Process for preparation of chiral pyrrolotriadimenol

Abstract

The present invention relates to a novel process for the preparation of chiral pyrrolotriadimenol of formula (I) wherein X is a halogen atom, n is an integer of 1, 2 or 3, and wherein the helical bond (II) represents (III) or (IV) or a mixture of enantiomers. The chiral pyrrolotriadimenol of formula (I) is a multipurpose intermediate for the preparation of a compound having the potential for use as an active pharmaceutical ingredient in a medicament.

Description

[0001] The present invention relates to a novel process for the preparation of chiral pyrrolotriazolols of formula I,

[0002]

[0003] wherein X is a halogen atom and n is an integer of 1, 2 or 3, and the helical bond

[0004] “ ”

[0005] represents “ ” or a mixture of enantiomers.

[0006] The chiral pyrrolotriazolols of formula I are versatile intermediates for the preparation of compounds having potential to act as active pharmaceutical ingredients in medicine. For example, the chiral pyrrolotriazolols of formula I can be used as intermediates for the preparation of compounds having potential to act as gamma-secretase modulators as disclosed in International Patent Publication WO 2020 / 120521, for the preparation of compounds having potential to act as melanocortin 4 receptor antagonists as disclosed in International Patent Publication WO 2021 / 250541, or for the preparation of compounds having potential to act as ubiquitin specific processing protease 1 as disclosed in International Patent Publication WO 2021 / 247606.

[0007] The present invention further relates to novel chiral pyrrolotriazolols of formula I,

[0008]

[0009] wherein X is a halogen atom and n is an integer of 1, 2 or 3, and the helical bond

[0010] “ ”

[0011] represents “ ” or a mixture of enantiomers.

[0012] It is an object of the present invention to find a suitable process for the preparation of such versatile chiral pyrrolotriazolol compounds of formula I.

[0013] This object can be achieved with a process as outlined below.

[0014] A process for the preparation of chiral pyrrolotriazolols of formula I,

[0015] ​​

[0016] Where X is a halogen atom, n is an integer of 1, 2 or 3, and therein a helical bond

[0017] “ "

[0018] represent" "or" "Or a mixture of enantiomers,"

[0019] The method includes taking a ketone of formula V,

[0020]

[0021] Where X and n are as described above,

[0022] a) Reduction with a metal complex catalyst in the presence of a reducing agent, or

[0023] b) Reduction with oxidoreductase

[0024] Chiral alcohols of formula I are formed.

[0025] The following definitions are set forth to illustrate and define the meaning and scope of various terms used to describe the present invention.

[0026] Term "C" 1-6 "-alkyl" refers to a branched or straight monovalent saturated aliphatic hydrocarbon group with one to six carbon atoms, preferably one to four, and more preferably one to two carbon atoms. The term is further illustrated by examples of groups such as methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, sec-butyl or tert-butyl, pentyl and its isomers, or hexyl and its isomers.

[0027] Term "C" 1-6 "-alkoxy" refers to a C-type compound as defined above, which is attached to an oxygen atom. 1-6 -alkyl group.

[0028] The term "halogen" refers to fluorine, chlorine, bromine, or iodine, but specifically to chlorine and bromine.

[0029] The term "aryl" refers to an aromatic carbon ring, such as a benzene ring or a naphthalene ring, preferably a benzene ring.

[0030] The term "heteroaryl" refers to an aromatic 5- to 6-membered monocyclic or 9- to 10-membered bicyclic ring, which may include 1, 2 or 3 heteroatoms selected from nitrogen, oxygen and / or sulfur, such as pyridinyl, pyrazolyl, pyrimidinyl, benzimidazolyl, quinolinyl and isoquinolinyl.

[0031] Spiral key

[0032] “ "

[0033] represents "R" or "S" and thus indicates the chirality of the molecule, but also a mixture of enantiomers. represents "R" or "S" and thus indicates the chirality of the molecule, but also a mixture of enantiomers.

[0034] When a chiral carbon is present in a chemical structure, it is meant that all stereoisomers associated with this chiral carbon are encompassed by the structure as pure stereoisomers as well as mixtures thereof.

[0035] The process of the present application can be illustrated with the following Scheme 1

[0036] Scheme 1

[0037]

[0038] wherein X is a halogen and n is 1, 2 or 3, and the helix bond

[0039] represents "R" or "S" and thus indicates the chirality of the molecule, but also a mixture of enantiomers.

[0040] represents "R" or "S" and thus indicates the chirality of the molecule, but also a mixture of enantiomers. represents "R" or "S" and thus indicates the chirality of the molecule, but also a mixture of enantiomers.

[0041] The reduction can be accomplished by:

[0042] a) reduction with a metal complex catalyst in the presence of a reducing agent, or

[0043] b) reduction with an oxidoreductase.

[0044] The ketone of formula V can be prepared according to the following Schemes 2 and 3

[0045] Scheme 2

[0046]

[0047] The process comprises in a first step,

[0048] a) reacting a 3,5-dihalo-1 H-1,2,4-triazole of formula II,

[0049]

[0050] wherein X is a halogen atom, by

[0051] a 1 ) Michael addition using an acrylate IIIa,

[0052]

[0053] wherein R is C​​​1-4 alkyl, or by

[0054] a 2 ) alkylating with an alkyl haloformate of the formula IIIb,

[0055]

[0056] wherein R is C 1-4 alkyl, n is an integer of 1, 2 or 3, and X is halogen,

[0057] to a 3,5-dihalo 1,2,4-triazole-carboxylate of the formula IV,

[0058]

[0059] wherein R, n and X are as described above; and in a second step,

[0060] b) ring closure of the 3,5-dihalo 1,2,4-triazole-carboxylate of the formula IV with an organometallic reagent.

[0061] 3,5-dihalo-1 H-1,2,4-triazole starting compounds, preferably 3,5-dibromo-1 H-1,2,4- triazole, are commercially available.

[0062] The Michael addition using acrylate IIIa and the alkylating using an alkyl haloformate of the formula IIIb are typically carried out in the presence of a base, which can be selected from organic bases such as triethylamine, N,N-diisopropylethylamine, tributylamine, and inorganic bases such as potassium carbonate or cesium carbonate.

[0063] Triethylamine is a preferred base for the Michael addition using acrylate IIIa and potassium carbonate is a preferred base for the alkylating using an alkyl haloformate of the formula IIIb.

[0064] Suitable acrylates IIIa are C 1-4 alkyl esters, preferably methyl esters, and suitable alkyl haloformates of the formula IIIb are bromo-C 1-4 alkyl esters, preferably methyl or ethyl esters.

[0065] The reaction can be carried out in a suitable solvent selected from alcoholic solvents (e.g. methanol, ethanol, isopropanol) or polar aprotic solvents (e.g. DMSO, acetonitrile, THF, MeTHF) at a reaction temperature between 30 °C and 170 °C, preferably in MeTHF at 70 °C.

[0066] N-alkylated N-heterocycles are reported in the literature (Gmach J. et al., Synthesis, 2016, 48, 2681-2704).

[0067] 3,5-dihalogen 1,2,4-triazole-carboxylates of the formula IV,

[0068]

[0069] wherein R is C 1-4 alkyl, n is an integer of 1, 2 or 3 and X is halogen, are novel compounds and thus constitute another embodiment of the present application.

[0070] In a preferred embodiment, X is bromine.

[0071] In another preferred embodiment, R is methyl or ethyl.

[0072] Particularly preferred 3,5-dihalogen 1,2,4-triazole-carboxylates of the formula IV are those, wherein

[0073] X = bromine, R = methyl and n is 1 ;

[0074] X = bromine, R = ethyl and n is 1 ;

[0075] X = bromine, R = methyl and n is 2;

[0076] X = bromine, R = ethyl and n is 2.

[0077] In step b), the 3,5-dihalogen 1,2,4-triazole-carboxylates of the formula IV can be subjected to a ring closure reaction with organometallic reagents such as organolithium or organomagnesium compounds.

[0078] Suitable organolithium compounds are n-hexyllithium, n-butyllithium, phenyllithium or methyllithium.

[0079] Suitable organomagnesium compounds are methylmagnesiumbromide, ethylmagnesiumchloride, ethylmagnesiumbromide, isopropylmagnesiumbromide or isopropylmagnesiumchloride.

[0080] The reaction can be carried out in a suitable solvent selected from 2-methyltetrahydrofuran, tetrahydrofuran, toluene or methyl-tert-butyl-ether at a reaction temperature between 0 °C and 100 °C.

[0081] The reaction requires quenching with an acid. Suitable quenching acids are acetic acid or citric acid.

[0082] Suitable reaction techniques are batch reactor devices, plug flow reactor devices or, as preferred technique, continuous stirred tank reactor (CSTR) devices.

[0083] The annelation can be done according to the following scheme 3:

[0084] Scheme 3:

[0085]

[0086] The annelation or the expansion of the cyclic ketone can occur by using diazo compounds (e.g. diazomethane or trimethylsilyldiazomethane) in the presence of a promoter such as a Lewis acid (e.g. BF3.Et20, AlMe3).

[0087] In step d 1 ) this leads to the annelation of the bicyclic 5-membered cyclic ketone Va to the bicyclic cyclohexyl ketone Vb.

[0088] In step d 2 ) the bicyclic 6-membered cyclic ketone Vb can be annelated to the 7-membered bicyclic ketone under similar experimental conditions.

[0089] The annelation of cyclic or acyclic ketones under different conditions is reported in the literature (Candeias et al., Chem, Rev, 2016, 2937-2981).

[0090] a) reduction of the ketone of formula V with a metal complex catalyst:

[0091] Suitable metal complex catalysts for the reduction of the ketone of formula V are ruthenium or iridium complex catalysts.

[0092] They can be selected from various ruthenium or iridium catalyst complexes as outlined below and include isomers and mixtures thereof.

