at 6000rpm, 4°C for 5 minutes. Plasma was collected for DMPK and ELISA assays and stored at —80°C until use. Brains were dissected out and divided along the midline, weighed and stored at -80°C until further use.

Method for Analysis of Plasma3 Brain and CSF Samples ation of Acetonitrile Working Solutions Test compound was prepared as a 1 mg free base/ml. on in DMSO, ed and sonicated for 5 min. The 1 mg/mL DMSO solution was diluted to 10 and ug/mL acetonitrile stocks, by adding 10 uL to 990 uL acetonitrile and 30 pL to 970 uL acetonitrile. respectively. The 10 and 30 ug/mL acetonitrile stocks were then serially diluted 1:9 (v/v) (100 uL stock into 900 uL acetonitn'le) to give the following solutions: 0.003, 0.01. 0.03, 0.1, 0.3, l, 3, 10 and 30 ug/mL itrile.

Pre aration of Plasma Standards. Blanks and Sam les 2012/050833 Control male e Dawley rat plasma and the study plasma samples were stored at —80°C until the day of analysis when they were thawed at room temperature.

Control plasma was centrifuged (2,000g for 10 min) and aliquoted (90 nL) into eppendorf tubes for preparation of standards and blank samples. Study samples were previously ted (100 nL) into eppendorf tubes immediately following collection of the plasma.

An aliquot (10 uL) of the appropriate acetonitrile stock was added to the control plasma (to give a final volume of 100 uL) to give the required calibration standards covering the range 1 — 3000 ng/mL. Double blank and blank s were prepared by adding 10 uL of acetonitrile to 90 uL of blank plasma.

Preparation of Brain Standards. Blanks and s Control male Sprague Dawley rat brain and the study brain samples were weighed after tion and stored at —80°C until the day of is when they were thawed at room temperature. Once thawed brains were d with water (4 mL per gram of tissue) and homogenised using a mechanical homogeniser. An t (100 uL) of each study sample was taken into Micronics tubes ready for analysis and sufficient aliquots (90 pL) of control brain homogenate prepared for preparation of standards and blanks.

An aliquot (10 uL) of the appropriate acetonitrile stocks was added to the control brain homogenate (to give a final volume of 100 uL) to give the required calibration standards covering the range 1.5 — 5000 ng/g. Double blank and blank samples were prepared by adding 10 til. of acetonitrile to 90 uL of blank brain homogenate.

Extraction of Plasma and Brain Samples. Standards and Blanks Each plasma and brain homogenate sample, standard and blank (100 uL) was extracted with an aliquot (300 nL) of acetonitrile (containing 0.1% formic acid and 100 ng/mL of an appropriate internal standard). Double blanks were extracted with an aliquot (300 uL) of acetonitrile containing 0.1% formic acid). All samples, standards 3O and blanks were then vortex mixed and centrifuged (2000g for 15 min). An aliquot (50 uL) of the resulting supernatant was then taken into a 2 mL p well plate and diluted with acetonitrilezwater (50:50 v/v) (150 uL) ready for analysis by a specific LC- MS/MS method.

Preparation of CSF Samples. Standards and Blanks Control male Sprague Dawley rat CSF and the study CSF samples were stored at -80°C until the day of analysis when they were thawed at room ature. An aliquot (50 uL) of each study sample was taken into Micronics tubes ready for analysis ~28- and sufficient ts (45 pL) of control CSF prepared for ation of standards and blanks.

An aliquot (5 uL) of the appropriate acetonitrile stocks was added to the control CSF (to give a final volume of 50 nL) to give the required calibration standards covering the range 1 ~ 1000 ng/mL. Double blank and blank samples were prepared by adding 5 uL of acetonitrile to 45 pL of blank CSF.

Extraction of CSF Samples. Standards and Blanks Each CSF sample, standard and blank (50 uL) was extracted with an aliquot (150 nL) of acetonitrile (containing 0.1% formic acid and 100 ng/mL of an riate internal standard). Double blanks were ted with an aliquot (150 pL) of acetonitrile containing 0.1% formic acid. All samples were then vortex mixed and an aliquot (50 pL) of each was then further diluted in 150 nL of acetonitrilezwater (50/50 v/v) in a 2 mL 96-deep well block ready for LC~MSIMS analysis.

All samples were then analysed using a Waters Acquity UPLC coupled to a Waters Xevo TQ mass spectrometer.

LC Conditions: Column: Acquity UPLC BEH C18, 1.7 um, 2.1 x 50 mm, maintained at 40°C Mobile Phase: A = 95% Water : 5% MeOH containing 0.01M Ammonium e B : 5% Water : 95% MeOH containing 0.01M Ammonium acetate Gradient: Flow rate: 0.6 mL/min; injection volume 5 uL; mpler temperature 6°C LC flow was diverted to waste for the first 0.3 min of each injection MS/MS transitions were optimised automatically by Waters QuanOptimise software.

Amxloid detection DEA! NaCl Extraction of Afl peptides from rat brain: 2012/050833 100ml of chilled 0.2% diethyl amine (DEA) in 50 mM NaCl (pH 10) was freshly prepared and lull/25mg brain tissue was added to each hemisphere (i.e. 40x brain volume). The brains were immediately homogenized using a Polytron PT 1200 for 1.5 minutes and samples left to incubate on ice for one hour after homogenisation. 3ml of the homogenate was transferred to a polyallomer tube (Beckman 3) and spun at 133000 x g (55,0001pm) for 45 min at 4° C. The supernatant was then neutralised to pH 8—8.3 by adding 1/10 volume 0.5M Tris/HCl, pH 6.8. The samples can be used fresh or snap frozen on dry~ice and stored at —80°C until ed for is 1 0 Human] Rat fiAmyloid (40‘! ELISA (Wake Kit} The Wako AB40 ELISA kit (Code No. 294-62501) uses the monoclonal antibody BNT77, raised against epitope AB(l 1-28) and the monoclonal antibody BA27, which specifically s the C-terminal portion of AB40. This kit is used for the quantitative determination of human or rat ABM-40) and also N—terminally truncated l 5 AB40 s (AB(x—40)) in biological matrices such as tissue culture medium, tissue homogenate, CSF and plasma.

For analysis, plasma and brain samples are diluted 1:1 with the standard diluent contained in the kit and CSF samples are d 1:8 with the standard diluent contained in the kit. The assay is carried out according to manufacturers instructions and 2 0 samples are analysed in duplicate. Data is analysed using Microsoft Excel 2003 and statistical analysis is carried out. using Genstat. 9Lb Edition.

Thus, when Comparative Example 4 was administered at a dose of g p.0. and plasma, brain and CSF samples were collected 2, 4, 6 and 8 hours post-dose the following concentrations were measured (Table 2): Table 2: Data for Comarative Exam le 4 (h) (nM) (11M) (11M) (11M) (nM) III--0.“-24 1162 O7 0.14 _--- M--- 1 Calculated by multiplying the [Pl] by P1 bu. 2 Calculated by multiplying the [Br] by Br Fu.

From the above study, Comparative Example 4 showed a 59% and 64% ion of A1340 in the brain at 4 and 6 hours respectively; and a 76% and 70% reduction of AB40 in the CSF at 4 and 6 hours respectively.

WO 98213 Certain compounds of the present invention have been assessed in vivo in the rat to corroborate the levels of CNS penetration; these data are presented in the tables below.

Surprisingly, it has been found that compounds of the present ion show increased CNS penetration in the rat relative to compounds from W02009/091016 by any of the aforementioned recognixed s of determining CNS penetration. Thus, the compounds of the t invention may show improved profiles in that they more readily target the site of action, the brain, and therefore may show improved efficacy or efficacy at lower concentrations or doses or sed peripherally mediated side effects, by way of preferential CNS partitioning, or a combination of any or all of these aspects.

Thus, when Example 8 of the present invention was administered at a dose of 10mg/kg p.o. and plasma, brain and CSF samples were collected 2, 4, 6 and 8 hours post-dose the following concentrations were ed (Table 3): Table 3: Data for Examtle 8 (h) (HM) (11M) (HM) (HM) (HM) mama- _-----E- 1 Calculated by multiplying the [Pl] by Pl F‘u. 2 ated by multiplying the [Br] by Br Fu. 3. Not quantifiable. Concentrations close to or below the lower limit of quantification and could not be accurately quantified. 2O 4. Not determined.

From the above study, Example 8 showed a 68% and 72% reduction of A340 in the brain at 4 and 6 hours respectively; and an 82% and 74% reduction of A340 in the CSF at 4 and 6 hours respectively.Thus compounds of the present invention show decreased ng efflux relative to previous disclosures whilst trating efficacy in the CNS. The efficacy is thus achieved with lower circulating plasma concentrations.

When Example 1 of the present invention was administered at a dose of 10mg/kg p.o. and plasma, brain and CSF samples were collected 2, 4, 6 and 8 hours post-dose, the following concentrations were measured (Table 4): Table 4: Data for Examle l (11) (11M) (11M) (HM) (11M) (HM) 2012/050833 _-_-__--- _-_-__-_- 1 Calculated by multiplying the [Pl]]by P1 Fu. 2 ated by multiplying the [Br] by Br Fu.

From the above study, Example 1 showed a 64% and 70% reduction of AB40 in the brain at 4 and 6 hours respectively; and a 80% and 85% reduction of A340 in the CSF at 4 and 6 hours respectively. Thus compounds of the present ion show decreased ng efflux relative to previous inventions whilst demonstrating efficacy in the CNS. The efficacy is thus achieved with lower circulating plasma concentrations.

Method For Determination of Plasma Protein Binding (PPB) and Brain Tissue Binding (BTB) Compound Preparation nds were dissolved in DMSO to give a 1 mg free base/mL solution, before further dilution to 100 ug/mL in acetonitrile (100 uL of 1 mg/mL into 900 uL acetonitrile).

Matrix Preparation On the morning of dialysis, control male Sprague Dawley rat plasma and brain, previously stored at —80°C were thawed at room temperature. Plasma was checked for pH and if necessary adjusted to 7.4 with 1M HCl. Plasma was then centrifuged (2000 g for 10 min) and the brains d with 2 mL of ate Buffered Saline (pH 7.4) per gram of tissue and homogenised using a mechanical homogeniser. An aliquot (10 uL) of the 100 ug/mL acetonitrile compound solution was then added to 1 mL of plasma and brain homogenate and vortex mixed to give a final compound concentration of 1 ug/mL in matrix.

RED Plate Preparation The Rapid Equilibrium Dialysis (RED) plate (Thermo Scientific) was prepared in accordance with the manufacturers guidelines ie. the base plate was soaked in 20% (v/v) l for 10 min and then rinsed twice with deionised water before being allowed to dry. The base plate was then filled with the appropriate number of disposable inserts (n=3 per compound) o Scientific) and matrix containing 1 ug/mL compound added into the matrix chamber of the inserts (200 uL) and an aliquot (350 uL) of PBS added to the buffer chamber. The plate was then covered with an adhesive and incubated in air at 37°C for 6 h with 130 rpm agitation.

Sampling Following the 6 h incubation the seal was removed and an aliquot (50 uL) taken from the PBS chambers and dispensed into Micronics tubes. Also, an aliquot (50 uL) was removed from the matrix chambers and placed into separate Micronics tubes.

Plasma and brain was then matrix matched with 50 pL of drug—free PBS and the PBS samples with 50 pL of the corresponding drug—free matrix, to give equal final compositions and volumes (100 nL).

Sample Analysis s were vortex mixed and an aliquot (300 nL) of acetonitrile ning 0.1% formic acid and 100 ng/mL of an appropriate internal standard added. Samples were then mixed and centrifuged (2000 g for 15 min) and an aliquot of the supernatant (100 pL) removed into a p well plate and d with an equal volume of water ready for analysis by LC~MS/MS. The following data was obtained for the following compounds in the above assay (Table 5).

Table 5: Rat PPB(%> Rat Plasma fu RatBTB(%) Comparative 65 .0 0.350 93 .3 0067 Exam le 4 0.049 0.013 0.104 0.022 Data represents the Mean of n=3 replicates fu = fraction unbound From the data presented herein above it will be apparent to those skilled in the art that the compounds of Examples 1 and 8 achieve a similar reduction of brain AB40 to that of Comparative Example 4, but with a lower plasma tration and free plasma concentration. This is advantageous and indicates that the compounds of the invention will have r or better efficacy at lower concentrations than the compounds of W02009/091016, and consequently will be less likely to cause unwanted peripherally mediated side effects, such as‘cardiovascular effects, phospholipidosis, liver toxicity, renal toxicity and gastrointestinal toxicity.

Assessment of effects on QTc interval in guinea pigs Male -Hartley guinea pigs were weighed and anaesthetised using 4% isoflurane in carbogen. hesia was maintained at 1.5% isoflurane and the animals were kept under anaesthesia for the duration of the study. Xylazine at Zing/kg i.m. was administered into the hind limb as a bradycardic agent to enable detection of QR: prolongation by the software.

The carotid artery and r vein were cannulated with lines ning heparinised saline and a 3-lead ECG connected and monitored using LabChart Pro software. Animals were allowed to stabilise for 30 minutes after the completion of the surgical procedure, before the initiation of an iv. vehicle (5%DMSO/90% MilliQ/5% O. IN HCl) infusion from time zero (infusion rate = 0.2 ml/kg/min). At 10 mins, an arterial blood sample was ted for PK analysis (lSOul; all collection syringes were heparinised). At 12 mins, drug infusion was started @ 2.0 mg/kg/10min iv. The dose was increased to 6.0 mg/kg/10min, then 20mg/kg/10min i.v., with a 10 minutes infusion period and two minutes blood sampling at each dose. After the final dose, a blood sample was taken and a second vehicle infusion initiated. Eight minutes later a terminal blood sample was ted for plasma PK analysis and the animal killed by a Schedule 1 method.

QTC (B azett’s) changes were analysed using rt Pro software. QTc was ged up to the highest dose/concentration tested, which corresponded to an unbound plasma concentration of 9503nM for Example 8 and 296nM for Example 1. ment of effects on QTc interval in beagle dogs Male beagle dogs were weighed and ed with sodium thiopental for induction of anesthesia. Anaesthesia was ined with mixture of l-l.5% isoflurane and oxygen and the animals were kept under artificial respiration and anesthesia using isoflurane for the duration of the study.

The carotid artery and saphenous vein were cannulated with lines containing nised saline and a LII ECG connected and monitored using polygraph system.

Animals were allowed to stabilise for 30 minutes before the infusion of compound solution and the infusion via cannula was started from time zero with l mg/kg/lOmin.

The dose was increased to 3 mg/kg/ 10min, then 10 mg/kg/10min. An arterial blood sample was collected after every dosing for PK analysis.

QTc was unchanged up to the highest dose/concentration tested, which corresponded to an unbound plasma concentration of 4128 nM for Example 8 and 1329 3O nM for Example 1.

Next, methods for preparing the compound of the formula (I) or a pharmaceutically able salt thereof according to the present invention will be described.

A. General Preparation Method A: A-(14) In the formula, X, Y and A are as defined above.

General Preparation Method A is a method for preparing a compound A—(lS) which corresponds to compound (1) according to the present invention from a compound A—( l) as a raw material through multiple steps of Step A—(i) to Step A—(xiv).

The compound A-(l) is commercially available.

Step A-gi): This step is a step of ing a nd A-(Z) by opening the epoxide A—(l) with a sulfonium ylide to generate an ediate allylic alkoxide which is then alkylated to give the compound A-(2). Those skilled in the art will appreciate that this -35_ transformation can be conducted in one pot or as two individual reactions. Those skilled in the art will appreciate the benefits and drawbacks of a one pot reaction compared to conducting two separate reactions and choose the best method for their requirements accordingly.

Specifically, the epoxide A-(l) can be opened by the anion of lrimethylsulfonium iodide and resultant loss of dimethylsulfide to give the ponding allylic alkoxide. Trimethylsulfonium iodide can be deprotonated with a suitable base, for example butyl lithium. The solvent used in the reaction is not particularly limited insofar as it does not ere with the reaction. es of suitable ts include THF. Those skilled in the art will iate that the word solvent in this instance is used to denote the liquid in which the reaction is effected and that the reagents may not be dissolved. Preferably the reaction should be conducted below room ature, preferably ~30 — 20°C. Upon on the reaction may be warmed to room temperature to facilitate reaction. The reaction time is not particularly limited and is usually 5 minutes to 24 hours, preferably 1 — 6 hours.

Those skilled in the art will appreciate that the alkoxide generated from this reaction can be reacted with an alkylating agent directly, such as tert—butyl bromoacetate, and that this reaction may proceed with or without additional solvents. If additional solvents are required to facilitate reaction, then solvents such as DMF or NMP are le. The reaction temperature is not particularly limited. Suitable reaction temperatures e room temperature to 80°C, preferably room temperature. The reaction time is not particularly limited and is usually 5 s to 1 week, ably 1 - 48 hours.

Those skilled in the art will appreciate that the intermediate alkoxide could be quenched, isolated and purified then subjected to independent alkylation conditions.

This reaction can be performed under the same conditions as those usually used in O— alkylation reaction of an alcohol compound (such as the conditions described in Tetrahedron Lett. 46 (2005) 45, 7751—7755). In this reaction, the compound A—(2) can be obtained by adding a base such as sodium hydride to a solution of the the intermediate alcohol in THF to prepare an alkoxide, and then reacting the alkoxide with the tert~buty1 bromoacetate, for example. The solvent used in the on is not particularly limited insofar as it does not inhibit the reaction and allows the starting material to be dissolved therein to a certain . Examples of the solvent include solvents such as THE, DMF and dimethyl sulfoxide. The reaction can be performed by g l to 3 lents of an appropriate base to act in the ce of such a solvent. Examples of the base used include sodium hydride, potassium hydride and t- butoxypotassium. The reaction time is not particularly limited and is usually 0.5 to 72 -36~ PCTIEP2012/050833 hours, and preferably 0.5 to 12 hours. The reaction temperature is usually —20°C to 50°C.

A more preferable result such as an improved yield may be achieved by adding a salt such as tetrabutylammonium iodide in this reaction.

Step A-gii): This step is a two step sequential reaction to obtain compound A—(3) from compound A—(2) by deprotecting the ester group then forming a Weinreb amide.

Specifically, the teit-butyl ester of compound A—(2) can be deprotected under the same conditions as those generally used in deprotection of a tert-butyl ester nd (such as the conditions described in a document such as T. W. Greene and P. G. M.

Wuts, "Protective Groups in Organic Chemistry, Third Edition", John Wiley & Sons (1999), p. 404—408). In this reaction, the compound A~(2) can be reacted with an appropriate acid in a le solvent, such as formic acid, as solvent and acid, for example. The solvent used in the on is not particularly limited insofar as it does not inhibit the reaction and allows the starting material to be dissolved therein to a n extent. The reaction time is not particularly limited and is usually 0.5 to 72 hours, and preferably 0.5 to 24 hours. The reaction temperature is usually ice—cold ature to 60°C.

The ediate acid can then be transformed to the [Weinreb amide (Tetrahedron Lett. 1981, 22, 3815) by reaction of N,O~dimethylhydroxylamine hydrochloride under standard amide formation conditions, ie by condensing the intermediate acid with N,O-dimethylhydroxylamine hydrochloride using a sing agent. Alternatively, this step is a step of obtaining a compound A—(3) by condensing the intermediate acid with N,O-dimethylhydroxylamine hydrochloride by acylation reaction.

The condensation reaction of the intermediate acid with NO- dimethylhydroxylamine hydrochloride using a condensing agent can be performed under the same conditions as those y used and described in the following documents. Examples of the known method e those in Rosowsky et al.; J . Med.

Chem, 34 (1), 227—234 (1991), wska et al.; Heterocycles, 32 (10), 1968-1972 (1991), and Romero et al.; J. Med. Chem, 37 (7), 998—1014 (1994).

The N,O—dimethy1hydroxylamine hydrochloride may be a free form or a salt.

The solvent in this on is not particularly limited insofar as it does not t the reaction. Examples of the solvent e tetrahydrofuran, oxane, ethyl acetate. methyl acetate, dichloromethane, chloroform, N,N-dimethylformamide, toluene, acetonitrile and xylene. Examples of the condensing agent include CD1 (N,N‘—carbonyldiimidazole), Bop (1H—1,2,3-benzotriazol— l— PCT/EP20121050833 yloxy(tri(dimethylamino))phosphonium hexafluorophosphate), WSC (1—ethyl—3—(3- dimethylaminopropyl)carbodiimide hydrochloride), DCC (N.N- ohexylcarbodiimide), diethylphosphoryl cyanide, PyBOP (benzotriazol—l- yloxytris(pyirolidino)phosphonium hexafluorophosphate) and EDC-HCl (1—ethyl-3—(3- dimethylaminopropyl)carbodiimide hydrochloride). Suitable conditions e an agent to activate the acid, such as N,N’-carbonyl diimidazole. One equivalent to a large excess of N,O-dimethylhydroxylamine hloride is used with respect to the intermediate acid. One equivalent to a large excess of an c base such as triethylamine may be added where necessary.

The reaction time is not particularly limited and is usually 0.5 to 72 hours, and preferably 0.5 to 24 hours. The reaction temperature varies according to the raw material used, the solvent and the like and is not particularly limited. Ice—cold ature to solvent reflux temperature is acceptable, ice cold to room temperature is preferable.

Step A-giii}: This step is a step of obtaining a compound A-(4) by on of an organometallic (aryllithium reagent or a Grignard reagent) reagent with compound A— (3) as described in Tetrahedron Lett. 1981, 22, 3815.

The reaction in this step can be med under the same conditions as those bed in Tetrahedron Lett. 1981, 22, 3815, for example.

The aryllithium reagent (including heterocyclic) or the Grignard t (including heterocyclic) can be ed by a method known to a person skilled in the art. Specifically, the corresponding phenyl lithium reagent or phenyl magnesium (Grignard) reagent can be prepared by halogen—metal exchange between an aryl halide nd and a commercially available organometallic reagent such as an alkyllithium reagent such as n—, sec— or tert—butyllithium or a Grignard reagent such as isopropylmagnesium bromide, or metallic magnesium, for e.

The solvent used in this step varies according to the starting material and the 3O reagent used, and is not particularly limited insofar as it does not inhibit the reaction, allows the starting al to be ved therein to a certain extent, and is always inert during the reaction. Preferable examples of the solvent include organic solvents such as diethyl ether, tetrahydrofuran, 1,4—dioxane, 1,2—dimethoxyethane, e and toluene, and mixed solvents thereof. The reaction time is not particularly limited and is usually 0.1 to 48 hours, and preferably 0.1 to 12 hours. The reaction temperature varies according to the starting material, the reagent used and the like, and is preferably maintained to be low, for example, at -78 - -60°C. -38— Step A—t iv 1: This step is a step of obtaining a compound A—(5) by oximation of the compound A-(4).

