NZ623858B2 - 5-(3-aminophenyl)-5-alkyl-5,6-dihydro-2h-[1,4]oxazin-3-amine derivatives - Google Patents

Abstract

The disclosure relates to 5-(3-aminophenyl)-5-alkyl-5,6-dihydro-2-[1,4]oxazin-3-amine derivatives (formula I) as inhibitors of beta-secretase, also known as beta-site amyloid cleaving enzyme, BACE, BACE 1, Asp2, or memapsin2. The invention is also directed to pharmaceutical compositions comprising such compounds and the use of such compounds and compositions for the prevention and treatment of disorders in which beta-secretase is involved, such as Alzheimer's disease (AD), mild cognitive impairment, senility, dementia, dementia with Lewy bodies, cerebral amyloid angiopathy, multi-infarct dementia, Down's syndrome, dementia associated with stroke, dementia associated with Parkinson's disease and dementia associated with beta-amyloid. Example compounds include: (5R,6R*)-6-(Difluoromethyl)-5-{2-fluoro-5-[(3-methoxypyridin-2-yl)amino]phenyl}-5-methyl-5,6-dihydro-2H-1,4-oxazin-3-amine (5R,6R)-6-fluoro-5-[2-fluoro-S-(pyrimidin-2-ylamino)phenyl]-5-methyl-5,6-dihydro-2H-1,4-oxazin-3-amine (5R,6R)-5-{2-fluoro-5-[(3-methoxypyridin-2-yl)amino]phenyl}-5-methyl-6-(trifluoromethyl)-5,6-dihydro-2H-1,4-oxazin-3-amine (5R,6R)-5-[2-fluoro-5-(pyrimidin-2-ylamino)phenyl]-5-methyl-6-(trifluoromethyl)-5,6-dihydro-2H-1,4-oxazin-3-amine. uch compounds and the use of such compounds and compositions for the prevention and treatment of disorders in which beta-secretase is involved, such as Alzheimer's disease (AD), mild cognitive impairment, senility, dementia, dementia with Lewy bodies, cerebral amyloid angiopathy, multi-infarct dementia, Down's syndrome, dementia associated with stroke, dementia associated with Parkinson's disease and dementia associated with beta-amyloid. Example compounds include: (5R,6R*)-6-(Difluoromethyl)-5-{2-fluoro-5-[(3-methoxypyridin-2-yl)amino]phenyl}-5-methyl-5,6-dihydro-2H-1,4-oxazin-3-amine (5R,6R)-6-fluoro-5-[2-fluoro-S-(pyrimidin-2-ylamino)phenyl]-5-methyl-5,6-dihydro-2H-1,4-oxazin-3-amine (5R,6R)-5-{2-fluoro-5-[(3-methoxypyridin-2-yl)amino]phenyl}-5-methyl-6-(trifluoromethyl)-5,6-dihydro-2H-1,4-oxazin-3-amine (5R,6R)-5-[2-fluoro-5-(pyrimidin-2-ylamino)phenyl]-5-methyl-6-(trifluoromethyl)-5,6-dihydro-2H-1,4-oxazin-3-amine.

Description

-(3-AMINOPHENYL)—5-ALKYL-5,6~DIHYDRO-2H—[l,4]OXAZINAMINE DERIVATIVES FIELD OF THE INVENTION The present invention relates to novel 5-(3—aminophenyl)-5—alkyl-5,6-dihydro- 4]oxazin—3-amine derivatives as inhibitors of beta—secretase, also known as beta- site amyloid ng enzyme, BACE, BACE], Asp2, or memapsinZ. The invention is also directed to pharmaceutical compositions comprising such compounds, to processes for preparing such compounds and compositions, and to the use of such compounds and compositions for the prevention and treatment of disorders in which beta—secretase is involved, such as Alzheimer's disease (AD), mild ive impairment, senility, dementia, dementia with Lewy , cerebral amyloid angiopathy, multi-infaret dementia, Down's syndrome, dementia associated with stroke, dementia associated with Parkinson's disease and dementia associated with beta-amyloid.

BACKGROUND OF THE INVENTION Alzheimer's Disease (AD) is a neurodegenerative disease associated with aging.

AD patients suffer from cognition deficits and memory loss as well as behavioral problems such as anxiety. Over 90% ofthose afflicted with AD have a sporadic form of the disorder while less than 10% of the cases are al or hereditary. In the United States, about 1 in 10 people at age 65 have AD while at age 85, 1 out of every two individuals are affected with AD. The average life ancy from the initial diagnosis is 7—10 years, and AD patients e extensive care either in an assisted living facility which is very costly or by family members. With the increasing number of elderly in the population, AD is a growing medical concern. Currently available therapies for AD merely treat the symptoms of the disease and include acetylcholinesterase inhibitors to improve ive properties as well as anxiolyties and antipsychotics to control the behavioral problems associated with this ailment.

The hallmark pathological features in the brain ofAD patients are neurofibillary tangles which are generated by hosphorylation of tau protein and amyloid plaques which form by aggregation of beta—amyloid 1—42 (Abeta 1—42) peptide. Abeta 1-42 forms oligomers and then fibrils, and ultimately d plaques. The oligomers and fibrils are ed to be especially neurotoxic and may cause most of the neurological damage associated with AD. Agents that t the formation of Abeta 1-42 have the potential to be e-modifying agents for the treatment of AD, Abeta 1—42 is generated from the amyloid precursor protein (APP), sed of 770 amino acids. The N-terminus of Abeta 1-42 is cleaved by beta-secretase (BACE), and then gamma-secretase cleaves the ninal end. In addition to Abeta 1-42, gamma- secretase also tes Abeta 1-40 which is the predominant cleavage t as well as Abeta 1—38 and Abeta 1-43. These Abeta forms can also aggregate to form oligomers and fibrils. Thus, inhibitors ofBACE would be expected to prevent the formation of Abeta 1-42 as well as Abeta 1—40, Abeta 1-38 and Abeta 1-43 and would be potential therapeutic agents in the treatment ofAD.

WO—2011/009943 (Novartis) discloses unsubstituted and 2~substituted oxazine derivatives and their use as BACE inhibitors for the treatment of neurological disorders. WO—2011/020806 ann—LaRoche) discloses 2,6—unsubstituted 3— amino—S—phenyl~5,6—dihydro—2H—[l,4]oxazine derivatives having BACEl and /or BACE2 inhibitory ties.

Y OF THE INVENTION The present invention is directed to 5,6—dihydro-2H—[l ,4]oxazinylamine derivatives of Formula (I) O R2 1 R3 \ H HZN N N\Ar and the tautomers and the stereoisomerie forms thereof, wherein R1 is fluoro, fluoromethyl, difluoromethyl or trifluoromethyl; R2 is hydrogen or trifluoromethyl; or R1 and R2 form a nt radical =CF2; R3 is hydrogen, C1-3alkyl, cyclopropyl, mono- or polyhalo-C1_3alkyl; R4 is hydrogen or fluoro; Ar is homoaryl or heteroaryl; wherein homoaryl is phenyl or phenyl substituted with one, two or three tuents selected from the group consisting of halo, cyano, C1_3alkyl, C1_3all Illustrative of the invention is a pharmaceutical composition comprising a pharmaceutically acceptable carrier and any ofthe compounds described above. An ration ofthe invention is a pharmaceutical composition made by mixing any of the compounds bed above and a ceutically able carrier. Illustrating the ion is a process for making a ceutical composition comprising mixing ofthe compounds described above and a pharmaceutically acceptable carrier.

Excmplifying the invention are methods of treating a disorder mediated by the beta—secretase enzyme, sing administering to a t in need thereof a therapeutically effective amount of any ofthe compounds or ceutical compositions described above.

Further exemplifying the invention are methods of inhibiting the beta-secretase enzyme, comprising administering to a subject in need f a therapeutically effective amount of any of the compounds or pharmaceutical compositions described above.

An example of the invention is a method of treating a disorder selected from the group consisting ofAlzheimer's disease, mild ive impairment, senility, dementia, dementia with Lewy bodies, cerebral amyloid angiopathy, multi-infarct dementia, Down‘s syndrome, dementia associated with stroke, dementia associated with Parkinson's e and dementia associated with beta—amyloid, preferably Alzheimer's disease, comprising administering to a subject in need thereof, a therapeutically effective amount of any of the compounds or ceutical compositions described above.

Another e of the invention is any of the compounds bed above for use in treating: (a) Alzheimer‘s Disease, (b) mild cognitive impairment, (0) senility, (d) dementia, (e) dementia with Lewy bodies, (f) Down's syndrome, (g) dementia associated with stroke, (h) dementia associated with Parkinson’s disease and (i) dementia associated with beta-amyloid, in a subject in need thereof.

DETAILED PTION OF THE INVENTION The present invention is directed to compounds of formula (I) as defined hereinbefore, and pharmaceutically acceptable salts and so lvates thereof. The compounds of formula (I) are inhibitors of the beta-secretase enzyme (also known as 2012/074349 beta-site ng , BACE, BACEl , Asp2 or memapsin 2), and are useful in the treatment of Alzheimer's disease, mild cognitive impairment, senility, dementia, dementia associated with stroke, dementia with Lewy bodies, Down‘s me, dementia associated with son‘s disease and dementia associated with beta- amyloid, preferably Alzheimer's disease, mild cognitive impairment or dementia, more preferably Alzheimer's disease.

In an embodiment of the ion, R1 is fluoro, difluoromethyl or trifluoromethyl; R2 is hydrogen or trifluoromethyl; or R1 and R2 form a divalent radical =CF2; R3 is C1_3alkyl, cyclopropyl, mono~ or polyhalo—C1.3alkyl; R4 is hydrogen or fluoro; Ar is heteroaryl; wherein heteroaryl is pyridyl or pyrimidyl, each ally substituted with one, two or three substituents selected from the group consisting of halo, cyano, C1_3alkyl, C2_3alkynyl, C1_3alkyloxy, mono— and polyhalo-Clgalkyl, mono- and polyhalo— C1-3alkyloxy, and C1.3alkyloxyC1_3alkyloxy; and the addition salts and the solvates thereof.

In another embodiment, R1 is fluoro, difluoromethyl or trifluoromethyl, and R2 is hydrogen; or R1 is fluoro and R2 is trifluoromcthyl; or R1 and R2 form a divalent radical =CF2; R3 is methyl or cyclopropyl; R4 is hydrogen or fluoro; Ar is aryl; wherein heteroaryl is pyridyl substituted with methoxy, or pyrimidyl; and the addition salts and the solvates thereof.

In another embodiment the carbon atom substituted with R3 has the R configuration.