[0093]

[0094]

[0095]

[0096] wherein, for each individual structure and independently from each other,

[0097] R 1 are independently from each other C 1-6 -alkyl, C 4-6 -cycloalkyl, phenyl or heteroaryl, optionally substituted with one or more C 1-6 -alkyl or C 1-6 -alkoxy;

[0098] R 2 independently of one another are hydrogen, C 1-6 -alkyl, C 4-6 -cycloalkyl, phenyl or heteroaryl, which are optionally substituted by one or more C 1-6 -alkyl or C 1-6 -alkoxy, or two R 2 together form a ring bridged by -(CH2)4- units;

[0099] R 3 independently of one another are hydrogen or C 1-6 -alkyl, C 4-6 -cycloalkyl, phenyl or heteroaryl, which are optionally substituted by one or more C 1-6 -alkyl or C 1-6 -alkoxy;

[0100] R 4 independently of one another are hydrogen, C 1-6 -alkyl, C 4-6 -cycloalkyl, phenyl or heteroaryl, which are optionally substituted by one or more C 1-6 -alkyl or C 1-6 -alkoxy, or two R 4 together form a ring bridged by -O-(CH2) x -O- units;

[0101] R 5 independently of one another are hydrogen, C 1-6 -alkyl, C 4-6 -cycloalkyl, phenyl or heteroaryl, which are optionally substituted by one or more C 1-6 -alkyl or C 1-6 -alkoxy, or two adjacent R 4 together with R 5 form a ring bridged by -(CH)4- units or -O-(CH2)x-O- units;

[0102] R 6 independently of one another are hydrogen or C 1-6 -alkyl;

[0103] X is a coordinating ligand or a counter anion selected from halogen, C 1-6 -alkoxy, tetrahaloborate, tetrakis(3,5-bis(trihalo-C 1-6 -alkyl)phenyl)borate, acetylacetonate, hexahalophosphate, p-toluenesulfonate, methanesulfonate or trihalomethanesulfonate;

[0104] Y is oxygen or -CH2-;

[0105] x is 1, 2 or 3;

[0106] and when Q 1 is nitrogen and Q 2 is carbon, the dotted ring represents an aromatic ring;

[0107] and when Q 1 and Q 2 are sulfur, the dotted ring represents a cycloalkane ring.

[0108] In preferred embodiments, Formula Xh, Xk, Xm or Xn is selected.

[0109] Preferred substituents for Xh and Xk are:

[0110] R 1 are independently of each other C 1-6 -alkyl, phenyl, optionally substituted by one or more C 1-6 -alkyl or C 1-6 -alkoxy;

[0111] R 2 are independently of each other hydrogen, phenyl, optionally substituted by one or more C 1-6 -alkyl, or two R 2 together form a ring bridged by a -(CH2)4- unit;

[0112] R 3 are independently of each other hydrogen or C 1-6 -alkyl, phenyl, optionally substituted by one or more C 1-6 -alkyl;

[0113] X is a coordinating ligand or counter anion selected from halogen, tetrafluoroborate ion, tetra(3,5-bis(trifluoromethyl)phenyl)borate ion, hexafluorophosphate ion or triflate ion.

[0114] Preferred substituents for Xm and Xn are:

[0115] R 1 are independently of each other phenyl, optionally substituted by one or more C 1-6 -alkyl or C 1-6 -alkoxy;

[0116] R 6 are independently of each other hydrogen or C 1-6 -alkyl;

[0117] X is a coordinating ligand selected from halogen

[0118] and when Q 1 is nitrogen and Q 2 is carbon, the dotted line ring represents an aromatic ring.

[0119] In another preferred embodiment, the substituents for Xhand Xkare:

[0120] R 1 are independently of each other methyl, phenyl, which is optionally substituted by one or more C 1-6 -alkyl groups;

[0121] R 2 are independently of each other hydrogen, phenyl, or both R 2 together form a ring bridged by a -(CH2)4- unit;

[0122] R 3 are independently of each other hydrogen or C 1-6 -alkyl;

[0123] X is a coordinating ligand or counter anion selected from chloride or triflate.

[0124] Further preferred substituents for Xm and Xn are:

[0125] R 1 is phenyl, which is optionally substituted by one or more C 1-6 -alkyl groups;

[0126] R 6 are independently of each other hydrogen, tert-butyl or methyl;

[0127] X is chloride

[0128] and when Q 1 is nitrogen and Q 2 is carbon, the dotted line ring represents an aromatic ring.

[0129] Suitable catalysts are typically commercially available, for example from Jiuzhou Pharma, Sinocompound or Johnson Matthey or from catalog suppliers such as for example Strem or Sigma Aldrich.

[0130] The reduction of the ketone of formula V can be performed in the presence of a reducing agent and a suitable organic solvent.

[0131] One option for the reducing agent is the use of a mixture of formic acid and a trialkylamine, preferably triethylamine or an alkali metal formate, such as sodium formate, in the presence of a tetraalkylammonium halide, such as tetrabutylammonium bromide (TBAB).

[0132] The mixing ratio can vary, for example between 1 and 5 equivalents of formic acid and 1 and 5 equivalents of triethylamine or, for example, 5 equivalents of sodium formate and 0.5 equivalents of TBAB.

[0133] In general, an organic solvent selected from, for example, ethanol, toluene, acetonitrile, 2-methyltetrahydrofuran or propylene carbonate can be used.

[0134] The reaction temperature is chosen between 10°C and 100°C, preferably between 20°C and 50°C.

[0135] In a further option, the reduction takes place in the presence of hydrogen at a hydrogen pressure of 1 bar to 100 bar, preferably 60 bar to 80 bar, and at a reaction temperature of 10°C to 100°C, preferably 40°C to 60°C.

[0136] Suitable organic solvents are aliphatic alcohols, such as ethanol.

[0137] Some catalysts require a base for activation.

[0138] Suitable bases are: inorganic bases selected from alkali metal or alkaline earth metal-carbonates, or alkali metal or alkaline earth metal-bicarbonates or phosphates or hydrogenophosphates or dihydrogenophosphates or acetates or formates; or organic bases selected from amines, alkali metal alcoholates or amidines. In general, organic bases are preferred. Typical representatives of organic bases are potassium tert-butoxide or 1,8-diazabicyclo[5.4.0]undec-7-ene (DBU), 1,4-diazabicyclo(2.2.2)octane (DABCO) and 7-methyl-1,5,7-triazabicyclo[4.4.0]dec-5-ene (MTBD), most preferred is potassium tert-butoxide.

[0139] The chiral pyrrolotriazole alcohol of formula I can be isolated from the reaction mixture by evaporation of the solvent. Subsequent column chromatography and / or product crystallization allows the chiral alcohol of formula I to be obtained in good yield, high purity and high enantiomeric excess.

[0140] b) reduction of the ketone of formula V with an oxidoreductase:

[0141] Suitable oxidoreductases are selected from: enzymes capable of reducing the ketone of formula V,

[0142]

[0143] wherein X is a halogen atom, n is an integer of 1, 2 or 3, and those oxidoreductases forming chiral pyrrolo-triazolols of formula I,

[0144]

[0145] wherein X and n are as described above, and wherein the helix bond

[0146]

[0147] represents "R" or "S" or a mixture of enantiomers. with an enantiomeric excess of at least 90%, preferably at least 95%, more preferably at least 98%.

[0148]

[0149] Asymmetric reductions are usually catalyzed by oxidoreductases in the presence of NADH or NADPH as cofactor, which is regenerated in situ. NAD + or NADP + is added to the reaction, not its reduced counterpart. The substrate to cofactor ratio (S / C) of NAD + or NADP + is usually kept in the range between 5 and 1000, preferably between 10 and 500.

[0150] The oxidized cofactor is usually regenerated continuously with a secondary alcohol as co-substrate. Typical co-substrates can be selected from 2-propanol, 2-butanol, pentane-1,4-diol, 2-pentanol, 4-methyl-2-pentanol, 2-heptanol, hexane-1,5-diol, 2-heptanol or 2-octanol, preferably 2-propanol. Preferably, the cofactor is regenerated by a co-substrate under the same enzyme catalyzing the target reaction. In another preferred embodiment, the acetone formed when 2-propanol is used as co-substrate is continuously removed from the reaction mixture.

[0151] ​​​​Also well known is the cofactor regeneration via additional enzymes oxidizing their natural substrates and providing reduced cofactors. For example secondary alcohol dehydrogenase / alcohols; glucose dehydrogenase / glucose; formate dehydrogenase / formate; glucose-6-phosphate dehydrogenase / glucose-6-phosphate; phosphite dehydrogenase / phosphite; hydrogenase / molecular hydrogen etc. Moreover, electrochemical regeneration methods are known and chemical cofactor regeneration methods including metal catalysts and reducing agents are also suitable. In particular, when glucose dehydrogenase / glucose is used for cofactor regeneration, the pH has to be maintained by controlled addition of base to neutralize the formed gluconic acid (byproduct of oxidation of the reduced nicotinamide cofactor regeneration). The substrate to cofactor ratio (S / GDH) is usually kept in the range between 5 and 1000, preferably between 10 and 200.

[0152] The preferred microbial oxidoreductases are derived from yeasts, bacteria, or from mammalian cells.

[0153] The oxidoreductases can be applied in the form of isolated enzyme(s) or as whole cells, optionally in immobilized form, by one of the numerous conventional methods described in the literature.

[0154] In a particular embodiment of the present application, the asymmetric reduction is carried out in aqueous medium in the presence of an organic co-solvent which can be selected, for example, from glycerol, 2-propanol, dimethyl sulfoxide, diethyl ether, tert-butyl methyl ether, diisopropyl ether, dibutyl ether, toluene, 2-methyltetrahydrofuran, ethyl acetate, butyl acetate, heptane, hexane or cyclohexene, or from mixtures thereof.

[0155] The presence of an organic co-solvent is particularly advantageous because a homogeneous suspension can be formed which allows the simple separation of the desired ketone of formula V by filtration.

[0156] The reaction concentration (concentration of the ketone of formula V and of the chiral alcohol of formula I in the reaction mixture) is usually kept in the range between 1% and 25% w / v, preferably between 4% and 20% w / v.

[0157] A buffered salt is used in the reaction, which can be selected, for example, from potassium phosphate buffer, Tris HC1 buffer, bicine buffer, HEPES buffer, PIPES buffer. The pH of the reaction is kept in the range between 6.0 and 9.0, preferably between 6.5 and 7.5.

[0158] The reaction temperature is usually kept in the range between 10°C and 50°C, preferably between 20°C and 30°C.

[0159] The substrate to enzyme ratio (S / E) is generally kept in the range between 5 and 1000, preferably between 10 and 200.