The reaction in this step can be performed under the same conditions as those y used in oximation reaction of a carbonyl compound such as the conditions described in Org. Lett. 9 (2007) 5, 753—756, Tetrahedron: Asymmetry 5 (1994) 6, 1018- 1028 and Tetrahedron 54 (1998) 22, 5868—5882.

Specifically, the compound A—(S) can be obtained by reacting the compound A— (4) with hydroxylamine or a hydroxylamine salt (such as hydroxylamine hydrochloride or ylarnine sulfate) in the presence of a base or in the absence of a base, for example.

The t used in this reaction is not particularly limited insofar as it does not inhibit the reaction. Preferable examples of the solvent include organic solvents such as ethanol, methanol, tetrahydrofuran, l,4~di0xane, 1,2—dimethoxyethane and dichloromethane, and mixtures of these solvents and water. Examples of the base used include sodium e, ne, sodium ide, cesium hydroxide, barium hydroxide and 2,6—lutidine. The reaction time is not particularly limited and is usually minutes to 24 hours, and ably 5 minutes to 12 hours. The reaction temperature is usually -20°C to solvent reflux temperature, and more preferably 0°C to solvent reflux temperature.

Step A—tv}: This step is a step of obtaining a nd A—(6) by a thermal intramolecular cycloaddition of the alkenyl oxime A-(5).

The reaction is conducted in the presence of an ve, for example hydroquinone.

The solvent used in this reaction is not particularly limited insofar as it does not inhibit the reaction. Suitable reaction solvents include high boiling solvents such as xylenes. The reaction temperature is not particularly limited and is usually 80 — 200°C 3O or t reflux temperature. The reaction time is not particularly limited and is usually 0.5 to 48 hours, and preferably 0.5 to 24 hours.

Step A~tvi): This step is a step of obtaining a compound A—(7) by subjecting the compound A-(6) to reductive ge reaction of the N-O bond.

The reductive ge reaction of the N-O bond can be performed under the conditions using zinc—acetic acid, a metal catalyst such as hydrogen-platinum oxide, or lithium aluminum e, for example. _39- The reaction using zinc such as zinc—acetic acid can be performed under the same conditions as those described in J. Org. Chem. 2003, 68, 1207—1215 and Org. Lett. 7 (2005) 25, 742, for example. Examples of the acid used e acetic acid, formic acid and hydrochloric acid. The solvent used in the reaction is not particularly limited r as it does not inhibit the reaction and allows the starting al to be ved therein to a certain extent. Examples of the solvent include methanol, ethanol, 1,4-dioxane, THF and water. The above acid may also be used as a solvent.

The reaction temperature is usually —20°C to solvent reflux temperature, and preferably ice—cold temperature to solvent reflux temperature. The reaction time is not particularly limited and is usually 5 minutes to 48 hours, and preferably 5 minutes to 24 hours.

The reaction using a metal catalyst such as hydrogen-platinum oxide can be performed under the same conditions as those bed in Tetrahedron: try 5 (1994) 6, 1018—1028 and Tetrahedron, Vol. 53, No. 16, pp 5752-5746, 1997, for example. The compound A-(7) can be obtained by hydrogenating the compound A—(6) using platinum oxide as a catalyst in a t such as methanol, for example.

The reaction using lithium um hydride can be performed under the same conditions as those described in Bull. Chem. Soc. Jpn., 66, 2730—2737 (1993), for example. The compound A-(7) can be obtained by reducing the compound A-(6) using lithium aluminum hydride in a solvent such as ether, for example.

Step A—tviig: This step is a step of ing a compound A—(8) from the compound A-(7).

The thiourea derivative A—(8) can be obtained from the compound A~(7) by a method known to a person skilled in the art.

The compound A-(8) can be obtained in this step by reacting the compound A- (7) with benzoyl isothiocyanate in a solvent such as dichloromethane or toluene. This reaction can be performed under the same conditions as those described in J. Med.

Chem. 1990, 33, 2393-2407, for example. The solvent used in the reaction is not 3O particularly limited insofar as it does not inhibit the reaction and allows the ng material to be ved n to a certain extent. Examples of the t e dichloromethane, chloroform, toluene, 1,4-dioxane and THF. The reaction temperature is usually -20°C to solvent reflux temperature, and preferably ice—cold temperature to solvent reflux temperature. The reaction time is not particularly limited and is usually 5 minutes to 48 hours, and preferably 5 minutes to 24 hours.

Step A-gviiig: PCT/EP20121'050833 This step is a method of obtaining a compound A—(9) by cyclizing the compound A—(S).

In this reaction, the compound A—(8) can be cyclized under various conditions to obtain the compound A—(9) by activating the alcohol of compound A-(8).

For example, the nd A—(9) can be obtained in this reaction by heating the compound A-(8) in a solvent such as methanol in the presence of an acid such as concentrated hydrochloric acid, for example. The t used in the on is not particularly limited insofar as it does not inhibit the reaction and allows the starting material to be dissolved therein to a n extent. es of the solvent include solvents such as methanol, ethanol, l—propanol and water, mixed solvents thereof, and acids used as a solvent. The on can be performed by using one equivalent to a large excess of an riate acid to act in the presence or absence of such a solvent.

Examples of the acid used include concentrated hydrochloric acid, hydrobromic acid, sulfuric acid, trifluoroacetic acid, methanesulfonic acid, trifluorornethanesulfonic acid and es thereof. The reaction time is not particularly limited and is usually 0.5 to 72 hours, and preferably 0.5 to 24 hours. The reaction temperature is usually ice-cold temperature to solvent reflux temperature.

Alternatively, the compound A-(9) can be obtained by reacting the compound A- (8) with trifluoromethanesulfonic anhydride in a solvent such as dichloromethane in the presence of a base such as ne. The solvent used in the reaction is not particularly limited insofar as it does not inhibit the reaction and allows the starting material to be dissolved therein to a certain extent. Those skilled in the art will appreciate that a solvent is not always required and the the on may also be conducted in the absence of a solvent, for example when the base is pyridine. Examples of the solvent include solvents such as dichloromethane, chloroform, l,2—dichloroethane, THF, 1,2- dimethoxyethane and toluene, and mixed solvents f. The reaction can be performed using 1 to 20 equivalents of an appropriate base in such a solvent.

Examples of the base used include pyridine, 2,6—lutidine, sodium carbonate, potassium carbonate and mixtures thereof. The reaction time is not particularly limited and is 3O y 0.5 to 24 hours, and preferably 0.5 to 12 hours. The reaction temperature is usually —78°C to room ature.

Step A—[ixgz This step is a method of ing the compound A-(lO) by deprotecting the protecting group of the compound A-(9). The compound A-(lO) can be obtained under deprotection conditions known to a person skilled in the art.

When the protecting group is a benzoyl group, the compound A—(lO) can be obtained in this reaction by heating the nd A—(9) in a solvent such as methanol PCT/EP20121’050833 in the presence of a base such as DBU. for e. This reaction can be performed under the same conditions as those described in Synth. Commun. 32 (2), 265-272 (2002), for example. The solvent used in the reaction is not particularly limited insofar as it does not inhibit the reaction and allows the starting material to be dissolved therein to a certain extent. es of the t include ts such as methanol, ethanol and anol. The reaction can be performed using 1 to 20 equivalents of an appropriate base in such a solvent. Examples of the base used include DBU. The reaction time is not particularly limited and is usually 0.5 to 24 hours, and preferably 0.5 to 12 hours. The reaction temperature is usually room temperature to solvent reflux temperature.

Alternatively, compound A—(lO) can be ed in this reaction by heating compound A-(9) with an inorganic base such as potassium carbonate, for example.

The solvent used in the reaction is not particularly limited insofar as it does not inhibit the on and allows the starting material to be dissolved therein to a certain extent.

Examples of the solvent include solvents such as methanol, ethanol and l-propanol.

The reaction can be med using 1 to 20 lents of an appropriate base in such a solvent, and preferably a slight excess is used. Examples of the base used include potassium carbonate. The reaction time is not particularly limited and is usually 0.5 to 24 hours, and preferably 0.5 to 12 hours. The reaction temperature is usually room ature to t reflux temperature, and preferably 50 — 100°C. Those skilled in the art will appreciate that the the selected solvent will limit the reaction temperature by its reflux temperature. Examples of le solvents include refluxing methanol.

Step A—gx}: This step is a step of obtaining the compound A—(l l) by nitration reaction of the compound A-(lO). In this nitration reaction, the compound A—(l 1) can be obtained from the compound A—(lO) by a method known to a person skilled in the art.

Examples of the nitrating agent used in the reaction e potassium nitrate/concentrated sulfuric acid, fuming nitric acid/concentrated sulfuric acid and fuming nitric acid/acetic anhydride. Suitable solvents for the reaction include trifluoroacetic acid. The reaction temperature is not particularly limited and is usually -20°C to room temperature, and preferable reaction temperatures include 0 — 10°C.

Step A—gxi}: This step is a step of obtaining a compound A—(12) by xycarbonylation of the amino group of the compound A—(l l).

The reaction can be performed under the same conditions as those generally used in t—butoxycarbonylation of an amino compound such as the conditions described in a document such as T. W. Greene and P. G. M. Wuts, "Protective Groups in Organic Chemistry, Third Edition". John Wiley & Sons (1999), P. 518-525. The compound A— (12) can be obtained by reacting the compound A-(l l) with di-tert-butyl dicarbonate using in a solvent such as tetrahydrofuran, for example. Alternative solvents include acetonitrile and DMF. Those d in the art will appreciate that a base may also be added to the reaction mixture, although is not essential. le examples of a base e, but are not limited to triethylamine and diisopropylethylamine. The reaction temperature is not particularly limited and is usually to room ature to reflux, and preferably room temperature to 60°C.

Step A-txii): This step is a step of obtaining a compound A-(13) from the compound A—(12).

The compound A-(13) is synthesized by reducing the nitro compound A-(l2) by a sis method known to a person skilled in the art. Examples of the method include reduction by catalytic hydrogenation using a noble metal catalyst such as Raney nickel, palladium, ium, rhodium or platinum. Other reducing ts include tin chloride, for example. Examples of the solvent include alcoholic solvents such as methanol, l and l-propanol, preferably ethanol. The reaction time is not particularly limited and is usually 0.5 to 24 hours, and preferably 0.5 to 18 hours. The 2O reaction temperature is usually room temperature. ative reduction reaction ions e reaction with iron with an additive such as ammonium chloride or hydrochloric acid, in an alcoholic solvent such as ethanol, at an appropriate reaction temperature, for example 65°C.

Step A—t xiii): This is a step of obtaining a compound A—(l4) from the compound A—(l3) by condensing compound A—(l3) with a carboxylic acid and a condensing agent. The condensation reaction can be performed under the same conditions as those usually used and described in the following documents. Examples of the known method include 3O those in Rosowsky et al.; J. Med. Chem, 34 (1), 227~234 , Brzostwska et al.; Heterocycles, 32 (10), 19684972 (1991), and Romero et al.; J. Med. Chem, 37 (7), 998—1014 .

The compound A-(13) may be a free form or a salt.

The solvent in this on is not particularly limited insofar as it does not inhibit the reaction. Examples of the solvent include tetrahydrofuran, 1,4—dioxane, ethyl acetate, methyl acetate, dichloromethane, chloroform, N,N-dimethylf01mamide, toluene, acetonitrile and xylene. Examples of the condensing agent include CDl (N,N'—carb0nyldiimidazole), Bop (1H— 1 ,2,3—benzotriazol— l— PCT/EP20121050833 yloxy(tri(dimethylamino))phosphonium orophosphate), WSC ( l—3—(3- ylaminopropy1)carbodiimide hydrochloride), DCC (N,N- dicyclohexylcarbodiimide), diethylphosphoryl cyanide, PyBOP (benzotriazol-lyloxytri (pyrrolidino)phosphonium hexafluorophosphate) and EDC . HCl (1—ethyl(3 - dimethylaminopropyl)carbodiimide hydrochloride). Suitable conditions include an agent to activate the acid, such as N,N’~carbonyl diimidazole. One equivalent to a large excess of the acid may be used with respect to the compound A-(13). One equivalent to a large excess of an organic base such as triethylamine or N,N— diisopropylethylamine may be added where necessary.

The reaction time is not ularly limited and is usually 0.5 to 72 hours, and preferably 0.5 to 24 hours. The reaction temperature varies according to the raw material used, the solvent and the like and is not particularly limited. Ice-cold temperature to solvent reflux temperature is acceptable, ice cold to room ature is preferable.

Alternatively, the compound A—( 14) can be obtained by ting the desired carboxylic acid to the ponding acid chloride and then ng the acid chloride with the compound A-( 13). The acid chloride can be synthesized by a means known to a person skilled in the art. For example the desired carboxylic acid may converted to the corresponding acid chloride by reaction with thionyl chloride in the presence or absence of a solvent, for example dichloromethane, N,N’-dimethylimidazolineone, NMP or DMF. One to two equivalents or a large excess of thionyl chloride may be used with respect to the desired carboxylic acid. The reaction temperature is ~30°C to reflux, and preferably ~10°C to room temperature. The acid chloride may also be formed by treating the acid with oxalyl chloride in a t such as dichloromethane in the presence of DMF. The reaction temperature is —30°C to room temperature, and preferably —10°C to room temperature Alternatively, the compound A—( 14) can be obtained by converting the desired carboxylic acid to a mixed acid anhydride and then reacting the mixed acid anhydride with the compound A-(l3). The mixed acid anhydride can be synthesized by a means 3O known to a person skilled in the art. The synthesis is performed by reacting the desired carboxylic acid with a chloroformate such as ethyl chloroformate in the presence of a base such as ylamine, for e. One to two lents of the chloroformate and the base are used with respect to the desired carboxylic acid. The reaction ature is —30°C to room temperature, and preferably —20°C to room temperature.

The step of sing the mixed acid anhydride with the compound l-(13) is performed by reacting the mixed acid anhydride with the compound l-(l3) in a solvent such as dichloromethane, tetrahydrofuran or N,N—dimethylformamide, for example.

One lent to a large excess of the desired carboxylic acid is used with respect to the compound A—(l3).

The reaction time is not particularly limited and is usually 0.5 to 48 hours, and preferably 0.5 to 12 hours. The reaction temperature is -20°C to 50°C, and preferably ~20°C to room temperature.

Alternatively, the compound A-(l4) can be obtained by converting the d carboxylic acid to an active ester and then reacting the active ester with the compound A—(l3). The step of obtaining the active ester is performed by reacting the desired carboxylic acid with an active ester synthesis reagent in a t such as 1,4—dioxane, tetrahydrofuran or N,N—dimethylformamide in the presence of a condensing agent such as DCC, for example. Examples of the active ester synthesis reagent e N- hydroxysuccinimide. One to 1.5 lents of the active ester synthesis reagent and the condensing agent are used with respect to the compound A-(l3). The reaction time is not particularly limited and is usually 0.5 to 48 hours, and ably 0.5 to 24 hours.

The reaction temperature is -20°C to 50°C, and preferably -20°C to room temperature.

The step of sing the active ester with the compound A—(l3) is performed by reacting the active ester with the compound A—(l3) in a solvent such as dichloromethane, tetrahydrofuran or N,N-dimethylformamide, for example. One equivalent to a large excess of the active ester is used with respect to the compound A- (13). The reaction time is not particularly limited and is y 0.5 to 48 hours, and preferably 0.5 to 24 hours. The reaction temperature is -20°C to 50°C, and preferably —20°C to room temperature.

Step A-txiv}: This step is a step of obtaining the compound A-(15 ) by deprotection of the t— butoxycarbonyl group of the compound .

The reaction can be performed under the same conditions as those generally used in deprotection reaction of a t~butoxycarbonyl group such as the conditions described in a document such as T. W. Greene and P. G. M. Wuts, "Protective Groups in Organic Chemistry, Third n", John Wiley & Sons (1999), P. 518-525. The nd A- (15) can be obtained by reacting the compound 1—(14) with a strong acid, for example trifluoroacetic acid in the presence or absence of a solvent. Suitable solvents include dichloromethane. Alternative acids include hydrochloric acid in suitable solvents, such as dichloromethane or dioxane, for example.

The reaction temperature is normally ice cold to 80°C, preferably room temperature. The reaction time is not particularly limited and is usually 5 minutes to 48 hours, and preferably 5 minutes to 12 hours.

B. General Preparation Method B: In the formula, X, Y and A are as defined above.

General Preparation Method B is an alternative method for preparing a compound A—(15) which ponds to nd (1) according to the present invention from a compound A—(l 1) as a raw material through multiple steps of Step B— (i) to Step .

The compound A—(l 1) may be prepared as described in General Preparation Method A or the examples.

X N02 F B-(i) o NH2 '——" A—(11) Step B—(i): This step is a step of obtaining a compound A—(_16) from the compound A—( 1 1).

The compound A—(16) is synthesized by reducing the nitro compound A-(l l) by a sis method known to a person skilled in the art. Examples of the method e ion by catalytic hydrogenation using a noble metal catalyst such as Raney nickel, palladium, ium, rhodium or platinum. Other reducing reagents include tin chloride, for example. Examples of the solvent include alcoholic solvents such as ol, ethanol and l—propanol, preferably ethanol. The reaction time is not particularly limited and is usually 0.5 to 24 hours, and preferably 0.5 to 18 hours. The reaction temperature is usually room temperature. Alternative reduction on conditions include reaction with iron with an additive such as ammonium chloride or hydrochloric acid, in an alcoholic solvent such as ethanol, at an appropriate reaction temperature, for example 65°C.

Step B—gii): This is a step of obtaining a compound A-(lS) from the compound A-(16) by condensing compound A-(l3) with a carboxylic acid and a condensing agent. The sation reaction can be performed under the same conditions as those usually used and described in the following documents. Examples of the known method include those in Rosowsky et al.; J. Med. Chem, 34 (1), 227-234 , Brzostwska et al.; Heterocycles, 32 (10), 1968-1972 (1991), and Romero et al.; J. Med. Chem., 37 (7), 998—1014 (1994).

The compound A—(15 ) can be obtained by converting the desired carboxylic acid to the corresponding acid chloride and then reacting the acid chloride with the compound . The acid chloride can be synthesized by a means known to a person skilled in the art. For example the desired carboxylic acid may converted to the corresponding acid chloride by reaction with thionyl chloride in the presence or absence of a solvent, for example dichloromethane, N,N’—dimethylimidazoline-2—one, NMP or DMF. One to two equivalents or a large excess of thionyl chloride may be used with respect to the d carboxylic acid. Those skilled in the art will appreciate that the choice of reaction conditions employed may affect the outcome of the reaction, for example the conditions may affect whether the acid chloride reacts with the aniline or the isothiourca moieties. Those skilled in the art will iate that the reaction of thionyl chloride with a carboxylic acid results in the concomitant formation of 1 equivalent of hydrochloric acid in addition to the formation of the desired acid chloride.

Those d in the art will appreciate that the current conditions do not employ a method of removing the thus formed hydrochloric acid. The hloric acid formed in this reaction may or may not affect the selectivity of the on which may or may not result in a ial outcome. The reaction time is not particularly limited and is usually 0.5 to 48 hours. and preferably 0.5 to 12 hours. The reaction temperature is - °C to reflux, and preferably -10°C to room temperature. The acid chloride may also be formed by treating the acid with oxalyl chloride in a solvent such as dichloromethane in the presence of DMF, The reaction temperature is -30°C to room temperature, and preferably —10°C to room temperature.

C. General Preparation Method C: General Preparation Method C is an alternative method for preparing a compound A—(Z) which is a synthetic ediate of the compound (1) according to the present invention from a compound A-(l7) as a raw material through Step C-(i).

The nd A-(l7) is cially available.

C—(i)i W0 —” 070K A-(17) F A-(2a) $.04: This step is a step of obtaining a compound A—(Za) from A-(l?) by adding a trifluoromethyl anion to the compound A—(17) to generate an intermediate c alkoxide or ediate trimethylsilyl ether which is then alkylated to give the compound A—(Za). Those skilled in the art will appreciate that this transformation can be ted in one pot or as two individual reactions. Those skilled in the art will .47~ appreciate the benefits and drawbacks of a one pot reaction compared to ting two separate reactions and choose the best method for their ements accordingly.

Specifically, acrolein A-(l7) can react with a trifluoromethyl anion which can be generated by the action of fluoride on reagents such as (trifluoromethyl)trimethylsilane to generate the corresponding c alkoxide or allylic timethylsilyl ether. The solvent used in the reaction is not particularly limited r as it does not interfere with the reaction. Examples of suitable solvents include THF. Acceptable temperature ranges for the reaction include -10°C to solvent reflux, preferably below room ature. Those skilled in the art will iate that n chemical reactions can be exothermic and that control measures should be put in place to control these exotherms. Those skilled in the art will also appreciate that the reaction rm may be controlled by allowing the solvent to reflux. Suitable precursors to generate oromethyl anion e, but are not limited to, (trifluoromethyl)trimethylsilane (Rupert’s t, Chem Rev 1997, 97, 757) and le fluoride sources include, but are not limited to, tetrabutylammonium fluoride (TBAF), tetrabutylammonium difluorotriphenylsilicate (TBAT) and caesium fluoride.

Although the initial reaction temperature may be below room temperature it is acceptable to allow the reaction temperature to reach solvent reflux during the course of the reaction to facilitate reaction. The reaction time is not particularly d and is usually 5 minutes to 24 hours, preferably 1 — 6 hours.

Those skilled in the art will appreciate that the alkoxide generated from this reaction can be reacted with an alkylating agent directly, such as tert—butyl bromoacetate, and that this reaction may proceed with or without additional solvents. If additional solvents are required to facilitate reaction, then ts such as DMF or NMP are suitable. The reaction temperature is not particularly limited. Suitable reaction temperatures include room temperature to 80°C, preferably room temperature. The reaction time is not particularly limited and is usually 5 minutes to 1 week, preferably 1 ~ 48 hours. Alternatively, the alkylation reaction can be conducted under phase transfer conditions, for example by adding an aqueous base, for e aqueous 3O sodium hydroxide. Those skilled in the art will appreciate that when these conditions are d the use of a phase transfer st is ed. Suitable phase transfer catalysts include, but are not limited to, tetrabutylammonium hydrogen sulfate.

Those skilled in the art will appreciate that the intermediate alkoxide could be quenched, isolated and purified then subjected to independent alkylation conditions.