In another embodiment the compound of formula (I) is selected from (5R,6S)cyclopropyl—6-fluoro-S- {3-[(3-methoxypyridin—2—y1)amino]phenyl} (trifluoromcthyl)—5 ydro-2H— 1 ,4-oxazin—3 -aminc; (5R,6R)fluoro {2-fluoro[(3~methoxypyridin—2-yl)amino]phenyl} hyl-5 ,6- dihydro—2H-l ,4-oxazin-3 -amine, (5R)(difluoromethylidene)-5 - {2-fluoro[(3-methoxypyridinyl)amino]phenyl } - 5—methyl-5,6~dihydro—2H— l ,4—oxazinamine; WO 83556 (5R,6R*)—6-(Difluoromethyl) {2-fluoro[(3-methoxypyridin—2-yl)amino]phenyl} -5 - methyl-5,6-dihydro-2H—1,4-oxazin-3~amine; (5R,6R)fluoro[2-fluoro(pyrimidin~2—ylamino )phenyl]methyl—5,6~dihydro- ZH-l ,4-oxazin-3—amine; )-5~ {Z-fluoro-S- [(3-methoxypyridin yl)amino]phenyl} —5 —methyl-6—(trifluoromethyl)-5 ,6-dihydro-2H— 1 ,4—oxazinamine; (5R,6R)—5-[2—fluoro—5-(pyrimidinylamino)phenyl]-5—methyl(trifluoromethyl)—5 ,6- dihydro—ZH-l ,4-0xazin-3 —amine; (5R,6R)—6—fluoro—5- ro-5{(3-methoxypyrazin—Z~yl)amino]phenyl} -5—methyl—5,6— dihydro-ZH— l ,4—oxazin-3 -amine; and (5R,6R)—5— {2—fluoro—5~[(3-methoxypyrazin-Z-yl)amino]phenyl} -5—methyl—6- (trifluoromethyl)-5,6-dihydro~2H— 1 ,4-oxazin—3 -amine.

DEFINITIONS "Halo" shall denote fluoro, chloro and bromo; "C1_3all The term "subject" as used herein, refers to an , ably a mammal, most preferably a human, who is or has been the object of treatment, observation experiment.

The term "therapeutically effective amount" as used herein, means that amount ofactive compound or pharmaceutical agent that elicits the biological or medicinal response in a tissue system, animal or human that is being sought by a researcher, veterinarian, medical doctor or other clinician, which includes alleviation of the symptoms of the disease or disorder being treated.

As used herein, the term "composition" is intended to encompass a t comprising the specified ients in the specified amounts, as well as any product which results, directly or indirectly, from combinations of the specified ingredients in the ed amounts.

Hereinbefore and hereinafter, the term "compound of formula (1)" is meant to include the addition salts, the solvates and the stereoisomers thereof.

The terms "stereoisomers" or "stereochemically isomeric forms" hereinbefore or hereinafter are used interchangeably.

The ion includes all stereoisomers of the compound of Formula (I) either as a pure stereoisomer or as a mixture of two or more stereoisomers.

Enantiomers are stereoisomers that are non-superimposable mirror images of each other. A 1:1 mixture of a pair of enantiomers is a racemate or racemic mixture.

Diastereomers (or diastereoisomers) are stereoisomers that are not enantiomers, i.e. they are not related as mirror images. If a compound contains a double bond, the substituents may be in the E or the Z configuration. If a compound contains a tituted cycloalkyl group, the substituents may be in the cis or trans configuration.

Therefore, the invention includes omers, diastereomers, racemates, E isomers, Z isomers, cis isomers, trans isomers and mixtures thereof.

The absolute ration is specified according to the Cahn-lngold-Prelog system. The configuration at an asymmetric atom is specified by either R or S. ed compounds whose absolute configuration is not known can be designated by (+) or (—) ing on the direction in which they rotate plane zed light.

When a specific stereoisomer is identified, this means that said stereoisomer is substantially free, i.e. ated with less than 50%, preferably less than 20%, more preferably less than 10%, even more preferably less than 5%, in particular less than 2% and most preferably less than 1%, of the other isomers. Thus, when a compound of formula (I) is for instance specified as (R), this means that the compound is substantially free of the (S) isomer; when a compound of formula (I) is for ce specified as E, this means that the compound is substantially free ofthe Z isomer; when a compound of a (I) is for instance specified as cis, this means that the compound is ntially free of the trans isomer.

The compounds of Formula (I) co—exist in a dynamic equilibrium with the tautomers of Formula (I-a).

R R1 O R2 0 R2 t 3 R R3 _a mi, .

R4 R4 (1-8) For use in ne, the salts of the nds of this invention refer to non- toxic "pharmaceutically acceptable salts". Other salts may, however, be useful in the preparation of compounds ing to this invention or of their pharmaceutically acceptable salts. Suitable ceutically acceptable salts of the compounds include acid addition salts which may, for e, be formed by mixing a solution of the nd with a solution of a pharrnaceutically acceptable acid such as hydrochloric acid, sulfuric acid, ic acid, maleic acid, succinic acid, acetic acid, benzoic acid, citric acid, tartaric acid, carbonic acid or phosphoric acid. rmore, where the compounds of the invention carry an acidic moiety, suitable pharmaceutically acceptable salts thereofmay include alkali metal salts, e.g., sodium or potassium salts; alkaline earth metal salts, e.g., calcium or magnesium salts; and salts formed with suitable c ligands, e. g., quaternary ammonium salts.

Representative acids which may be used in the preparation of pharmaceutically acceptable salts include, but are not limited to, the ing: acetic acid, 2,2- dichloroactic acid, acylated amino acids, adipic acid, alginic acid, ascorbic acid, L- aspartic acid, benzenesult‘onic acid, benzoic acid, 4- acetamidobenzoic acid, (+)- camphoric acid, camphorsulfonic acid, capric acid, caproic acid, caprylic acid, cinnamic acid, citric acid, cyclamic acid, ethane-1,2-disulfonic acid, ethanesulfonic acid, 2—hydr0xy—ethanesulfonic acid, formic acid, fumaric acid, galactaric acid, ic acid, glucoheptonic acid, D—gluconic acid, D-glucoronic acid, L—glutamic acid, beta— utaric acid, glycolic acid, hippuric acid, hydrobromic acid, hydrochloric acid, (+)—L—lactic acid, (i)-DL—lactic acid, lactobionic acid, maleic acid, (—)—L-malic acid, c acid, (i)~DL—mandelic acid, methanesulfonic acid, naphthalene—Z—sulfonic acid, naphthalene-1,5- disulfonic acid, l-hydroxy-2—naphthoic acid, nicotinic acid, nitric acid, oleic acid, orotic acid, oxalic acid, palmitic acid, pamoic acid, phosphoric acid, L- pyroglutamic acid, salicylic acid, 4-amino-salicylic acid, sebacic acid, stearic acid, succinic acid, sulfuric acid, tannic acid, (+)-L—tartaric acid, thiocyanic acid, p—toluenesulfonic acid, trifluoromethylsulfonic acid, and undecylenic acid.

Representative bases which may be used in the preparation of pharmaceutically acceptable salts include, but are not limited to, the following: ammonia, L-arginine, benethamine, benzathine, calcium hydroxide, choline, dimethylethanolamine, dicthanolaminc, dicthylaminc, 2-(dicthylamino)-cthanol, cthanolaminc, cthylcnc- diamine, N—methyl-glucamine, hydrabamine, lH—imidazole, L-lysine, magnesium hydroxide, 4-(2-hydroxyethyl)-morpho line, piperazine, potassium ide, l-(2- hydroxyethyl)-pyrrolidine, secondary amine, sodium hydroxide, anolamine, tromethamine and zinc hydroxide.

The names ofthe compounds ofthe present invention were generated according to the nomenclature rules agreed upon by the Chemical Abstracts Service (CAS) using Advanced Chemical Development, 1110., software ame product version 1001; Build 15494, 1 Dee 2006) or according to the nomenclature rules agreed upon by the International Union of Pure and Applied try (IUPAC) using ed Chemical Development, Inc., software (ACD/Name product version 10.010.14105, October 2006). In case oftautomeric forms, the name of the depicted tautomen'c form of the structure was generated. The other non—depicted tautomeric form is also included within the scope of the present invention.

Preparation of the compounds Experimental procedure 1 The final compounds according to Formula (1), can be prepared by reacting an intermediate compound of Formula (II) with a compound of a (III) according to reaction scheme (1), a reaction that is performed in a suitable reaction-inert solvent, such as, for e, isopropanol or 1,4-dioxane, in the presence of a suitable acid, such as, for example, H2804 or HCl, under thermal conditions such as, for example, heating the reaction mixture at 100 °C, for example for 16 ho urs.This conversion can also be performed in the presence of a Pd-complex st such as, for example, tris(dibenzylideneacetone)dipalladium(0) [CAS 513643] in a suitable reaction— inert solvent, such as, for example, 1,4-dioxane, ethanol or mixtures of inert solvents, in the presence of a suitable base, such as, for example, aqueous K3PO4, N32CO3 or CszC03 and a le ligand such as, for example, 1,1’—bis(dipheny1phosphino)— ferrocene [CAS 121508], under thermal conditions such as, for example, heating the reaction mixture at 160 0C under ave irradiation until completion of the on, for example 1 hour. In reaction scheme (1), all variables are defined as in Formula (I) and W is halo.

Ar- ., R2 R’ HzNj: R1 RX\ (111)1{i:RHqN X\ I RAT X1\X3 X1\ 3 2,,X (II-a) x (1) x Reaction Scheme 1 Experimental procedure 2 The final compounds ing to Formula (I-b) wherein R3 is difluoromethyl and R4 is an hydrogen, can be ed by catalytic enation of an intermediate compound of Formula (I-a) according to reaction scheme (2). Said sion may be conducted by treatment of the intermediate nd of Formula (I—a) with hydrogen in the presence of a suitable catalyst such as, for example, palladium on carbon, a suitable catalyst poison, such as, for example, thiophene, in a suitable reaction—inert solvent, such as, for example, ethyl acetate. The mixture is stirred under hydrogen atmosphere, at a suitable temperature, typically room temperature, at a suitable pressure, such as, for example, atmospheric pressure, for example for 16 hours. In on scheme (2), all variables are defined as in Formula (I). / 5F "reduction" HZN N H2NR1j::1{: I XS/Lxmw Xl\ {/XS (1-3) X' (Lb) Xll\X on Scheme 2 Experimental procedure 3 l5 The intermediate compounds of Formula (ll-a) and (ll-b) can generally be prepared following the reaction steps shown in the reaction scheme (3) below. wo 2013/083556 - 10 _ 2 R3 2 3 R1 o R4 F R’ o R4 \ R54 \ R54 6 HZN N X\ W HZN N X\ NHR I I X1\ {/X) XK ¢X3 (II-b) X- (II-a) x~ 2 R3 R2 R3 Z R3 R1io R4 D RI R 0 R4 R‘ E o R4 R54 HzNR6 R54 5 (R6-H) j; R54 3 £11 |X\ NHR (V) s E |X\ s E lX\ NO X]\ 7¢X3 Xl\ 74x3 X1\ 4X3 (IV-b) X" (IV-a) X" (IV-c) X2 I B I B I B 2 R3 R2 R3 2 3 R‘ R j: R4 R1 R o R4 R1 D o R4 R5 R5 j: R5 X w H NR6 4 6 6 0 N 2 o N X NHR (R =H) o N X4 No2 H \ I (V) H \ | H \ q I X1\ 9X3 Xl\ 9X) X1\ ¢X3 (VI-b) x2 (v1-2) X2 (VI-c) X2 o R‘ HRZ C (VII) C (VII) T C halo halo R3 3 a (VII) R R" HO R4 HO R4 HO E R4 R5 R5 <—-—6 R5 HzN X: W HZN X: NHR6 (R "H) HEN X: No2 { I ~ I X1\ 24x3 X1\ 2¢X° X1\ ¢X3 (VIII—b) X (VIII-a) X (anc) x2 Reaction Scheme 3 A: thioamide—to—amidine conversion B: amide-to-thioamide conversion ation) C: cyclization D: Buchwald-Hartwig type coupling (when W is Halo) E: nitro-to-amino reduction (when R6 is H) F: Bromo-to-amine conversion (when R6 is H) The amidine derivatives in the above reaction scheme may be conveniently prepared from the corresponding thioamide derivatives ing art-known thioamide- to-amidine conversion procedures (reaction step A). Said conversion may conveniently be conducted by treatment of the said thioamides with an ammonia source such as, for example, ammonium chloride or aqueous ammonia, in a suitable reaction-inert solvent such as, for example, water or methanol and the like, under thermal conditions such as, for example, heating the reaction mixture at 60 °C, for example for 6 hours.