[0160] At the end of the reaction (usually conversion > 90%), the product is conventionally worked up by extraction.

[0161] Depending on the ketone substrate, the preferred catalyst / cofactor / co-substrate system can vary.

[0162] For the formation of (S)-2-bromo-6,7-dihydro-5H-pyrrolo[l,2-b][l,2,4]triazol-7-ol of formula (S)-Ia, the following oxidoreductases have proven useful.

[0163] The NADPH-dependent oxidoreductases can be selected from the KRED-NADPH-130, KRED-P1-C01, KRED-P2-D11, KRED-P2-D12, KRED-463 types from Codexis Inc, or the NADH-dependent oxidoreductases can be selected from the ADH-109, ADH-132 and ADH-172 types from c-LEcta.

[0164] For the formation of (R)-2-bromo-6,7-dihydro-5H-pyrrolo[l,2-b][l,2,4]triazol-7-ol of formula (R)-Ia, the following oxidoreductases have proven useful.

[0165] The NADPH-dependent oxidoreductases can be selected from the ADH-61 type from Johnson Matthey, or the NADH-dependent oxidoreductases can be selected from the KRED-NADH-110 type from Codexis Inc.

[0166] For the formation of (S)-2-bromo-5,6,7,8-tetrahydro-[l,2,4]triazolo[l,5-a]pyridin-8-ol of formula (S)-Ib, the following oxidoreductases have proven useful.

[0167] The NADPH-dependent oxidoreductases can be selected from the ADH-153 type from Johnson Matthey, or the NADH-dependent oxidoreductases can be selected from the ADH-109, ADH-110, ADH-132 and ADH-172 types from c-LEcta.

[0168] For the formation of (R)-2-bromo-5,6,7,8-tetrahydro-[l,2,4]triazolo[l,5- a]pyridin-8-ol of the formula (R)-lb, the following oxidoreductases have proven useful.

[0169] The NADPH-dependent oxidoreductases can be selected from the KRED-425 type from Codexis Inc., from the ADH-19, ADH-20 and ADH-61 type from Johnson Matthey.

[0170] For the formation of (S)-2-bromo-6,7,8,9-tetrahydro-5H-[l,2,4]triazolo[l,5- a]azepin-9-ol (S)-lc, the following oxidoreductases have proven useful.

[0171] The NADPH-dependent oxidoreductases can be selected from the ADH-153 type from Johnson Matthey, from the KRED-P2-Dl 1, KRED-464 type from Codexis Inc., or the NADH-dependent oxidoreductases can be selected from the ADH-109, ADH-110, ADH-132 and ADH-172 type from c-LEcta.

[0172] For the formation of (R)-2-bromo-6,7,8,9-tetrahydro-5H-[l,2,4]triazolo[l,5- a]azepin-9-ol (R)-lc, the following oxidoreductases have proven useful.

[0173] The NADPH-dependent oxidoreductases can be selected from the KRED-425 type from Codexis Inc., from the ADH-19, ADH-20 and ADH-61 type from Johnson Matthey.

[0174] chiral pyrrolotriazolols of the formula

[0175]

[0176] wherein

[0177] X is a halogen atom and

[0178] n is an integer of 1, 2 or 3, and the helix bond

[0179] “ ”

[0180] represents “ ” or “ ” or a mixture of enantiomers.

[0181] novel compounds and thus constitute another embodiment of the present application.

[0182] In a preferred embodiment,

[0183] X is chlorine or bromine, preferably bromine, and

[0184] n is 1, 2 or 3, and the helical bond

[0185] “ ”

[0186] represents “ ” or “ ” or a mixture of enantiomers.

[0187] In another preferred embodiment,

[0188] a) X is bromine and n is 1, and the helical bond “ ” represents “ ”,

[0189] b) X is bromine and n is 1, and the helical bond “ ” represents “ ”,

[0190] c) X is bromine and n is 2, and the helical bond “ ” represents “ ”,

[0191] d) X is bromine and n is 2, and the helical bond “ ” represents “ ”,

[0192] e) X is bromine and n is 3, and the helical bond “ ” represents “ ”,

[0193] f) X is bromine and n is 3, and the helical bond “ ” represents “ ”.

[0194] Example

[0195] Abbreviations:

[0196]

[0197]

[0198] Ru-catalyst

[0199]

[0200]

[0201]

[0202] All solvents, reagents and compounds are purchased and used without further purification unless otherwise stated. All catalysts and ligands are commercially available, for example from Jiuzhou Pharma, Sinocompound or Johnson Matthey or catalog suppliers such as for example Strem or Sigma Aldrich.

[0203] Analytical methods:

[0204] NMR methods:

[0205] 1 H-NMR spectra were measured in CDCl3or DMSO-d6 solutions at 25 °C on a Bruker AV 600 MHz, 400 MHz or 300 MHz spectrometer equipped with a DCH cryoprobe, chemical shifts (δ) reported in ppm (δ = 0 ppm) using trimethylchlorosilane as internal standard.

[0206] LC / MS methods:

[0207] LC / MS methods determine the conversion of II to IVa to IVc and IVa to IVc to Va to Vc and the purity of IVa to IVc and Va to Vc:

[0208] System: UPLC, Waters, photodiode array detector (PDA, Waters). Evaporative light scattering detector (ELSD, VWR 90 LT). LC: Stationary phase: Agilent Zorbax Eclipse Plus C18 RRHT, L = 30 mm, ID = 2.1 mm, 1.8 pm, eluent: A) water 0.1% formic acid; B) MeCN 0.07% formic acid. Pump program: 97 A:3 B, gradient to 13 A:87 B over 2 min, including UV spectrum. Run time: 2 min. Flow: 1 mL / min. Column oven temperature: 50 °C; injection volume: 2 pi. MS: single quadrupole (Waters SQD01). m / z 150-900. Detection: PDA 210 to 400 nm. Retention time: II: 0.65 min, IVa: 0.93 min, IVb: 1.08 min, IVc: 1.15 min, Va: 0.70 min, Vb: 0.61 min, Vc: 0.69 min

[0209] HPLC method:

[0210] a) HPLC method to determine conversion of Va to Ia and purity and enantiomeric excess of Ia:

[0211] System: Agilent 1290. Stationary phase: Chiralpak IBN-3, L = 150 mm, ID = 4.6 mm, 3 pm, eluent: A) n-heptane, B) EtOH. Pump program: 75 A:25 B for 10 min. Run time: 10 min. Flow: 1 mL / min. Column oven temperature: 25 °C; injection volume: 10 pi. Detection: DAD 205 nm. Retention time: Va: 13.80 min, (S)-Ia: 3.72 min, (R)-Ia: 4.36 min.

[0212] b) HPLC method to determine conversion of Vb to Ib and purity and enantiomeric excess of Ib:

[0213] System: Agilent 1290. Stationary phase: Chiralpak IBN-3, L = 150 mm, ID = 4.6 mm, 3 µm. Eluent: A) n-Heptane, B) EtOH. Pump program: 95 A: 5 B for 10 min, gradient to 45 A: 55 B over 5 min, then hold for 2 min, gradient to 95 A: 5 B over 0.1 min, then hold for 2.9 min. Run time: 20 min. Flow: 1 mL / min. Column oven temperature: 20 °C; Injection volume: 6 µL. Detection: DAD 210 nm. Retention time: Vb: 17.0 min, (S)-Ib: 9.0 min, (R)-Ib: 9.7 min.

[0214] c) HPLC method to determine conversion of Vc to Ic and purity and enantiomeric excess of Ic:

[0215] System: Agilent 1290. Stationary phase: Chiralpak AD-3, L = 150 mm, ID = 4.6 mm, 3.0 µm. Eluent: A) n-Heptane, B) EtOH. Pump program: 90 A: 10 B for 10 min. Run time: 10 min. Flow: 1 mL / min. Column oven temperature: 25 °C; Injection volume: 10 µL. Detection: DAD 205 nm. Retention time: Vc (keto / enol forms): 5.70 min and 6.60 min, (S)-Ic: 3.47 min, (R)-Ic: 3.81 min.

[0216] Optical rotation method:

[0217] Optical rotation values were obtained using an Anton Paar MCP-500 instrument. Optical rotation was measured in methanol at 20 °C at the indicated concentration.

[0218] Reaction scheme

[0219] Synthesis of chiral alcohols Ia to Ic

[0220] Scheme 4:

[0221]

[0222] Ring expansion of Va and Vb

[0223] Scheme 3:

[0224]

[0225] Step A: Synthesis of esters IVa to IVc

[0226] Example 1:

[0227] a) Synthesis of methyl 3-(3,5-dibromo-1,2,4-triazol-1-yl)propionate (IVa)

[0228]

[0229] 3,5-Dibromo-1H-1,2,4-triazole (40.00 g, 176.32 mmol, 1.00 equivalent) was suspended in MeTHF (90 mL). Triethylamine (5.35 g, 52.90 mmol, 0.30 equivalent) was added to the suspension. After stirring for 5 min at room temperature, the solution was incubated at 70 °C. Methyl acrylate (15.94 g, 16.78 mL, 185.14 mmol, 1.05 equivalents) was added over 4 h, and the solution was stirred further at 70 °C for 20 h. The solution was cooled to 20 °C, washed with 2 M HCl aqueous solution (27.24 g, 0.30 equivalents), followed by washing with water (20 mL). The organic phase was evaporated under vacuum to give the title compound (55.60 g, purity 92.0%, yield 92.7%) as a colorless to pale yellow oil.

[0230] LC / MS: 313.9, 315.9 [M+H] + , ESI pos.

[0231] 1 H-NMR (300 MHz, DMSO-d6): δ 4.37 (t, J = 6.49 Hz, 2H), 3.60 (s, 3H), 2.94 (t, J = 6.49 Hz, 2H).