This reaction can be performed under the same conditions as those usually used in O— alkylation reaction of an alcohol compound (such as the conditions bed in Tetrahedron Lett. 46 (2005) 45, 7751-7755). In this reaction, the compound A—(Za) can be obtained by adding a base such as sodium hydride to a solution of the the ~48- intermediate l in THF to prepare an alkoxide, and then reacting the alkoxide with the tert—butyl bromoacetate, for e. The solvent used in the reaction is not particularly limited insofar as it does not inhibit the reaction and allows the starting material to be dissolved therein to a n . Examples of the solvent include ts such as THE DMF and dimethyl sulfoxide. The reaction can be performed by g l to 3 equivalents of an appropriate base to act in the presence of such a solvent. Examples of the base used include sodium hydride, potassium hydride and t— butoxypotassium. The reaction time is not particularly limited and is usually 0.5 to 72 hours, and preferably 0.5 to 12 hours. The reaction temperature is usually —20°C to 50°C.

A more preferable result such as an improved yield may be achieved by adding a salt such as tetrabutylammonium iodide in this reaction.

Those d in the art will appreciate that this reaction generates a new chiral centre in the compound A-(Za) and that compound A—(Za) is the same as compound A- (2) except that compound A-(Z) is enatiomerically pure s compound A—(Za) is racemic. It will be appreciated by those skilled in the art the enantiomerically pure compound A—(Z) and the racemic compound A—(Za) are indistinguishable by analytical techniques such as NMR and liquid chromatography, however they are distinguishable by chiral HPLC. Those skilled in the art will appreciate that the most appropriate methods to obtain the desired mer from the compound A-(Za) or a more advanced synthetic intermediate or final compound. Appropriate methods and appropriate stages of enantiomeric purification include those as detailed in the examples.

In a further aspect, the present invention provides a process of preparing a compound of formula (I), which comprises ng a nd of formula A—(lé), n X is defined in formula (I), with a compound of formula (II) wherein A and Y are as defined in formula (I), or a C 1-6 alkyl ester, acid anhydride or acid halide thereof, to yield a compound of formula (I), and optionally converting the compound to a further compound of formula (I) or forming a pharmaceutically acceptable salt thereof.

A Y A Y X NH2 ;I/ X NWfJ;N:I/ 0 (II) o F F N NHz N NH \ 2 o I 0 \Y FSC H A-ue) H The reaction of A-(l 6) and (H) may conveniently be conducted in a solvent (such as tetrahydrofuran, 1,4—dioxane, ethyl acetate, methyl acetate, dichloromethane, chloroform, N,N~dimethylformamide, toluene, itrile or xylene) at a temperature _49_ in the range of —30 °C to 100 °C. In one embodiment of the invention, compound (11) may conveniently take the form of an acid halide (e.g. chloride) as may be prepared by reacting the acid with a suitable reagent (e.g. thionyl chloride) It will be iated by those skilled in the art that in the process of the present invention certain functional groups such as hydroxyl, carboxyl or amino groups in the starting reagents may need to be protected by protecting . Thus the preparation of the compounds of formula (I) may additionally involve incorporation and removal of one or more protecting groups. The protection and deprotection of functional groups is for example described in T. W. Greene and P. G. M. Wuts, "Protective Groups in Organic Chemistry, Third Edition", John Wiley & Sons (1999).

The present invention will be described more specifically below with reference to Examples, Preparation Examples and Test Example. However, the present ion is not d thereto. The abbreviations used in Examples are conventional abbreviations known to a person d in the art. Some abbreviations are shown below: LCMS, LC/MS & LC—MS (liquid chromatography/mass spectrometry); MS (mass spectrometry); MDAP (mass directed auto purification); NMR (nuclear magnetic resonance); 3, d, t, dd. m. br (singlet, doublet, triplet, doublet of doublets, multiplet, broad); Ph, Me, Et, Pr, Bu, Bn (phenyl, , ethyl, propyl, butyl, benzyl); THF (tetrahydrofuran); DCM (dichloromethane); DMF (N,N-dimethylformamide); h, hr, hrs (hours); EDC & EDAC (N-3—(dimethylaminopropyl)—N’ethylcarbodiimide hydrochloride); DMAP (4—N,N—dimethylaminopyridine); DMSO (dimethylsulfoxide); UV (ultraviolet); RT & rt (room ature); Rt (retention time); min & mins (minutes); EtOAc (ethyl acetate); EtZO (diethyl ether); MeCN (acetonitrile); EtOH (ethanol); MeOH (methanol); PhCH3 & PhMe (toluene); tlc (thin layer tography); TFA (trifluoroactic acid); NaOH (sodium hydroxide); HCl (hydrochloric acid); NMP (N-rnethylpyrrolidinone or l-methyl—Z-pyrrolidinone); HPLC (high performance liquid chromatography); TBAF butylammonium e); BuLi (n—butyl lithium); PyBOP: benzotriazol—l~yloxytris(pyrrolidino)phosphonium hexafluorophosphate; Pdgdb‘cl}: tris(dibenzylideneacetone)dipalladium; Pd(t-Bu3P)2: bis(tri-t—butylphosphine)palladium; TEA: trifluoroacetic acid; pTLC: preparative thin— layer chromatography; HRMS (high resolution mass spectrometry); Tr or Trt (trityl or triphenylmethyl). 1H NMR spectra were recorded on a Bruker AM series spectrometer operating at a (reported) frequency of 400 MHz. Chemical shifts in proton r magnetic resonance spectra are recorded in 5 units (ppm) relative to tetramethylsilane and coupling constants (J) are ed in Hertz (Hz). Patterns are designated as s: t, d: doublet, t; triplet, br; broad.

The "room ature" in the following Examples and Preparation Examples lly refers to about 10°C to about 35°C. "%" indicates wt% unless otherwise Chemical names were generated from chemical structures using ChemBioDraw Ultra 11.0 and 12.0.

Description of Figures: Figure 1 is a Typical Chromatogram from a Chiral HPLC Isolation of Compound l—(20).

Preparation e 1 Inthesis of tert—but l 4aS.SS.7aS -7a— 5—amino~2—fluoro hen 1 -5— trifluorometh 1 - 4a,5.7 7a—tetrah dro-4H—furo 3.4—d l 3 n-2~ 1 carbamate 1— 1‘3 1-(2) S Inthesis of Kerr-but 1 ZS -1 1.1-trifluorobut—3-en—2— 1 ox acetate To a suspension of trimethylsulfonium iodide (110 g) in THE (500 mL) at -30°C was added lithium hexamethyldisilazide (530 mL, 1N in THF) portionwise over 45 mins.

After stirring at —20°C for 20 mins, (S)—2—trifluoromethyloxirane (37.97 g) was added at the same temperature over 15 mins, and the mixture was allowed to warm to RT and stirred for 3 h. The slurry was then added portionwise to an ice-cold solution of ten- butyl bromoacetate (10568 g) in NMP (200 mL). The resulting mixture was allowed to warm to RT and stir for 2 days, before dilution with EtOAC (1 L). The organic layer was washed with sodium onate (sat, aq., 4 x 400 mL), dried over MgSO4 and evaporated. The residue was purified by silica gel column chromatography (5% EtOAc in hexanes) to obtain the title compound (70.1 g) which was used in the subsequent step without purification. 1H—NMR (400 MHZ, CDC13) 5 (ppm): 1.30 (s, 9 H) 3.83 — 3.96 (m, 2 H) 4.14 - 4.21 (m, l H) 5.34 - 5.48 (m, 2 H) 5.56 — 5.71 (m, l H) PCTIEP2012/050833 vl )oxy )acetamide zert—Butyl {[(ZS)-l,1,1-trifluorobut—3~en~2~ylloxy}acetate (70.1 g, crude) was dissolved in ice—cold formic acid (200 mL). The mixture was allowed to warm to RT and stir overnight. The reaction mixture was then concentrated under reduced pressure, toluene (200 mL) was added, the mixture concentrated, before a second addition of toluene (200 mL) and concentration to an oil. The e was dissolved in DCM (600 mL), cooled in an ice-bath, and N,N "carbonyl azolc (35 g) was added portionwise over 20 mins. After stirring for 45 mins, N, O-dimethyl hydroxylamine hloride (22 g) was added, and the reaction mixture was allowed to warm to RT and stir overnight. Saturated NaHC03 (500 mL) and brine (250 mL) were then added, and the mixture extracted with EtOAc (3 x 750 mL). The combined organic ns were dried over MgSO4 and evaporated. The residue was purified by silica gel column chromatography (1% to 30% EtOAc in hexanes) to obtain the title compound (25.17 g). 1H—NMR (400 MHZ,CDC13) 5 (ppm) 3.21 (s, 3 H), 3.71 (m, 3 H), 4.36 - 4.51 (m, 3 H), .54 — 5.69 (m, 2 H), 5.84 (ddd, J=l7.7, 10.4, 7.3 Hz, 1 H) l-(4) S nthesis of S '2-fluoro hen 1 ' 1.1 l—trifluorobut-3—en-2— 1 ox ethanone A on of n-butyllithium in hexane (2.50 M; 90 mL) was added dropwise over 25 mins to a solution of 2-bromofluorobenzene (40.35 g) in THF (250 mL) under a N3 atmosphere at -78°C. The reaction solution was allowed to warm to -60°C and stir for 60 min. (S)—N-Methoxy-N—methyl~2—((l,1,1-trifluorobut—3-en~2—yl)oxy)acelamide (40 g) in THF (25 mL) was added dropwise to the reaction on, and after stirring at -60°C for 2 h, aqueous NH4Cl (100 mL) was added to the reaction solution, followed by warming to RT. Brine (200 mL) was added to the reaction solution, and the mixture was extracted with EtOAc (3 x 400 mL). The ed organic portions were dried over MgSO4, ated, and the residue was purified by silica gel column chromatography (1% to 10% EtOAc in hexanes) to obtain the title compound (33.59 g). 1H—NMR (400 MHz, CDC13) 5 (ppm): 4.40 (pentet, J 6.3 Hz, 1 H) 4.81 — 4.87 (m, 2 H), .54 — 5.69 (m, 2 H), 5.86 (ddd, J 4, 7.3 Hz, 1 H) 7.12 — 7.22 (m, 1 H) 7.24 ~ 7.34 (m, l H) 7.54 - 7.63 (m, l H) 7.94 - 8.02 (m, l H). 1—(5) S s of S —l— 2-fluoro hen l -2— l.l.l—trifluorobut—3—en—2- lox , ethanone oxime (S)-l-(2-Fluorophenyl)—2-((1,1,l—trifluorobuten-2—y1)oxy)ethanone (41.22 g) was dissolved in anhydrous methanol (400 mL) and hydroxylamine hydrochloride (14.0 g) and sodium acetate (19.0 g) were added. The reaction mixture was heated to 50C for 90 min, then cooled to RT, concentrated in vacuo and the residue d by silica gel chromatography (2% to 15% EtOAC in hexanes) to afford the title compound as a mixture of geometric isomers (40.54 g). 1H—NMR (400 MHZ, CDC13) 5 (ppm): 4.04 — 4.15 (m, 0.8 H), 4.18 — 4.26 (m, 0.2 H), 4.44 - 4.57 (m, 0.4 H) 4.79 - 4.90 (m, 1.6 H) .37 — 5.56 (m, 2 H) 5.64—5.78 (m, 1 H) 7.03 - 7.54 (m, 2 H), 7.90 — 7.26 (m, 2 H) 7.33 (br s, 0.2 H), 8.51 (br s, 0.8 H). 1—(6) Synthesis of g3aR.4S.6aS)-6a-g2-fluorophenyl )~4— 3O 1trifluoromethyl)hexahydrofuro]3 ,4—c lisoxazole (S)-l—(2-fluorophenyl)—2—((1,1,l—trifluorobuten-2—yl)oxy)ethanone oxime (40.54 g) was dissolved in xylenes (400 mL) and hydroquinone (4.0 g) was added. The reaction mixture was heated to reflux (heating block temperature 140°C) for 22 h, then cooled and evaporated. The residue was purified by silica gel column chromatography (1% to 30% EtOAc in hexanes) to obtain the title nd (28.76 g). 1H-NMR (400 MHZ, CDClg) 8 (ppm): 3.71-3.81 (m, l H), 4.04 — 4.35 (m, 4 H), 4.51-4.62 (m, 1 H), 5.38-5.54 (in, l H), 7.07 - 7.26 (m, 2 H), 7.32 - 7.42 (m, 1 H), 7.54-7.67 (in, 1 H). 1—(7) 2S.3R 4S —4-amino-4— 2—fluoro hen 1 —2— orometh l tetrah drofuran—3— (3aR,4S,6aS)-6a-(2-fluorophenyl)(trifluoromethyl)hexahydrofuro[3,4—c]isoxazole (28.76 g) was dissolved in acetic acid (200 mL) and the solution was cooled to 0°C.

Zinc (50 g) was added, and the on mixture was allowed to warm and stir at RT for 16h. The reaction mixture was then diluted with EtOAc (500 mL) and filtered h celite, washing with a further 500 mL of EtOAc. The combined c portions were ated, ved in chloroform (200 mL), and ammonia (28% aq., 250 mL) was added slowly. The layers were separated, and the aqueous n was further extracted with chloroform (2 x 250 mL). The combined organic extracts were dried over anhydrous MgSO4 and evaporated to afford the title compound (31.12 g) which was used in the subsequent step without further cation. 1H-NMR (400 MHz, CDClg) 5 (ppm): 2.93 (ddd, J=7.7, 4.9, 2.5 Hz, 1 H), 3.84 (dd, J=12.4, 4.8 Hz, 1 H), 4.05 (dd, J=9.2, 3.2 Hz, 1 H), 4.17 (dd, J=12.4, 2.3 Hz, 1 H), 4.31 (d, J=9.3 Hz, 1 H), 4.72 (quin, J=7.3 Hz, 1 H), 7.13 (ddd, J=13.1, 8.8, 1.3 Hz, 1 H), 7.22 (td, J=7.6, 1.3 Hz, 1 H), 7.31 - 7.40 (m, 1 H), 7.51 (Cd, J=8.0, 1.6 Hz, 1 H) l~(8) S s of N- ’ ’(3S 4R SS ’2—fluoro hen 1 —4- h drox meth l —5— (trifluoromethyl 1tetrahydrofuranyl Zearbamothioyl zbenzamide Benzoyl isothiocyanate (19.0 mL) was added to a solution containing ((2S,3R,4S) amino(2—fluorophenyl)—2—(trifluoromethyl)tetrahydrofuran—3—y1)methanol (28.72 g) in DCM (150 mL), and the mixture was stirred at RT for 18 h. Sodium bicarbonate (sat, aq., 200 mL) was then added, the mixture extracted with EtOAc (3 x 300 mL), dried over MgSO4 and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (5% to 30% EtOAc in hexanes) to obtain the title compound (37.07 g). 1H-NMR (400 MHZ, CDC13) 8 (ppm): 3.22 (dd, J=8.l, 4.5 Hz, 1 H), 3.31 (td, J=8.0, 3.0 Hz, 1 H), 3.94 — 4.07 (m, l H), 4.31 - 4.46 (m, 1 H), 4.53 (d, J=9.9 Hz, 1 H), 4.83 (d, J=9.9 Hz, 1 H), 6.97 ~ 7.14 (m, 1 H), 7.22 (td, J=7.7, 1.3 Hz, 1 H), 7.31 — 7.45 (m, 1 H), 7.49 — 7.61 (m, 2 H), 7.61 — 7.70 (m, 1H), 7.75 (td, 128.1, 1.5 Hz, 1 H), 7.79 — 7.93 (m, 2 H), 8.90 (s, l H), 11.85 (s, 1 H) 1—(9) S nthesis of N- 4aS.SS,7aS)-7a~ 2—fluoro hen 1 —5— trifluorometh 1 -4a.5.7.7a- tetrahvdro—4H—furol 3.4—d l 1,3 [thiazin—Z-yl )benzamide N—(((3S,4R,SS)-3~(2-Fluorophenyl)-4—(hydroxymethyl) (trifluoromethylfietrahydrofuranyl)carbamothioyl)benz.amide (31.1 g) was dissolved in pyridine (150 mL), and the mixture cooled to -20°C. Trifluoromethanesulfonic anhydride (14.0 mL) was added dropwise over 30 min and the reaction was allowed to warm to 0°C. After stirring for 2 h, the reaction was quenched by the addition of PCT/EP20121'050833 ammonium chloride (sat, aq., 400 mL) and ted with EtOAc (3 x 500 mL). The combined organic ts were dried over MgSO4, concentrated in vacuo and purified by silica gel column chromatography (2% to 30% EtOAc/hex) to obtain the title compound (18.50 g). 1H NMR (400 MHz, CDC13) 5 ppm 2.86 (dd, J=13.9, 3.5 Hz, 1H), 3.25 (d, J=13.6 Hz, 1 H), 3.61 (br. s., 1 H), 4.00 ~ 4.10 (m, l H), 4.66 (d, J=8.8 Hz, 1 H), 4.78 — 4.87 (m, 1 H), 7.12 - 7.60 (m, 6 H), 7.68 — 7.73 (m, 1 H), 7.99 — 8.16 (br. s., 2 H), 8.62 — 8.66 (m, 1 H) 1—(10) S nthesis of N— 4aS.SS.7aS —7a— 2—fluoro henvl —5- trifluorometh l 421 5.7,7a— tetrahvdro—4H—furol 3.4-d l 1,3 lthiazin—2-y11benzamide N—((4aS,SS,7aS)-7a-(2—F1uorophenyl)~5-(trifluoromethy1)~4a,5.7,7a—tetrahydro-4H— furo[3,4—d][1,3]thiazin-Z-yl)benzamide (21.5 g) was dissolved in ol (160 mL), 1,8~diazabicyclo[5.4.0]undec—7-ene (16.29g) was added, and the solution was heated to reflux (heating block temperature 80°C). After 16 h, the reaction mixture was trated under reduced pressure, and the residue purified by silica gel column chromatography (10% to 60% EtOAc in hexanes) to afford the title compound (13.82 g). 1H NMR (400 MHZ, CDC13) 5 ppm 2.85 (dd, J=l3.6, 3.8 Hz, 1 H), 3.14 (dd, , 3.2 Hz. 1 H). 3.33 — 3.45 (m. 1 H), 3.92 (dd, J=8.1, 2.0 Hz, 1 H), 4.49 (br. s., 2 H). 4.63 - 7.22 (m, - 4.76 (m, 2 H), 7.08 (ddd, J=12.6, 8.1, 1.0 Hz, 1H), 7.13 1 H), 7.25 - 7.36 (m, 1 H), 7.44 (td, J=8.0, 1.9 Hz, 1 H) l-(l l) S nthesis of .7aS —7a— 2—fluoro—5-nitro hen 1 -5— trifluorometh l - .7.7a—tetrah —furo 3.4—d 1.3 thiazin—2-arnine N-((4aS,SS,7aS)—7a—(2—Fluorophenyl)~5~(trifluorornethyl)—4a,5,7,7a—tetrahydr0—4H- furo[3,4—d][l,3]thiazin—2-yl)benzamide (5.15 g) was dissolved in TFA (75 mL), and the solution was cooled to 0°C. Sulfuric acid (cone, 20 mL) was added, followed by fuming nitric acid (2 mL) dropwise over 20 mins. After stirring at 0°C for 90 mins, the on mixture was poured onto ice (200 g) and basified to pH 12 with 6N NaOH (aq.).

After allowing the ice to melt, the mixture was extracted with EtOAc (3 x 500 mL), and the combined organic portions dried over MgSO4 and evaporated to afford the title compound (22.1 g, purity approx. 71%) which was used in the subsequent step without purification. 1H NMR (400 MHZ, CDC13) 8 ppm 2.89 (d, J=3.8 Hz, 1 H), 3.09 (br. s., 1 H), 3.28 - 3.54 (in, l H), 3.80 — 4.03 (m, l H), 4.50 — 4.70 (m, 3 H), 4.71 — 4.86 (m. l H), 7.21 — 7.30 (m. 1 H), 8.18 — 8.28 (m, 1 H), 8.45 (dd, J=6.8. 2.8 Hz, 1 H) 1-(12) Synthesis of tart-but l 4aS.SS.7aS ~7a- 2-fluoronitro hen 1 —5- trifluorometh 1 -4a 5.7.7a-tetrah dro-4H-furo 3.4—d 1 3 thiazin l carbamate (4aS,SS,7aS)-7a-(2—Fluoronitrophenyl)—5-(trifluoromethyl)-4a,5,7,7a—tetrahydro—4H— furo[3,4—d][1.3]thiazin—2—amine (20.6 g, crude) was dissolved in THE (300 mL), di—tert— butyl dicarbonate (12 g) was added portionwise over 20 mins and the reaction mixture was heated to 60°C. Further portions of di-tert-butyl dicarbonate (10 g) were added until starting material was consumed by TLC. The reaction mixture was cooled and sodium bicarbonate (sat., aq.. 200 mL) and brine (200 mL) were added. The mixture was then extracted with EtOAc (3 x 500 mL) and the combined organic portions were dried over MgSO4 and evaporated. The residue was purified by silica gel column tography ( 10% to 25% EtOAc in hexanes) to afford the title compound (16.62g). 1H NMR (400 MHZ, CDCig) 5 ppm 1.55 (s. 9 H). 2.73 — 2.84 (m, l H), 2.92 e 3.05 (m, 1 H). 3.43 — 3.55 (m, 1 H), 3.81 — 3.94 (m, 1 H). 4.57 (d, J=8.3 Hz, 1 H), 4.73 ~4.83 (m. 1 H), 7.19 - 7.39 (m, 2 H), 8.20 — 8.29 (m, 1 H), 8.32 (d, J=6.8 Hz, 1 H) l—(13) S nthesis of tert-but 1 4aS,SS 7218 —7a— 5—amino—2—fluoro hen 1 —5- trifluorometh 1 -4a.5 7.7a—tetrah dro-4H—furo 3,4-d 1.3 n—2— 1 carbamate lert—B utyl ((4aS ,SS,7aS)-7a-(2—fluoronitrophenyl)—5—(trifluoromethyl)—4a,5,7,7a- tetrahydro—4H—furo[3,4~d][l,3]thiazin—2—y1)carbamate (16.61 g) was ved in ethanol (250 mL) and tin chloride dihydrate (25.0 g) was added. After stirring at RT for 18 h. the on was poured onto NaOH (2N aq., 300 mL) and celite® (~50 g) was added.