Alternatively, ediate compounds of Formula (II-a) wherein R6 is hydrogen in the above reaction scheme (3) can be prepared from the corresponding intermediate compounds of Formula (II-b) via copper catalyzed type coupling procedure ion step F). Said ng may be conducted by treatment of said intermediate compounds of Formula (II-b) with sodium azide in a le reaction- inert solvent, such as, for example, DMSO, in the presence of a mixture of le bases, such as, for e, dimethylethylenediamine and NazCO3, and a copper catalyst such as, CuI, under thermal conditions such as, for example, g the reaction mixture at 1 l0 °C, until completion of the reaction, for example l hour.

The thioamide derivatives in the above reaction scheme (3) can be prepared fi‘om amide derivatives following art-known thionation procedures (reaction step B).

Said conversion may conveniently be conducted by treatment of the said amides with a thionation agent such as, for example, phosphorous pentasulfide or 2,4-bis-(4-methoxy- )—l,3-dithia—2,4—diphosphetanc 2,4—disulfidc [Lawcsson’s reagent, CAS 19172— 47—5], in a reaction inert t such as, for example, tetrahydrofuran or oxane and the like, under thermal conditions such as, for example, heating the reaction e at 50 °C, for example for 50 minutes.

The amide derivatives in the above reaction scheme (3) can be prepared from the beta-aminoalcohol derivatives ofFormula (VIII) and intermediate compounds of Formula (VII) following art-known cyclization procedures (reaction step C). Said cyclization may conveniently be conducted by treatment of the said beta-aminoalcoho ls with an intermediate nd of Formula (VII) in the presence of a base, such as potassium tert-butoxide, or a mixture of bases such as potassium tert—butoxide/NN— diisopropylethylamine a on inert solvent, such as for example tetrahydrofuran and the like, at -80 °C to 100 °C, preferably -15 °C to 25 °C for 30 minutes to 100 hours, preferably 1 hour to 24 hours.

Additionally ediate compounds of Formula (IV-a) and (VI-a) in the above reaction scheme (3) can be prepared from the corresponding intermediate compounds of Formula (IV-b) and (VI-b) following art-known Buchwald-Hartwig type coupling ures (reaction step D). Said coupling may be conducted by treatment of intermediate compounds of Formula (IV-b) and (VI-b) with an intermediate compound ofFormula (V) in a suitable reaction-inert solvent, such as, for example, ethanol or mixtures of inert solvents such as, for example, 1,2-dimethoxyethane/water/ethanol, in the presence of a suitable base, such as, for example, aqueous K3PO4 or CszC03, a Pd-complex catalyst such as, for example, [1,1 ’-bis(diphenylphosphino)ferrocene]- dichloropalladiumfll) [CAS 72287—26~4] or trans-bis(dicyclohexylamine)palladium diacetate [DAPCy, CAS 6283398] under thermal conditions such as, for example, heating the on mixture at 80 0C, for example for 20 hours or for example , heating the reaction mixture at 130 0C, for example for 10 minutes under microwave irradiation.

Additionally intermediate nds of Formula (IV-a) and (VI—a) in the above on scheme (3), wherein R6 = H, can be prepared from the corresponding intermediate nds of Formula (IV-c) and (VI—c) following own nitro-to- amino reduction procedures (reaction step E). Said reduction may conveniently be conducted following art-known catalytic hydrogenation procedures. For example, said reduction may be carried out by stirring the reactants under a hydrogen atmosphere and in the presence of an appropriate catalyst such as, for example, palladium-on—charcoal, platinum-on-charcoal, nickel and the like catalysts. Suitable solvents are, for e, water, alkanols, cg. methanol, ethanol and the like, esters, c.g. ethyl acetate and the like. In order to enhance the rate of said ion reaction it may be ageous to elevate the temperature and/or the pressure of the reaction mixture.

Undesired further hydrogenation of certain functional groups in the reactants and the reaction ts may be prevented by the addition of a catalyst poison such as, for example, thiophene and the like, to the reaction mixture.

The intermediate nds of Formula (VII), (VIII-a), (Vm—b) and (VIII-c) can generally be ed following art-known Strecker type procedures described in literature, followed by standard chemical ormations of the cyano group.

Experimental procedure 4 The intermediate compounds of Formula (IX—a), (IX-b) and (IX-c) can generally be prepared following the reaction steps shown in the reaction scheme (4) below.

W0 2013/083556 _ 13 _ HZN V —-—> o 1I \r} (v11) H \ 1I H X \Xzox 11 3 X \XZI’X X \Xz’zx IX-c (XII) (XI) ( ) 0 H G O F Rs _. 1 4 R5 0 g X\ W | 1g]: X: W X1\ 3 ¢X I 3 (X-b) x2 X\X2"X (IX-b) Reaction Scheme 4 G: ation H: chlorination I: trifluoromcthylation J: ion K: cyclization Intermediate compounds of Formula (IX-a) and (1X -b) in the above on scheme (4) can be ed from an intermediate compound of Formula (X—a) and (X -b) following art-known fluorination procedures (reaction step G). Said conversion may be conducted by treatment of the intermediate compounds of Formula (X—a) and (X -b) in the presence of a fluorinating agent such as for example diethylaminosulphur trifluoride (DAST) in a le reaction inert solvent, such as for example dichloro- methane. The reaction mixture is stirred at suitable temperature, for example 0" C for the required time to achieve completion ofthe reaction, for example 20-40 minutes.

Intermediate compound of Formula (IX—c) in the above reaction scheme (4) can be prepared from intermediate compounds ofFormula (X-a) following art—known chlorination procedures (reaction step H). Said conversion may be conducted by ent of the intermediate compound of a (X-a) with a le chlorinating agent such as, for example, thionyl chloride, in the presence of a base such as, for example, pyridine in a reaction-inert solvent, such as, for example, dichloromethane.

The reaction mixture is d at suitable temperature, for example 0 °C for the required time to e completion of the reaction, for example 30-60 minutes.

Intermediate compounds of Formula (X-a) of the above reaction scheme (4) can be prepared from intermediate compounds of Formula (XI) following art-known trifluoromethylation procedures (reaction step I). Said conversion may be conducted by treatment of the intermediate compound of Formula (X1) in the presence of utyl ammonium fluoride (TBAF), with a trifluoromethylating agent such as, for example, (trifluoromethyl)trimethyl silane, in a suitable reaction—inert solvent, such as, for example, tetrahydrofiiran. The reaction mixture is stirred at suitable temperature, for example room ature for the required time to achieve completion of the reaction, for example two hours. ediate compounds ofFormula (X—b) in the above reaction scheme (4) can be prepared from ediate compounds of Formula (X1) following art-known reduction procedures (reaction step I). Said conversion may be conducted by treatment ofthe ediate compound of Formula (XI) with a reducing agent such as, for example, diisobutylaluminium hydride, in a suitable reaction-inert solvent, such as for example tetrahydrofuran. The reaction mixture is d at suitable temperature, lly from -78° C to room temperature for the ed time to achieve completion ofthe reaction, for example two hours. lnterrnediate compounds of a (X1) in the above reaction scheme (4) can be prepared from intermediate nds of Formula (XII) following art—known two- step eyelization procedures (reaction step K). Said conversion may be conducted by first, treatment of the intermediate compounds ula (Xll) with an intermediate compound of a (Vll), such as, for example, chloroaeetylchloride in the presence of a base such as, for example, NaOH, in a suitable mixture of inert solvents such as, for example, water and l,4—dioxane or water and THF. The pH of the reaction mixture is adjusted to a suitable pH value, for e, 10-1 1, by addition of a suitable base such as, for example, NaOH. The reaction mixture is stirred at a le temperature, for example, 0 °C to 25 °C for the required time to achieve completion of the reaction, for example 1-4 hours. The obtained crude residue can subsequently be eyelised to provide the intermediate (XI) by the addition of a le base such as, for example, K2C03, C32C03, NN—diisopropylethylamine or NaHCOg, in a suitable reaction-inert solvent, such as for example, aeetonitrile or DMF, The reaction mixture is stirred under thermal conditions such as, for example, heating the reaction mixture at 25 °C to 80°C for 2-24 hours or for example, heating the reaction mixture at 140 °C for 15-30 minutes under microwave ation. This conversion can also be performed in the absence of a base in a suitable reaction-inert solvent, such as for example, aeetonitrile or DMF, at a suitable ature, typically 40 °C to 110 °C, for a period of, for example, 24-48 hours.

Experimental procedure 5 The intermediate compounds ofFormula (XIII) and (XIV) can generally be prepared from intermediate compounds ofFormula (IX-c) ing aIt~known reductive dehalogenation procedures (reaction step L). Said conversion may be conducted by treatment ofthe intermediate of Formula (IX-c) with a suitable zine reagent, such as, for example, zine dust or zinc copper couple in a suitable solvent, such as acetic acid, at a suitable temperature, typically from room temperature to 80 °C, for the required time to achieve tion ofthe reaction, for example 1—1 6 hours. This conversion affords a mixture of the intermediate compounds of Formula (XIII) and (XIV) in different ratio depending on the reaction conditions and the reactants. 0 C53‘ O 0 CF3 / 1 F Rs —»L + Rs 4 N X W X4 W O 11%} X\ Wr O O N H \ | I H I Y 3 3 1 (IX-c) XK {/X3 1 (XIII) X\X2’/X (XIV) x~ X\X2’/X Reaction Scheme 5 PHARMACEUT[CAL COMPOSITIONS The present ion also provides itions for ting or treating diseases in which inhibition ofbeta-secretase is beneficial, such as Alzheimer's disease (AD), mild cognitive impairment, senility, dementia, dementia with Lewy bodies, Down's syndrome, dementia associated with stroke, dementia associated with Parkinson's disease and dementia associated with beta-amyloid. Said compositions sing a therapeutically effective amount of a compound according to formula (I) and a pharrnaeeutieally acceptable carrier or t.

While it is le for the active ingredient to be administered alone, it is preferable to present it as a pharmaceutical ition. Accordingly, the present invention further provides a pharmaceutical composition comprising a compound according to the present invention, together with a ceutically acceptable carrier or t. The carrier or diluent must be "acceptable" in the sense ofbeing compatible with the other ingredients of the composition and not deleterious to the ents thereof.