[0232] b) Large-scale synthesis of methyl 3-(3,5-dibromo-1,2,4-triazol-1-yl)propionate (IVa):

[0233] Dissolve 3,5-dibromo-lH-l,2,4-triazole (131.0 kg, 577 mol, 1.00 equiv) in 2-MeTHF (160 kg). Add triethylamine (17.1 kg, 170 mmol, 0.30 equiv) to the light suspension and warm the mixture to IT 70 °C. Add methyl acrylate in 2-MeTHF (51.9 kg, 602 mol, 1.05 equiv in 110 kg 2-MeTHF) over a 4 hour period. Further stir the reaction mixture at IT 70 °C for 24 h. Confirm conversion by GC analysis (typically > 98.5a%). Wash the reaction mixture at 20 °C with HC1 (25.2 kg 25% HC1 + 60 kg water) and at 35 °C with water (65 kg). Azeotropically distil the organic phase (200 to 100 mbar) to remove water until < 200 ppm is reached (about 650 kg 2-MeTHF). Dilute the dried organic phase with 2-MeTHF and toluene (total 1528 kg) to obtain the title compound as about 8 wt% solution.

[0234] Example 2:

[0235] Synthesis of ethyl 4-(3,5-dibromo-l,2,4-triazol-l-yl)butanoate (IVb)

[0236]

[0237] Dissolve 3,5-dibromo-lH-l,2,4-triazole (24.70 g, 108.88 mmol, 1.00 equiv) in MeCN (250 mL) and DMF (25 mL). To this solution add potassium carbonate (37.62 g, 272.19 mmol, 2.50 equiv). After stirring at room temperature for 5 min, add ethyl 4-bromobutanoate (23.36 g, 17.30 mL, 119.77 mmol, 1.10 equiv) and stir the suspension at 75 °C for 3.5 h, at which time LC / MS indicates the reaction has reached complete conversion. Allow the suspension to cool, filter it, and wash the filter cake several times with EtOAc (250 mL) to ensure the product is collected in solution. Evaporate the solution under vacuum to remove all solvents (including DMF). Purify the crude product by silica gel chromatography (eluent: EtOAc / n-heptane, 5% to 40%) to obtain the title compound as a colorless oil (32.80 g, 93.0% purity, 86.6% yield).

[0238] LC / MS: 341.9, 343.9 [M+H] + , ESI pos.

[0239] 1 H-NMR (300 MHz, CDCl3): δ 4.24 (t, J = 6.9 Hz, 2H), 4.15 (q, J = 7.3Hz, 2H), 2.41 - 2.34 (m, 1H), 2.25 - 2.14 (m, 2H), 1.27 (t, J = 7.2 Hz, 3H).

[0240] Example 3:

[0241] Synthesis of ethyl 5-(3,5-dibromo-l,2,4-triazol-l-yl)pentanoate (IVc)

[0242]

[0243] Dissolve 3,5-dibromo-lH-l,2,4-triazole (20.00 g, 88.16 mmol, 1.00 equiv) in THF (70.53 mL). Add potassium carbonate (12.43 g, 89.92 mmol, 1.02 equiv) to the solution and after stirring at room temperature for 5 min, add ethyl 5-bromopentanoate (18.80 g, 13.79 mL, 89.92 mmol, 1.02 equiv) and stir the suspension at 60 °C for 20 h. Allow the suspension to cool to room temperature and filter off the solids through a funnel with a fritted disc (G3) and wash the solids with THF (100 mL). Evaporate the solution at 40 °C under vacuum (200-10 mbar) to yield the title compound (33.00 g, 90% purity, 94.9% yield) as a colourless oil.

[0244] LC- MS 355.9 / 357.9 [M+H]+, ESI pos.

[0245] 1 H NMR (400 MHz, DMSO-d6) δ ppm 1.11 - 1.24 (m, 3 H) 1.45 - 1.59 (m, 2H) 1.73 - 1.87 (m, 2 H) 2.27 - 2.40 (m, 2 H), 3.98 - 4.11 (m, 2 H) 4.11 -4.22 (m, 2 H) ppm.

[0246] Step B: Synthesis of ketones Va to Vc

[0247] Example 4:

[0248] a) Synthesis of 2-bromo-5,6-dihydropyrrolo[l,2-b][l,2,4]triazol-7-one (Va)

[0249]

[0250] In a continuous stirred tank reactor (CSTR) setup, methyl 3-(3,5-dibromo-l,2,4- triazol-l-yl)propanoate (20.00 g, 63.91 mmol, 1.00 equiv) was dissolved in MeTHF (357 mL) to give charge A solution (0.173 M). Charge A was dosed into CSTR1 (IT -60 °C, Tres 10 min) in parallel with n-hexyllithium (2.5 M in n-heptane, 22.93 g, 83.08 mmol, 1.30 equiv, 0.41 mL / min). The reaction mixture was transferred from CSTR1 to CSTR2 (IT + 10 °C, Tres 10 min) where a citric acid solution (30 wt% in water, 127.82 mmol, 2.00 equiv, 0.987 mL / min) was added. The biphasic reaction mixture was continuously (within 80 min) transferred from CSTR2 to batch reactor 3. After the reaction, ca. 480 mL of biphasic reaction mixture was collected in batch reactor 3. The aqueous phase was separated and the organic phase was washed with water (60 mL). The organic phase was concentrated in vacuo to ca. 50 mL volume. n-Heptane (15 mL) was added and the suspension was cooled to 0 °C. The solid was filtered off by means of a fritted funnel (G3) and the solid was washed with an ice-cold mixture of MeTHF / n-heptane (1 :2 v / v, 25 mL), dried under vacuum to yield the title compound as a dark brown solid (5.86 g, 91.0% purity, 45.4% yield). A (4.59 mL / min) with n-hexyllithium (2.5 M in n-heptane, 22.93 g, 83.08 mmol, 1.30 equiv, 0.41 mL / min) into CSTR1 (IT -60 °C, Tres 10 min). The reaction mixture was transferred from CSTR1 to CSTR2 (IT + 10 °C, Tres 10 min) where a citric acid solution (30 wt% in water, 127.82 mmol, 2.00 equiv, 0.987 mL / min) was added. The biphasic reaction mixture was continuously (within 80 min) transferred from CSTR2 to batch reactor 3. After the reaction, ca. 480 mL of biphasic reaction mixture was collected in batch reactor 3. The aqueous phase was separated and the organic phase was washed with water (60 mL). The organic phase was concentrated in vacuo to ca. 50 mL volume. n-Heptane (15 mL) was added and the suspension was cooled to 0 °C. The solid was filtered off by means of a fritted funnel (G3) and the solid was washed with an ice-cold mixture of MeTHF / n-heptane (1 :2 v / v, 25 mL), dried under vacuum to yield the title compound as a dark brown solid (5.86 g, 91.0% purity, 45.4% yield). CSTR2 (IT + 10 °C, Tres 10 min) where a citric acid solution (30 wt% in water, 127.82 mmol, 2.00 equiv, 0.987 mL / min) was added. The biphasic reaction mixture was continuously (within 80 min) transferred from CSTR2 to batch reactor 3. After the reaction, ca. 480 mL of biphasic reaction mixture was collected in batch reactor 3. The aqueous phase was separated and the organic phase was washed with water (60 mL). The organic phase was concentrated in vacuo to ca. 50 mL volume. n-Heptane (15 mL) was added and the suspension was cooled to 0 °C. The solid was filtered off by means of a fritted funnel (G3) and the solid was washed with an ice-cold mixture of MeTHF / n-heptane (1 :2 v / v, 25 mL), dried under vacuum to yield the title compound as a dark brown solid (5.86 g, 91.0% purity, 45.4% yield). CSTR2 (IT + 10 °C, Tres 10 min) where a citric acid solution (30 wt% in water, 127.82 mmol, 2.00 equiv, 0.987 mL / min) was added. The biphasic reaction mixture was continuously (within 80 min) transferred from CSTR2 to batch reactor 3. After the reaction, ca. 480 mL of biphasic reaction mixture was collected in batch reactor 3. The aqueous phase was separated and the organic phase was washed with water (60 mL). The organic phase was concentrated in vacuo to ca. 50 mL volume. n-Heptane (15 mL) was added and the suspension was cooled to 0 °C. The solid was filtered off by means of a fritted funnel (G3) and the solid was washed with an ice-cold mixture of MeTHF / n-heptane (1 :2 v / v, 25 mL), dried under vacuum to yield the title compound as a dark brown solid (5.86 g, 91.0% purity, 45.4% yield).

[0251] LC / MS: 201.9, 203.9 [M+H] + , ESI pos.

[0252] 1H-NMR (300 MHz, DMSO-d6): δ 4.48 (t, J = 5.19 Hz, 2H), 3.24 (t, J =5.19 Hz, 2H).

[0253] b) Scale-up synthesis of 2-bromo-5,6-dihydropyrrolo[1,2-b][1,2,4]triazol-7-one (Va):