The resulting mixture was filtered through more celite® and extracted with EtOAc (2 x 500 mL). The combined c ns were dried over MgSO4 and evaporated to afford the title compound (15.52 g). This material could be used crude but a portion was purified by silica gel column chromatography (20% to 50% EtOAc in hexanes) to afford pure material (recovery 79%). 1H NMR (400 MHZ, CDC13) 5 ppm 1.53 (s, 9H), 2.77 (d, 1:144 Hz, 1H), 3.09 (br. s., 1H), 3.46 (br. s., 1H), 3.62 (br. s., 2H), 3.87 (br. 3., 1H), 4.61 (d, J=8.6 Hz. 1H), 4.71 (br. 3., 1H), 6.61 (br. s., 2H), 6.85 — 6.95 (m, 1H) ation Example 2 S nthesis of ten—but l 4aS.SS.7aS ~7a- 5~amino-2—fluoro hen 1 trifluorometh l - 4a 5.7.7a-tetrah dro-4H—furo 3.4—d 1.3 thiazin—2— 1 carbamate 2— 10 —56— W0 20121098213 PCTIEP2012/050833 l ‘9“ " F " F o O O FFW FW W Frm F F 2-(2) F F 249) 2—(1) S nthesis of S —1— 2.3—difluoro hen l 1. l . l ~trifluorobut-3—en—2— l)ox ethanone A solution of llithium in hexane (2.50 M, 13.5 mL) was added dropwise over 20 mins to a solution ning l—bromo-2,3—diflu0robenzene (6.50 g) in Eth (50 mL) under a N2 atmosphere at -78°C. The reaction solution was allowed to stir for 60 mins.

(S)—N-Methoxy-N-methyl-2—((1,l,l—trifluorobut—3—en~2~yl)oxy)acetamide) (5 .20 g) in EtZO (10 mL) was then added dropwise to the reaction solution, and after stirring at - 78°C for l h, aqueous NH4Cl (50 mL) was added to the on solution, followed by warming to RT. NaHC03 (sat. aq., 100 mL) was added to the reaction solution, and the mixture was extracted with EtOAc (3 x 100 mL). The combined organic portions were dried over MgSO4, evaporated, and the residue was purified by silica gel column chromatography (1% to 10% EtOAc in hexanes) to obtain the title nd (3.91 g). 1H NMR (400 MHz, CDC13) 5 ppm: 4.33 - 4.43 (m, 1 H), 4.80 — 4.84 (m, 2 H), 5.55 — .67 (m, 2 H), 5.76 - 5.94 (m, l H), 7.18 — 7.28 (m, 1 H), 7.37 — 7.47 (m, l H), 7.70 (ddt, J=7.9, 6.0. 1.7 Hz, 1 H) 2—(2) S nthesis of S —l— 2.3—difluoro hen 1 1.1,l-trifluorobut—3—en—2— Vl )oxyjethanone oxime WO 98213 (S)~1—(2,3—difluorophenyl)—2-((1,1,l-trifluorobut—3—en—2—yl)oxy)ethanone (3.91 g) was dissolved in anhydrous ol (40 mL) and hydroxylamine hydrochloride (1.25 g) and sodium acetate (1.68 g) were added. The reaction mixture was heated to 50°C for 90 min, then cooled to RT, concentrated in vacuo and the residue purified by silica gel chromatography (2% to 20% EtOAc in hexanes) to afford the title compound as a mixture of ric isomers (4.10 g). 1H—NMR (400 MHz, CDCI3) 5 (ppm): 4.04 - 4.26 (m 1 H), 4.43 — 4.55 (m, 0.4 H) 4.80 — 4.89 (m, 1.6 H) 5.39 - 5.55 (m, 2 H) 5.64— .80 (m, 1 H) 7.05 — 7.30 (m, 3 H), 7.76 (br s, 0.2 H), 8.30 (br s, 0.8 H). 2—(3) S nthesis of 48 —6a- 2.3-difluoro hen l —4- 'trifluorometh l hexah drofuro 3 4- clisoxazole (S)(2,3-difluorophenyl)—2-((1,1,1—trifluorobuten-2—yl)oxy)ethanone oxime (4.10 g) was dissolved in xylenes (40 mL) and hydroquinone (380 mg) was added. The on mixture was heated to reflux (heating block temperature 140°C) for 20 h, then cooled and evaporated. The residue was purified by silica gel column chromatography (1% to 25% EtOAc in hexanes) to obtain the title compound (3.16 g). 1H NMR (400 MHZ, CDC13) 6 ppm 3.77 (br. s., 1H), 3.99 - 4.16 (m, 1H), 4.16 - 4.22 (m, 1H), 4.22 — 4.44 (m, 2H), 4.51 (d. J=9.9 Hz, 1H). 5.44 (s, 1H), 7.07 — 7.24 (m, 2H). 7.38 (br. s.. 1H) 2-(4) Synthesis of {128.3 R,4S 2aminog 2,3-difluorophenyl 3 gtrifluoromethyl )tetrahydrofuran—3—yl )inethanol (4S)-6a—(2,3-diflu0rophenyl)—4—(trifluoromethyl)hexahydrofuro[3,4—c]isoxazole (3.16 g) was dissolved in acetic acid (20 mL) and the reaction mixture cooled to 0°C. Zinc (5.0 g) was added, and the reaction was allowed to warm and stir at RT for 20 h. The reaction mixture was then diluted with EtOAc (50 mL) and filtered through celite®, washing with a further 100 mL of EtOAc. The combined organic portions were ated, dissolved in CHC13 (20 mL), and ammonia (28% aq., 25 mL) was added slowly. The layers were separated, and the aqueous portion was further extracted with CHC13 (2 x 25 mL). The combined organic ts were dried over anhydrous MgSO4 and evaporated to afford the title compound (3.12 g) which was used in the subsequent step without further purification. 1H NMR (400 MHZ, CDClg) 5 ppm 2.93 (ddd, J=7.8, 5.1, 2.8 Hz, 1 H), 3.85 (dd, J=l2.4, 5.1 Hz, 1 H), 4.03 (dd, J=9.1, 2.8 Hz, 1 H), 4.14 (dd, J=12.3, 2.7 Hz, 1 H), 4.35 (d, J=9.1 Hz. 1 H), 4.68 (quin, J=7.3 Hz, 1 H), 7.09 - 7.25 (m, 2 H), 7.25 - 7.34 (m, 1 H) 2—(5) S s of N- . . gtrifluoromethyl )tetrahvdrofuran-3—yl mothioyl )benzamide Benzoyl isothiocyanate (2.0 mL) was added to a solution containing ((23,3R,4S)—4~ amino—4-(2,3-difluorophenyl)-2—(trifluoromethyl)tetrahydrofuran-3 -yl)methanol (3 .12 g) in DCM (20 mL), and the mixture was d at RT for 18 h. Sodium bicarbonate (sat, aq., 50 mL) was then added, the mixture extracted with EtOAc (3 x 75 mL), dried over MgSO4 and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (5% to 40% EtOAc in hexanes) to obtain the title nd (4.18 g). 1H NMR (400 MHZ, CDC13) 5 ppm 3.12 (dd, 1:7.6, 4.3 Hz, 1 H), 3.18 - 3.29 (m, l H), 4.03 (ddd, J=12.3, 7.2, 4.5 Hz, 1 H), 4.35 - 4.49 (m, 1 H), 4.59 (d, J=9.9 Hz, 1 H), 4.81 (d, J=9.6 Hz, 1 H), 7.07 — 7.23 (m, 2 H), 7.49 (t, J=7.2 Hz, 1 H), 7.56 (t, J=7.7 Hz, 2 H), 7.67 (t, J=7.5 Hz, 1 H), 7.88 (d, J=7.l Hz, 2 H), 8.92 (s, 1 H), 11.89 (s, 1 H) 2-(6) S nthesis of N—( 4aS,SS 7aS)-7a- 2,3-difluoro hen l trifluorometh l - 4a 5.7,7a—tetrah dro-4H-furo 3.4-d 1.3 thiazin—Z— l benzamide N—(((3S,4R,58)(2,3-Dif1uorophenyl)—4—(hydroxymethyl) (trifluoromethyl)tetrahydrofuran—3—yl)carbamothioyl)benzamide (2.99 g) was dissolved in ne ( 14 mL), and the mixture cooled to —20°C. Trifluoromethanesulfonic anhydride (1.55 mL) was added dropwise over 10 min and the reaction mixture was allowed to warm to -10°C. After stirring for 2 h, a further portion of trifluoromethanesulfonic anhydride (1.0 mL) was added dropwise over 10 min, the reaction was stirred for a further 2 h, and was then ed by the on of ammonium chloride (sat., aq., 50 mL) and extracted with EtOAc (3 x 100 mL). The combined organic extracts were dried over MgSO4, concentrated in vacuo and purified by silica gel column tography (5% to 20% EtOAc/hex) to obtain the title compound (1.20 g). 1H NMR (400 MHZ, CDC13) 8 ppm 2.86 (d, J=10.6 Hz, l H), 3.20 (br. 3., 1H), 3.55 (br. s.. l H), 4.04 (br. s., l H), 4.65 (d, J=8.8 Hz, 1 H), 4.81 (br. s., 1H), 7.06 — 7.24 (m, 3 H), 7.40 - 7.64 (m, 3 H), 7.82 — 8.21 (m, 2 H) 2—(7) S nthesis of 4aS.5S.7aS ~7a~ 2.3-difluoro hen l) trifluorometh l —4a.5 ,7,7a- vdro-4H-fnrol 3.4-d [ 1,3 lthiazin—2—arnine N—((4aS,SS,7aS)-7a—(2,3—Difluorophenyl)(trifluoromethyl)~4a,5,7,7a-tetrahydro-4H— furo[3,4—d][1,3]thiazinyl)benzamide (2.00 g) was dissolved in methanol (250 mL), l,8—diazabicyclo[5.4.0]undec—7—ene (1.53g) was added, and the solution was heated to reflux (heating block temperature 80°C). After 3 h, the reaction mixture was concentrated under reduced pressure, d with water (100 mL) and extracted with EtOAc (3 x 100 mL). The combined organic portions were dried over MgSO4, ated, and the residue purified by silica gel column chromatography (0% to 50% EtOAc in hexanes) to afford the title compound (1.42 g). 1H NMR (400 MHz, CDClg) WO 98213 8ppm 2.86 (dd, J=13.6, 3.8 Hz, 1 H), 3.15 (dd, J=13.8, 3.2 Hz, 1 H), 3.27 - 3.42 (m, 1 H). 3.93 (dd, J=8.2. 1.9 Hz, 1 H), 4.39 - 4.78 (m, 4 H), 6.96 — 7.25 (m. 3 H) 2—(8) S nthesis of 4aS.SS.7aS —7a— 2 3-difluoronitro hen l trifluorometh l)- 4a.5.7 7a—tetrah dro-4H—furo 3 4—d 1.3 thiazin—2—amine (4aS,SS,7aS)-7a—(2,3—Dilluoropheny1)—5—(trifluoromethyl)—4a,5,7,7a~tetrahydro—4H- furo[3,4-d][1,3]thiazin—2—amine (1.42 g) was dissolved in TFA (6 mL), and the solution was cooled to 0°C. Sulfuric acid (cone, 1 mL) was added, followed by fuming nitric acid (0.30 mL) dropwise over 20 mins. After stirring at 0°C for 1 h, the reaction e was poured onto ice (50 g) and basified to pH 12 with 2 N NaOH (aq.). After allowing the ice to melt, the mixture was extracted with EtOAc (3 x 75 mL), and the combined organic portions dried over MgSO4 and evaporated to afford the title compound (1.91 g, purity . 80%) which was used in the subsequent step without cation. 1H NMR (400 MHZ, CDCl3) 5 ppm 2.88 (dd, J=13.8, 3.9 Hz, 1 H), 3.11 (dd, J=13.6, 2.8 Hz, 1 H), 3.37 (dt, J=7.4, 3.5 Hz, 1H), 3.93 (d, J=7.8 Hz, 1 H), 4.53 - 4.83 (m, 4 H), 8.09 (ddd, J=9.0, 6.3, 2.9 Hz, 1 H), 8.27 (dt, J=5.2, 2.6 Hz, 1 H) 2—(9) Synthesis of ut l "4aS 58.7aS —7a— 2.3—difluoro~5-nitro hen l -5— trifluorometh l-4a.5.7.7a~tetrah dro-4H-furo 3.4—d 1.3 thiazin—2- lcarbamate (4aS,SS,7aS)-7a-(2,3-Difluoronitrophenyl)(trifluoromethyl)—4a,5,7,7a—tetrahydro- 4H—furo[3,4—d][1,3]thiazin—2—amine (1.91 g, crude) was ved in THF (10 mL), di— tert—butyl dicarbonate (1.3 g) was added portionwise over 20 mins and the reaction mixture was heated to 65°C. After 3 h, the reaction mixture was cooled and sodium bicarbonate (sat, aq., 50 mL) was added. The mixture was then extracted with EtOAc (3 x 750 mL) and the combined c portions were dried over MgSO4 and evaporated. The e was purified by silica gel column chromatography (0% to % EtOAc in hexanes) to afford the title compound (1.43g crude, in a mixture with the bis-boc version). 2—(10) S nthesis of [err—but 1 42.8 SS 7213 -7a- . (t1fi1uoromethy1)-4a,5 27,7a—tetrahydro—4H~furo[ 3 ,4-d H 1,3 lthiazin—2—y1 )carbamate The crude mixture obtained in preparation example 2—(9) containing Iert—Butyl ((4aS,SS,7aS)—7a-(2,3—difluoro—5~nitrophenyl)(trifluoromethyl)—4a,5,7,7a—tetrahydro— o[3,4—d][1,3]thiazin—2—yl)carbamate) (1.43 g) was dissolved in ethanol (25 mL) and tin chloride dihydrate (2.50 g) was added. After stirring for 18 h, the solution was poured onto NaOH (2N aq., 100 mL) and extracted with EtOAc (3 x 100 mL). The combined organic portions were dried over MgSO4, evaporated and purified by silica gel column cinematography (0% to 30% EtOAc in hexanes) to afford firstly the bis—boc ~60~ PCT/EP20121050833 product (730 mg), and secondly the title nd (300mg). 1H NMR (400 MHz, CDC13) 25me 1.53 (5,9 H), 2.76 (dd. 1:139, 3.8 Hz, 1H), 3.08 (d. J=13.6 Hz. 1 H). 3.36 - 3.45 (m, 1 H), 3.71 (br. s., 2 H), 3.90 (d, J=8.3 Hz, 1 H), 4.57 (d, J=8.6 Hz, 1 H), 4.68 - 4.79 (m, 1 H), 6.30 - 6.37 (m, 1 H), 6.42 - 6.49 (m, 1 H) Pre n exam 1e 3: S nthesis of 5—ethox razine—2-carboxv1ic acid 3- 2 /N Cl r /0\w):\N]/ N O O VOYENJ/ HOWE:j/O O O 3-(1) 3(2) S nthesis of eth 15-ethox razine-2—carbox late 3— 1 A stirred solution of methyl 5~chloropyrazine—Z—carboxylate (0.50 g) in ethanol (10 mL) was cooled to 0°C. and sodium ethoxide (21% w/w solution in ethanol, 1 mL) was added over 10 mins. After allowing to warm to RT and stir for 2 h, water (100 mL) was added and the mixture extracted with EtOAc (2 x 150 mL). The combined organic portions were dried over MgSO4 and evaporated to afford the title compound. (0.65 g, purity approx. 85%). 1H NMR (400 MHz, CDClg) 8 ppm 1.45 (t, J=7.1 Hz, 3 H), 1.46 (t, .1271 Hz, 3 H), 4.48 (q, J=7.1 Hz, 2 H), 4.49 (q, 1271 Hz, 2 H), 8.28 (d, J=1.3 Hz, 1 H), 8.88 (d, J=1.3 Hz, 1 H) Synthesis of 5—ethoxypyrazine—2—carboxylic acid 3—(2) Ethyl xypyrazinc—Z—carboxylatc (0.65g, approx. purity 85%) was dissolved in dioxan (3 mL) and water (3 mL) was added, followed by lithium hydroxide drate (255 mg, portionwise over 10 mins). After stirring at RT for 24h, 13th (25 mL) and NaHC03 (sat, aq., 25 mL) were added. The layers were separated and the c layer was extracted with NaOH (1 N, aq., 25 mL). The combined aqueous portions were ied with 6N HCl to pH 2, and the mixture extracted with EtOAc (3 x 40 mL). The combined EtOAc extracts were dried over MgSO4 and evaporated to afford the title compound as an off—white powder. 1H NMR (400 MHz. CDC13) 5 ppm 1.47 (t, J=7.1 Hz, 3 H), 4.53 (q, J=7.1 112,2 H), 8.16 (d, J=1.2 Hz, 1H), 8.98 (d, 121.2 Hz, 1 H) Pre aration exam 1e 4: S nthesis of 5—ethox razine-2—carboxvlic acid ’4— 3 O O F F F F HZNYLN[ka /N])K / EX _., :\N —-—-—> \N/N]X /O HO \N 0 o O 0 4(1) 4-(2) 4-(3) Meth t l razinecarbox late 4- 1 S—Acetylpyrazine-Z—carboxamide (3.275 g) was dissolved in methanolic HCl (1.25 N, 150 mL) and the reaction mixture was heated to reflux and d overnight. After cooling, sodium bicarbonate was added and the mixture was extracted with EtOAc.

The EtOAc layer was dried over MgSO4 and evaporated to afford the title compound (3.79 g, approx purity 90%). 1H NMR (400 MHZ, CDCl3) 5 ppm 2.78 (s, 3 H), 4.10 (s, 3 H), 9.33 (d, J=15 Hz, 1 H), 9.36 (d, J21.5 Hz, 1 H) Methyl 5—1 1,1—difluoroethvl )pyrazinecarboxylate 4-12 1 Methyl 5-acetylpyrazine-Z-carboxylate (300 mg, approx purity 90%) was ved in DCM (15 mL) and cooled to 0°C under nitrogen. Bis(2-methoxyethyl)aminosulfur trifluoride (0.61 mL) was added dropwise and the reaction mixture allowed to warm to RT and stir overnight. Sodium bicarbonate (sat, aq.) was added cautiously and the mixture extracted with DCM. The organic portions were dried over MgSO4. evaporated and purified by silica gel chromatography (35% EtOAc in hexane) to afford the title compound (155 mg) as a white solid. 1H NMR (400 MHZ, CDClg) 5 ppm 2.00 (t, J=18.8 Hz, 3 H), 4.01 (s, 3 H), 8.98 (d, J=1.5 Hz, 1 H), 9.24 ((1, 1:1.5 Hz, 1 H) 5—; 1,1—Difluoroethyl )pyrazine—2-carboxy1ic acid 4—1 3 1 Methyl 5-(1,l-difluoroethyl)pyrazine~2~carboxylate (0.65 g, approx. purity 85%) was dissolved in dioxan (2 mL) and water (:2 mL) was added, followed by lithium hydroxide monohydrate (54 mg, portionwisc). After stirring at RT for 90mins, the e was concentrated to 2 mL and Eth (20 mL) added. The mixture was then extracted with NaOH (1 N, aq., 20 mL), and the aqueous portions ed with 6N HCl to pH 2.

The aqueous portion was then extracted with EtOAc, dried over MgSO4 and evaporated to afford the title nd as a white solid (119 mg). IH NMR (400 MHZ, CDClg) 5 ppm 2.11 (t, J=18.8 Hz, 3 H), 9.01 (d, J=1.3 Hz, 1 H), 9.47 (d, J=1.3 Hz, 1 H) Preparation example 5: Synthesis of S—(‘fluoromethyl)pyrazine—2—carboxylic acid 15—13)) O ‘0 )1) [N ”YEN”)2 ”fig“ F —> | I _’ o \ HO \ / \(ENEAFfiN N O O 0 (1) 5-(2) 5-(3) Meth 15— h drox meth 1 razine—Z—carbox late 5- l To a solution of methyl 5—formylpyrazine—2—carboxylate (2.47 g) in THF (20 mL) was added sodium borohydride (170 mg) portionwise over 10 mins. After stirring for 1 11, methanol (10 mL) was added. The reaction mixture was stirred for a further 20 mins, and then HCl (1 N, aq., 20 mL) and brine (20 mL) were added. The mixture was extracted with EtOAc (3 x 40 mL) and the combined organic portions dried over MgSO4 and evaporated to afford the title compound (1.31 g). 1H NMR (400 MHz, 5ppm 4.07 (s, 3 H), 4.98 (br. s., 2 H), 8.80 (s, l H), 9.27 (s, 1 H) Methyl 5—(fluoromethVl)pvrazine—2-carboxylate 5—g 2) To a solution of methyl roxymethyl)pyrazine—2-carboxylate (0.64 g) in THF (20 mL) was added triethylamine (2.30 g) and the solution was cooled to 0°C.

Triethylarnine trihydrofluoride (1.22 g) was then added followed by orobutanesulfonyl fluoride (2.28 g) di‘opwise over 5 mins. After warming to RT and stirring for 2 h, NaHC03 (sat, aq., 100 mL) was added, and the mixture was extracted with EtOAc (2 x 50 mL). The combined organic portions were dried over MgSO4, evaporated, and purified by silica gel chromatography (5% to 50% EtOAc in hexane) to afford the title compound (94 mg). 1H NMR (400 MHZ, CDCl3) 5 ppm 4.07 (s, 3 H), 5.67 (d, J=46.5 Hz. 2 H), 8.89 (s. 1 H), 9.28 (s, 1 H) 5- Fluorometh 1) razine-Z-carbox lic acid 5- 3 Methyl 5—(f1uoromethyl)pyrazine—Z-carboxylate (94 mg) was dissolved in dioxan (1 mL) and water (1 mL) was added, followed by lithium hydroxide monohydrate (60 mg).

After stirring at RT for 18 h, EtZO (20 mL) was added and the mixture was then ted with NaOH (1 N, aq., 2 x 20 mL). The aqueous portions were acidified with 6N HCl to pH 1, extracted with EtOAc (2 x 40 mL), the combined organic portions dried over MgSO4 and evaporated to afford the title nd as a white solid (71 mg). 1H NMR (400 MHZ, CDCig) 5 ppm 5.70 (d, J=46.2 Hz. 2 H), 8.85 (s, 1 H), 9.40 (s, 1 H) Pre aration exam 1e 6: S nthesis of 5—difluorometh l razine-Z—carbox lic acid '6-‘5 ~63- PCT/EP20121050833 ”MINTN/ , O XYLD//O rv/ O 6-(1 ) 5(2) F F ,Nf0 fixm—waiemmflj/kF /N]/KF O O 543) -(4) 6—(5) S nthesis of t-but lS—meth l Irazinecarbox late 6- l A boron trifluoride-diethyl ether x (91.7 tLL) was added dropwise to a suspension of 2-methylpyrazine—5-carb0xylic acid (1 g) and tert-butyl 2,2,2— trichloroacetimidate (4.75 g) in THF (20 mL) under ice~cooling. The reaction on was warmed to RT, followed by stirring for 2 h. A saturated NaCl solution and EtOAc were added to the reaction solution, and the organic layer was separated. The organic layer was dried over anhydrous MgSO4. and the insoluble matter was separated by filtration. The filtrate was trated and purified by silica gel column chromatography to obtain the title compound (1.4 g). 1H—NMR (CDC13) 8 (ppm): 1.65 (s, 9H), 2.65 (s, 3H), 8.57 (d, J: 1.2 Hz, 1H), 9.10 (d, J = 1.6 Hz, 1H).