WO 83556 The pharmaceutical compositions of this invention may be prepared by any methods well known in the art of pharmacy. A therapeutically effective amount of the particular compound, in base form or addition salt form, as the active ingredient is combined in intimate admixture with a pharmaceutically acceptable carrier, which may take a wide variety of forms depending on the form of preparation desired for administration. These pharmaceutical compositions are desirably in y dosage form suitable, preferably, for systemic administration such as oral, percutaneous or eral stration; or l administration such as via inhalation, a nose spray, eye drops or via a cream, gel, shampoo or the like. For example, in preparing the compositions in oral dosage form, any of the usual pharmaceutical media may be ed, such as, for example, water, glycols, oils, alcohols and the like in the case of oral liquid preparations such as suspensions, syrups, elixirs and solutions: or solid carriers such as starches, sugars, kaolin, lubricants, binders, disintegrating agents and the like in the case ofpowders, pills, capsules and tablets. Because of their ease in administration, tablets and es represent the most advantageous oral dosage unit form, in which case solid pharmaceutical carriers are obviously employed. For parenteral compositions, the carrier will usually comprise e water, at least in large part, though other ingredients, for example, to aid solubility, may be included.

Injectable solutions, for e, may be prepared in which the carrier comprises saline solution, glucose solution or a mixture of saline and glucose solution. Inj e suspensions may also be prepared in which case appropriate liquid carriers, suspending agents and the like may be employed. In the compositions le for percutaneous administration, the carrier optionally comprises a penetration enhancing agent and/or a suitable wettable agent, optionally combined with suitable additives of any nature in minor proportions, which additives do not cause any significant deleterious effects on the skin. Said additives may facilitate the stration to the skin and/or may be helpful for preparing the desired compositions. These compositions may be administered in various ways, e.g., as a transdermal patch, as a n or as an ointment.

It is especially advantageous to formulate the aforementioned pharmaceutical compositions in dosage unit form for ease of administration and uniformity of dosage.

Dosage unit form as used in the cation and claims herein refers to physically te units suitable as unitary dosages, each unit containing a predetermined quantity ofactive ingredient calculated to produce the desired therapeutic effect in association with the required pharmaceutical carrier. Examples of such dosage unit forms are tablets (including scored or coated tablets), capsules, pills, powder packets, wafers, _ 17 _ injeetable solutions or suspensions, teaspoonfuls, tablespoonfuls and the like, and segregated multiples thereof.

The exact dosage and frequency of administration s on the particular compound of formula (I) used, the particular condition being treated, the severity of the ion being treated, the age, weight, sex, extent of disorder and general physical condition ofthe particular patient as well as other medication the individual may be taking, as is well known to those skilled in the art. Furthermore, it is evident that said effective daily amount may be lowered or increased depending on the se of the treated subject and/0r depending on the evaluation of the physician prescribing the compounds of the instant invention. ing on the mode of administration, the pharmaceutical ition will comprise from 0.05 to 99 % by weight, preferably from 0.1 to 70 % by weight, more preferably from 0.1 to 50 % by weight of the active ingredient, and, from 1 to 99.95 % by weight, preferably from 30 to 99.9 % by weight, more preferably from 50 to 99.9 % by weight of a pharmaceutically acceptable carrier, all percentages being based on the total weight of the ition.

The present compounds can be used for systemic administration such as oral, aneous or parenteral administration; or topical administration such as Via inhalation, a nose spray, eye drops or via a cream, gel, shampoo or the like. The compounds are preferably orally administered. The exact dosage and frequency of administration depends on the particular compound according to formula (I) used, the particular condition being treated, the ty ofthe condition being treated, the age, weight, sex, extent of disorder and general physical condition of the particular patient as well as other medication the individual may be taking, as is well known to those skilled in the art. Furthermore, it is evident that said effective daily amount may be lowered or increased depending on the response of the treated subj eet and/or depending on the evaluation of the ian ibing the compounds of the instant invention.

The amount of a compound of Formula (I) that can be combined with a carrier material to produce a single dosage form will vary depending upon the disease treated, the mammalian species, and the particular mode of administration. However, as a general guide, suitable unit doses for the compounds of the t invention can, for example, preferably contain between 0.1 mg to about 1000 mg ofthe active nd.

A preferred unit dose is n 1 mg to about 500 mg. A more preferred unit dose is n 1 mg to about 300mg. Even more preferred unit dose is between 1 mg to about 100 mg. Such unit doses can be administered more than once a day, for example, 2, 3, 4, 5 or 6 times a day, but preferably 1 or 2 times per day, so that the total dosage for a 70 kg adult is in the range of 0.001 to about 15 mg per kg weight of subject per administration. A preferred dosage is 0.01 to about 1.5 mg per kg weight of subject per administration, and such therapy can extend for a number ofweeks or months, and in some cases, years. It will be understood, however, that the specific dose level for any particular patient will depend on a variety of factors including the ty of the specific compound employed; the age, body weight, general health, sex and diet of the individual being d; the time and route of administration; the rate of excretion; other drugs that have previously been administered; and the ty of the particular disease undergoing therapy, as is well understood by those of skill in the area.

A typical dosage can be one 1 mg to about 100 mg tablet or 1 mg to about 300 mg taken once a day, or, multiple times per day, or one time—release capsule or tablet taken once a day and containing a proportionally higher content of active ingredient. The time-release effect can be obtained by capsule als that dissolve at different pH values, by capsules that release slowly by c pressure, or by any other known means of controlled release.

It can be necessary to use s outside these ranges in some cases as will be apparent to those skilled in the art. Further, it is noted that the clinician or treating physician will know how and when to start, interrupt, adjust, or terminate therapy in conjunction with individual patient se.

For the compositions, methods and kits provided above, one of skill in the art will understand that preferred nds for use in each are those compounds that are noted as preferred above. Still further preferred compounds for the compositions, s and kits are those compounds provided in the non—limiting es below.

EXPERIMENTAL PART after, the term "mp." means melting point, "aq." means aqueous, "rm." means reaction mixture, "rt." means room temperature, ‘DIPEA’ means N,N— diisopropylethylamine, "DIPE" means diisopropylether, ‘THF’ means tetrahydrofuran, ‘DMF’ means dimethylformamide, ‘DCM’ means dichloromethane, "EtOH" means ethanol ‘EtOAc’ means cetate, "AcOH" means acetic acid, "iPrOH" means isopropanol, "iPrNHz" means isopropylamine, "MeCN" means acetonitrile, "MeOH" means methanol, "Pd(OAc)z" means palladium(II)diacetate, "rac" means racemic, ‘sat.’ means saturated, ‘SFC’ means supercritieal fluid chromatography, ‘SFC—MS’ means supercritieal fluid chromatography/mass spectrometry, "LC-MS" means liquid chromatography/mass spectrometry, "GCMS" means gas chromatography/mass spectrometry, "HPLC" means high-performance liquid chromatography, "RP" means ed phase, "UPLC" means ultra-performance liquid chromatography, "le means retention time (in minutes), "[M+H]+" means the protonated mass of the free base of the nd, "DAST" means diethylaminosulfur trifluoride, "DMTMM" means 4-(4,6— dimethoxy—l,3,5-triazin—2—y1)methylmorpholinium chloride, "HATU" means 0-(7- azabenzotriazol—1-yl)-N,N,N’,N’—tetramethyluronium orophosphate, "Xantphos" means. (9,9-dimethyl—9H-xanthene-4,5—diyl)bis[diphenylphosphine], "TBAT" means tetrabutyl ammonium triphenyldifluorosilicate, "TFA" means trifuoroacetic acid, "EtZO" means diethylether, "DMSO" means dimethylsulfoxide, "MeCN" means acetonitrile.

For key intermediates, as well as some final compounds, the absolute configuration of chiral centers (indicated as R and/or S) were ished Via comparison with samples ofknown configuration, or the use of analytical techniques suitable for the determination of absolute configuration, such as VCD (vibrational cicular dichroism) or X-ray crystallography. When the absolute configuration at a chiral center is unknown, it is arbitrarily designated R*, A. Pre aration of the intermediates Example A1 Pre aration of intermediate 1. [\ll/ \I/O Titanium(lV) isopropoxide (202 mL, 658 mmol) was added to a stirred mixture of ethyl 2-(3—bromo-phenyl)oxo-acetate [(CAS 62123-80—2), 80 g, 329 mmol] and (S)- yl-Z-propanesulfinamide (47.9 g, 395 mmol) in n-heptane (740 mL). The mixture was stirred at 80 °C for 4 hours. The mixture was cooled to room temperature, and water was added. The resulting mixture was filtered over a diatomaceous earth pad and rinsed with n—heptane. The organic layer was separated, dried (MgSO4), filtered and trated in vacuo. The residue was purified by flash column tography (silica gel; eluent: n—heptane/EtOAc 100/0 to 50/50). The desired ons were collected and trated in vacuo to yield intermediate 1 (91 g, 74% yield). 2012/074349 Example A2 Pre aration of intermediate 2.

HN’SsO \fO R Cyclopropylmagnesium bromide (l M, 300 mL, 300 mmol) was added dropwise to a stirred solution ofintermediate 1 (91 g, 243 mmol) in DCM (1500 mL) at -40 °C. The e was stirred at this temperature for 30 min, and then the reaction was quenched by the addition of a sat. aq. NH4Cl solution, followed by water. The mixture was extracted with DCM. The organic layer was separated, dried (MgSO4), d and the solvents evaporated in vacuo to yield intermediate 2 (100 g, 99% yield), which was used as such in the next step.

Examp.le A3 Pre aration of intermediate 3.

HN’S\\O A 1M aq. NaOH solution (750 mL, 750 mmol) was added to a on of crude intermediate 2 (100 g, 240 mmol) in MeOH (400 mL). The resulting mixture was stirred at reflux for 4 hours. The mixture was cooled to r.t., and then partitioned between water and EtOAc. The aqueous layer was separated and neutralized by the addition of a 1M aq. HCl solution (750 mL), and then extracted with DCM. The c layer was separated, dried ), filtered and the solvents evaporated in vacuo. The residue was triturated with DIPE/MeCN, and the resulting solids were filtered off and dried in vacuo to yield intermediate 3 (37 g, 41% yield). ° (589 OLD: +3759 nm, 0 0.564 w/V %, MeOH, 20 OC). The absolute configuration was determined by X—ray diffraction.

Example A4 Preparation of intermediate 4.

O Hydrochloric acid salt Intermediate 3 (37 g, 99 mmol) was stirred in 4M HCI solution in dioxane (74 mL) and 1,4-dioxane (75 mL) at room temperature for 30 min. To the resulting suspension, DIPE was added, and the precipitate was filtered off and dried in vacuo to yield intermediate 4 (28 g, 92% yield). ° (589 OLD: —68.89 mm, c 0.646 W/v %, MeOH, 20 OC) Example A5 Preparation rmediate 5.

HN—L/C' A 1M aq. NaOH solution (182.6 mL, 182.6 mmol) was added to a solution of ' intermediate 4 (28 g, 91.3 mmol), and the mixture was cooled on an ice-bath. To this mixture, a solution of chloroacetylchloride (21.8 mL, 274 m0 1) in THF (280 mL) was added dropwise at 15 °C over one hour, while simultaneously adding a solution of a % aq. NaOH solution to maintain the pH around 10-11. After completion of the reaction, cone. aq. HCl solution was added lly to the reaction mixture until pH 2.

The mixture was partly concentrated in vacuo, and the ing itate was filtered off washed with DIPE, and dried in vacuo to give intermediate 5 (26 g, 82% yield). 0 (589 01.1): —6.49 nm, 0 0.5855 w/V %, MeOH, 20 OC) Example A6 Pre aration of intermediate 6.