[0254] Methyl 3-(3,5-dibromo-1,2,4-triazol-1-yl)propanoate (ca. 8.00 wt% in 2-MeTHF / toluene) (Feed A) (212 g / min) was dosed in parallel with n-hexyllithium (2.5 M in n-heptane, 18 g / min, 1.20 equiv) into CSTR1 (IT -60 °C, Tres 10 min). The reaction mixture was transferred from CSTR1 to CSTR2 (IT 0 to +10 °C, Tres 10 min) where a citric acid solution (30 wt% in water, 50 g / min, 1.5 equiv) was added. The biphasic reaction mixture was continuously transferred from CSTR2 to batch reactor 3. The process was run until all Feed A was consumed. The aqueous phase was separated and the organic phase was washed with water (130 kg). The organic phase was concentrated in vacuo (up to JT 35 °C) to ca. 300 L volume (target 20 to 25 wt% 2-MeTHF). The resulting suspension was cooled to -10 °C and further stirred at -10 °C for at least 2 hours. The solid was filtered off through a funnel (G3) with a fritted disc and the solid was washed with ice-cold toluene (260 kg), dried under vacuum to yield the title compound as a brown solid (65.4 kg, 90% assay, isolated yield 50.5%). 10 min) into CSTR2 (IT 0 to +10 °C, Tres 10 min) where a citric acid solution (30 wt% in water, 50 g / min, 1.5 equiv) was added. The biphasic reaction mixture was continuously transferred from CSTR2 to batch reactor 3. The process was run until all Feed A was consumed. The aqueous phase was separated and the organic phase was washed with water (130 kg). The organic phase was concentrated in vacuo (up to JT 35 °C) to ca. 300 L volume (target 20 to 25 wt% 2-MeTHF). The resulting suspension was cooled to -10 °C and further stirred at -10 °C for at least 2 hours. The solid was filtered off through a funnel (G3) with a fritted disc and the solid was washed with ice-cold toluene (260 kg), dried under vacuum to yield the title compound as a brown solid (65.4 kg, 90% assay, isolated yield 50.5%). 10 min) into CSTR2 (IT 0 to +10 °C, Tres 10 min) where a citric acid solution (30 wt% in water, 50 g / min, 1.5 equiv) was added. The biphasic reaction mixture was continuously transferred from CSTR2 to batch reactor 3. The process was run until all Feed A was consumed. The aqueous phase was separated and the organic phase was washed with water (130 kg). The organic phase was concentrated in vacuo (up to JT 35 °C) to ca. 300 L volume (target 20 to 25 wt% 2-MeTHF). The resulting suspension was cooled to -10 °C and further stirred at -10 °C for at least 2 hours. The solid was filtered off through a funnel (G3) with a fritted disc and the solid was washed with ice-cold toluene (260 kg), dried under vacuum to yield the title compound as a brown solid (65.4 kg, 90% assay, isolated yield 50.5%). 10 min) into CSTR2 (IT 0 to +10 °C, Tres 10 min) where a citric acid solution (30 wt% in water, 50 g / min, 1.5 equiv) was added. The biphasic reaction mixture was continuously transferred from CSTR2 to batch reactor 3. The process was run until all Feed A was consumed. The aqueous phase was separated and the organic phase was washed with water (130 kg). The organic phase was concentrated in vacuo (up to JT 35 °C) to ca. 300 L volume (target 20 to 25 wt% 2-MeTHF). The resulting suspension was cooled to -10 °C and further stirred at -10 °C for at least 2 hours. The solid was filtered off through a funnel (G3) with a fritted disc and the solid was washed with ice-cold toluene (260 kg), dried under vacuum to yield the title compound as a brown solid (65.4 kg, 90% assay, isolated yield 50.5%).

[0255] Example 5:

[0256] 2-bromo-6,7-dihydro-5H-[1,2,4]triazolo[1,5-a]pyridin-8-one (Vb)

[0257]

[0258] In a CSTR setup, ethyl 4-(3,5-dibromo-1,2,4-triazol-1-yl)butanoate (31.00 g, 90.91 mmol, 1.00 equiv) was dissolved in MeTHF (482 mL) to give a Feed A solution (0.182 M). The Feed A (6.00 mL / min) was dosed in parallel with n-hexyllithium (2.5 M in n-heptane, 30.11 g, 109.09 mmol, 1.20 equiv, 0.54 mL / min) into CSTR1 (IT -60 °C, Tres 10 min). The reaction mixture was transferred from CSTR1 to CSTR2 (IT +10 °C, Tres 10 min) where an acetic acid solution (10 wt% in water, 118.18 mmol, 1.30 equiv, 0.88 mL / min) was added. The biphasic reaction mixture was continuously (over 80 min) transferred from CSTR2 to batch reactor 3 (594 mL of biphasic reaction mixture in total). The aqueous phase was separated and the organic phase was washed with water (60 mL). The organic phase was concentrated in vacuo to a volume of ~50 mL. n-Heptane (15 mL) was added and the suspension was cooled to 0 °C. The solid was filtered off through a funnel (G3) with a fritted disc and the solid was washed with an ice-cold mixture of MeTHF / n-heptane (1:5 v / v, 25 mL), dried under vacuum to yield the title compound as a light yellow solid (4.20 g, 98.0% purity, 21.4% yield). CSTR2 (IT +10 °C, Tres 10 min) where an acetic acid solution (10 wt% in water, 118.18 mmol, 1.30 equiv, 0.88 mL / min) was added. The biphasic reaction mixture was continuously (over 80 min) transferred from CSTR2 to batch reactor 3 (594 mL of biphasic reaction mixture in total). The aqueous phase was separated and the organic phase was washed with water (60 mL). The organic phase was concentrated in vacuo to a volume of ~50 mL. n-Heptane (15 mL) was added and the suspension was cooled to 0 °C. The solid was filtered off through a funnel (G3) with a fritted disc and the solid was washed with an ice-cold mixture of MeTHF / n-heptane (1:5 v / v, 25 mL), dried under vacuum to yield the title compound as a light yellow solid (4.20 g, 98.0% purity, 21.4% yield). CSTR2 (IT +10 °C, Tres 10 min) where an acetic acid solution (10 wt% in water, 118.18 mmol, 1.30 equiv, 0.88 mL / min) was added. The biphasic reaction mixture was continuously (over 80 min) transferred from CSTR2 to batch reactor 3 (594 mL of biphasic reaction mixture in total). The aqueous phase was separated and the organic phase was washed with water (60 mL). The organic phase was concentrated in vacuo to a volume of ~50 mL. n-Heptane (15 mL) was added and the suspension was cooled to 0 °C. The solid was filtered off through a funnel (G3) with a fritted disc and the solid was washed with an ice-cold mixture of MeTHF / n-heptane (1:5 v / v, 25 mL), dried under vacuum to yield the title compound as a light yellow solid (4.20 g, 98.0% purity, 21.4% yield).

[0259] LC / MS: 215.9, 217.9 [M+H] ESI pos. + , ESI pos.

[0260] 1 H-NMR (300 MHz, DMSO-d6): δ 4.40 (t, J = 6.1, 2H), 2.77 (t, J = 6.5Hz, 2H), 2.35 (pent., J = 6.5, 6.1 Hz, 2H).

[0261] Example 6:

[0262] Synthesis of 2-bromo-5,6,7,8-tetrahydro-[1,2,4]triazolo[1,5-a]azepin-9-one (Vc)

[0263]

[0264] In a CSTR apparatus as outlined, 5-(3,5-dibromo-l,2,4-triazol-l-yl)pentanoic acid ethyl ester (30.80 g, 86.75 mmol, 1.00 equiv) was dissolved in MeTHF (570.2 mL) to give a charge A solution (0.130 M). Charge A (4.00 mL / min) was dosed in parallel with n-hexyllithium (2.5 M in heptane, 41.64 mL, 104.1 mmol, 1.20 equiv, 0.25 mL / min) into CSTR 1 (IT -60 °C, Tres 10 min). The reaction mixture was transferred from CSTR 1 to CSTR 2 (IT + 5 °C, Tres 10 min) where an acetic acid solution (10 wt% in water, 127.82 mmol, 2.00 equiv, 0.625 mL / min) was added. The biphasic reaction mixture was continuously (over 150 min) transferred from CSTR 2 to a batch reactor 3 (a total of 731 mL of biphasic reaction mixture was collected). The aqueous phase was separated and the organic phase was washed with water (60 mL). The organic phase was concentrated in vacuo to give crude 2-bromo-5,6,7,8-tetrahydro-[l,2,4]triazolo[l,5-a]azepin-9-one. The product was purified by SiliaSep™ HP 25-40 pm silica gel column chromatography (EtOAc / n-heptane) followed by re-purification by C18-silica gel chromatography (water / MeCN) to provide the title compound as a white solid (315 mg, purity 95%, yield 3.1%).

[0265] LC / MS: 229.98, 231.97 [M+H]+, ESI pos.

[0266] 1 H NMR (600 MHz, CDCl3) δ ppm 4.48 - 4.62 (m, 2 H), 2.89 - 2.99 (m, 2H), 2.16 - 2.28 (m, 2 H), 2.02 - 2.12 (m, 2 H) ppm.

[0267] Step D: Synthesis of ketones Vb to Vc

[0268] Example 7:

[0269] Synthesis of 2-bromo-6,7-dihydro-5H-[l,2,4]triazolo[l,5-a]pyridin-8-one (Vb)

[0270]

[0271] To a solution of 2-bromo-5,6-dihydropyrrolo[l,2-b][l,2,4]triazin-7-one (0.50 g, 2.48 mmol, 1.00 equiv) dissolved in DCM (10 mL) at -78 °C was added 2 M trimethylaluminum in 2 M toluene (1.49 mL, 2.97 mmol, 1.20 equiv) dropwise, followed by the addition of trimethylsilyldiazomethane in diethyl ether (1.36 mL, 2.72 mmol, 1.10 equiv) to the reaction mixture at -78 °C. The reaction was stirred at -20 °C for 3 h (when LC / MS indicated formation of product), and 1 M HC1 solution was added. The solution was extracted with DCM twice. The organic layer was dried over MgS04and concentrated to dryness. The crude material was purified by silica gel flash chromatography to obtain the title compound as a light brown solid (55 mg, 98.0% purity, 9.6% yield).

[0272] LC / MS: 215.96, 217.98 [M+H] + , ESI pos.

[0273] 1 H-NMR (300 MHz, CDCl3): δ 4.47 (m, 2H), 2.86 (m, 2H), 2.48 (m, 2H).

[0274] Example 8:

[0275] Synthesis of 2-bromo-5,6,7,8-tetrahydro-[l,2,4]triazolo[l,5-a]azepin-9-one (Vc)

[0276]

[0277] Dissolve 2,6-di-tert-butyl-4-methylphenol (2.24 g, 10.18 mmol, 2.20 equiv) in DCM (10 mL) at room temperature and add 2 M trimethylaluminum in 2M of toluene (2.78 mL, 5.55 mmol, 1.20 equiv). Stir the mixture for 1 h and after cooling to -78°C add 2-bromo-6,7-dihydro-5H-[1,2,4]triazolo[1,5-a]pyridin-8-one (1.00 g, 4.63 mmol, 1.00 equiv) dissolved in dichloromethane (10 mL) followed by TMS-diazomethane 2M in diethylether (2.55 mL, 5.09 mmol, 1.10 equiv). Stir the reaction at -78°C for 2 h until LC / MS indicates formation of product, therefore add 1M HCl solution. Extract the reaction with DCM twice and dry the organic layer over MgS04and concentrate to dryness under vacuum. Purify the crude material by flash chromatography on silica gel (40 g, EtOAc in n-heptane 0-20%). Further purify the obtained compound by flash chromatography on C18 (50 g, water in MeCN 10% to 95%) to obtain the title compound as a light brown solid (106 mg, purity 95.0%, yield 10.0%).