S nthesis of t-but 15- E dimeth lamino-vin l - A mixture of t—butyl ylpyrazine—Z—carboxylate (1.35 g), DMF (25 mL) and N,N— dimethylformamide dimethylacetal (25 mL) was stirred at 130°C for 5 h. The reaction solution was cooled to RT and diluted with EtOAc. The mixture was washed with a saturated NaCl solution three times. The organic layer was dried over anhydrous MgSO4, and the insoluble matter was separated by filtration. The filtrate was trated and the residue was purified by silica gel column chromatography to obtain the title compound (648 mg). lH—NMR (CDC13) 8 (ppm): 1.63 (s, 9H), 3.00 (s, 6H), 5.16 (d, J = 12.8 Hz, 1H). 7.72 (d, J = 12.8 Hz, 1H), 8.16 (d, J = 1.2 Hz, 1H), 8.81 (d, J :16 Hz, 1H).

Synthesis of t—butyl ylpyrazine-Z-carboxylate 6—131 Sodium periodate (1.67 g) was added to a solution of t-butyl 5-((E)-2—dimethylamino- Vinyl)—pyrazine—2—carboxylate (645 mg) in 50% THF-water (26 mL), and the mixture was stirred at RT for 4 h. A saturated NaHC03 solution and EtOAc were added to the reaction solution, and the organic layer was separated. The organic layer was washed with a saturated NaCl solution and dried over anhydrous MgSO4. The insoluble matter was separated by filtration and the filtrate was concentrated. The residue was purified by silica gel column tography to obtain the title compound (249 mg). 1H- ~64— NMR (CDC13) 8 (ppm): 1.68 (s, 9H), 9.25 (d, J = 1.2 Hz, 1H), 9.36 (d, J = 1.6 HZ, 1H), .2 (s. 1H).

Synthesis of t-butyl 5-difluoromethylpyrazine—Z-carboxylate 6—141 [Bis(2—methoxyethyl)amino]sulfur trifluoride (662 uL) was added dropwise to a solution of l 5-formylpyraaine-Z-carboxylate (249 mg) in CH2C13 (12 mL) under a N2 atmosphere under ice-cooling. The reaction solution was stirred for 2 h while gradually returning to RT. A saturated Nal-IC03 solution and EtOAc were added to the reaction solution, and the organic layer was separated. The c layer was washed with a saturated NaCl solution and dried over anhydrous MgSO4. The insoluble matter was separated by ion and the filtrate was concentrated. The residue was purified by silica gel column chromatography to obtain the title compound (175 mg). 1H-NMR (CDC13) 5 (ppm): 1.67 (s, 9H), 6.75 (t, J =: 54.4 Hz, 1H), 9.02 (d, J = 0.8 Hz, 1H), 9.25 (d, J = 0.8 Hz, 1H).

Synthesis of 5~difluoromethy1pyrazine—2-carboxylic acid 6-(5) Trifluoroacetic acid (1 mL) was added to a solution of t—butyl 5- difluoromethylpyrazine-Z-carboxylate (175 mg) in dichloromethane (1 mL), and the mixture was stirred at RT for 5 h. Ether and 5 N NaOH were added to the reaction on. The aqueous layer was separated and made acidic with 5 N hloric acid.

EtOAc was added to the s layer, and the organic layer was separated. The organic layer was dried over anhydrous Mg8 04, and the insoluble matter was separated by filtration. The filtrate was concentrated to obtain the title compound (100 mg).

H—NMR (CDC13) 5 (ppm): 6.80 (t, J = 54.4 Hz, 1H), 9.02 (s, 1H), 9.47 (s, 1H).

Pre aration exam le 7: S nthesis of 5-c ano ridine—2-carbox lic acid 7—2 / CN / \.fl / ——* 1 ’Ho i / /O \N N N 1 O O 7~(1) 7-(2) S nthesis of meth l 5—c ano idine—2—carbox late 7 l A mixture of methyl opyridine~2~carboxy1ate (2.8 g) and copper cyanide (3.6 g) in NMP (30 mL) was heated with ng at 170°C for 1.5 h. Water was added to the reaction solution at RT, and the insoluble matter was removed by filtration. The filtrate was extracted with EtOAc. The extract was washed with a saturated NaCl solution and then dried over anhydrous MgSO4. The drying agent was removed by filtration and the filtrate was concentrated under reduced pres sure. The resulting crude t was purified by silica gel column chromatography (EtOAc—heptane system) to obtain the title compound (920 mg). 1H-NMR (400 MHZ, CDC13) 5 (ppm): 4.06 (s, 3H), 8.16 (dd, J = 2.0, 8.0 HZ. 1H), 8.27 (d, J = 8.0 HZ. 1H), 9.01 (d, J = 2.0 Hz. 1H).

Synthesis of 5-cyanopyridine-2—carboxylic acid 7-12) A solution of the compound of Preparation Example 13~(1) (920 mg) and a 5 N NaOH on (2.26 mL) in ethanol (30 mL) was stirred at RT for 10 min. 5 N hydrochloric acid (5.2 mL) was added to the reaction solution at RT, followed by extraction with EtOAc. The extract was dried over ous MgSO4. The drying agent was removed by filtration and the filtrate was concentrated under reduced pressure to obtain the title compound (800 mg). 1H—NMR (400 MHZ, DMSOd6) 5 (ppm): 8.18 (d. J = 8.0 HZ, 1H), 8.51 (dd, J = 2.0, 8.0 HZ, 1H), 9.12918 (m, 1H).

Pre aration exam 1e 8: S nthesis of 5- methox methvl razine—2-carbox lic acid 8— (2)) N N /N / fOMe / for/1e (Kn/E I /O \N HO \N / N o 0 (1) 8-(1) 8—(2) S s of meth 15- methox meth 1 razine—2—carboxvlate 8- 1 The compound obtained in preparation example 5—( 1) (279 mg) was dissolved in DMF and the solution was cooled to 0°C. Sodium hydride (60% in mineral oil, 70 mg) was added, followcd by thanc (250 mg). After 2 days, water (25 mL) was added, and the solution extracted with EtOAc (100 mL). The aqueous layer was saturated with NaCl, and r ted with EtOAc (2 x 50 mL). The combined organic portions were dried over MgS O4, evaporated, and purified by silica gel chromatography (30% to 50% EtOAc in hexanes) to afford the title compound (55 mg, approx. purity 65%). 1H NMR (400 MHZ, CDClg) 8 ppm 3.55 (s, 3 H), 4.05 (s, 3 H), 4.72 (s, 2 H), 8.84 (d, J=1.0 HZ, 1 H), 9.25 (d, J=1.0 Hz, 1 H) S nthesis of 5— methox meth l razine—2—carbox lic acid 8— 2 Methyl 5—(methoxymethyl)pyraZine—2—carboxylate, 8(1), (55 mg, crude) was ved in 1,4—dioxane (1 mL) and water (1 mL) was added followed by lithium hydroxide monohydrate (50 mg). After stirring at RT for 1 h, water (20 mL) was added and the mixture was extracted with ether (20 mL). The aqueous portion was acidified to pH 2 and extracted with EtOAc (2 x 25 mL). The combined EtOAc layers were dried over MgSO4 and evaporated to afford the title compound (19 mg). 1H NMR (400 MHZ, CDC13) Sppm 3.58 (s. 3 H), 4.77 (s, 2 H), 8.80 (br. s., 1 H), 9.38 (br. 3., 1 H) -66— Pre aration exam 1e 9: S—Methox razine-2—carbox lie acid '1. MeOH, H2804 2. NaOMe N OMe 2. NaOH I Nj/C' | HO HO /j/ N/ N O O -Chloropyrazineearboxylic acid (5.0 g, 0.032 mol) was charged to a round-bottom flask equipped with a thermocouple, overhead stirrer and reflux ser. Methanol (37.5 mL, 0.926 mol) was charged followed by cone. sulfuric acid (0.2 mL, 0.004 mol).

The 3—neck flask was equipped with a heating mantle, and then the reaction mixture was heated to ca. 65.0 °C (T al). The reaction e continued to stir at ca. 65.0 “C (T internal) for ca. 4 h. The reaction mixture cooled to ca. 25.8 °C (T internal).

Methanol (12 mL, 0.31 mol) was charged and the slurry continued to stir at ca. 22.3 °C (T internal) for ca. 15 min then cooled to ca. 10.0 0C (T internal) under an atmosphere of nitrogen. 25% Sodium methoxide in methanol (1 :3, Sodium methoxidezMethanol, 7.7 mL) was charged to flask while temperature remained below 30.0 °C (T al).

The reaction e was adjusted to 20.4 0C (T internal). After 30 min, sodium hydroxide (2.0 g, 0.04 mol) and water (37.5 mL, 2.08 mol) were combined to form a on, and then the solution was charged to the reaction mixture. Water (50.0 mL, 2.78 mol) was charged and then the reaction mixture was heated to 40.0 °C (T internal) for ca. 60 mins. The heating mantle was removed, and then the reaction mixture cooled to ca. 25.4 °C (T internal). 38% aq. HCl Solution (38:62, hydrogen chloridezwater, 4.0 mL) was added at a rate (ca. 5 min.) such that the temperature remained below 30.0 °C (T internal). The thick slurry was stirrred for l h at ca. 21.4 °C (T internal), and then filtered over a sintered . The solids were rinsed with water (10.0 mL, 0.555 mol) and dried under vacuum overnight to afford 5— methoxypyrazine-2—carboxy1ic acid ). 1H NMR (500 MHz, DMSO) 5 13.24 (1H, br s), 8.79 (1H, d, J = 1.2 Hz), 8.37 (1H, d, J: 1.2 Hz), 3.98 (s, 3H); 13c NMR (125 MHZ, DMSO) 5 165.36, 161.88, 143.88, 136.82, , 54.69.

General procedure for the coupling of anilines prepared in ation Examples 1-113) and 2— 10 with ar learbox lic acids: Pre aration of N— 3— 421853.733 —2—amino—5— trifluorometh 1 ~4a.5.7 7a-tetrah dro—4H—furo 3 4—d 1,3 thiazin—7a— fluorophenyl1—5-methoxypvrazinecarboxarnide (Example 1 1 ~67- N¢\Iro\ O \ N N NH2 O ‘Y te rt-Butyl ((4aS ,SS,7aS)—7a—(5—aminofluorophenyl)-5—(trifluoromethyl)-4a,5,7,7a— ydro—4H—furo[3,4-d][1,3]thiazin-2—yl)carbamate (Preparation Example 1, 56 mg) was dissolved in DCM (2 mL) and 5—methoxypyrazine—2-carboxylic acid (40 mg), N,N- diisopropylethylamine (80 mg) and (1H~benzotriazolyloxy)tripyrrolidin- l- yl)phosphonium hexafluorophosphate (135 mg) were added. The on e was stirred at RT for 18 h, and sodium bicarbonate (sat, aq., 25 mL) was added. The mixture was extracted with EtOAc (2 x 40 mL), the combined c portions were dried over MgS O4, evaporated and purified by silica gel chromatography (EtOAc/hexanes gradient) to afford the amide (40 mg) as a white solid. The amide was dissolved in DCM (2 mL) and TFA (1 mL) was added. After stirring at RT for 2 h, the reaction mixture was evaporated and sodium bicarbonate (sat, aq., 25 mL) was added. The mixture was extracted with EtOAc (2 x 25 mL), and the combined organic portions dried over MgSO4 and evaporated to afford the title compound as a white solid (34 mg). 1H NMR (400 MHZ, CDC13) 5 ppm 2.78 — 2.87 (In, 1 H), 3.11 — 3.21 (m, 1 H), 3.38 — 3.48 (m, 1 H), 3.82 - 4.06 (m, 4 H), 4.48 — 4.72 (m, 2 H), 6.96 — 7.10 (m, l H), 7.52 (d, J=4.8 Hz, 1 H), 7.87 (d, J=8.8 Hz, 1 H), 8.09 (s, 1 H), 8.95 (s, 1 H), 9.46 (br. s., 1 H) Exam 1e 2: N— 3— 4218 SS 7aS amino-5— trifluorometh 1 ~4a.5 7,7a—tetrah dm—4H- furo 3 4~d 1.3 thiazin—7a—vl)—4-fluoro hen l c ano icolinamide 0 ‘\ N NH2 0 E F FFH Synthesized from l'ert—butyl [(4aS,5S,7aS)—7a-(_5—aminofluorophenyl)—5- trifluoromethyl-4a,5 ,7,7a-tetrahydro—4H—furo[3 ,4-d] [1,3 ] n-Z-yl]carbamate and 5- cyanopyrazine-Z-carboxylic acid according to the general procedure. 1H NMR (400 MHz,CDC13)6 ppm 2.82 (dd, 1213.6, 3.5 Hz, 1 H), 3.14 — 3.21 (m, 1 H), 3.37 — 3.45 ~68~ (m, 1 H), 3.91 (d, J=9.1 Hz, 1 H), 4.50 — 4.71 (m, 2 H), 7.08 (dd, J=11.9, 8.8 Hz, 1 H), 7.46 — 7.57 (m, 1 H). 7.92 (dt. J=8.8, 3.4 HZ, 1 H), 8.17 (dd, J=8.3, 2.0 Hz, 1 H). 8.34 (d.

J=8.1Hz, 1H), 8.81 - 8.92 (m, 1 H) Exam 1e 3: N— 3— 43853735 ~2—amino-5— orometh 1 —4a.5.7.7a-tetrah Ciro—4H— carboxamide N NH2 O \Y Synthesized from rert—butyl [(4aS,SS,7aS)—7a— (5-amino-2—fluoropheny1)—5- romethyl-4a,5 ,7,7a-tetrahydro—4H—furo[3 ,4—d] [1 ,3]thiazin—2-y1]carbamate and 5— difluoromethyl—pyrazine—Z—carboxylic acid according to the general procedure. 1H NMR (400 C13)5 ppm 2.81) (dd, J=13.8, 3.7 Hz, 1 1-1), 3.11 (dd, J=13.6, 2.8 Hz, 1 H), 3.31 — 3.41 (m, 1 H), 3.86 (d, J=8.3 Hz, 1 H), 4.57 (d, J=8.3 Hz, 1 H), 4.64 (dt, 1:147, 7.1 Hz, 1 H), 4.75 (br s, 2 H), 6.69 (t, J=56.3 Hz, 1 H), 7.07 (dd, J=11.6, 8.8 Hz, 1 H), 7.57 (dd, J:7.1, 2.8 Hz, 1 H), 7.87 (dt, J=8.5, 3.6 Hz, 1 H), 8.85 (s, 1 H), 9.45 (s, 1 H), 9.58 (br. 3., 1 H) Example 4: N—g 3;((4as,SS.7aS)-2—amino-5—(tn'f1uoromethy1)-4a,5,7.7a—tetrahydro—4H- furof3,4—d1[1.3]thiazin—7a—V1)—4—fluoropheny1)~5—(‘trifluoromethyl)picolinamide N/l F O \ F F Synthesized from tert—butyl [(4aS,SS,7aS)-7a— (5—amino—2—fluorophenyl)—5- trifluoromethyl~4a,5,7,7a-tetrahydro-4H-furo[3,4-d] [1,3]thiazin—2-y1]carbamate and 5- trifluoromethyl—pyridine—2—carboxylic acid according to the general procedure. 1H NMR (400 MHz, CDC13) 8 ppm 2.88 (dd, J=13.6, 3.8 Hz, 1 H), 3.20 (dd, J=13.6, 3.0 Hz, 1 H), 3.37 - 3.53 (m, 1H), 3.94 (dd, J:8.3. 2.3 Hz, 1 H), 4.40 - 4.89 (m, 4 H), 7.14 (dd, J=11.9, 8.8 Hz, 1 H), 7.66 (dd, J=6.8, 2.8 Hz, 1 H), 7.98 (ddd, 1:8.8, 4.0, 3.0 Hz, 1 H). 8.20 (dd, J=8.2, 1.6 Hz, 1 H), 8.45 (d, J=8.3 Hz, 1 H), 8.90 (s, 1 H). 9.95 (s. 1 H) Exam 1e 5: N— 3- 4aS.SS 7aS -2—amino trifluorometh 1 -4a.5 7.7a-tetrah dro—4H— Synthesized from utyl [(4aS,SS,7aS)-7a- (5-amino-2—fluoropheny1)—5— trifluoromethyl—4a,5 ,7,7a-tetrahydr0-4H-fur0[3,4—d] [1,3]thiazin-2—y1]carbamate and 5- methyl-pyrazine-Z—carboxylic acid according to the general procedure. 1H NMR (400 MHz, CDC13) 5 ppm 2.72 (s, 3 H), 2.88 (dd, J=13.6, 3.8 Hz, 1 H), 3.21 (dd, 1:136, 3.0 Hz, 1 H), 3.42 — 3.49 (m, 1 H), 3.94 (d, J=8.3 Hz, 1 H), 4.39 - 4.80 (m, 4 H), 7.14 (dd, J=11.9, 8.8 HZ, 1 H), 7.62 (dd, J=7.1, 2.8 Hz, 1H), 7.93 - 7.99 (m, 1 H), 8.46 (d, J=1.0 Hz, 1 H), 9.39 (d, J=1.3 Hz, 1 H), 9.65 (s, 1 H) Exam 1e 6: N— 3- 4aS.SS.7aS no-5— trifluorometh 1 —4a 5.7.7a—tetrah dro-4H— furo 3,4-d 1.3 thiazin—7a— ' ' ' Synthesized from terz—butyl [(4aS,SS,7aS)-7a— (5~mnino—2—fluor0phenyl)—5- trifluoromethy1-4a,5 ,7,7a-tetrahydr0-4H—fu1‘0[3,4—d] [1,3]thiazin—2-yl]carba1nate and 5- 2 O methyl-pyridine-Z-carboxy1ic acid according to the l procedure. 1H NMR (400 MHZ, CDC13) 5 ppm 2.37 (s, 3H), 2.78 (dd, J:13.6, 3.8 Hz, 1H), 3.12 (dd, 1:136, 2.8 Hz, 1H), 3.28 — 3.40 (m, 1H), 3.87 (d, J=8.1 Hz, 1H), 4.28 — 5.02 (m, 4H), 7.02 (dd, , 8.8 Hz, 1H), 7.55 (dd, J=7.1, 2.8 Hz, 1H), 7.63 (dd, J=8.0, 1.4 Hz, 1H), 7.88 (ddd, J=8.8, 4.0, 2.9 Hz. 1H), 8.10 (d, J=8.1 Hz, 1H). 8.31 — 8.40 (m, 1H). 9.90 (s, 1H) Exam le 7: N- 3— 4aS 58.7aS ~2—amino trifluorometh 1 -4a.5.7.7a-tetrah dro-4H- furo 3 4-d 1.3 thiazin«7a— PCTI’EP2012/050833 O \ N\ NH2 0 I F FFH Synthesized from terr—butyl [(4aS,SS,7aS)—7a- (5-amin0-2—fluor0pheny1)—5— trifluoromethyl—4a,5 ,7,7a~tetrahydro~4H—furo[3 ,4~d] [1,3]thiazin—2-y1]carbamate and 5- ethyl—pyridine—Z-carboxylie acid according to the general ure. 1H NMR (400 MHZ, CDC13) 8 ppm 1.33 (t, J=7.6 Hz, 3 H), 2.78 (q, J=7.6 Hz, 2 H), 2.88 (dd, J=13.5, 3.7 Hz. 1 H), 3.22 (dd. J=13.6. 2.8 Hz, 1 H), 3.42 — 3.48 (m. 1 H), 3.96 (d, J=7.3 Hz, 1 H), 4.44 — 4.95 (m, 4 H), 7.12 (dd, J=11.6, 8.8 HZ, 1 H), 7.62 (dd, J=6.9, 2.7 Hz, 1 H), 7.74 (dd, 128.1, 1.8 Hz, 1 H), 7.98 (dt, 1:8.7, 3.5 Hz, 1 H), 8.21 (d, J=8.1 Hz, 1 H), 8.44 - 8.48 (m, 1 H), 10.00 (s, 1 H) Exam 1e 8: N— 3— 4aS.5S.7aS no—5— trifluorometh 1 —4a 5.7.7a-tetrah dro—4H- furo 3 4-d 1.3 thiazin-7a—v1)-4—fluoro hen 1 —5— fluorometh 1 razine—2-carboxamide Synthesized from tert—butyl [(4aS,SS,7aS)~7a— (5—amino—2—f1uoropheny1)—5— trifluoromethyl—4a,5 ,7,7a—tetrahydro—4H—furo[3 ,4—d] [1,3]thiazin-2—yl]carbamate and 5— fluoromethyl-pyrazine—2—carboxylic acid ing to the general procedure. Details of an actual preparation are as fel1ows:~tert—Buty1 ((4aS,SS,7aS)-7a—(5—amin0—2— fluorophenyl)—5—(triflu0romethy1)-4a,5,7,7a—tetrahydro-4H-furo[3,4—d][1,3]thiazin-2— yl)carbamate (500 mg) was dissolved in DCM (10 ml.) and 5—fluoromethy1-pyraz.ine—2— carboxylic acid (223 mg), N,N~diisopropylethylamine (521 mg) and (1H—benzolriazel- 1—yloxy)tripyrrolidin—1-y1)phosphonium hexafluorophophate (750 mg) were added. The reaction mixture was stirred at RT for l h, and sodium bicarbonate (sat., aq., 50 mL) was added. The mixture was extracted with EtOAc (2 x 75 mL), the combined c portions were dried over MgS 04. evaporated and ed by silica gel chromatography (0% to 30% EtOAc/hcxanes gradient) to afford the amide (613 mg) as a white solid.