Br R0 HNWJ Intermediate 5 (0.7 g, 202 mmol) and NaHC03 (0.34 g, 4.04 mrnol) were ved in DMF (17 mL), and the reaction mixture was stirred at 80 0C for 2 hours. The mixture was lly concentrated under reduced pressure, cooled to r.t. and then filtered over diatomaceous earth . The filtrate was concentrated in vacuo, and the residue was purified by flash column chromatography (silica gel; eluent: n-heptane/EtOAc 100/0 to 50/50). The desired fractions were collected and concentrated in vacuo to yield intermediate 6 (0.54 g, 86% yield). ° (589 0.1) : -15.68 nm, e 0.37 w/v %, MeOH, 20 °C) Example A7 Pre aration of intermediate 7.

Br R0 To a solution of intermediate 6 (4.2 g, 13.54 mmol) in THF (55 mL) was added TBAT (0.73 g, 1.35 mmol). Then, (trifluoromethyl)trimethyl silane (4.0 mL, 27 mmol) was added dropwise, and the r.m. was stirred at room temperature for 2 hours. The mixture was quenched with s NaCl, extracted with EtOAc, the organic phase was separated, dried (MgSO4) and concentrated in vacuo.. The resulting oil was purified by column chromatography (silica gel; : DCM/EtOAc lOO/O to 0/100). The desired fractions were collected and concentrated in vacuo to yield intermediate 7 (3 g, 58% yield) as a mixture of cis and trans s, which was used as such in the next step.

Example A8 Pre aration of intermediate 8.

Br R0 Intermediate 7 (3 g, 7.9 mmol) was dissolved in DCM (20 mL) and DAST (1.16 mL, 9.5 mmol) was added dropwise at r.t. The reaction e was stirred at r.t. for 1 hour and then the rm. was concentrated under d re. The residue was partitioned between DCM and an aq. sat. NaHCO; on. The organic layer was separated and the aqueous layer was extracted with DCM. The combined organic layers were dried (MgSO4), filtered, and the solvent evaporated in vacuo. The e was purified by flash column chromatography (silica gel; eluent: n-hcptanc/EtOAc 100/0 to 0/100).

The desired fractions were collected and concentrated in vacuo to yield intermediate 8 (2 g, 66% yield) as a mixture of cis and trans isomers, which was used as such in the next step.

Example A9 Pre n of intermediate 9.

Br R0 HNm) P285 (1.16 g, 5.23 mmol) was added to a solution of intermediate 8 (2 g, 5.23 mmol) in THF (43 mL) at room temperature. The mixture was stirred at 70 0C for 3 hours.

Then the mixture was cooled to room temperature, d off and the organic solvent evaporated in. vacuo. The crude product was purified by flash column chromatography (silica gel; eluent: n-heptane/DCM 80/100 to . The desired fractions were collected and evaporated in vacuo to yield ediate 9 (1.6 g, 77% yield) as a mixture of cis and trans isomers.

Example A10 Pre aration of intermediate 10 and 11. s F R NV N? N H2 NH2 intermediate 10 intermediate 11 Intermediate 9 (4.2 g, 10.55 mmol), was added to a mixture of7N ammonia in MeOH (16 mL) and an aq. NH4OH solution (40 mL), and the reaction mixture was stirred at 140 °C for 1 hour under microwave irradiation. Then the solvent was evaporated and the residue was ved in DCM, dried (MgSO4), filtered, and the solvent evaporated in. vacuo. The residue was purified by flash column tography (silica gel; eluent: n—heptane/EtOAc 100/0 to 50/50). The desired fractions were collected and concentrated in vacuo to yield intermediate 10 (2.44 g, 61% yield) and intermediate 11 (0.7 g, 17% yield).

Example A1 1 Pre aration ofintermediate 12.

H2N o WO 83556 2012/074349 Intermediate 10 (2.44 g, 6.4 mmol) was combined with NaN; (1.04 g, 16 mmol), CuI (1.52 g, 8.0 mmol) and NazCO3 (1.357 g, 12.8 mmol) in DMSO (92 mL) and the on was degassed. After that, N,N’-dimethylethylenediamine (1.2 mL, 11.2 mmol) was added and the mixture was heated at 110 °C until completion of the reaction, about 6 hours. The reaction mixture was poured in DCM. Ammonium ide (28% in water) was added and the organic layer was separated and washed three times with ammonium hydroxide solution. Then the organic layer was dried (Mg2804), filtered and concentrated in vacuo to yield intermediate 12 (2 g, 98% yield).

Example A12 Preparation of intermediate 13.

B F Trimethylsilylcyanide (30.7 mL, 230 mmol) was added to a stirred solution of 5- bromo—Z-fluoroacetophenone (25 g, 115 mmol) and NH4C1 (18.5 g, 345 mmol) in NH;/Me0H (150 mL). The mixture was stirred at room temperature for 3 days. Then the solvent was evaporated in vacuo and the residue was taken up in EtOAc (80 mL).

The solid was filtered and the e was evaporated in vacuo to yield intermediate 13 (27.9 g, quant. yield) which was used in the next step without further purification.

Example A13 Preparation ofintermediate 14.

H2N o B F Intermediate 13 (27 g, 111 mmol) was dissolved in HCl (37% in 1120) (130 mL) and acetic acid (130 mL) and the e was refluxed for 16 hours. After cooling to room temperature, the mixture was concentrated in vacuo. Water was added and the aqueous layer was extracted with EtOAc. The aqueous layer was basified with aq. NaOH solution (25%) to pH 7. The aqueous layer was partially concentrated in vaczto. The mixture was cooled down in an ice bath and the itate was filtered off, washed with water and then Et20 and dried in vacuo to yield intermediate 14 (18 g, 62% yield) as a white solid. _ 25 _ Example A14 ation of intermediate 15.

HgN o B F A mixture of intermediate 14 (15 g, 57.2 mrnol) in a solution of 10% H2804 in methanol (330 mL) was refluxed for 48 h. The rm. was concentrated in vacuo. Water was added and the on was basified to pH 8 with sat. aq. NHC03 solution. The aqueous layer was then extracted with EtOAc. The organic layer was separated, dried (MgSO4), filtered and concentrated in vacuo to yield intermediate 15 (15 g, 95% yield). e A15 Preparation ofintermediate 16.

H2N o B F Intermediate 15 (10 g) was separated into the corresponding enantiomers by preparative SFC on (Chiralpak® Daicel AD 30 x 250 mm). Mobile phase (C02, MeOH with 0.2% iPrNHz) to yield intermediate 16 (4.2 g, 42% yield). 0 (365 OLD: —10.1 mm, c 0.762 w/V %, MeOH, 20 °C).

Example A16 Preparation of intermediate 17.

H2N o B F THF (150 mL) was added to a solution of intermediate 16 (40 g, 145 mmol) in NaOH (1 M in H20, 360 mL). The mixture was stirred for 4 hours at room ature. The mixture was concentrated in. vacuo to afford intermediate 17 (42 g) as a white solid, which was used as such in the next reaction step.

W0 83556 Example A17 Pre aration of intermediate 18.

Cl—>/~NH o R Br F To a cooled solution rmediate 17 (41.3 g, 145 mmol) in H20 (150 mL), a solution of chloroacetyl chloride (24 mL, 304.5 mmol) in 1,4-dioxane (75 mL) was added dropwise. Simultaneously, NaOH (5M in H20, 29 mL) was added to adjust the pH at 10—1 1. The organic layer was separated, and the aqueous layer extracted with R20. Then the s layer was acidified with HCl (6 M, in H2O) until pH 2. The precipitated white solid was collected by filtration, washed with H20 and dried to yield intermediate 18 (42 g, 86% yield).

Example A18 Pre aration of intermediate 19.

HNJS ediate 18 (42 g, 124 mmol) and NaHCO3 (20.8 g, 248 mmol) were dissolved in DMF (1000 mL), and the reaction mixture was stirred at 80 °C for 3 hours. The mixture was partially concentrated under reduced pressure, cooled to r.t. and then d over diatomaceous earth . The filtrate was concentrated in vacuo, and the residue was purified by flash column chromatography (silica gel; eluent: MeOH/DCM 0/100 to /95). The desired fractions were ted and concentrated in vacuo to yield intermediate 19 (36 g, 96% yield).

Example A19 Pre aration of intermediate 20.

HNJK, BrwoOH A solution of intermediate 19 (10 g, 21.5 rnmol) in THF (105 mL) was cooled to —78 °C under N2 atmosphere. Then, diisobutylaluminium hydride (43 mL, 43 mmol) was slowly added. The reaction mixture was stirred for 2 hours allowing it to slowly warm up to room temperature. The reaction mixture was cooled down to 0 °C and quenched by slow addition of aqueous 1N HCl solution. The mixture was then extracted with EtOAc, the organic layers were separated, dried (NaZSO4), d and the solvents ated in vacuo to yield intermediate 20 (6.6 g, quant. yield, mixture tereoisorners 80/20) which was used as such in the next reaction step.

Example A20 Pre aration of intermediate 21.

HNJfi BrwoF Intermediate 20 (6.3 g, 20.7 mmol) was dissolved in DCM (841r mL) and the reaction was cooled down to 0 OC. Then DAST (3 mL, 24.9 mmol) was added se. After 20 min at 0 °C the reaction mixture was quenched with aq. sat. NaHC03 solution and extracted with DCM. The combined organic layers were dried (MgSO4), filtered, and the solvent evaporated in vacuo. The crude product was suspended from DIPE, filtered off and dried under vacuum at 60°C to yield ediate 21 (4.2 g, 66% yield, mixture of diastereoisomers 80/20).

Example A21 Pre aration of intermediate 22.

HNJS BrwoF Intermediate 22 was synthesized following the same approach described in the Example A9. Starting from intermediate 21 (4.2 g, l3.7 mmol) intermediate 22 was obtained (3 g, 68% yield, mixture of diastereoisomers 60/40).

Example A22 Pre aration ofintermediate 23 and 24.

NH2 NH2 F F F F intermediate 23 intermediate 24 WO 83556 ~28- Intermediate 22 (6 g, 18.6 mrnol) was dissolved in 7N ammonia in MeOH (300 mL) and the on mixture was stirred at 60 °C for 18 hours. The solvent was evaporated and additional 7N ammonia in MeOH was added (300 mL) and the mixture was stirred at 60 °C for an additional 18 hours. Then the solvent was evaporated and the crude product purified by column chromatography (silica gel; eluent: 7 M solution of ammonia in methanol/DCM 0/100 to 10/90). The desired fractions were collected and concentrated in vacuo to yield intermediate 23 (3.7 g, 65% yield) and intermediate 24 (0.6 g, 11% yield).

Example A23 Prc aration of ediate 25. "dug?"F Intermediate 23 (1.6 g, 5.24 mmol) was combined with NaN; (0.85 g, 13 mmol), Cul (1.25 g, 6.5 mmol) and Na2C03 (1.1 g, 10.5 mmol) in DMSO (75 mL) and the reaction was degassed. After that, N,N’-dimethylethylenediamine (1 mL, 9.1 mmol) was added and the mixture was heated at 110 °C until completion of the reaction, about 4 hours.