[0278] LC / MS: 229.98, 231.97 [M+H] + , ESI pos.

[0279] 1 H-NMR (300 MHz, CDCl3): δ 4.54 (m, 2H), 2.93 (m, 2H), 2.23 (m, 2H),2.09 (m, 2H).

[0280] Step C: Synthesis of alcohols la to lc via metal catalyzed reduction

[0281] Example 9.1:

[0282] Synthesis of (S)-2-bromo-6,7-dihydro-5H-pyrrolo[1,2-b][1,2,4]triazol-7-ol ((S)-Ia)

[0283]

[0284] In a glovebox (< 1 ppm O2), 2-bromo-5,6-dihydropyrrolo[l,2-b][l,2,4]triazol-7-one (500 mg, 2.48 mmol, 1.00 equiv) and Ru-461 (78.73 mg, 123.76 µmol, 0.05 equiv) were weighed into a 50 mL Schlenk flask and dissolved in MeCN (12 mL). A pre-mixed mixture of formic acid (224.06 uL, 5.84 mmol, 2.36 equiv) and triethylamine (651.20 mg, 6.44 mmol, 2.60 equiv) was then added to the solution and rinsed with MeCN (2 mL). The flask was sealed with a septum and removed from the glovebox. The flask was connected to an argon line and the yellow reaction mixture was stirred in an oil bath at Tj 31 °C for 20 h to achieve full conversion (LC / MS analysis). The yellow reaction mixture was cooled to room temperature and the solvent was removed to dryness under vacuum to yield the crude product. The crude product was purified by column chromatography with EtOAc / n-hexanes to yield 385 mg of the title compound as a white solid. The crude product was suspended in EtOAc (2 mL) and the mixture was stirred at room temperature for 10 min before n-hexanes (4 mL) were added to yield a suspension. The suspension was stirred at room temperature for 30 min before cooling down to 0 °C with an ice bath and stirring at 0 °C for another 30 min. The solids were filtered off through a funnel (G3) with a fritted disc and the solids were washed with an ice-cold mixture of EtOAc / n-hexanes (1 :2 v / v, 5 mL) and dried under vacuum to yield the title compound as an off-white solid (374 mg, purity >99%, yield 74.0%, (S):(R)-Ia = >99.95:0.05).

[0285] MS (EI + ): m / z 203.9770 [M+H] + .

[0286] 1H-NMR (600 MHz, DMSO-d6) δ 6.00 (br s, 1H), 5.06 (dd, J = 7.5, 3.7Hz, 1H), 4.24 (dddd, J = 10.9, 8.5, 4.9, 0.9 Hz, 1H), 4.05 (ddd, J = 10.9,8.6, 5.0 Hz, 1H), 2.93 (dddd, J = 13.4, 8.5, 7.7, 5.0 Hz, 1H), 2.33 (dddd, J= 13.5, 8.6, 4.8, 3.9 Hz, 1H).

[0287] [α] D 20 : +10.08° (MeOH, c = 1.107).

[0288] Example 9.2:

[0289] Synthesis of (R)-2-bromo-6,7-dihydro-5H-pyrrolo[1,2-b][1,2,4]triazol-7-ol ((R)-Ia)

[0290]

[0291] In a glove box (< 1 ppm O2), 2-bromo-5,6-dihydropyrrolo[1,2-b][1,2,4]triazol-7- one (100 mg, 495.03 pmol, 1.0 equiv) was weighed into a 10 mL Schlenk flask and suspended in MeCN (0.5 mL). Then, formic acid (57 mg, 47.5 pL, 1.24 mmol, 2.5 equiv) and triethylamine (62.6 mg, 86.1 pL, 618.8 pmol, 1.25 equiv) were added, followed by Ru-466 (3.15 mg, 4.95 pmol, 0.01 equiv) and acetonitrile (0.5 mL). The Schlenk flask was sealed with a septum, removed from the glove box, and the reaction mixture was stirred at Tj 32 °C for 19 h (LC / MS indicated complete conversion). The yellow reaction mixture was cooled to room temperature and the solvent was removed to dryness under vacuum to yield the crude product. The crude product was purified by column chromatography with EtOAc / n-heptane to yield 92 mg of the title compound as a white solid (purity >99%, yield 90.1%, (S):(R)-Ia = 0.5:99.5).

[0292] [α] D20 -11.0° (MeOH, c = 1.053).

[0293] Examples 9.3 to 9.11

[0294] Synthesis of (S)- or (R)-2-bromo-6,7-dihydro-5H-pyrrolo[l,2-b][l,2,4]triazol-7-ol (la)

[0295]

[0296] Reduction of Va (5 mg, 25 μmol) to la under the conditions as listed in Table E9, analogous to Example 9.1 with the HTE setup (48 to 96 well plate format).

[0297] Table E9 a

[0298]

[0299] a Conditions: 5 mg scale, 0.5 mol% of catalyst, 0.5 M concentration, 40 °C.

[0300]

[0301] b Conditions: 5 mg scale, 0.5 mol% of catalyst, solvent (10 V). c 2 mg scale, 2 mol% of catalyst, solvent (10 V). d 5 mg scale, 2 mol% of catalyst, KOtBu (0.3 equiv.).

[0302] Example 10.1:

[0303] Synthesis of (S)-2-bromo-5,6,7,8-tetrahydro-[l,2,4]triazolo[l,5-a]pyridin-8-ol ((S)-Ic)

[0304]

[0305] In a glove box (< 1 ppm O2), a Schlenk vessel was charged with 2-bromo-6,7-dihydro-5H- [1,2,4]triazolo[1,5-a]pyridin-8-one (100 mg, 462.9 µmol, 1.0 equiv), toluene (2 mL), formic acid (106.5 mg, 88.8 µL, 2.1 mmol, 5.0 equiv), trimethylamine (234.2 mg, 322.6 µL, 2.3 mmol, 5.0 equiv) and stirred for 2 min to give a solution. To this solution was added Ru-461 (14.72 mg, 23.14 µmol, 0.05 equiv), the Schlenk vessel was sealed with a septum and removed from the glove box. The flask was connected to an argon line and the yellow reaction mixture was stirred in an oil bath at Tj 42°C for 4 h to achieve full conversion (LC / MS analysis, (S):(R)-Ib: 98.6:1.4). The yellow reaction mixture was cooled to room temperature and the solvent was removed to dryness under vacuum to yield crude product (114 mg). The crude product was purified by column chromatography with EtOAc / n-heptane to yield the title compound as a white solid (72 mg, purity 95.0%, yield 71.0%, (S):(R)-Ib = 98.7:1.3).

[0306] 1 H-NMR (600 MHz, DMSO-d6): δ 5.85 (br s, 1H), 4.70 (t, J = 5.1, 1H), 4.14 (s, 1H), 4.00 (ddd, J = 12.9, 7.9, 5.2 Hz, 1H), 2.08 - 2.17 (m, 1H), 1.96 - 2.03 (m, 1H), 1.88 - 1.94 (m, 1H), 1.80 - 1.85 (m, 1H).

[0307] [α] D 20 : +0.525° (MeOH, c = 1.047).

[0308] Example 10.2:

[0309] (R)-2-bromo-5,6,7,8-tetrahydro-[1,2,4]triazolo[1,5-a]pyridin-8-ol

[0310] Synthesis of (R)-Ib

[0311]

[0312] In a glove box (< 1 ppm O2), a pressure vessel was charged with 2-bromo-6,7-dihydro-5H- [1,2,4]triazolo[1,5-a]pyridin-8-one (100 mg, 462.9 µmol, 1.0 equiv), KOtBu (1.00 mg, 9.30 µmol, 0.02 equiv) and Ir-15 (6.74 mg, 9.3 µmol, 0.02 equiv) and ethanol (1.5 mL) to give an orange suspension. The pressure vessel was sealed and pressurized with 7 bar of argon and removed from the glove box. The pressure vessel was connected to a hydrogenation line, the line and the pressure vessel were flushed with hydrogen. The pressure vessel was then pressurized with 70 bar of hydrogen and stirred at Tj 52 °C for 21 h. The pressure vessel was then brought to ambient temperature and the pressure was released. The pressure vessel was opened and sampled for analysis ((S):(R)-Ib = 5:95). The reaction mixture was transferred to a round bottom flask and the solvent was removed under vacuum to yield the crude title compound 108 mg as a light red oil. The crude product was purified by column chromatography with EtOAc / n-heptane to yield the title compound as a white solid (71 mg, purity 99.0 percent, yield 77.0 percent, (S):(R)-Ib = 5.3:94.7).

[0313] MS (EI + ): m / z 217.0 [M-H] + .

[0314] [α] D 20 : -3.836° (MeOH, c = 1.040).

[0315] Examples 10.3 to 10.10:

[0316] Synthesis of (S)- or (R)-2-bromo-5,6,7,8-tetrahydro-[1,2,4]triazolo[1,5-a]pyridin-8-ol ((S)- / (R)-Ib)

[0317]

[0318] Vb (2 mg, 8.7 µmol) was reduced to Ib under the conditions as listed in Table E10, analogous to Example 10.1 employing a HTE setup (48 to 96 well plate format).

[0319] Table E10:

[0320]

[0321] a Conditions: 2 mg scale, solvent (20 V), 40 °C.

[0322]

[0323] b Conditions: 2 mg scale, EtOH (12.4 V), KOtBu (0.02 eq), 50 °C.

[0324] Example 11.1:

[0325] Synthesis of (S)-2-bromo-6,7,8,9-tetrahydro-5H-[1,2,4]triazolo[1,5- a]azepin-9-ol ((S)-Ic)

[0326]

[0327] In a glovebox (< 1 ppm O2), 2-bromo-5,6,7,8-tetrahydro-[1,2,4]triazolo[1,5- a]azepin-9-one (50 mg, 217.3 pmol, 1.00 eq) and Ru-461 (1.38 mg, 2.17 pmol, 0.01 eq) were weighed into a 10 mL Schlenk flask and dissolved in MeCN (1 mL). Then, a pre- made mixture of MeCN (0.5 mL), formic acid (25 mg, 20.8 pL, 543.3 pmol, 2.5 eq) and triethylamine (27.50 mg, 37.8 pL, 271.67 pmol, 1.25 eq) was added to the solution. The flask was sealed with a septum, removed from the glovebox, and the yellow reaction mixture was stirred in an oil bath at Tj 33 °C for 22 h to achieve full conversion (LC / MS analysis). The yellow reaction mixture was cooled to room temperature and the solvent was removed to dryness under vacuum to yield the crude product. The crude product was purified by column chromatography with EtOAc / n-hexane to yield the title compound as a white solid (45 mg, purity >99%, yield 89.2%, (S):(R)-Ic = 93.6:6.4).