The amide was dissolved in DCM (2 mL) and TFA (1 mL) was added. After stirring at RT for 2 h, the reaction e was evaporated and sodium bicarbonate (sat, aq., 25 mL) was added. The mixture was extracted with EtOAc (3 x 25 mL), and the ed organic portions dried over MgSO4 and evaporated to afford the title compound as a white solid 1H NMR (400 MHz, CDC13) 5 ppm 2.89 (dd, J=13.8, 3.7 Hz, 1 H), 3.21 (dd, J=13.9, 2.5 Hz, 1 H), 3.40 - 3.51 (m, 1 H), 3.96 (d, J=7.3 Hz, 1 H), 4.42 — 4.85 (111,4 H), 5.69 (d, J: 46.5 Hz, 2 H), 7.15 (dd, J=11.9, 8.8 Hz, 1 H), 7.64 (dd, 127.1, 2.8 Hz, 1 H), 7.94 - 8.00 (m, 1 H), 8.77 (s, 1 H), 9.47 (s, 1 H), 9.68 (s, 1 H) Exam 1e 9: N— 3- 4318.53 7213 —2—amino-5— trifluorometh 1 —4a.5.7.7a—tetrah dro—4H— furo 3 4—d 1.3 thiazin-7a-V1)—4—f1uoro hen l —5-rnethox icolinamide N/ O\ O \ N NH2 O Y sized from tert-bulyl [(4aS,SS,7aS)-7a- (5—amino—2—fluorophenyl)—5— trifluoromethyl-4a,5 ,7,7a-tetrahydro-4H-furo[3,4-d] [1,3]thiaziny1]carbamate and 5- methoxypyridme-Z—carboxylic acid according to the general procedure. 1H NMR (400 MHz, CDC13) 8 ppm 2.88 (dd, J=13.8, 3.7 Hz, 1 H), 3.23 (dd, J=13.8, 2.4 Hz, 1 H), 3.46 (d, J=7.3 Hz, 1 H), 3.89 - 4.02 (m, 4 H), 4.54 — 5.00 (m, 4 H), 7.11 (dd, , 8.8 Hz, 1 H), 7.36 (dd, J=8.8, 2.8 Hz, 1 H), 7.61 (dd, J=7.1, 2.8 Hz, 1 H), 7.97 (dt, J=8.5, 3.6 Hz, 1 H), 8.23 — 8.30 (m, 2 H), 9.86 (s, 1 H) Exam 1e 10: N— 3- 4218 58.7213 ~2~amino—5~ trifluoromelh 1 ~4a.5 7.7a—telrah ~ furo 3.4—d 1.3 thiazin-7a4-fluoro hen 1 ~5-ethox razine—2-carboxarnide Synthesized from tert—butyl [(4aS,SS,7aS)-7a- (5 —amino-2—fluorophenyl)—5- trifluoromethyl—4a,5 ,7,7a~tetrahydro-4H—furo[3,4-d] hiazin—2-y1]carbamate and 5- ethoxypyrimidine-Z-carboxylic acid according to the general procedure. 1H NMR (400 MHz, CDC13) 5 ppm 1.47 (t, J=7.1 Hz, 3 H), 2.87 (dd, 1:136, 3.5 Hz, 1 H), 3.20 (d, J=13.6 Hz, 1 H), 3.40 ~ 3.50 (m, 1 H), 3.94 (d, J=7.8 HZ, 1 H), 4.46 — 4.95 (In, 6 H), 7.11 (dd. J=11.5. 9.0 Hz, 1 H), 7.54 - 7.63 (m, 1 H), 7.90 - 8.01 (m, 1 H). 8.13 (s, 1 H). 9.00 (s, 1 H), 9.52 (br. 3., 1 H) Exam 1e 11: N— 3- 421853.738 ~2—amino—5— trifluoromethvl)-4a.5.7.7a—tetrah (ho-4H— furo 3.4—d 1.3 lhiazin-7a—v1)-4—fluoro hen 1 1.1—difluoroeth 1 razine—Z- carboxamide Synthesized from tert-buty1 [(4aS,SS,7aS)-7a— (5 —amino—2—fluorophenyl)—5 - trifluoromethy1—4a,5 ,7,7a—tetrahydro-4H—furo[3,4—d][1,3]thiaziny1]carbamate and 5- (1,1-difluorocthy1)pyrazinc—Z-carboxylic acid according to the l procedure. 1H NMR (600 MHz, CDC13) 5 ppm 2.03 (t, J=18.8 Hz, 3 H), 2.82 (d, 1212.0 Hz, 1 H), 3.14 (d, 1:124 Hz, 1 H), 3.35 — 3.46 (m, 1 H), 3.77 - 4.07 (m, 1 H), 4.20 - 4.93 (m, 4 H), 7.08 (dd, 1211.7, 9.0 Hz, 1 H), 7.56 (d, J=4.5 Hz, 1 H), 7.80 - 7.93 (m, 1 H), 8.87 (s, 1 H), 9.43 (s, 1 H), 9.59 (br. 3., 1 H) 3- 4218 55.7215 —2—amino-5— trifluorometh 1)-4a.5 7.7a—tetrah — furgL3,4-dH léLthiaZin-h—V1)—4-fluorophgny1)—5—('trif1uoromethy_l)pyrazine—Z— OVVN N NH2 O ‘Y F FH F sized from tert—butyl [(4aS,SS,7aS)-7a- (5—amino—2—fluor0pheny1)—5— trifluoromethy1-4a,5,7,7a-tetrahydro-4H-furo[3,4—d] [1,3]thiaziny1]carbamate and 5- trifluoromethylpyrazine-Z—carboxylic acid according to the general procedure. 1H NMR (400 MHz, CDC13) 5 ppm 2.80 (dd, J=13.6, 3.8 Hz, 1 H), 3.11 (dd, J=13.8, 2.9 Hz, 1 H), 3.30 - 3.44 (m, 1 H), 3.87 (d, J28.3 Hz, 1 H), 4.25 — 5.14 (m, 4 H), 7.07 (dd, , 8.8 HZ, 1 H), 7.57 (dd, J=6.8, 2.8 Hz, 1 H), 7.86 (dt, J=8.4, 3.6 HZ, 1 H), 8.89 (s, 1H). 9.53 (S, 2 H) Exam le 13: N— 3- 4213 SS 7213 amino trifluoromethvl)-4a.5.7.7a—tetrah dro-4H- carboxamide sized from tert—butyl [(4aS,SS,7aS)~7a— (5-amino-2—f1uorophenyl)—5- trifluoromethyl—4a,5,7,7a~tetrahydro-4H-furo[3,4—d] [1,3]thiazin—Z—yl]carbamate and 5- (methoxymethyl)pyrimidine-2—carboxylic acid according to the general procedure. 1H NMR (400 MHZ, CDC13) 5 ppm 2.79 (dd, J=13.6, 3.8 Hz, 1 H), 3.11 (dd, , 2.8 Hz, 1 H), 3.32 — 3.39 (m, 1 H), 3.49 (s, 3 H), 3.84 (d, J=8.3 Hz, 1 H), 4.34 - 4.73 (m, 6 H), 7.05 (dd, J=11.6, 8.8 Hz, 1 H), 7.55 (dd, J=6.8, 2.8 Hz, 1 H), 7.88 (dt, J=8.4, 3.6 Hz, 1 H), 8.63 (s, 1 H), 9.34 (d. J=1.0 Hz, 1 H), 9.60 (s, 1 H) Exam 1e 14: N— 3- 4aS 58.7aS —2-amino—5— trifluorometh l ~4a.5—dih dro—4H— 2H3 )methyloxy [Eyrazine—Z— carboxamide Synthesis of 14—(2) 5—]g2H3)methyloxy[pyrazine-Z-carboxylic acid 11N\ IfN\ oco3 N\ 00:33 *r —‘"’ Me02C N [330020 N/ II H020 N Part (I): (2H_3)methyl 5—[(2H; meth 10x razine-Z—carbox late Freshly cut sodium metal (160 mg) was added portionwise over 10 mins to (2H3)methan(2H)ol (5 mL) and the on was stirred until the sodium had dissolved.

This solution was then added to methyl 5-chloropyrazine—2-carboxylate (1.02 g) in (2H3)methan(2H)ol (5 mL) and the solution was allowed to stir at RT for 1 hr. The solution was then concentrated under reduced pressure to a volume of about 2 mL, and water (50 mL) was added. The mixture was ted with EtOAc (2 x 50 mL), the combined organic portions were dried over MgSO4 and evaporated to afford the title compound (745 mg). 1H NMR (400 MHz, CDC13) 5 ppm 8.30 (d, J=1.3 Hz, 1 H), 8.91 (d, J=1.3 Hz, 1 H) PCTIEP2012/050833 Part (II ): 5-1t2H;)methyloxylgyrazine-Z—carboxylic acid To a stirred solution of (2H3)methyl 5-[(2H3)methyloxy]pyrazinecarboxy1ate in 1,4- dioxane (5 mL) was added water (5 mL) followed by lithium hydroxide monohydrate (300 mg). After stirring for 1 hr, the reaction mixture was trated under d pressure to about 5 mL and extracted with diethyl ether (25 mL). The organic layer was extracted with 1N NaOH (aq., 10 mL), and the combined aqueous portions were acidified to pH 2 with 6N hydrochloric acid. After cooling in a fridge, the mixture was filtered to afford the title compound as a pale brown powder (660 mg). 1H NMR (400 MHZ, CDC13) 8 ppm 8.21 (d. J=1.3 Hz, 1H), 9.01 (d, J=1.3 Hz, 1 H). 10.12 (br s., 1 H) furo 3.4 2H; )methyloxy I ne-2—carboxamide NWo/ jiH OWN H NYNHz tert—Butyl ((4aS,SS,7aS)—7a—(5—amino-2—1'luorophenyl)—5-(trifluoromethyl)—4a,5,7,7a— tetrahydro—4H—furo[3,4—d][1,3]thiazin—2—yl)carbamate (100 mg) was dissolved in DCM (2 mL) and 5—[(ZH3)methyloxy]pyrazine—2—carboxylic acid (55 mg), N,N— diisopropylethylamine (112 mg) and nzotriazol-1—yloxy)tripyrrolidin—1— yl)phosphonium hexafluorophosphate (180 mg) were added. The reaction mixture was stirred at RT for 18 h, and sodium bicarbonate (sat, aq., 25 mL) was added. The mixture was extracted with EtOAc (2 x 40 mL), the combined c portions were dried over MgSO4, ated and purified by silica gel chromatography (2 % to 25% EtOAc in hexanes) to afford the amide (127 mg) as a white solid. The amide was dissolved in DCM (2 mL) and TFA (1 mL) was added. After stirring at RT for 2 h, the reaction mixture was evaporated and sodium bicarbonate (sat, aq., 25 mL) was added.

The mixture was extracted with EtOAc (2 x 40 mL), and the combined organic portions dried over MgSO4 and ated to afford the title compound as a white solid (104 mg). 1H NMR (400 MHZ, CDC13) 6 ppm 2.87 (dd, J=13.6, 3.8 Hz, 1 H), 3.21 (dd, J=13.6. 2.8 Hz, 1 H). 3.39 - 3.53 (m, 1H), 3.95 (d, J=8.3 Hz, 1 H), 4.65 (d, J=8.3 Hz, 1 H), 4.72 (quin, .1272 Hz, 1 H), 4.87 (br s, 2 H), 7.12 (dd, J=11.9, 8.8 Hz, 1 H), 7.60 (dd, 126.9, 2.7 Hz, 1 H), 7.95 (dt, J=8.5, 3.6 Hz, 1 H), 8.16 (d, J=1.0 Hz, 1 H), 9.02 (d, J=1.0 Hz, 1 H), 9.52 (s, 1 H) PCT/EP20121050833 Exam 1e 15: N— 3— 4aS SS 7218 amino~5— trifluorometh 1 —4a.5 7 7a—tetrah dro—4H- furo 3,4-d 1.3 thiazin-7a— 1 -4.5-difluoro hen 1 —5— difluorometh 1 carboxamide Synthesized from tert—butyl [(4aS,SS,7aS)-7a— (5~amino-2,3~dif1u01'ophenyl)~5— trifluoromethyl—Arafi,7,7a—tetrahydro-4H—furo[3 ,4—d] [1,3]t111azin~2—y1]carbmnate and 5— difluoromethyl-pyrazine-2—c2u'b0xy11c acid according to the general procedure. 1H NMR (400 MHZ, CDC13) 8 ppm 2.90 (dd, J213.8, 3.7 Hz, 1 H), 3.19 (dd, J213.8, 2.7 Hz, 1 H), 3.31 — 3.49 (m, 1 H), 3.95 (d, J=7.6 Hz, 1 H), 4.44 — 5.15 (m, 4 H), 6.81 (t, J255.8 Hz, 4 H), 7.22 — 7.35 (m, 1 H), 8.08 (ddd, J211.2, 6.8, 2.7 Hz, 1 H), 8.94 (s, 1 H), 9.53 (s, 1 H), 9.67 (s, 1 H) Exam 1e 16: N— 3- .7aS ~2—amin0 rometh 1 -4a.5.7.7a-tetrah dl‘O-4H- furo 3.4-d 1.3 thiazin-7a— 1 -4 5—difluoro hen 1 -5—methox razine—2—carboxamide OwNl F NH YNHg F Synthesized from utyl [(4aS,SS,7aS)—7a— (5-amino—2,3-difluor0phenyl)~5~ t1‘ifluoromethy1—4a,5 ,7,7a—tetrahydro—4H—furof3 ,4-d] hiazin—2-y1]carbamate and 5- methoxypyrazine-Z-cm‘boxylic acid according to the general procedure. 1H NMR (400 MHz, CDC13 + MeOD) 8 ppm 2.83 (dd, J=13.9, 3.8 Hz, 1 H), 3.14 (dd, , 3.0 Hz, 1 H), 3.29 — 3.39 (m, 1 H), 3.87 ((1, 128.3 Hz, 1 H), 4.04 (s, 3 H), 4.60 (d, J28.3 Hz, 1 H), 4.67 (quin, 126.3 Hz, 1 H), 7.11 ~ 7.21 (m, 1 H), 8.03 (ddd, J211.6, 6.9, 2.7 Hz, 1 H), 8.15 ((1, 121.3 Hz, 1H), 8.96 (d, J21.3 Hz, 1 H) Exam 1e 17: N— 3- 4218 SS 7218 amino—5- trifluorometh 1 -4a.5 7.7a-tetrah dro—4H- furo[3,4—dU 1.31thiazin—7a—yl)—4,5~difluoropheny1)methylpyrazinecarboxamide ~76— 2012/050833 Synthesized from tert—butyl [(4aS,SS,7aS)-7a- (5-amino—2,3—difluoropheny1)~5— oromethyl—4a,5,7,7a~tetrahydro—4H—furo[3 ,4—d] [l ,3]thia7.in-2~yl]carbamate and 5— methylpyrazine—2—carboxy1ic acid according to the general procedure. 1H NMR (400 MHZ, CDC13 + MeOD) 8 ppm 2.68 (s, 3 H), 2.84 (dd, J=13.6, 3.8 Hz, 1 H), 3.15 (dd, J=13.9, 3.0 Hz, 1 H), 3.30 - 3.42 (In, 1 H), 3.88 (d, J=10.4 Hz, 1 H), 4.61 (d, J=8.6 Hz, 1H), 4.68 (quin, J=7.2 Hz, 1 H), 7.13 ~ 7.25 (m, 1 H), 8.05 (ddd, J=11.6, 6.8, 2.8 Hz, 1 H), 8.46 (s, 1 H), 9.30 (d, J=1.3 Hz, 1 H) Exam 1e 18: N— 3- 4aS.SS.7aS amino—5- trifluorometh 1 —4a.5.7.7a—tetrah dro-4H- carboxamide Synthesized from tert—butyl [(4aS,SS,7aS)—7a- (5—amino—2,3-difluorophenyl)—5~ romethyl—4a,5,7,7a~tetrahydro—4H~furo[3,4-d][1,3]thiazin—2-y1]carbamate and 5» (fluoromethyl)pyrazine—2-carb0xylic acid according to the general procedure. 1H NMR (400 MHZ, CDC13) 8 ppm 2.89 (dd, J=13.6, 3.8 Hz, 1 H), 3.19 (dd, J=13.6, 3.0 Hz, 1 H), 3.43 (dd, J=7.5, 3.4 Hz, 1 H), 3.86 — 3.99 (m, 1 H), 4.39 — 4.67 (m, 3 H), 4.74 (quin, .1271 Hz. 1 H), 5.76 (d, J=45.5 Hz. 2 H), 8.10 (ddd, J=11.4, 6.8, 2.8 Hz, 1 H), 8.77 (s, 1 H), 9.46 (s, 1 H), 9.69 (s, 1 H) Alternate Preparations of the compounds of Examples 1 and 8 are described herein below. For these alternate prepartions 1H NMR and 13C NMR spectra were recorded on a Varian 400 MHz or 500 MHz instrument with VNMR 6.1C software. 2012/050833 Alternative Pre aration of N— 3— 4aS SS 7aS —2-amino—5— trifluorometh 1 -4a.5 7.7a— tetrah dr0—4H—furo 3.4-d 1.3 thiazin-7a- carboxamide {Example 1 l 1— 14 S nthesis of tart—But 12— 1.1.1-trifluorobut—3-en—2- lox acetate 0HC\/ OTMS aq'NaOH / odk k O ———————~—> F C/i\<¢9 CF3TMS cat. TBAF 3 l/Njg; BU4N H304 3 toluene 0 °C A on vessel was charged with toluene (3.2 L)_. THF (0.60 L) and acrolein (0.40 L, .985 mol) at It under nitrogen. (Trifluoromethyl)trimethylsi1ane (1.003 kg, 7.059 mol) was added at 17 °C. The reaction mixture was cooled to 2.5 °C and TBAF (0.01 M in THF, 0.400 L, 0.004 mol) was added over 2 h. During addition of TBAF, the temperature of the reaction mixture increased to 65 °C. The reaction mixture was cooled to 0 °C, and after 2h, tetra—n-butylammonium hydrogen sulfate (0.171 kg, 0.503 mol) was added, followed by tert-butyl cetate (0.987 kg, 5.064 mol). Sodium ide (50% wt in water, 4.2 kg, 52.6 mol) was added over 2h while maintaining the temperature under 10 0C. After 2h at 0-5 ”G, to the on mixture was added water (2.9 L) and methyl tert—butyl ether (6.0 L). The aq. phase was extracted one more time with methyl tert-butyl ether (6.0 L). The organic phases were combined and washed with 14% aq. NaCl (3 x 1.6 L). The organics were concentrated under vacuum to afford the title compound as an oil (1.150 kg, 94.5%) which was used in the subsequent stage without additional purification. 1H NMR (500 MHZ, CDC13) 5 ppm 5.86 — 5.74 (m, 1H), 5.59 (d, J = 17.5 Hz, 1H), 5.56 (d, J = 10.9 Hz, 1H), 4.37 — 4.30 (n1, 1H), 4.11 (d, J: 16.5 Hz, 1H), 4.06 (d, J: 16.4 HZ, 1H), 1.40 (s, 9H); 13C NMR (125 MHz, CDC13) 6 ppm 168.51, 128.49 ((1. J: 1.7 Hz), 123.86, 123.71 (q, J: 281.8 Hz), 82.22. 78.67 (q, J = 31.5 Hz), 66.60, 28.02. 1— 15 S nthesis ofNMethox ~N—meth 1-2— 1 1.1—t1i11uorobut—3—en2— 10x acetamide (RioJ< HCOOH (4V) OiNO //\E;3 MeONHMeHCI //\E;a CDLDCM stage 2 To a reactor containing tert—butyl 2~(1,1,1-trifluorobut—3—enyloxy)acetate (1.150 kg, 4.788 mol) was added formic acid (6.2 kg) at rt. The reaction e was heated to 55—60 °C for 4—5 h. The formic acid was evaporated under vacuum (T = 40-45 0C) and chased with toluene (2 x 3.0 L). To the e was added CH2C13 (2.0 L) and further concentrated under vacuum. To the resulting residue was added CHgClg (4.6 L) and the solution was cooled to 0 °C, followed by N,N-carbony1diimidazole g, 6.49 mol) in five portions. The mixture was stirred for 30 mins, and ND- dimethylhydroxylamine hydrochloride (0.67 kg, 6.72 mol) was added in portions while maintaining temperature below 10 °C. The reaction mixture was warmed to rt and stirred over 14 h. The reaction mixture was cooled to 3.2 °C and imidazole (100.7 g, 1.48 mol) was charged in two portions. The reaction mixture was warmed to rt and water (1.4 kg) was added, followed by methyl tert—butyl ether (14.0 L). The organic phase was washed with 2.0 N aq. HCl (1.0 L and 0.7 L), followed by sat. aq. NaHC03 (1.2 L) and sat. aq. NaCl (1.20 L). The organics were concentrated to afford the title compound as an oil (0.95kg, 87.2%). 1H NMR (500 MHz, CDCl3) 5 ppm 5.85 — 5.76 (m, 1H), 5.62 (d, J = 17.2 Hz, 1H), 5.56 (d, J = 10.4 Hz, 1H), 4.49 — 4.34 (m, 3H), 3.68 (s, 3H), 3.67 (s, 1H), 3.18 (s, 3H), 3.08 (s, 1H); 13C NMR (126 MHZ, CdCl3) 5 ppm 1699*, 1634*, 128.61, 123.87 ((1, J: 282.0 Hz), , 78.54 (q, J: 31.3 Hz), 66.12, 61.52, 60.56, 36.20, 32.24. Note: this compound is a 3:1 mixture of amide bond rotamers. $Carbony1 chemical shifts ted indirectly through 1H—13C HMBC (heteronuclear multiple-bond correlation).

HRMS Calculated for C8H13F3N03 [M+H]+228.0848; found 228.0836. 1— 16 S nthesis of 1~ 2—Fluoro hen 1 1.1.1-triflu0robut—3—en 10x ethanone W°$N°RAVK©01:3 I CF3 nBuLi, THF stage 3 To a solution 1—bromo—2-t1uorobenzene (0.967 kg, 5.527 mol) in THF (6.2 L) at —75 °C, was added n—butyllithium (2.50 M in hexane, 2.09 L, 5.22 mol) while maintaining temperature below —65 CC (ca. 100 min.) After 15 mins, a on of N—methoxy—N- methyl—2—(1,1,1~trifluorobut—3-en—2—yloxy)acetamide (0.949 kg, 4.178 mol) in THF (1.6 L) was added while ining temperature below ~65 °C (ca. 70 min). After 2.5 h at —78 0C, the reaction was quenched by on of sat. aq. NH4C1 (3.0 L) and methyl utyl ether (9.0 L). The reaction mixture was warmed to rt, the aq. phase was extracted again with methyl tert—butyl other (2.5 L). The organic phases were combined, washed with sat. aq. NaCl (2 x 0.3 L) and concentrated under vacuum to afford the title compound as an oil (1.007kg, 80.0%). 1H NMR (500 MHZ, CDCl3) 6 ppm 7.96 (td, J = 7.6, 1.8 Hz. 1H), 7.62 — 7.54 (m, 1H), 7.33 — 7.25 (m, 1H), 7.20 — 7.12 (m, 1H), 5.86 (ddd. J: 17.5, 10.4, 7.3 Hz, 1H), 5.60 (dd, J = 20.5, 13.8 Hz, 2H), 4.91 — 4.76 (m, 2H), 4.39 (dq, J = 12.8, 6.4 Hz, 1H); 13(3 NMR (125 MHz,CDC13) 8 ppm 193.55, 162.14 (d, JCF = 254.1 Hz), 135.36 (d, Jan: 9.2 Hz), 130.62 (d, JCF = 3.2 20121’050833 Hz), 128.49, 124.85 (d, JCF = 3.3 Hz), 123.89, 122.93, 122.72 ((1, JCF = 24.5 HZ), 116.50 ((1. JCF = 23.7 HZ), 78.97 (q. JCF = 31.4 Hz), 74.56 (d. .13: = 12.4 Hz).