The reaction mixture was poured in DCM. Ammonium hydroxide (28% in water) was added and the organic layer was separated and washed three times with ammonium hydroxide. Then the c layer was dried (MngO4), filtered and concentrated in vacuo. The crude product was purified by column chromatography (silica gel; eluent: 7 M solution of a in methanol/DCM 0/100 to 10/90). The desired fractions were collected and concentrated in vacuo to yield intermediate 25 (0.3 g, 24% yield). e A24 Pre aration of intermediate 26.

HNJfi or:F3 To a solution of intermediate 19 (11.6 g, 38.5 mmol) in THF (117 mL) was added TBAT (2.08 g, 3.85 mmol). Then, (trifluoromcthyl)trirncthyl silanc (12.5 mL, 84.6 mmol) was added dropwise, and the r.m. was stirred at room temperature for 20 minutes. The mixture was quenched with aqueous NaCl and ted with EtOAc. The combined organic layers were dried (MgSO4), d and concentrated in vacuo to -29_ yield intermediate 26 (14 g, 98% yield) as a mixture of cis and trans isomers, which was used as such in the next step.

Example A25 Pre aration of intermediate 27.

HNJfi Br R Intermediate 26 (14 g, 37.6 mmol) was dissolved in DCM (600 mL) and cooled to 0 °C and then l chloride (11.2 mL, 150 mmol) was added dropwisc . The reaction mixture was stirred for 30 min at 0 °C and then pyridine (18.2 mL, 225.7 mmol) was added. After 30 minutes the on was hydrolyzed with an aqueous 1N HCl solution and then extracted with DCM. The organic layers were separated, dried (MgSO4), filtered and evaporated in vacuo. The crude product was purified by flash column chromatography (silica gel; eluent: 7 M solution of ammonia in methanol/DCM 0/100 to 2/98). The desired fractions were collected and concentrated in. vacuo to yield intermediate 27 (6 g, 41% yield, mixture of diastereoisomers).

Example A26 Pre aration of intermediate 28.

F HN/LS F F ediate 27 (7 g, 17.9 mmol) and zinc copper couple (8.55 g, 66.3 mmol) were stirred in acetic acid (420 mL) at room temperature for 16 hours. The reaction mixture was filtered, washed with DCM and concentrated in vacuo. um hydroxide solution (28% in water) and DCM were added and the mixture was d at room temperature for one hour. The organic layer was separated and the s layer was extracted with DCM. The combined organic layers were dried (MgSO4), filtered and evaporated in. vacuo to yield intermediate 28 (6 g, 99% yield) as a white powder.

Example A27 Pre aration of intermediate 29.

F HN/Ufi F F P285 (5.95 g, 26.8 mmol) was added to a solution of intermediate 28 (6 g, 17.9 mmol) in THF (145 mL) at room temperature. The e was stirred at 70 °C for 90 minutes. Then the mixture was cooled to room temperature, filtered off and the organic solvent evaporated in vacuo to yield intermediate 29 (5.9 g), which was used as such in the next step.

Example A28 Pre aration of ediate 30.

FN¢R F F Intermediate 30 was synthesized following the same approach described in the Example A22. Starting from intermediate 29 (5.9 g, 16.8 mmol) intermediate 30 was obtained (4.04 g, 72% yield).

Example A29 Pre aration of intermediate 31.

F RN91 F F Intermediate 31 was synthesized following the same approach bed in the Example A23. Starting from intermediate 30 (3.6 g, 10.7 mmol) ediate 31 was obtained (1.52 g, 52% yield). - 31 _ e A30 Pre aration of intermediate 32.

HNJS Br R To a solution of intermediate 27 (3 g, 7.68 mmol) in acetic acid (136 mL), zinc (1.26 g, 19.2 mmol) was added. The reaction mixture was then stirred at 80 °C for 3 hours, after that the reaction was filtered hot and concentrated in vacuo. The residue was dissolved in DCM and washed with um hydroxide solution. The organic phase was separated, dried (MgSO4) and the solvent concentrated in vacuo.The crude product purified by column chromatography (silica gel; eluent: 7 M solution of ammonia in methanol/DCM 0/100 to 3/97). The desired fractions were collected and trated in vacuo to yield intermediate 32 (2.7 g, 99% yield).

Example A31 Pre aration of intermediate 33. 1-1ng Br R Lawesson’s reagent (6.82 g, 16.85 mmol) was added to a solution of intermediate 32 (6 g, 16.85 mmol) dissolved in THF (68 mL) at room temperature. The mixture was stirred at 60 °C for 4 hours. Then the mixture was cooled to room temperature, filtered off and the c solvent evaporated in vacuo. The crude product was purified by flash column chromatography a gel; eluent: heptanes/DCM 100/0 to . The d fi'actions were collected and evaporated in vacuo to yield intermediate 33 (6 96% yield) as a yellowish oil.

Example A32 Pre aration ofintermediate 34 and intermediate 35 NH2 NH2 Br R o F BrWOCF3 intermediate 34 intermediate 35 _32- Intermediate 33 (6 g, 16.1 mmol) was ved in 7N ammonia in MeOH (97 mL) and the reaction mixture was stirred at 80 °C for 24 hours. Then the solvent was evaporated and the crude product purified by column chromatography (silica gel; eluent: 7 M solution of ammonia in methanol/DCM 0/100 to 2/98). The desired fractions were collected and concentrated in vacuo to yield ediate 34 (3.4 g, 59% yield) and intermediate 35 (0.75 g, 13% .

Example A33 Pre n of intermediate 36.

HZN R R0 Intermediate 34 (3.4 g, 9.6 mmol) was combined with NaNs (1.56 g, 24 mmol), CuI (2.28 g, 12 mmol) and N32CO3 (2.03 g, 19.1 mmol) in DMSO (137 mL) and the reaction was degassed. After that, N,N’-dimethylethylenediamine (1.8 mL, 168 m0 1) was added and the e was heated at 110 °C until completion of the on, about 1 hour. The reaction mixture was filtered off and the filter cake was washed with EtOAc. Water and EtOAc were added to the filtrate and the mixture was acidified by addition of HCl (1M in H20). The organic layer was then separated and the aqueous layer washed with EtOAc. Then the water layer was basified with an aq. ammonia solution and extracted again with EtOAc. The combined organic layers were dried, (NaZSO4) d and concentrated in vacuo to yield intermediate 36 (2.5 g, 90% yield). an: ~94.91° (0589 mm, c 0.393 w/V %, MeOH, 20 0C) B. Preparation of the final compounds Example Bl Preparation of compound 1:_(5R.6S)cyclopropyl—6-flu0ro13-[L3—methoxypyridin— 2-yl)aminolphenylj (trifluoromethyl)-5 ,6—dihydro-2H— 1 .4—0xazin—3—amine {11;R ca, \ / 0\ Intermediate 12 (0.07 g, 0.221 mmol) was dissolved in isopropanol (5 mL) and 2- bromo—3—methoxypyridine (0.083 g, 0.441 mmol) and sulfiiric acid (0.108 g, 1.1 mmol) were added. The mixture was d for 40 hours at 80 °C. The mixture was allowed to cool down to room temperature. DCM and sat. aq. NaHC03 solution were added. The organic layers was separated, dried (MgSO4), filtered and the solvents evaporated in vacuo. The crude product was d by flash column chromatography (silica gel; eluent: 7 M solution of ammonia in methanol/DCM 0/100 to 10/90). The desired fractions were collected and concentrated in vacuo. The residue was then dissolved in DIPE and converted to the HCl salt by addition ofHCl in isopropanol. The ing solid was filtered and dried in vacuo to yield compound 1 (0.025 g, 25% yield) as hydrochloric salt. c B2 Preparation of compound 2: (5R 6R)-6—fluoro-5— {2—fluoro~5—[(3-methoxypyridin—2- ylzammino]ph2enL}_5——-_5_,methyl6——dihydro-2H—14oxazin—3——amine (30'? F Intermediate 25 (0.3 g, 1.24 mmol) was dissolved in isopropanol (15 mL) and 2- bromo—3—methoxypyridine (0.467 g, 2.49 mmol) and sulfuric acid (0.61 g, 6.22 mmol) were added. The mixture was stirred for 40 hours at 80 °C. The mixture was d to cool down to room temperature. DCM and sat. aq. NaHCOg solution were added. The c layers was separated, dried (MgSO4), filtered and the solvents evaporated in. vacuo. The crude t was purified by preparative HPLC on (RP Vydac Dena1i® C18 - 10pm, 200 g, 5 cm). Mobile phase (0.25% NH4HCO3 solution in water, CH3CN) to yield compound 2 (0.044 g, 10% yield).

Example B3 Preparation of compound 3: (51?)—6~(difluoromethy_lidene)-_5- {2-fluoro[(3-metho_xy~ pflidinyl)amino]phenyl } methyl—5:6—dihydro—2H-l fixazinfi-amine "2%/ N O i30HfiS/FN F Intermediate 31 (0.35 g, 1.29 mmol) was dissolved in isopropanol (15 mL) and 2—bromo-3—methoxypyridine (0.485 g, 2.58 mmol) and sulfuric acid (0.34 mL, 6.45 mmol) were added. The mixture was stirred for 72 hours at 80 °C. The e was allowed to cool down to room temperature. DCM and sat. aq. NaHC03 solution were added. The organic layers was ted, dried (MgSO4), filtered and the solvents evaporated in vacuo. The crude product was purified by flash column chromatography (silica gel; eluent: 7 M solution of ammonia in methanol/DCM 0/100 to 2/98). The desired fractions were collected and concentrated in vacuo. The residue was suspended from DIPE/heptanes, filtered and dried under high vacuum to yield compound 3 (0.279 g, 57% yield) as a white powder.

Example B4 Preparation of compound 4: (5R,6R*);6~(Difiuoromcthvl)-5~_{2-fluoro—5-|(3-mcthoxy— pyridin—Z—yl)amino]phenyl} —5~methyl—546—dihydro-2H—l 14—oxazin-3—amine Compound 3 (0.228 g, 0.603 mmol) was dissolved in EtOAc (4 mL) and palladium on carbon (10%) (0.064 g, 0.06 mmol) and thiophene (0.4% on in THF, 0.8 mL, 0.041 mmol) were added. The mixture was hydrogenated at rt and atmospheric pressure for 16 hours. The catalyst was filtered off and the ts evaporated in vacuo. The crude product was purified by flash column chromatography (silica gel; eluent: 7 M solution of ammonia in methanoL/DCM 0/100 to 2/98). The desired fractions were collected and concentrated in vacuo. The residue was suspended from DIPE/heptanes, filtered and dried under high vacuum to yield compound 4 (0.074 g, 32% yield). phenyl |methyl-5,6~dihydro~2H— 1A—oxazin—3—amine >/—\ H F Intermediate 25 (0.048 g, 0.199 mmol) was dissolved in 1,4-dioxane (2 mL) and 2- yrimidine (0.032 g, 0.199 mmol) and 4M HCl on in e (0.1 mL, 0.4 mmol) were added. The mixture was d for 16 hours at 100 0C. The mixture was allowed to cool down to room temperature. DCM and sat. aq. NaHCO; solution were added. The organic layers was separated, dried (MgSO4), filtered and the ts evaporated in vacuo. The crude product was purified by flash column chromatography a gel; eluent: 7 M solution of ammonia in methanol/DCM 0/100 to 5/95). The desired fractions were collected and concentrated in vacuo to yield compound 5 (0.013 g, 20% yield).