[0328] MS (EI + ): m / z 231 (M + ).

[0329] 1H-NMR (600 MHz, CDCl3): δ 4.93 (dd, J = 8.8, 2.6 Hz, 1H), 4.34 - 4.46 (m, 1H), 4.07 - 4.18 (m, 1H), 2.67 - 3.68 (m, 1H), 2.13 - 2.28 (m, 1H), 1.95- 2.04 (m, 1H), 1.72 - 1.94 (m, 4H).

[0330] [α] D 20 : +11.296° (MeOH, c = 0.108)

[0331] Example 11.2:

[0332] Synthesis of (R)-2-bromo-6,7,8,9-tetrahydro-5H-[1,2,4]triazolo[1,5- a]azepin-9-ol ((R)-Ic)

[0333]

[0334] In a glove box (< 1 ppm O2), 2-bromo-5,6,7,8-tetrahydro-[1,2,4]triazolo[1,5- a]azepin-9-one (60 mg, 260.8 µmol, 1.0 equiv) and Ru-466 (1.66 mg, 2.61 µmol, 0.01 equiv) were weighed into a 10 mL Schlenk tube and dissolved in MeCN (1.2 mL). Then, a pre-made mixture of MeCN (0.6 mL), formic acid (30 mg, 25 µL, 652 µmol, 2.5 equiv) and triethylamine (33 mg, 45.4 µL, 326 µmol, 1.25 equiv) was added to the solution. The flask was sealed with a septum, removed from the glove box, and the yellow reaction mixture was stirred in an oil bath at Tj 32°C for 19 h to achieve full conversion (LC / MS analysis). The yellow reaction mixture was cooled to room temperature and the solvent was removed to dryness under vacuum to yield the crude product. The crude product was purified by column chromatography with EtOAc / n-heptane to yield the title compound as a white solid (56 mg, purity >99%, yield 92.5 percent, (S):(R)-Ic = 6.2:93.8).

[0335] [α] D 20 : - 1.669° (MeOH, c = 1.067).

[0336] Examples 11.3 to 11.10:

[0337] Synthesis of (S)- and (R)-2-bromo-6,7,8,9-tetrahydro-5H- [1,2,4]triazolo[1,5-a]azepin-9-ol ((S)- / (R)-Ic)

[0338]

[0339] Vc (2 mg, 8.7 µmol) was reduced to Ic under the conditions as listed in Table E11, analogous to Example 11.1 with the HTE setup (48 to 96 well plate format).

[0340] Table E11:

[0341]

[0342] a Conditions: 2 mg scale, catalyst (5 mol%), solvent (10 V), 40 °C, 16 h.

[0343]

[0344] b Conditions: 2 mg scale, EtOH (12.4 V), KOtBu (0.02 eq), 50 °C, 20 h.

[0345] Step C: Synthesis of alcohols la to lc via enzyme catalyzed reduction

[0346] Examples 12.1 to 12.10:

[0347] Synthesis of (S)- and (R)-2-bromo-6,7-dihydro-5H-pyrrolo[1,2-b][1,2,4]triazol-7-ol ((S)- / (R)-Ia)

[0348]

[0349] To identify oxidoreductases capable of reducing 2-bromo-5,6-dihydropyrrolo[1,2-b][1,2,4]triazol-7-one to (S)- or (R)-2-bromo-6,7-dihydro-5H-pyrrolo[1,2-b][1,2,4]triazol-7-ol, a set of oxidoreductases was screened.

[0350] The reaction was prepared in 100 mM potassium phosphate buffer at pH 7. NADP + (1 g / L), NAD +(1 g / L), a redox enzyme as outlined in Table E12 (2 g / L), D-glucose (100 mM), glucose dehydrogenase GDH-105 (Codexis) (0.1 g / L) prepared from water were added to the reaction. The substrate of the reaction was dissolved in dimethyl sulfoxide at a concentration of 100 g / L and was distributed in the reaction at a final concentration of 5 g / L. The final volume of the reaction was 0.5 mL.

[0351] The reaction was incubated for 18 hours at room temperature. The reaction was quenched by the addition of 1 volume of acetonitrile. The reaction was then centrifuged at 3220 relative centrifugal force (rcf) for 5 minutes and the clear solution was subsequently transferred to a new glass vial for analysis.

[0352] The selection of the optimal enzymes for the conversion is reported in Table E12:

[0353] Table E12:

[0354]

[0355] Example 12.11:

[0356] Synthesis of (S)-2-bromo-6,7-dihydro-5H-pyrrolo[l,2-b][l,2,4]triazol-7-ol ((S)-Ia)

[0357]

[0358] Similar to Example 12.6, a reaction solution of 25.5 mL of an aqueous buffer containing a redox enzyme KRED-P2-D11 from Codexis (3 mg), D-(+)-glucose monohydrate (9 mmol), commercial glucose dehydrogenase GDH-105 from Codexis (15 mg), and an oxidation co-factor NADP + (15 mg) from Roche Diagnostics (100 mM potassium phosphate buffer, pH 6.5) was prepared under gentle stirring. The reaction solution was incubated and stirred for 5 min at ambient temperature (23 °C). The reduction was started later by the addition of 1.5 g (7.43 mmol) of 2-bromo-5,6-dihydropyrrolo[l,2-b][l,2,4]triazol-7-one (Va) in 3 mL of toluene. The pH was adjusted and kept constant during the reaction by dosing NaOH 1 M via a Metrohm pH Stat (Metrohm 902 Titrando).

[0359] The complete conversion (IPC: >99.9 area % product) was achieved at constant pH in 8 h at ambient temperature, consuming 7.43 mL of NaOH. Toluene was removed from the reaction by evaporation under reduced pressure. The reaction was then filtered through filter paper. After filtration, sodium carbonate (20 g) and 2-methyltetrahydrofuran (100 mL) were added to the reaction, mixed vigorously, and the phases were allowed to spontaneously separate. The separated aqueous phase was again extracted with 2-methyltetrahydrofuran (100 mL), and the combined phases were dried over MgS04, filtered, and evaporated under vacuum at 40 °C to yield the title compound as an off-white solid (1.15 g, purity >95%, yield 76.0%, (S):(R)-Ia >99.9:0.1%, (R)-Ia and Va not detected).

[0360] LC / MS: 203.98 (M+H) + ESI pos.

[0361] 1 H-NMR (600 MHz, DMSO-d6) δ 5.99 (br s, 1H), 5.05 (dd, J = 6.9, 3.2Hz, 1H), 4.23 (dddd, J = 10.8, 8.5, 4.9, 0.9 Hz, 1H), 4.04 (ddd, J = 10.9, 8.6, 5.0 Hz, 1H), 2.93 (dddd, J = 13.4, 8.5, 7.8, 5.0 Hz, 1H), 2.32 (dddd, J = 13.4, 8.6, 4.9, 3.8 Hz, 1H).

[0362] Example 12.12:

[0363] Synthesis of (R)-2-bromo-6,7-dihydro-5H-pyrrolo[l,2-b][l,2,4]triazol-7-ol ((R)-Ia)

[0364]

[0365] Similar to Example 12.12, 32.1 mL of a solution containing the oxidoreductase KRED-NADH-110 (250 mg) from Codexis, D-(+)-glucose monohydrate (27.2 mmol), the commercial glucose dehydrogenase GDH-105 (250 mg) from Codexis, and the oxidative cofactor NADP from Roche Diagnostics was prepared under gentle stirring. + The reaction solution was prepared in an aqueous buffer (100 mM potassium phosphate buffer, pH 6.5) of 250 mg. The reaction solution was incubated at ambient temperature (23 °C) with stirring for 5 min. Reduction was then initiated by adding 5 g (24.75 mmol) of 2-bromo-5,6-dihydropyrrolo[1,2-b][1,2,4]triazol-7-one (Va) in 7.5 mL of toluene. The pH was adjusted and kept constant during the reaction by feeding 1 M NaOH via a Metrohm pH Stat (Metrohm 902 Titrando). Complete conversion was achieved at constant pH over 8 h at ambient temperature (IPC: >99.9 area % product), consuming 12.38 mL of NaOH. Toluene was removed from the reaction by evaporation under reduced pressure. The reaction mixture was then filtered through filter paper. After filtration, sodium carbonate (20 g) and 2-methyltetrahydrofuran (100 mL) were added to the reaction mixture, which was vigorously mixed and allowed to spontaneously separate the phases. The separated aqueous phase was extracted again with 2-methyltetrahydrofuran (100 mL), and the combined phases were dried over MgSO4, filtered, and evaporated under vacuum at 40 °C to produce the title compound as a grayish-white solid (1.64 g, purity >95%, yield 32.0%, (S):(R)-Ia = 1.4:98.6, Va not detected).

[0366] LC / MS: 203.98 (M+H) + ESI pos.

[0367] 1H NMR (600 MHz, DMSO-d6) δ ppm 5.99 (br s, 1 H), 5.05 (br dd, J=6.9,3.2 Hz, 1 H), 4.23 (dddd, J=10.8, 8.5, 4.9, 0.9 Hz, 1 H), 4.04 (ddd, J=10.8,8.6, 5.0 Hz, 1 H), 2.93 (dddd, J=13.4, 8.5, 7.8, 5.0 Hz, 1 H), 2.32 (dddd, J=13.4, 8.6, 4.9, 3.8 Hz, 1 H)。

[0368] Example 12.13:

[0369] Large scale synthesis of (S)-2-bromo-6,7-dihydro-5H-pyrrolo[l,2-b][l,2,4]triazol-7-ol ((S)-la)

[0370] Similar to Example 12.11, 11.75 kg KH2PO4 and 11.76 K2HPO4, 385 g NADP + The reaction was prepared by mixing 1.54 kg of KRED-P2-D11 from Codexis, 1.54 kg of Na2HPO4, 1.54 kg of NaOH, and 1.54 kg of KH2PO4 in 1,100 L of water. Subsequently, 240 L of isopropanol was added in the reactor. The pH of the solution was 7.0 and the solution was incubated at 22 °C. The reaction was started by adding 77 kg (442 mol) of 2-bromo-5,6-dihydropyrrolo[l,2-b][l,2,4]triazol-7-one (Va) in 8 portions of 9.8 to 9.6 kg at small intervals. The reaction was complete after 12 hours (IPC: >99.9% conversion, 97.9% la, (S):(R)-la > 99.9:0.1%).