HRMS Calculated for C12H10F4Og [M+H]+ 263.0695; found 263.0709.

NHN20H HCI, H02 0 F N F aOAc | Aro MeOH AD CF3 CF3 stage 4 To a reactor was added ylarnine hydrochloride (0.34 kg, 4.95 mol), sodium acetate (0.47 kg, 5.70 mol) and MeOH (2.68 L). To this suspension was charged a solution of 1-(2-fluorophenyl)—2—(l.1,l~trifluorobut—3—en-2—yloxy)ethanone (0.998 kg. 3.806 mol) in MeOH (1.8 L) and the reaction mixture was heated to 40—50 °C. Upon tion (ca. 2 h) the reaction mixture was cooled to rt, and filtered over Celite (0.5 wt) and rinsed with EtOAc (3.0 L). The filtrate was concentrated under vacuum and to the resulting residue was added methyl rert-butyl ether (6.3 L), water (0.94 L) and sat. aq. NaHC03 (2.5 L). The organic phase was washed once with water (1.6 L) and sat. aq. NaCl (0.1 L). The organic phase was concentrated under vacuum to afford the title compound as an oil (1.03 kg, 95.0%). 1H NMR (500 MHz, CDC13) 6 ppm 7.49 — 7.35 (m. 2H), 7.24 — 7.06 (m. 2H), 5.78 — 5.65 (m, 1H), 5.54 — 5.40 (m, 2H). 4.89 — 4.81 (m, 1H), 4.53 (d, J :: 12.6 Hz, 1H), 4.47 (d, J = 12.6 Hz, 0.5H), 4.27 — 4.18 (m, 11-l), 4.13 — 4.05 (m, 0.5H).

HRMS Calculated for C12H11F4N02 278.0804; found 278.0780.

Note: 1—(2—Fluorophenyl)-2—(1,1,1-trifluorobut~3~en~2~yloxy)ethanone oxime exists as an equilibrium of structural isomers, which accounts for the less-than-whole—number al values. 1—18 S ntheis of ‘3aR* 4S*.6aS* —6a-'2—fluoro hen 1—4— rometh l hexah drofuro 3.4-c isoxazole | . F A0 hydroqumone 23: O ‘0 CF3 xylenes 130 OC H To a solution of 1—(2—fluorophenyl)(1,1,1—trifluorobut~3~en-2—yloxy)ethanone oxime (1.085 kg, 3.328 mol) in xylenes (6.9 L) was added hydroquinone (862g, 0.8 mol) at rt.

The solution was heated to 128 "C (internal temperature) for 18 h. The solution was cooled to rt, and hexanes (7.0 L) was added, followed by 4.0 M aq. HCl (2.4 L). The —80— PCTIEP2012/050833 reaction mixture was stirred for l h, and filtered. To the solid was added water (2.0 L), methyl utyl ether (7.0 L) and 25% wt. aq. NaOH (0.4 L). The aq. layer was extracted once with methyl tert-butyl ether (7.0 L), the organics were combined, washed with 27% aq. NaCl (2.0 L) and trated under vacuum to a black oil (512.0g, 56%). 1H NMR (500 MHz,CDC13)8 ppm 7.64 - 7.52 (m, 1H), 7.39 — 7.31 (m, 1H), 7.19 (td, J = 7.7, 1,2 Hz, 1H), 7.11 (ddd, J: 11.9, 8.2, 1.0 Hz, 1H), 4.54 (d. J: 10.1 Hz, 1H), 4.34 —4.23 (m, 1H), 4.26 — 4.17 (m, 1H). 4.16 (d, J = 10.2 Hz, 1H), 4.10 (d, J = 8.5 Hz, 1H), 3.71 (d, J = 20.2 Hz, 1H); 13C NMR (125 MHz, CDC13) 8 ppm 160.59 ((1, JCF = 247.0 Hz), 130.50 (d, JCF = 8.7 Hz), , 124.69 (d, JCF = 3.3 Hz), 124.45 (q, JCF =: 281.8 Hz), 124.43 ((1, JG}: = 11.9 Hz), 116.66 (d, JCF = 22.7 Hz), 83.70 (q, JCF = 32.1 Hz), 78.17 ((1, JCF = 3.1 Hz), 77.63. 54.53.

HRMS Calculated for C13H11F4N02 [M+H]+ 278.0804; found 278.0802. 1-(192 Synthesis of) 28*.3R* 48* (trifluoromethyl )tetrahydrofuran~3~yl )methanol F F a Zn, AcOH 0 ‘o THF, water 0 F3C H F3C OH racemic Zinc (389.2 g, 5.95 mol) was placed in a on vessel, and water was added (893 mL).

Acetic acid (135 mL, 2.38 mol) was added while maintaining the temperature below °C. After 15 min, 6a-(2—fluoropheny1)(trifluoromethyl)hexahydrofuro[3,4— c]isoxazole (550.0 g, 1.98 mol) was added as a on in THF (665 mL). The reaction mixture was stirred over 16 h at rt. Methylene chloride (1.89 L) was added. followed by 28% aq. NH4OH (552 mL) while the temperature was kept below 30 °C.

The mixture was stirred for 30 min, and then filtered over Celite (80 g) g with methylene chloride (378 mL). The aq. layer was extracted with methylene chloride (1.89 L). The organics were combined, washed with sat. aq. NaCl (1.0 L) and concentrated under vacuum to afford an oil (502 g, 90.6%). The crude residue was used in the following step without additional purification.

HRMS Calculated for C12H13F4N02 [M+H]+ 280.0961; found 280.0972. 1120) Synthesis of (125,312,413 2Amino-4—g 2-fluorophcnyl )-2~ (trifluoromethyl )tetrahydrofuranyl )methanol {25.35 )—2.3 -bis( benzoyloxy zsuccinate O 932 ' OH F HO D-DBTA NHz 63.7. o EtOH water 0 'IIII ..,,| F03 F30 OH To a solution of 4—amino—4—(2—fluorophenyl)-2—(trifluoromethyl)tetrahydrofuran—3— yl)methanol (0.502 kg, 1.798 mol) in ethanol (4.865 L) was added dibenzoyl-D-tartaric acid (0.642 kg, 1.798 mol). The resulting suspension was heated to 67 0C. Water (94.0 mL, 5.2 mol) was added over 15 min while maintaining temperature >66 0C.

The resulting solution was cooled to to 45 °C while precipitation occurred. The slurry was reheated to 60 0C, and then cooled to ambient temperature at 5 °C/hour. The slurry was ed, and the solid was rinsed with premixed and cooled solution of ethanol (950 mL) and water (20 mL). The solid was dried until constant weight under vacuum (370 g, 97.6% 99). 1H NMR (500 MHZ, CD3OD) 6 ppm 8.13 (d, J = 7.2 Hz, 4H), 7.66 4 7.58 (m, 3H), 7.54 — 7.45 (m, 5H), 7.36 ~ 7.20 (m, 2H), 5.92 (s, 2H), 4.79 - 4.66 (m, 1H), 4.40 — 4.28 (m. 1H), 4.04 (dd. J = 12.1. 3.4 Hz, 1H), 3.92 (dd. J = 12.1. .4 Hz, 1H), 3.30 — 3.24 (m, 1H); 13C NMR (125 MHZ, DMSO) 5 ppm 169.61, 165.81, 160.23 ((1, J: 246.1 Hz), 133.00, 131.34 ((1, J: 9.1 Hz), 129.65, 129.55, 128.08, 127.97 ((1, J: 3.5 Hz), 124.95 (d, J = 3.3 Hz), 116.56 (d, J = 23.5 Hz), 77.48 (q, JCF = 31.0 Hz), 76.33, 73.20, 65.61 (d, J = 3.1 Hz), 57.11.

HRMS ated for C12H13F4N02 [M+H]+ 280.0961; found 2800967 (for amino alcohol).

The absolute stereochemistry of the title nd was assigned by comparison with a sample prepared starting from oenriched (S)—2-(t1ifluoromethyl)oxirane.

Chiral HPLC parameters: Equipment, ts, and Mobile Phase: E I ui I ment: HPLC column: Chiralcel OD, 4.6 x 250 mm, 10 um, Daicel Chemical Industries, Ltd., catalog no. 14025.

Solvent Delivery System: Agilent 1100 HPLC ternary pump, low pressure mixing with in—line de- asser, or e uivalent.

Autosampler: Agilent 1100 autosampler, 0.1 to 100 uL range, or euivalent.

Detector: Ailent 1100 variable wavelength detector or ec nt.

Chromatographic Software: Agilent ChemStation software version A.09.03 or higher for HPLC, Waters Em-ower 2 Build 2154 or - Iuivalent.

Volumetric Glassware Class A.

Volumetric 1 nette Class A. ~82— or Calibrated El pendorf ad'ustable volume, or euivalent.

Analytical balance, ca-able of wei-hin ' i 0.1 mg.

Reagents: Heotane: HPLC ; ade, Baker (catalo; no. 9177—03) or e uivalent.

IIx7 Pro nanol HPLC grade, Baker (catalo; no. 9095—03) or e uivalent.

Trleth lamme drich (catalo no. T0886) or - - uivalent. | Mobile Phase: Add 70 mL 2-propanol and 930 mL heptane (measured separately with a 100 mL and lOOO—mL ted cylinders) and 1.0 mL triethylamine (measured with volumetric glass pipette) to an appropriate flask and mix. Degas in—linc duiing use.

Diluting Solution: 2-Propanol HPLC Parameters: HPLC column: Chiralcel OD, 4.6 x 250 mm, 10 um, Daicel Chemical Industries, Ltd, catalo no. 14025.

Temerature: Howraew Gradient: _ ion Volume: Detection: Data ac-uisition time: Total run time: 30 min Column Maximum Pressure: 35 Bar Needle Wash: 2- ro n anol >“Flow rate ma be ad'usted i 0.2 ml/min to obtain s-ecified retention times.

Retention Times for es and ties: Compound Peak Retention Time (Relative Retention Time, RRT) .6 min i 10% (RRT 1.00) -83— (Relative Retention Time, RRT) 19.2 min iNH2 (RRT 0.93) F36 H (Enantiomer) A Typical Chromatogram from a Chiral HPLC Isolation of Compound l—(20) is ted in Figure 1. 'u, EtOAc, 1N NaOH cliH F30 H F3C 0“ To chiral salt ((25, 3R, 4S)—4-Amino—4—(2~fluoropheny1)—2- oromethylfietrahydrofurany1)methanol (ZS, 3S)-2,3 -bis(benzoyloxy)succinate (0.361 kg, 0.556 mol) was added BtOAc (1.08 L) and the suspension was cooled to — 3 0C. 1.0 N aq. NaOH (1.30 L) was added over 20 mins while maintaining T < 5 °C.

After 5 mins, benzoyl isothiocyanate (80.0 mL, 594 mmol) was added over 8 mins while maintaining T < 5 0C. After 1 h, EtOAc (722 mL) was charged. The aq. layer was d, and the organics were washed with sat. aq. NaHC03 (361 mL) and sat. aq.

NaCl (361 mL). The organics were filtered over celite (90 g) and rinsed with EtOAc (360 mL). The organics were concentrated under vacuum to afford a residue which was re-dissolved into CH2C12 (1.1 L) and concentrated to afford the title compound as yellow foam (261 g, 99% yield accounting for residual solvents) which was used in the following step. 1H NMR (500 MHZ, DMSO) 8 ppm 12.04 (5, 21-1), 11.20 (3, 21-1), 7.95 (d, J = 7.4 Hz, 2H), 7.69 — 7.60 (m, 1H), 7.56 — 7.42 (m, 2H), 7.37 — 7.28 (m, 1H), 7.24 — 7.12 (m, 2H). 5.59 (t, J = 4.5 Hz, 1H), 5.03 (d, J : 9.7 Hz, 1H), 4.92 (d, J z 9.7 Hz, 1H), 4.75 — 4.63 (m, 1H), 3.92 — 3.74 (m, 2H), 2.77 — 2.66 (m, 1H); 13C NMR (125 MHz, DMSO) 8 ppm 179.98, 167.85, 159.75 (d, la: 2 245.0 Hz), 133.44, , 129.88, 129.81, 129.04, 128.85, 126.31 ((1, JCF = 9.8 Hz), 124.36, 116.83 (d, JCF = 23.4 Hz), 76.11 (q, JCF = 31.0 Hz). 74.37 (d, JCF = 6.1 Hz), 68.77 (d, JCF = 3.4 Hz), 57.03, 52.23. l-lRMS Calculated for C20H13F4N303S [MM-11+ 441.0896; found 441.0818. 1— 22 S nthesis of N— 2—fluoro hen 1 —5— trifluorometh 1)-4a.5.7.‘7a- tetrahvdro-4H-furol 3 .4—d H 1,3 thiazin—2—yl zbenzarnide F H H F N H \H/ ‘Bz szo, pyridine ‘82 o o S \‘Sf CHZCIZ H H F3C OH FSC A solution of N—((3S,4R,5S)—3—(2—fluorophcnyl)—4—(hydroxymcthy1)-5—(trifluoromcthyl)— tetrahydrofuran—3—ylcarbamothioyl)benzamide (258.3 g, 583.8 mmol) in CH2C12(1.55 L) was cooled to —19.4 °C. ne (118 mL, 1.46 mol) was added while maintaining temperature at —20 OC, and then the reaction mixture was cooled to -24 °C. In another en purged vessel, CH2C12 (258 mL) was added followed by trifluoromethanesulfonic anhydride (108.0 mL, 642.2 mmol). The resulting solution was added to the reaction mixture over 30 min, while maintaining temperature < - 19.7 °C. Upon completed addition, the reaction mixture was stirred for 30 min at — °C to -15 °C, and then warmed to -11 °C over 20 min. Saturated aq. NH4C1 (646 mL) and water (390 mL) was added. The mixture was warmed to t ature and the aq. layer was removed. The organics were washed with premixed saturated aq. 1.5 NH4C1 (646 mL) and water (390 mL). The aq. layers were combined, and extracted once with CH2C12 (520 mL). The organics were combined, and concentrated under vacuum to afford a light orange foam (250 g, 100%). The residue was used in the next stage t purification. 1H NMR (500 MHZ, CDC13) 5 ppm 8.03 (d, J = 6.7 Hz, 2H), 7.52 (t, J: 7.0 Hz, 1H), 7.48 — 7.31 (m, 4H), 7.20 (t, J = 7.4 Hz. 1H), 7.12 (dd, J = 12.0, 8.4 Hz, 1H), 4.82 — 4.73 (m, 1H), 4.60 (d, J = 8.9 Hz, 1H), 4.03 (d, J = 8.3 Hz, 1H), 3.57 (d, J: 2.7 Hz, 1H), 3.20 (d, J: 13.6 Hz, 1H), 2.81 (dd, J: 13.8, 2.5 Hz, 1H); 13C NMR (125 MHz, CDC13) 8 ppm 171.50, 159.57 ((1, JCF : 247.2 Hz), 134.62, 132.49, 130.65 ((1, Jcp J: 8.8 Hz), 129.77, 128.51, 128.45, 125.14 (q, JCF = 281.8 Hz), 124.97 ((1, JCF : 3.0 Hz), 124.66 (d, JCF : 10.3 Hz), 117.05 ((1, JCF = 23.5 Hz), 66.81 ((1, JG}: 2 5.2 Hz), 38.90, 23.20.

HRMS Calculated for C20H16F4N2033 [M+H]+ 425.0947; found 425.0945. 1— 23 S nthesis of 4aS.SS.7aS -7a— 2—fluoro hen l —5-(trifluorometh l —4a.5.7.7a- tetrah -furo 3.4-d 1.3 thiazin-Z-amine F H N\ N‘Bz NH2 K2303 0 Y O s MeOH H 3 F3C —85— W0 20121098213 To a solution of N—((4aS,55,7aS)-7a-(2—fluorophenyl)—5—(tiifluoromethyl)—4a,5,7,7a- tetrahydro—4H—furo[3.4—d][1.3]thiazinyl)benzamide (250.2 g, 589.5 mmol) in methanol (1.25 L) was added K2003 (81.5 g, 590.0 mmol). The suspension was heated to 65 °C for 6 h. Upon cooling to ambient temperature, the solvent was evaporated under vacuum. To the resulting residue, was added 1.0 N aq NaOH (1.18 L) and THF (502 mL). The heterogeneous mixture was heated to 45 °C for 1 h. The mixture was cooled to ambient ature, and EtOAc (1.38 L) was added. The aqueous layer was ted with EtOAc (0.75 L). The organics were combined, washed with saturated aq. NaHCO3 (500 mL) and saturated aq. NaCl (500 mL). The cs were concentrated under vacuum to afford the title compound as a brown oil (184.1 g, 91.6 % yield accounting for residual solvents). 1H NMR (500 MHZ, DMSO) 6 ppm 7.49 — 7.42 (m, 1H), 7.40 — 7.33 (m, 1H), 7.26 — 7.15 (m, 2H), 6.26 (s, 2H), 4.77 — 4.54 (1n, 1H), 4.40 (d, J = 8.0 Hz, 1H). 3.80 (dd, J = 7.9, 2.3 Hz, 1H), 3.24 — 3.17 (m, 1H), 3.00 (dd, J = 13.9, 3.2 Hz, 1H), 2.85 (dd, J: 13.9, 3.9 Hz, 1H); 13C NMR (125 MHz, DMSO) 5 ppm 159.75 (d, JCF = 245.1 Hz), 149.51, 131.31 ((1, J3: = 3.9 Hz), 130.13 (d, JCF = 8.8 Hz), 128.08 (d, JCF = 10.4 Hz), 128.28 (q, JCF = 282.1 Hz). 124.87 ((1, JCF = 3.0 Hz), 116.80 ((1, J = 23.8 Hz). 78.77, 76.80 (q, JCF = 30.8 Hz), 66.31, 36.37, 23.27.

HRMS Calculated for C13H12F4N20S [M+H]+ 321.0685; found 321.0677. 1. HNQ3 NO2 Hgso4 F TFA N NH2 F \Y NYNHZ HCI 0 2. HCI s o F e3 To a cooled vessel containing (4aS,5$,7aS)—7a—(2—fluorophenyl)—5—(trifluoromethyl)- 4a,5,7,7a~tetrahydro—4H—furo[3,4~d][1,3]thiazin—2-arnine (184.1 g, 574.8 mmol) was added trifluoroaeetic acid (0.954 kg) in ns while the temperature was maintained below 20 °C. The mixture was cooled to 3.5 °C and sulfuric acid (146 mL, 2.73 mol) was added over 20 min while the temperature was maintained below 5 °C. Fuming nitric acid (39.8 mL, 0.948 mol) was added over 30 min, while the temperature was maintained below 10 °C. After 1.5 h at 0—10 °C, the reaction e was slowly quenched by transferring into an aq. on of NaOH (575 g. 14.4 mol) in water (4.6 L) cooled to 5 °C. The resulting suspension was stirred for l h at 21 °C. The suspension was then filtered and the solid rinsed with cold water (920 mL). The solid was dried under vacuum until constant weight, and then dissolved into ethanol (1.05 L). —86~ The solution was heated to 35 °C, and conc. HCl (55.6 mL, 0.690 mol) was added while maintaining temperature below 40 °C. The suspension was then cooled to —5 OC. held for 1 hr and filtered. The solid was rinsed with cold ethanol (420 mL) and dried until constant weight to obtain the title compound (185.0 g, 87.3%). 1H NMR (500 MHZ, DMSO) 6 ppm 11.80 (s, 2H), 8.45 — 8.36 (m, 1H), 8.31 (dd, J: 6.6, 2.5 HZ, 1H), 7.66 (dd, J: 11.1, 9.3 HZ, 1H), 4.96 — 4.72 (m, 1H), 4.58 (d, J: 10.0 HZ, 1H), 4.27 (d, J = 9.9 HZ, 1H), 3.76 — 3.66 (m, 1H), 3.39 (dd, J : 14.9, 3.6 Hz, 1H), 3.24 (dd, J: 14.3, 4.6 HZ, 1H); 13C NMR (125 MHZ, DMSO) 8 ppm , 163.33 (d, JCF = 257.8 Hz), 144.58, 127.61 ((1, JCF= 11.6 Hz), , 124.10, 119.28 ((1, JCF = 26.5 Hz), 77.38 (q. .lcp = 31.5 Hz), 75.99. 65.88 ((1, Jcp= 4.8 HZ), 40.36, 23.98.

HRMS Calculated for C13H11F4N3038 [M+H]+ 36; found 366.0523.

N02 NH2 N NH2 HCl N\ NH2 Fe,HC| 0 Y o I EtOH F30 H F30 Ethanol (0.975 L) was added to iron powder (62.5g, 1.12 mol) under nitrogen atmosphere. Concentrated HCl (9.03 mL) was added at ambient temperature and the suspension was heated to 65 °C for 1.5 h. The suspension was then cooled to 50 °C, and sat. aq. NH4C1 (299 g) were added. The temperature of the reaction mixture was allowed to reach 50 °C_, and (4215,53,7aS)—'7a~(2-fluoronitrophenyl)—5- (trifluoromethyl)—4a,5 tetrahydro—4H-furo [3 ,4—d] [1 ,3]thiaZin-2—amine hloride (75.0 g, 187.0 mol) was added in portions while maintaining temperature below 68 °C. After 30 min, ethanol (0. 45 L) was added, and the reaction mixture was cooled to 20—25 0C over 1 h. The suspension was stirred for 2 h and filtered over Celite (75 g) rinsing with ethanol (0.972 L). The solution was concentrated under vacuum to a brown solid. Water (0.9 L) was added followed by 3.0 N NaOH (0.187 L, 560 mmol) while maintaining temperature below 35 °C. The resulting suspension was d for 1 h at 20-25 °C. The suspension was filtered, and the solid was rinsed with cold water (0.38 L). The solid was dried under vacuum at 40—45 °C over 24 h to obtain the title compound (57.7 g, 95.5 %). 1H NMR (500 MHZ, DMSO) 8 ppm 6.81 (dd. J : 12.5, 8.6 Hz, 1H), 6.62 (dd, J: 7.0, 2.9 Hz, 1H), 6.50 — 6.42 (m, 1H), 6.16 (5, 21-1). 4.96 (5, 21-1), 4.72 — 4.54 (m, 1H), 4.35 (d, J = 7.8 HZ, 1H), 3.74 (dd, J: 7.8, 2.5 Hz,1H), 3.18 — 3.08 (m, 1H), 3.01 (dd, J: 13.9, 3.0 HZ, 1H). 2.84 (dd, J: 13.8, 3.8 HZ, 1H); 13C NMR (125 MHZ, DMSO) 5 ppm 156.20 (d, JCF : 243.0 PCTfEP2012/050833 Hz), 148.73, 145.49, 127.86 ((1, Jcp = 11.0 Hz), 116.79 (d, JCF = 24.8 HZ), 116.10 ((1, JCF = 3.3 Hz), 114.10 (d. JCF = 8.0 Hz), 78.89, 76.5701.ch = 31.0 HZ). 66.35. 36.35. 23.11.