Example B6 Preparation of compound 6: QR,6R)—5-{.Lfluoro-S—fl3-methox ridin—Z- 1 amino phenyl } —5-methyl—6-(trifluoromethyl)—5 ,6-dihydro—2H—l .4-oxazinamine H2 5/ do"$05:.

\ . Intermediate 36 (0.1 g, 0.343 mmol) was dissolved in isopropanol (4 mL) and 2— bromo-3—methoxypyridine (0.129 g, 0.687 mmol) and sulfuric acid (0.09 mL, 1.72 mmol) were added. The mixture was stirred for 40 hours at 80 °C. The mixture was allowed to cool down to room temperature. DCM and sat. aq. NaHC03 solution were added. The organic layers was separated, dried ), filtered and the solvents evaporated in vacuo. The crude product was purified by flash column chromatography (silica gel; eluent: 7 M solution of ammonia in methanol/DCM 0/100 to 5/95) to yield nd 6 (0.013 g, 20% yield).

Example B7 Preparation of compound 7: (5R,6Q—5-[2-fluoro(pyrimidin—;ylaming)ghcnyl];5— methyl—64 trifluoromethyD—S16—dihydro—2H— L4-oxazin—3—amine Hz‘i/ H CF3 Intermediate 36 (0.1 g, 0.343 mmol) was ved in oxane (3.4 mL) and 2- bromopyrimidine (0.055 g, 0.343 mmol) and 4M HCl solution in dioxane (0.17 mL, 0.69 mmol) were added. The mixture was stirred for 16 hours at 100 °C. The mixture was allowed to cool down to room temperature. DCM and sat. aq. NaHCOs on were added. The organic layers was separated, dried (MgS O4), filtered and the solvents evaporated in vacuo. The crude product was purified by flash column chromatography (silica gel; eluent: 7 M solution of ammonia in methanol/DCM 0/100 to 5/95). The 2012/074349 —36- desired fractions were collected and trated. This crude was The crude product was d by preparative HPLC on (Chiralpal Diacel AS 20x250mm). Mobile phase (C02, MeOI—I with 0.2% iPrNH2) to yield compound 7 (0.036 g, 28% yield).

Example B8 Preparation of compound 8: (5RpR)—6-fluoro—5_-{_2-fluoro-5—i(3—methoxypyrazin yl )aminolphenyl} ~5-methyl—5.6-dihydro—2H~ 1 ,4-oxazin—3 ~arnine >/-\ N R0 0/ R H F NH" F \\/N Intermediate 25 (0.15 g, 0.622 mmol) was dissolved in 1,4—dioxane (6 mL). 2—Iodo—3— methoxypyrazine (0.12 g, 0.508 mmol), cesium carbonate (0.405 g, 1.244 mmol, 1,1'— phenylphosphino)ferrocene (0.052 g, 0.093 mmol) and tris(dibenzylideneacetone) dipalladiurn(0) (0.028 g, 0.031 mmol) were added. The reaction tube was sealed and the mixture was stirred at 160 °C for 1 hour under microwave irradiation. After cooling, the reaction mixture was diluted with DCM and filtered over dicalite. The filtrate was concentrated in. vacuo. The crude product was d by flash column chromatography (silica gel; eluent: 7 M solution of a in methanol/DCM 0/100 to 5/95). The desired fractions were collected and concentrated. This crude was r purified by preparative HPLC on (Chiralpal Diacel AS 20x250mm). Mobile phase (C02, MeOH with 0.2% iPrNHZ) to yield compound 8 (0.010 g, 5% yield).

Example B9 Preparation of compound 9: N o " H\(:§§ff The resulting oil was purified by flash column chromatography (silica gel; elueng : 7M solution of ammonia in methanoL’DCM 0/100 to 5/95). The desired fractions were collected and further purified by preparative SFC on (Chiralcel Diacel OD 20 x 250 mm). Mobile phase (C02, iPrOH with 0.2% iPrNHz), to yield compound 9 (0.006 g, Example B10 Pre aration of com ound 10: Br H F Intermediate 25 (0.482 g, 2 mmol) was ved in iPrOH (24 mL), then 2.3— dibromopyridine (0.948 g, 4 mmol) and sulfuric acid (0.533 mL, 10 mmol) were added.

The reaction mixture was stirred for 4 days at 80°C. The reaction was d to cool down, then DCM and a sat. aq. NaHC03 solution were then added. The phases were separated and the organic layer was dried and trated under reduced pressure. The resulting oil was purified by flash column chromatography (silica gel; elueng : 7M solution of ammonia in methanol/DCM 0/100 to 10/90). The desired fractions were collected and further d by preparative SFC (Chiralpak Daicel AS 20 microhm 500 gr). Mobile phase (C02, iPrOH with 0.2% 1PI'NH2), to yield compound 10 (0.135 g, 17%) Example B11 Preparation of compound 11 H2in N o N R \\ H§ After cooling, the reaction mixture was diluted with DCM and filtered over dicalite.

The filtrate was concentrated in vacuo. The crude product was purified by flash column chromatography a gel; eluent: 7 M solution of a in methanol/DCM 0/100 to 5/95). The d fractions were collected and concentrated. This crude was further -38— purified by ative SFC on lpal Diacel AS 20x250mm). Mobile phase (C02, MeOI—I with 0.2% iPrNHz) to yield compound 11 (0.052 g, 11% yield).

Compounds 1 to 11 in tables 1-5 list the compounds that were prepared by analogy to one ofthe above Examples. in case no salt form is indicated, the compound was obtained as a free base. ‘Ex. No.’ refers to the Example number according to which protocol the compound was synthesized. "Co. No.’ means compound number.

Table l: —stereochemistr’ C5(R);C6(S) Single diastereoisomer Pure enantiomer X19 X29 X39 X4 stereochemistry C6(R) X1=CF 2 \ \ Single diastereoisomer X2=X3=X4=CH N H Pure enantiomer.

C- R ;C R) d X1=CF >(. .) Single diasteremsomer.

X2=X3:X4=CH Pure enantiomer Cs(R);C6(R) X1=CF Single diastereoisomer X2=X3=X4=CH Pure enantiomer Cs(R);Co(R) Single diastereoisomer Pure enantiomer _ 39 _ Table 3: C5(R) Pure enantiomer stereochemistrv C5(R);C6(R*) Single diastereoisomer Pure enantiomer Table 5: Cs(R);C6(R) X2=X3=X4=CH Single diastereoisomer Pure omer Cs(R);C6(R) Single diastereoisomer Pure enantiomer C5(R);C6(R) Single diastereoisomer Pure enantiomer C. Analytical Part LCMS For (LC)MS-charaeterization of the nds of the present invention, the following s were used.

General grocedure: The LC measurement was performed using an Acquity UPLC (Waters) system comprising a binary pump, a sample organizer, a column heater (set at 55 °C), a diode- array detector (DAD) and a column as specified in the respective s below. Flow from the column was split to a MS spectrometer. The MS detector was configured with an cleetrospray ionization source. Mass spectra were acquired by scanning from 100 to 1000 in 0.18 seconds using a dwell time of 0.02 seconds. The capillary needle voltage was 3.5 kV and the source temperature was maintained at 140 °C. Nitrogen was used as the nebulizer gas. Data acquisition was performed with a Waters—Mieromass MassLynx—Openlynx data system.

[Method 1 sed phase UPLC (Ultra Performance Liquid Chromatography) was carried out on a bridged ethylsiloxane/silica hybrid (BEH) C18 column (1.7 pm, 2.1 x 50 mm; Waters Acquity) with a flow rate of 0.8 ml/min. Two mobile phases (mobile phase A: lOmM ammonium e in H20/acetonitrile 95/5; mobile phase B: aeetonitrile) were used to run a nt condition from 95 % A and 5 % B to 5 % A and 95 % B in 1.3 minutes and hold for 0.2 minutes. An injection volume of 0.5 ul was used.

Cone voltage was 10 V for positive ionization mode and 20 V for negative ionization mode.

[Method 2: Reversed phase UPLC (Ultra mance Liquid Chromatography) was carried out on a bridged ethylsiloxane/silica hybrid (BEH) C18 column (1.7 pm, 2.1 x 50 mm; Waters Aequity) with a flow rate of 0.8 ml/min. Two mobile phases (10 mM ammonium e in H20/acetonitrile 95/5; mobile phase B: acetonitrile) were used to run a gradient condition from 95 % A and 5 % B to 5 % A and 95 % B in 1.3 minutes and hold for 0.3 minutes. An injection volume of 0.5 ul was used.

Cone voltage was 10 V for positive ionization mode and 20 V for negative ionization mode.

Method 3: Reversed phase UPLC (Ultra Performance Liquid Chromatography) was carried out on a d ethylsiloxane/silica hybrid (BEH) C18 column (1.7 um, 2.1 x 50 mm; Waters Acquity) with a flow rate of 0.8 ml/min. Two mobile phases (10 mM ammonium WO 83556 acetate in HZO/acetonitrile 95/5; mobile phase B: acetonitrile) were used to run a gradient condition from 95 % A and 5 % B to 5 % A and 95 % B in 1.3 minutes and hold for 0.3 minutes. An injection volume of 0.5 ul was used.

Cone voltage was 10 V for positive ionization mode and 20 V for negative ionization mode.

Melting Points Values are either peak values or melt ranges, and are obtained with experimental ainties that are commonly associated with this analytical method.

DSC823e (indicated by DSC in Table 6) lO For a number of compounds, melting points were determined with a e (Mettler-Toledo). Melting points were measured with a temperature gradient of ° C/minute. Maximum ature was 4000C.

Table 6: Analytical data — Rt means retention time (in minutes), [M+H]+ means the l5 protonated mass of the compound, method refers to the method used for (LC)MS.

Co. Nr. Rt [M+H]+ Method Melting Point 1 1.10 425 2 n.d. 2 0.81 349 2 n.d. 3 0.90 379 2 b r 4 0.78 381 2 136.93°C 0.67 320 1 n.d. 6 0.94 399 2 n.d. 7 0.8 370 2 n.d. 8 0.82 350 3 n.d. 9 0.81 344 1 n.d. 0.90 398 2 n.d. ii 0.87 394 3 n.d. n.d. means not determined, b.1‘. means broad range l Rotations Optical rotations were measured on a Perkin-Elmer 341 polarimeter with a sodium lamp and reported as follows: [ohm (c g/100 m1, solvent).

WO 83556 Table 7: Analytical data — Optical rotation values for enantiomerically pure com . ounds 0.456 DMF 20 365 0.1765 I DMF 20 1 +9.12 l 365 0.57 l DMF 20 For a number of compounds, lH NMR spectra were recorded on a Broker DPX~360, on a Bruker DPX-400 or on a Bruker Avance 600 spectrometer with rd pulse sequences, operating at 360 MHz, 400 MHz and 600 MHZ respectively, using CHLOROFORM-d (deuterated chloroform, CDC13) or DMSO-d6 (deuterated DMSO, dimethyl—d6 sulfoxidc) as solvents. Chemical shifts (8) are reported in parts per n (ppm) relative to tetramethylsilane (TMS), which was used as internal standard.