[0371] The isopropanol and acetone were removed at reduced pressure at 55 °C until 560 kg of distillate was removed. 1150 kg of water was added and the reaction was stirred for 30 minutes. The pH was then set to 2.0 by adding 53 kg of 20% aqueous sulfuric acid and the mixture was then stirred at 55 °C for 45 minutes to achieve enzyme precipitation.

[0372] The solution was filtered over a 3M™ Zeta Plus™ cartridge and the filter was then rinsed twice with a mixture of 231 kg of water and 11.5 kg of 20% aqueous sulfuric acid and also the solution was passed through the filter.

[0373] The filtered solution was heated at 100°C and water was distilled until about 720 kg of solution remained in the autoclave. The temperature was reduced to 25°C, 1,150 kg of 2-methyltetrahydrofuran was added to the reactor and compound la was extracted in the organic phase. The aqueous fraction was removed and the extraction process was repeated twice by adding 493 kg of 2-methyltetrahydrofuran. Subsequently, the 2-methyltetrahydrofuran solution of la was filtered through a Zetacarbon filter.

[0374] The 2-methyltetrahydrofuran solution was heated to 110°C and the solvent was removed under vacuum until 380 kg of solution remained in the reactor. The solution was cooled to 80°C and subsequently further cooled to 20°C at a rate of 20°C / hour. Then 237 kg of n-heptane was added to the reactor over 45 minutes and the solution was stirred for 2 hours. The suspension was then cooled to 0°C at a rate of 10°C / hour and stirred for 2 hours.

[0375] The suspension was centrifuged and the crystals were washed with 264 kg of n-heptane. Finally the crystals were dried at 50°C under full vacuum in an oven.

[0376] Compound la was isolated with a reaction yield of 84% (IPC purity 99.7%, (S):(R)-la > 99.9:0.1%) as off-white crystals.

[0377] Examples 13.1 to 13.9:

[0378] Synthesis of (S)- and (R)-2-bromo-5,6,7,8-tetrahydro-[l,2,4]triazolo[l,5- a]pyridin-8-ol ((S)- / (R)-lb)

[0379]

[0380] To identify oxidoreductases capable of reducing 2-bromo-6,7-dihydro-5H- [l,2,4]triazolo[l,5-a]pyridin-8-one to (8S) or (8R)-2-bromo-5,6,7,8-tetrahydro- [l,2,4]triazolo[l,5-a]pyridin-8-ol, a set of oxidoreductases was screened. The reactions were prepared in 100 mM potassium phosphate buffer at pH 6.5. NADP + (1 g / L), NAD +(S)- and (R)-2-bromo-6,7,8,9-tetrahydro-5H-[1,2,4]triazolo[1,5- a]azepin-9-ol ((S)- / (R)-Ic) were synthesized

[0381] Table E13:

[0382]

[0383] Examples 14.1 to 14.11:

[0384] (S)- and (R)-2-bromo-6,7,8,9-tetrahydro-5H-[1,2,4]triazolo[1,5- a]azepin-9-ol ((S)- / (R)-Ic) were synthesized

[0385]

[0386] To identify oxidoreductases capable of reducing 2-bromo-5,6,7,8-tetrahydro- [1,2,4]triazolo[1,5-a]azepin-9-one to (9S) or (9R)-2-bromo-6,7,8,9-tetrahydro-5H- [1,2,4]triazolo[1,5-a]azepin-9-ol, a panel of oxidoreductases was screened. Reactions were prepared in 100 mM potassium phosphate buffer at pH 6.5. NADP + (1 g / L), NAD +(1 g / L), oxidoreductase (0.08 g / L), D-glucose (100 mM), glucose dehydrogenase GDH-105 (Codexis) (0.02 g / L) were added to the reaction from stock prepared in water. The substrate for the reaction was dissolved in DMSO at a concentration of 40 g / L and dispensed in the reaction at a final concentration of 2 g / L. The final volume of the reaction was 0.5 mL. The reaction was incubated at room temperature for a time between 2 and 16 h. The reaction was quenched by the addition of 1 volume of MeCN. The reaction was then centrifuged at 3220 ref for 5 min and the clear solution was then transferred to a new glass vial for analysis. The selection of enzymes used for the conversion is reported in Table E14:

[0387] Table E14:

[0388]

Claims

1. A method for preparing chiral pyrrolotriazole of formula I, Where X is a halogen atom, n is an integer of 1, 2 or 3, and therein a helical bond " ” represent" "or" "Or a mixture of enantiomers," The method includes processing a ketone of formula V, Where X and n are as described above, a) Reduction with a metal complex catalyst in the presence of a reducing agent, or b) Reduction with oxidoreductase To form the chiral alcohol of formula I.

2. The method according to claim 1, wherein the metal complex catalyst is a ruthenium or iridium complex catalyst.

3. The method according to claim 1 or 2, wherein the metal complex catalyst is selected from ruthenium or iridium catalyst complexes of the following formula. in, For each individual structure and independent of each other, R 1 C, independent of each other 1-6 -alkyl, C 4-6 -cycloalkyl, phenyl, or heteroaryl, optionally via one or more C 1-6 -alkyl or C 1-6 -Alkyl substitution; R 2 Independently, they are hydrogen and C. 1-6 -alkyl, C 4-6 -cycloalkyl, phenyl, or heteroaryl, optionally via one or more C 1-6 -alkyl or C 1-6 -Alkoxy substitution, or two R 2 Together they form a ring bridged by -(CH2)4- units; R 3 They are either hydrogen or C, independent of each other. 1-6 -alkyl, C 4-6 -cycloalkyl, phenyl, or heteroaryl, optionally via one or more C 1-6 -alkyl or C 1-6 -Alkyl substitution; R 4 Independently, they are hydrogen and C. 1-6 -alkyl, C 4-6 -cycloalkyl, phenyl, or heteroaryl, optionally via one or more C 1-6 -alkyl or C 1-6 -Alkoxy substitution, or two R 4 Together they form a group consisting of -O-(CH2). x -O- is the ring of unit bridging; R 5 Independently, they are hydrogen and C. 1-6 -alkyl, C 4-6 -cycloalkyl, phenyl, or heteroaryl, optionally via one or more C 1-6 -alkyl or C 1-6 -alkoxy substitution, or two adjacent R 4 With R 5 Together they form a ring bridged by -(CH)4- units or -O-(CH2)xO- units; R 6 They are either hydrogen or C, independent of each other. 1-6 -alkyl; X is a coordinated ligand or counter anion selected from the following: halogen, C 1-6 -alkoxy group, tetrahaloboron ion, tetra(3,5-bis(trihalo-C) 1-6 -alkyl)phenyl)borate ion, acetylacetonate ion, hexahalophosphate ion, p-toluenesulfonate ion, methanesulfonate ion or trihalomethanesulfonate ion. Y represents oxygen or -CH2-; x is 1, 2, or 3; And when Q 1 It is nitrogen and Q 2 When the carbon element is used, the dashed ring represents an aromatic ring; And when Q 1 and Q 2 When the sulfur content is 6, the dashed ring represents a cycloalkane ring.

4. The method according to any one of claims 1 to 3, wherein the metal complex catalyst is selected from metal catalyst complexes of formula Xh, Xk, Xm or Xn.

5. The method according to any one of claims 1 to 4, wherein the reducing agent is a mixture of formic acid and trialkylamine or hydrogen.

6. The method according to claim 5, wherein the reaction with the mixture of formic acid and trialkylamine occurs in the presence of an organic solvent and at a reaction temperature of 10°C to 100°C.

7. The method according to claim 5, wherein the reaction with hydrogen occurs in an organic solvent at a hydrogen pressure of 1 bar to 100 bar and at a reaction temperature of 10°C to 100°C.

8. The method according to claim 1, wherein the selected oxidoreductase has the potential to reduce the ketone of formula V and form a chiral pyrrolotriazol of formula I with an enantiomer excess of at least 90%, preferably at least 95%, and more preferably at least 98%.

9. The method according to claim 1 or 8, wherein the enzymatic reduction is carried out in the presence of NADH or NADPH as a cofactor.

10. The method according to any one of claims 1, 8 or 9, wherein the cofactor is regenerated using a co-substrate.

11. The method according to claim 10, wherein the co-substrate is a secondary alcohol, preferably 2-propanol.

12. The method according to any one of claims 1, 8 to 11, wherein the enzymatic reduction is carried out in an aqueous medium at a temperature of 10°C to 50°C in the presence of an organic co-solvent.

13. The method according to claim 1, wherein the ketone of formula V can be prepared by: a) The 3,5-dihalo-1H-1,2,4-triazole of formula II, Where X is a halogen atom, through... a 1 Michael addition using acrylate IIIa, Where R is C 1-4 alkyl, or through a 2 Alkylation of alkyl esters of halogenated carboxylic acids of formula IIIb Where R is C 1-4 Alkyl group, where n is an integer of 1, 2, or 3 and X is a halogen. It is converted into the 3,5-dihalogen 1,2,4-triazole-carboxylic acid ester of formula IV. Where R, n, and X are as described above; and b) The 3,5-dihalogen 1,2,4-triazole-carboxylic acid ester of formula IV undergoes ring closure with an organometallic reagent to form a ketone of formula V. Where X and n are as described above.

14. A chiral pyrrolotriazole of formula I, Where X is a halogen atom and n is an integer of 1, 2 or 3, and the helical bond... " ” represent" "or" "Or a mixture of enantiomers." 15. The chiral pyrrolotriazol according to claim 14, wherein X is chlorine or bromine, preferably bromine, and n is 1, 2, or 3, and the helical bond... "represent" "or" "Or a mixture of enantiomers."

Images