HRMS Calculated for C13H13F4NgOS [M+H]+ 336.0794; found 336.0789.

The title compound was subjected to an Ames test. (Salmonella typhimurium tester strains TA98, TAlOO, TA1535 and TA1537 and ichia coli tester strain WP2 uv.

Mutation Research 1975, 3], 347; Mutation Research 1976, 38, 3; Proc. Nat. Acad. Sci.

USA 1976, 73, 950; Proc. Nat. Acad. Sci. USA 1975, 72, 5135) in the absence and presence of rat liver S9. The nd was negative up to the highest dose/concentration tested (SOOOug/plate). 1~ 26 S nthesis of N— 3— 4aS.SS 7aS amino—5~ trifluorometh l 7.7a~ tetrahvdro—4H—furo 3.4-d l thiazin—7a- l fluoro hen l —5-methox carboxamide N OMe N OMe HO ‘I I N/ N \N F o o NYNHZ N\ NH2 0 o Y S 80012, s F3C H DMI F3C H A suspension of oxypyrazine—Z-carboxylic acid (26.29 g, 0.17 mol) in N,N’— dimethylimidazoline—Z—one (160 mL) was stirred at ambient temperature for 15 min, then cooled to 2.2 0C. Thionyl chloride (14.7 mL, 0.202 mol) was added while maintaining temperature under 5 °C. The resulting suspension was stirred at 0—10 °C for 2 h while it transitioned to a clear solution. In another , (4aS,SS,7aS)—7a-(5- amino—Z—fluorophenyl)(trifluor0methyl)~4a,5 ,7,7a-tetrahydro—4H—furo[3 ,4- d][1,3]thiazin-2—amine (52.0 g, 0.155 mol) was dissolved into N,N’— dimethylimidazoline-Z-one (160 mL). The ing solution was added to the solution of acyl chloride while maintaining temperature below 10 °C. The reaction e was stirred for 30 min. Water (780 mL) was charged while maintaining temperature below 30 °C. The ion mixture was stirred for 30 min, and then EtOAc (780 mL) was added. To this mixture was added, 50% aq. NaOH (84.8 g) until the pH of the aqueous layer reached 11. The aq. layer was extracted with EtOAC (260 mL). The organics were ed, washed with sat. aq. NaCl (260 mL) and water (260 mL).

The organics were filtered over Celite pad (26 g) and rinsed with EtOAc (260 mL). The organics were concentrated under vacuum to afford a solid. To the solid was added 1- propanoi (728 mL), and the suspension was heated to 75 °C until a clear solution formed. The solution was cooled to ~10 °C and held for l h. The solid was filtered, rinsed with cold l-propanol (104 mL) and dried under vacuum (35 °C) until constant -88— weight to afford the title compound (62.1 g, 84.9 %). 1H NMR (500 MHZ, DMSO) 5 ppm 10.56 (s, 2H). 8.88 (d, J: 1.2 Hz. 1H), 8.39 (d, J: 1.2 Hz. 1H). 7.95 — 7.83 (m, 2H), 7.18 (dd, J = 12.0, 8.8 Hz, 1H), 6.25 (s, 2H), 4.76 — 4.60 (m, 1H), 4.36 (d, J: 8.1 Hz, 1H), 4.01 (s, 3H), 3.88 (dd, J: 7.9, 2.3 Hz, 1H), 3.23 — 3.11(m, 2H), 2.91 (dd, J = 13.8, 3.6 Hz, 1H); 13C NMR (125 MHz, DMSO) 5 ppm 162.11, 161.93, 156.13 ((1. Jcr : 242.9 Hz), 149.38. 142.01, 138.35, 135.09, 133.98, 128.53 (d, Jcp = 11.6 Hz), 126.06 (q, JCF : 282.0 Hz), 123.32, 121.93 (d, .13: = 8.6 Hz), 116.76 ((1,ch = 25.1 Hz), 78.86 ((1, JCF = 6.9 Hz), 76.94 (q, Jcp = 30.5 Hz), 66.37, 54.75, 36.44, 23.53.

HRMS Calculated for C19H17F4N5038 [M+H]Jr 472.1066; found 472.1052.

Specific optical rotation [0.]D +1105 (c 0.519, MeOH) Specific optical rotation parameters: ent: Polarimeter: Perkin Elmer, model 341 or equivalent.

Cell: Microglass cell, 100 mm pathlength, 1.0 mL ty, Perkin—Elmer Cat. # 047.

Balance: Calibrated analytical e capable of weighing i 0.1 Water Bath: NESLAB RTE 1121 Chiller or equivalent.

Volumetric glassware: Class A.

Quartz Standard ID number 098799. or equivalent.

Polarimeter: Perkin Elmer, model 341 or equivalent.

Reagents: Methanol: HPLC grade, Baker (catalog no. 9093—03) or equivalent Instrument parameters: Lamp: Na/Hal, Perkin-Elmer Cat. # 754.

Cell: Microcell (100mm), Perkin—Elmer Cat. #B004- 1693.

Cell Path: 100 mm (1 ter) Mode: OROT Wavelength: 5 89 nm Cell Temperature: 20°C Integration time: 2 seconds Aperture: MICRO Water bath temperature: 20 i 1°C —89- 2012/050833 Alternative Pre aration of N— 3— 4aS SS 7aS ~2—amino—5— trifluorometh 1 —4a 5 7 7a— tetrahvdro—4H—furo 3.4—d 1 3 thiazin-7a— 1 fluoro hen l fluorometh l) ’ 2—carboxamide Example 8) N L1H;/N NHz \H/E j/\F 1 HO / N F o O 2 S O EtOAc, TSP E F3C H —(Fluoromethyl)pyra7.ine—2-carboxylic acid (32.6 g, 1.05 equiv) and (4aS,SS,7aS)-7a- no~2—11uorophenyl)—5~(trilluoromethyl)-4a,5,7,7a-tetrahydro—4Hluro[3,4- d][1,3]thiazin—2—amine (70.0 g, 1.0 equiv)I were charged to a reactor and ethyl acetate (EtOAe, 630 mL) was added to the mixture to give a suspension. A solution of n- e phosphonic acid ide (T3P, 146 g, 1.10 equiv, 50 wt% in EtOAc) was added at ambient temperature while controlling the internal temperature below 30 °C.

The reaction mixture was stirred at 40-45 ”C >3 hours and monitored by HPLC. The on mixture was cooled to 15-20 “C and water (140 mL) was charged. After 10- 15 minutes charged 28% ammonium hydroxide (175 mL) while controlling the temperature below 30 °C. EtOAc (245 mLO was added and the reaction mixture was stirred for 30 minutes at ambient temperature. The aqueous phase was separated and back-extracted with EtOAc (490 mL). The organic phases were combined and washed with 15% aq. NaCl (140 mL) and water (140 mL). The organic layer was filtered over Celite (1.0 Wt) and rinsed with EtOAc (140 mL). The solution was trated under vacuum to obtain a beige solid (quantitative crude yield) which was recrystallized from l—propanol to afford N-(3~((4aS,SS,7aS)—2-amino-5—(trifluoromethyl)-4a_.5,7,7atetrahydro-4H-furo [3 ,4-d] [1,3]thiazinm7a—yl)~4-fluorophenyl)—5-(fluoromethyl)pyrazine~ 2-carboxamide as a white solid (70.0 g). 1H NMR (500 MHz, DMSO) 8 10.89 (s, 1H), 9.30 (s, 1H), 8.89 (s, 1H), 7.95 (dd, J = 7.3, 2.7 Hz, 1H), 7.94 — 7.89 (m, 1H), 7.21 (dd, J = 12.0, 8.8 Hz, 1H), 6.22 (s, 2H), 5.71 (d, J = 46.3 Hz, 2H), 4.77 — 4.61 (m, 1H), 4.37 (d, J = 8.1 Hz, 1H), 3.87 (dd, J = 8.0, 2.7 Hz. 1H), 3.20 (dt. .12 7.0. 3.5 Hz. 1H). 3.15 (dd. J: 13.9, 3.1 Hz. 1H). 2.91 (dd, J: 13.8, 3.8 Hz, 1H).13C NMR (126 MHz, DMSO) 5 161.32 (s), 155.82 (d. J = 243.4 Hz), 153.71 ((1, J = 18.7 Hz), 148.77 (s), 144.71 ((1, J: 1.9 Hz), 143.30 (s), 141.01 (d, J: 5.6 Hz), 134.36 ((1, J = 2.0 Hz), 128.20 ((1, J = 12.1 Hz), 125.57 (q, J = 2830 Hz), 12312 (d, PCT/EP20121’050833 J: 3.6 Hz), 121.64 ((1, J: 8.6 Hz), 116.35 ((1, J = 25.2 Hz), 82.55 (d, J = 165.8 Hz), 78.37 (3), 76.44 (q. J = 30.6 Hz), 65.89 ((1, J = 5.3 Hz). 35.89 (3). 23.01 (s).

HRMS Calculated for C19H17F5N50-23 [M+H]+ 474.1023; found 474.1032. ic Optical Rotation: [(111320 = +102.4 1A preparation of (4aS,SS,7aS)—7a—(5-amino-2—fluorophenyl)—5-(trilluoromethyl)~ 4a,5,7,7a-tetrahydro—4H—furo[3,4~d][1,3]thiazin~2—amine is described herein above in step 1—(25) in the ative preparation of N—(3—((4aS,SS,7aS)—2~amino—5— (trifluoromethyl)—4a,5 ,7,7a—tetrahydro—4H—furo [3 ,4—d] [1,3 ] thiazin—7a—yl)—4~ fluorophenyl)—5—methoxypyrazine—2-carboxamide (Example 1).

In. vitro ar assay: Quantification of AE peptide in culture of neurons from rat fetus brain (_ l) Rat primary neuronal culture y neuronal cultures were prepared from the cerebral cortex of embryonic day 18 Wistar rats (Charles River, UK). Specifically. the embryos were aseptically removed from nt rats under ether anesthesia. The brain was isolated from the embryo and immersed in HBSS (Sigma Aldrich #H9269) containing lOmM HEPES (Gibco #15630—056). The cerebral cortex was collected from the isolated brain under a stereoscopic microscope. The cerebral cortex fragments ted were enzymatically treated in an enzyme solution containing 0.05% trypsin—EDTA solution (GIBCO, #25300) at 37°C for 20 minutes to disperse the cells. The cells were then washed twice and then gently resuspended in Neurobasal medium (Gibco #21103) supplemented with 2% B27 supplement (GIBCO #17504—044). 0.5 mM amine (GIBCO #25030), 1x N2 (GIBCO #17502-048), ml Pen/Strep (GIBCO 15140— 122) and 5% heat inactivated FCS (PAA #A15-701). The cell dispersion was filtered through a 40—pin nylon mesh (BD Falcon #352340) to remove the remaining cell mass, and thus a neuronal cell suspension was obtained. The al cell sion was diluted with the medium above and then plated in a volume of 100 uL /well at an initial cell density of 3.25 X 105 cells/ml in poly- D~lysine coated 96—well culture plate (Greiner #655940). The plated cells were cultured in the culture plate at 37°C in 5% COg—95% air for 24hrs. The total amount of the medium was replaced with ‘assay Neurobasal medium’ (as above excluding heat inactivated PCS), and then the cells were cultured for a further five days.

WO 98213 (2) Addition of compound The drug was added to the culture plate on Day 6 of culture as follows. 8 point compound serial dilutions were generated in DMSO at a concentration of x1000 that of the final assay concentration (FAC). Compound solutions were then ed by adding 999u1 of ‘Assay Neurobasal media’ (as described in above section) to lul of DMSO compound stock. The total amount of the medium was removed from each of the cell plate wells, and l40uL/well of ‘Assay Neurobasal media’ was added followed by 60ul of compound solution. . The final DMSO tration was 0.1%. (3) Sampling The cells were cultured for either 1 or 3 days after addition of the compound for ABx—40 and ABX~42 assays respectively. lSOul of sample medium was collected and used as the ELISA sample. (4) Evaluation of cell survival Cell survival was ted using an Alamar assay according to the following procedure. After collecting the sample to be used in the ELISA assay, 50rd of 20% Alamar blue solution (Invitrogen #DALI 100) in assay Neurobasal media, was added to 50u1 of remaining sample within each well. Cells were then incubated at 37°C in 5% C02~95% air for 1hr.

Measurement. of fluorescence intensity for each well was the carried out at 0nrn using a Pherastar plus plate reader (BMG labtech). Upon measurement, wells having no cells plated and containing only the medium and Alamar solution were set as background (bkg). (5) Ali ELISA Human/Rat [3 d (42) ELISA Kit Wako (#290—62601) and Human/Rat [3 d (40) ELISA Kit Wako (#294-62501) from Wako Pure al Industries, Ltd. were used for AB ELISA. AB ELISA was carried out according to the protocols 3O recommended by the manufacturers, bed in the documents accompanying the kits.

The results were shown as percentage of the control groups and ICSO values for each compound were determined using four parameter logistic fit model using the XLFITS software package (IDBS).

The compounds of the present invention have an AB42 production reducing effect.

The compound of the general formula (I) or pharmaceutically acceptable salt thereof according to the present invention has an AB42 production reducing effect.

Thus, the present invention can particularly provide a prophylactic or therapeutic agent for a neurodegenerative disease caused by AB such as Alzheimer—type dementia or Down‘s syndrome.

As measured by the above in vitro assay, nd Examples 1 to 18 showed ICSO values of less than 0.1 uM as shown in Table 5: Table 5: Examgle ICso (11M) Examgle ICso (uM) 1 0.008 11 0.010 2 0.004 12 0.006 3 0.004 13 0.044 4 0.008 14 0.002 0.012 15 0.007 6 0.006 16 0.009 7 0.008 17 0.05 1 8 0.006 18 0.015 9 0.007 0.010

Claims (18)

1. A compound of formula (I): A Y Si TN/ l X NH F (0 N NH2 0 Y or a pharmaceutically able salt thereof, wherein X is hydrogen or fluorine; A is CH or N; Y is methyl, ethyl, monofluoromethyl, difluoromethyl, trifluoromethyl, 10 difluoroethyl, methoxy, ethoxy, methoxymethyl or -C EN.

2. A compound as claimed in claim 1 wherein X is hydrogen, or a ceutically acceptable salt thereof. 15

3. A compound as claimed in claim 1 or claim 2 wherein A is N, or a pharmaceutically acceptable salt thereof.

4. A compound as claimed in any one of claims 1 to 3 n Y is methyl, monofluoromethyl, difluoromethyl, trifluoromethyl or methoxy, or a pharrnaceutically 20 acceptable salt f.

5. A compound as claimed in any one of claims 1 to 4 wherein the compound is selected from: N-(3-((4aS,5S,7aS)-2—amino(trifluoromethyl)—4a,5,7,7a-tetrahydro-4H—furo[3,4- 25 d] [ l ,3 ]thiazin—7a—yl)—4-fluorophenyl)methoxypyrazine—2—carboxamide,- N—(3-((4aS,5S,7aS)amino-5—(trifluoromethyl)—4a,5,7,7a-tetrahydro-4H-furo[3,4- d] [ l ,3]thiazin-7a-yl)fluorophenyl)cyanopicolinamide ; N—(3-((4aS,5S,7aS)amino-S-(trifluoromethyl)-4a,5,7,7a-tetrahydro-4H-furo[3,4- d] [ l ,3]thiazin—7a-yl)fluorophenyl)—5-(difluoromethyl)pyrazine-2—carb0xamide ; N-(3 —((4aS,5S,7aS)amino(trifluoromethyl)-4a,5 ,7,7a—tetrahydro-4H—furo [3 ,4- d] [ 1 ,3 ]thiazin-7a—yl)fluorophenyl)(trifluoromethy1)picolinamide ; N—(3-((4aS ,5 S,7aS)—2-amino(trifluoromethyl)-4a,5,7,7a-tetrahydro-4H-furo [3 ,4- d][1,3]thiazin-7a-y1)fluorophenyl)methy1pyrazine—2-carboxamide ; N-(3-((4aS,5S,7aS)amin0(trifluoromethy1)-4a,5,7,7a-tetrahydro-4H-fur0 [3 ,4- d] [ 1 ,3 ] thiazin—7a—yl)fluorophenyl)-5—methy1picolinamidc ; N—(3-((4aS,5S,7aS)—2—amino-5—(trifluoromethyl)-4a,5,7,7a—tetrahydro-4H—furo[3 ,4— d][1 ,3 ]thiazin-7a—y1)fluoropheny1)ethy1picolinamide ; N—(3 -((4aS,5S,7aS)amino-5—(trifluoromethyl)—4a,5,7,7a—tetrahydro-4H-furo [3 ,4— 10 d] [ l ,3 ]thiazin—7a—yl)fluorophenyl)—5-(fluoromethyl)pyrazinecarboxamide ; N—(3-((4aS,5S,7aS)~2-amino(trifluoromethy1)-4a,5,7,7a-tetrahydro—4H-fi1ro [3 ,4— d] [l ,3]thiazin-7a—yl)fluorophenyl)—5—methoxypicolinamide ; N—(3—((4aS,5S,7aS)amino—5~(trifluoromethyl)—4a,5 ,7,7a-tetrahydro-4H—furo [3 ,4— d] [l ,3]thiazin—7a-yl)—4—fluoropheny1)—5-ethoxypyrazine—Z—carb0xamide ; 15 N—(3-((4aS,5S,7aS)—2-amino(trifluoromethyl)—4a,5,7,7a—tetrahydro-4H—fi1ro [3 ,4— d] [ l ,3 ] thiazin—7a-yl)fluorophenyl)—5—(l ,1~difluoroethyl)pyrazine—2-carboxamide ; N-(3 —((4aS ,5 S,7aS)amino(trifluoromethyl)—4a,5 ,7,7a—tetrahydro-4H-furo[3 ,4- d] [ l ,3 ] thiazin—7a—yl)~4~fluorophenyl)-5 uoromethyl)pyrazinecarboxamide ; N—(3-((4aS,5S,7aS)amino(trifluoromethy1)-4a,5,7,7a-tetrahydro-4H-fur0 [3 ,4- 20 d] [1 ,3 ]thiazin-7a-y1)—4-fluoropheny1)(methoxymethyl)pyrazine-Z-carboxamide: ; N— {3 —[(4aS,5 S,7aS)—2-amino—5-(triflu0romethyl)-4a,5-dihydro-4H—furo [3 ,4 d] [ l ,3]thiazin-7a(7H)-y1] —4—fluorophenyl} [(2H3)methy1oxy]pyrazine—2— carboxamide,- N—(3-((4aS ,5S,7aS)—2-amino—5-(trifluoromethyl)-4a,5,7,7a-tetrahydro—4H—furo [3 ,4— 25 d] [l ,3 ]thiazin—7a—y1)-4,5~difluorophenyl)—5—(difluoromethyl)pyrazine-2—carboxamide ; N—(3-((4aS,SS,7aS)—2—amino—5—(trifluoromethy1)—4a,5,7,7a-tetrahydro-4H—fi1ro [3 ,4- d] [l ,3 ]thiazin—7a—yl)-4,5—difluoropheny1)—5-methoxypyrazine—2—carboxamide ; N—(3 -((4aS,5 S,7aS)—2~amino—5-(trifluoromethyl)-4a,5,7,7a—tetrahydro-4H—furo[3 ,4— d] [ 1 ,3 ]thiazin—7a-yl)—4,5-difluorophenyl)-5—methy1pyrazine—2-carboxamide ; 3O N—(3-((4aS,5S,7aS)—2-amino—5-(trifluoromethyl)—4a,5,7,7a—tetrahydro-4H—fi1ro[3 ,4- d] [l ,3 in-7a—yl)~4,5-difluorophenyl)—5-(fluoromethyl)—pyrazine—2—carboxamide; or a ceutically acceptable salt thereof.

6. A compound as claimed in claim 1 which is N—(3-((4aS,SS,7aS)amino—5- 35 oromethyl)-4a,5,7,7a—tetrahydro-4H—furo [3 ,4-d][1,3]thiazin-7a-yl) fluorophenyl)meth0xypyrazine—2-carboxamide, or a pharmaceutically acceptable salt thereof.

7. A compound as claimed in claim 1 which is (4aS,5S,7aS)-2—amino (trifluoromethyl)-4a,5,7,7a—tetrahydro-4H-furo [3 ,4-d] [l ,3]thiazin-7a—y1)—4— fluorophenyl)-5—(fluoromethyl)pyrazine—2-carboxamide, or a pharmaceutically acceptable salt thereof.

8. A compound as claimed in any one of claims 1 to 7, or a ceutically able salt thereof, for use in therapy.

9. A compound as claimed in any one of claims 1 to 7, or a pharmaceutically 10 acceptable salt thereof, for inhibiting beta-site d—B sor protein ng enzyme 1 (BACEl).

10. A nd as claimed in any one of claims 1 to 7, or a pharmaceutically acceptable salt thereof, for ng a neurodegenerative disease, wherein the 15 neurodegenerative e is Alzheimer—type dementia (AD) or Down's syndrome.

11. Use of a compound as claimed in any one of claims 1 to 7, or a pharmaceutically acceptable salt thereof, for the manufacture of a medicament for the treatment or prevention of a neurodegenerative disease, wherein the neurodegenerative disease is 20 Alzheimer-type dementia (AD) or Down's syndrome.

12. A compound as claimed in any one of claims 1 to 7, or a ceutically acceptable salt thereof, for treating type 2 diabetes. 25

13. Use of a compound as claimed in any one of claims 1 to 7, or a pharmaceutically acceptable salt thereof, for the manufacture of a medicament for the treatment or prevention of type 2 diabetes.

14. A pharmaceutical composition comprising the compound as claimed in any one 30 of claims 1 to 7, or a pharmaceutically acceptable salt thereof, as an active ingredient in association with a pharmaceutically acceptable carrier.

15. A pharmaceutical product comprising, in combination, a first active ingredient which is a compound as claimed in any one of claims 1 to 7 or a pharmaceutically 35 acceptable salt thereof, and at least one further active ingredient useful in treating a neurodegenerative disease. —96-

16. The compound according to claim 1, substantially as herein described with reference to any one of the examples.

17. The use according to claim 11 or 13, ntially as herein described with reference to any one of the examples.

18. The pharmaceutical composition according to claim 15, substantially as herein described with reference to any one of the examples.