Table 8: C0. Nr. NMR result 1H NMR (360 MHz, DMSO-ds) d ppm 1.52 (s, 3 H) 3.87 (s, 3 H) 3.90 — 4.04 (m, 1 H) 4.06 - 4.21 (m, 2 H) 5.43 _ 5.75 (m, 1 H) .77 (br. s., 2 H) 6.73 (dd, 7:77, 5.1 Hz, 1 H) 6.99 (dd, 7:117, 4 g 8.8 Hz, 1 H) 7.19 (d, J=7.0 Hz, 1 H) 7.71 (d, J=4.0 Hz, ' 1 H) 7.85 (ddd, J=8.2, 3.7, 3.5 Hz, 1 H) 7.91 (dd, 3:70, 2.6 Hz, 1 H) 8.04 (s, 1 H) ' 1H NMR (360 MHz, CDClg-d): 5 1.62 (t, J=1.8 Hz, 3 H) 3.24 - 3.85 (m, 2 H) 3.95 — 4.28 (m, 2 H) 5.93 — 6.14 (m, 1 H) 6.68 (t, u] J=4.76 Hz, 1 H) 7.02 (dd, J=11.53, 8.96 Hz, 1 H) 7.32 (dd, J=6.95, 2.93 Hz, 1 H) 7.63 — 7.74 (m, 2 H) 8.37 (d, J=4.76 Hz, 2 1H NMR (360 MHz, CDClg-d): 6 1.67 (s, 3 H) 3.90 (s, 3 H) 4.22 (s, 2 H) 4.65 (q, J=8.40 Hz, 1 H) 6.68 (dd, J=8.05, 5.12 Hz, 1 H) 6.91 3 — 7.04 (m, 2 H) 7.08 (s, 1 H) 7.65 (dd, J=6.59, 2.93 Hz, 1 H) 7.80 (dd, J=5.12, 1.10 Hz, 1 H) 8.07 - 8.17 (m, 1H) _ 43 _ Co. Nr. NMR result 1H NMR (360 MHZ, , d): 5 1.68 (s, 3 H) 4.22 (s, 4 H) 4.63 (q, J=8.05 Hz, 1 H) 6.70 (t, J=4.76 Hz, 1 H) 7.01 (dd, J=11.34, 8.78 Hz, 1 H) 7.22 (s, 1 H) 7.70 (dd, J=6.59, 2.93 Hz, 1 H) 7.89 (dt, J=8.78, 3.66 Hz, 1 H) 8.40 (d, J=4.76 Hz, 2 H) , 1H NMR (360 MHz, , cues-d): 6 1.65 (t, J=1.83 Hz, 3 H) 3.97 — 4.30 (m, 5 H) 5.95 — 6.17 (m, 1 H) 6.95 _ 7.10 (m, 2 H) 7.39 (dd, J=6.95, 2.93 Hz, 1 H) 7.46 (d, J=2.93 Hz, 1 H) 7.64 (d, J=3.29 [ Hz, 1 H) 7.86 (ddd, J=8.78, 4.03, 2.93 Hz, 1 H) 1H NMR (360 MHZ, CDClg-d): d 1.21 (d, J=6.22 HZ, 3 H) 3.98 - 4.30 (m, 2 H) 5.91 - 6.15 (m, 1 H) 6.78 (dd, J=7.68, 4.76 Hz, 1 9 H) 6.95 — 7.11 (m, 2 H) 7.31 (dd, J=6.77, 2.74 Hz, 1 H) 7.64 _ 7.72 (m, 1 H) 7.75 — 7.83 (m, 1 H) 8.33 (dd, J=5.12, 1.83 HZ, 1 1H NMR (360 MHZ, CDClg-d): d 1.66 (t, J=1.65 HZ, 3 H) 3.97 — 4.30 (m, 2 H) 5.94 — 6.16 (m, 1 H) 6.62 (dd, J=7.68, 4.76 HZ, 1 H) 6.93 (s, 1 H) 7.04 (dd, J=11.34, 8.78 Hz, 1 H) 7.32 (dd, J=6.95, 2.93 Hz, 1 H) 7.65 - 7.78 (m, 2 H) 8.10 (dd, J=4.76, 1.46 Hz, 1 H) ‘ 1H NMR (360 MHZ, CDClg—a'): d ppm 1.67 (s, 3 H) 4.14 — 4.34 (m, 4 H) 4.65 (d, J=8.42 HZ, 1 H) 6.78 (dd, J=7.50, 4.94 Hz, 1 11 H) 6.98 - 7.08 (m, 2 H) 7.71 (dd, J=6.59, 2.93 HZ, 1 H) 7.78 (dd, I J=7.68, 1.83 Hz, 1 H) 7.94 (ddd, J=8.78, 4.03, 2.93 Hz, 1 H) i 8.37 (dd, .7=5.12, 1.83 Hz, 1 H) D. Pharmacological examples The compounds ed in the t ion are inhibitors of the beta—site APP-cleaving enzyme 1 (BACEl). tion of BACE1, an aspartic protease, is believed to be relevant for treatment of Alzheimer’s Disease (AD). The production and accumulation of beta—amyloid peptides (Abeta) from the beta—amyloid precursor protein (APP) is believed to play a key role in the onset and progression of AD. Abeta is produced from the amyloid precursor protein (APP) by sequential cleavage at the N- and C-termini of the Abeta domain by beta-secretase and gamma-secretase, respectively.

Compounds of Formula (I) are expected to have their effect substantially at BACEl by virtue oftheir ability to inhibit the tic activity. The behaviour of such inhibitors tested using a biochemical Fluorescence Resonance Energy Transfer (FRET) based assay and a cellular aLisa assay in SKNBEZ cells bed below and which are suitable for the identification of such compounds, and more particularly the nds according to Formula (I), are shown in Table 9 and Table 10.

Biochemical FRET based assay This assay is a Fluorescence Resonance Energy Transfer Assay (FRET) based assay. The substrate for this assay is an APP derived 13 amino acids peptide that contains the ‘Swedish’ Lys-Met/Asn—Leu mutation ofthe amyloid precursor protein (APP) beta-secretase cleavage site. This substrate also contains two fluorophores: (7-methoxyeoumarin—4-yl) acetic acid (Mca) is a fluorescent donor with tion wavelength at 320 nm and emission at 405 nm and 2,4-Dinitrophenyl (an) is a proprietary quencher acceptor. The distance n those two groups has been selected so that upon light excitation, the donor fluorescence energy is significantly quenched by the acceptor, through resonance energy transfer. Upon cleavage by BACE], the fluorophore Mca is separated fiom the quenching group an, restoring the full fluorescence yield of the donor. The increase in fluorescence is linearly related to the rate of proteolysis.

Briefly in a 384-well format recombinant BACEl protein in a final concentration of 1 ug/ml is incubated for 120 minutes at room temperature with 10 um substrate in tion buffer (40 mM Citrate buffer pH 5.0, 0.04 % PEG, 4 % DMSO) in the e or presence of compound. Next the amount of proteolysis is directly measured by fluorescence measurement at T=0 and T=120 (excitation at 320 nm and emission at 405 nm). s are sed in RFU (Relative scence Units), as difference between T120 and TO.

A best-fit curve is fitted by a minimum sum of squares method to the plot of %Controlmin versus compound concentration. From this an TCso value (inhibitory concentration g 50% inhibition of activity) can be obtained.

LC = Median ofthe low control values = Low control: Reaction without enzyme HC = Median ofthe High control values = High Control: on with enzyme %Effect = 100-[(sample-LC) / (HC-LC) *100] %Control = e /HC)* 100 %Controlmin = (sample-LC) / (BC—LC) * 100 The following ified compounds were tested essentially as described above and exhibited the following the activity: Table 9: Biochemical FRET based Biochemical FRET based Co. Nr. assay ; Elcso i 7 5.43 l 8 6.51 9 6.65 n.t. i 11 n.t.

Cellular aLisa assay in SKNBEZ cells In two OLLisa assays the levels of Abeta total and Abeta 1—42 produced and secreted into the medium of human neuroblastoma SKNBE2 cells are quantified. The assay is based on the human neuroblastoma SKNBE2 expressing the wild type Amyloid Precursor n (hAPP695). The compounds are diluted and added to these cells, incubated for 18 hours and then measurements ofAbeta 1-42 and Abeta total are taken. Abeta total and Abeta 1-42 are measured by sandwich OLLisa. OLLisa is a sandwich assay using biotinylated dy AbN/ZS attached to streptavidin coated beads and antibody Ab4G8 or cAb42/26 conjugated acceptor beads for the detection of Abeta total and Abeta 1-42 respectively. In the presence ofAbeta total or Abeta 1—42, the beads come into close ity. The excitation of the donor beads provokes the release of singlet oxygen les that trigger a cascade ofenergy transfer in the acceptor beads, resulting in light emission. Light emission is measured after 1 hour incubation ation at 650 nm and emission at 615 nm).

A best—fit curve is fitted by a minimum sum of squares method to the plot of %Controlmin versus compound concentration. From this an ICso value (inhibitory concentration causing 50 % tion of activity) can be obtained.

LC = Median of the low control values = Low control: cells preincubated without compound, without biotinylated Ab in the OtLisa HC = Median of the High control values = High Control: cells ubated without compound _ 46 _ %Effect = 100-[(sample—LC) / (HC—LC) * 100] %Control = e /HC)* 100 %Controlmin = (sample-LC) / (HC-LC) * 100 The following exemplified nds were tested essentially as described above and exhibited the following the activity: Table 10: Cellular aLisa assay in Cellular aLisa assay in SKNBEZ cells SKNBEZ cells C0. Nr.

Abeta 42 Abetatotal 11 mt n.t. I n.t. means not tested

Claims (2)

Claims

1. A compound of Formula (I) O R2 L 3 HZN N N\Ar or a tautomer or a stereoisomeric form thereof, wherein R1 is fluoro, fluorornethyl, difluoromethyl or trifluoromethyl; R2 is hydrogen or trifluoromethyl; or 10 R1 and R2 form a divalent radical =CF2; R3 is hydrogen, C1_3alkyl, cyclopropyl, mono- or polyhalo—C1_3alkyl; R4 is hydrogen or fluoro; Ar is homoaryl or heteroaryl; wherein homoaryl is phenyl or phenyl substituted with one, two or three substituents l5 ed from the group consisting of halo, cyano, C1_3alky1, kyloxy, mono- and lo-C1_3alkyl, mono-and polyhalo-C1_3alkyloxy; heteroaryl is selected from the group consisting of l, pyrimidyl, pyrazyl, zyl, furanyl, thienyl, pyrrolyl, pyrazolyl, imidazolyl, triazolyl, tetrazolyl, thiazolyl, isothiazolyl, thiadiazolyl, oxazolyl, isoxazolyl, and oxadiazolyl, each 20 optionally substituted with one, two or three substituents selected from the group consisting of halo, cyano, C1_3alkyl, C2-3alkyny1, C1_3alkyloxy, mono— and polyhalo—C1_ galkyl, mono- and polyhalo—C1.3alkyloxy, and C1.3alkyloxyC1_3alkyloxy; or an addition salt thereof. 25

2. The compound of claim 1 wherein R1 is fluoro, difluoromethyl or trifluoromethyl; R2 is hydrogen or trifluoromethyl; or R1 and R2 form a divalent radical =CF2; R3 is C1-3alkyl, cyclopropyl, mono- or polyhalo—C1-3alkyl; 30 R4 is hydrogen or fluoro; Ar is heteroaryl; n heteroaryl is pyridyl or pyrimidyl, each optionally substituted with one, two or three substituents ed from the group consisting of halo, cyano, C1_3alkyl, _ 48