HK1094189A - Aminopropanol derivatives as sphingosine-1-phosphate receptor modulators - Google Patents

Description

Aminopropanol derivatives as sphingosine-1-phosphate receptor modulators

The present invention relates to hydro-propanol derivatives, processes for their preparation, their use and pharmaceutical compositions containing them.

More specifically, the present invention provides compounds of formula I, in free form or in salt form:

wherein:

R₁is C_1-6An alkyl group; by hydroxy, C_1-2Alkoxy or C substituted by 1 to 6 fluorine atoms_1-6An alkyl group; c_2-6An alkenyl group; or C_2-6An alkynyl group;

x is O, CH₂C ═ O or a direct bond；

R₂Is optionally substituted C_1-7Alkyl, optionally substituted C_1-7Alkenyl, optionally substituted C_1-7Alkynyl, optionally substituted C_3-6Cycloalkyl, optionally substituted phenyl or optionally substituted heteroaryl, wherein C is substituted_1-7Alkyl radical, C_1-7Alkenyl radical, C_1-7Alkynyl or C_3-6Cycloalkyl has 1 to 5 substituents selected from the group consisting of: hydroxy, halogen, C_1-4Alkyl radical, C_1-4Alkoxy, C substituted by 1 to 5 halogen atoms_1-4Alkoxy radical, C_3-6Cycloalkyl radical, C_3-6Cycloalkoxy, C_3-6Cycloalkyl radical C_1-5Alkyl radical, C_3-6Cycloalkoxy group C_1-5Alkyl, cyano, optionally substituted phenyl, optionally substituted phenyloxy, optionally substituted heteroaryl, optionally substituted heteroaryloxy, optionally substituted heterocyclyl optionally linked through oxygen;

and wherein the phenyl, phenyloxy, heteroaryl, heteroaryloxy, heterocyclyl optionally linked through oxygen may be substituted independently of each other by 1 to 5 substituents selected from the group consisting of: hydroxy, halogen, C_1-4Alkyl, C substituted by 1 to 5 fluorine atoms_1-4Alkyl radical, C_1-4Alkoxy, C substituted by 1 to 5 fluorine atoms_1-4Alkoxy, cyano, phenyl and phenyl substituted with 1 to 5 substituents selected from the group consisting of: hydroxy, halogen, C_1-4Alkyl, C substituted by 1 to 5 fluorine atoms_1-4Alkyl radical, C_1-4Alkoxy, C substituted by 1 to 5 fluorine atoms_1-4Alkoxy and cyano; or phenyl, phenyloxy, heteroaryl, heteroaryloxy, heterocyclyl which are optionally linked via oxygen, may be fused independently of one another with a heterocyclyl;

R₃is Z-X₂Wherein Z is CH₂CHF or CF₂And X₂Is OH or a group of the formula (a)

Wherein Z₁Is a direct bond, CH₂、CHF、CF₂Or O, and R₆And R₇Independently of one another, H, C optionally substituted by 1, 2 or 3 halogen atoms_1-4Alkyl or benzyl;

and R is₄And R₅Independently of one another, H, C optionally substituted by 1, 2 or 3 halogen atoms_1-4An alkyl or acyl group;

the fused rings a and b are independently of each other C_5-6Cycloalkyl, aryl, heterocyclyl or heteroaryl.

The alkyl group or alkyl moiety may be straight or branched. When the alkyl group is substituted with a hydroxyl group, it is preferably substituted on the terminal carbon atom. The alkenyl group may be, for example, a vinyl group. Alkynyl may be, for example, propyn-2-yl. The acyl group may be R-CO, wherein R is C_1-6Alkyl radical, C_3-6Cycloalkyl, phenyl or phenyl C_1-4An alkyl group. Halogen may be fluorine, chlorine or bromine, preferably fluorine or chlorine. Aryl may be phenyl or naphthyl.

Heteroaryl may be a 5 to 8 membered aromatic ring containing 1 to 4 heteroatoms selected from N, O and S, for example pyridyl, pyrimidinyl, pyrazinyl, furyl, oxazolyl, isoxazolyl, thienyl, thiazolyl, isothiazolyl, pyrrolyl, imidazolyl or pyrazolyl.

Heterocyclyl means a 3 to 8, preferably 5 to 8 membered saturated or unsaturated heterocyclic ring containing 1 to 4 heteroatoms selected from N, O and S, such as tetrahydrofuranyl, tetrahydropyranyl, aziridinyl, piperidinyl, pyrrolidinyl, piperazinyl.

Examples of fused rings a and b are, for example, naphthyl; a benzoxazolyl group; a benzothiazolyl group; a benzofuranyl group; an indolyl group; (ii) an indazolyl group; or N-substituted-indazolyl radicals such as N-C_1-4Alkyl-indazolyl, or e.g. N-aryl-indazolyl, wherein aryl is optionally substituted aryl such as benzyl or benzyl optionally substituted by e.g. methoxy or nitro.

The compounds of formula I may be present in free form or in salt form, for example as addition salts with, for example, inorganic acids, such as hydrochloride, hydrobromide or sulfate, and organic acids, such as acetate, fumarate, maleate, benzoate, citrate, malate, methanesulfonate or benzenesulfonate. Compounds of formula I and salts thereof in the form of hydrates or solvates are also part of the invention.

When the compound of formula I has an asymmetric center in the molecule, various optical isomers can be obtained. The invention also includes enantiomers, racemates, diastereomers and mixtures thereof. For example, with R₁、R₃And NT₄R₅The central carbon atom of (a) may have the R or S configuration. Compounds having the following three-dimensional configuration are generally preferred.

Further, when the compounds of formula I include geometric isomers, the present invention includes cis-compounds, trans-compounds, and mixtures thereof. Similar considerations apply to starting materials containing the above-mentioned asymmetric carbon atoms or unsaturated bonds, such as the compounds of formulae II, III or IV shown below.

In the compounds of the formula I, the following meanings are preferred, alone or in any subcombination:

x is O or a direct bond;

2.R₁is CH₃Or CH₂-OH；

3.R₂Is C_1-7Alkyl, substituted C_1-7Alkyl, substituted phenyl, phenylalkyl, substituted phenylalkyl, biphenyl, substituted biphenyl, heteroaryl or substituted heteroaryl,

e.g. C_3-7Alkyl or C substituted by 1 to 5 fluorine atoms_3-7An alkyl group;

e.g. phenyl or substituted phenyl, e.g. substituted by hydroxy, C_1-4Alkyl radicals such as trifluoromethyl, C_1-4Alkoxy, halogen or cyano-substituted phenyl;

e.g. biphenyl or substituted biphenyl, e.g. by C_1-4Alkyl, substituted C_1-4Alkyl radicals such as trifluoromethyl, C_1-4Alkoxy, halogen or cyano substituted biphenyl;

e.g. substituted C_1-4Alkyl such as trifluoromethyl, cyano or phenyl substituted heteroaryl;

for example thienyl substituted by phenyl or furyl substituted by phenyl;

4.R₃is CH₂-OH or CH₂-OPO₃H₂；

5.R₄And R₅Are both hydrogen;

a and b are independently of each other aryl or heteroaryl, preferably a and b together form 2, 6-or 2, 7-disubstituted naphthyl, 2, 5-or 2, 6-disubstituted benzoxazolyl, 2, 5-or 2, 6-disubstituted benzothiophenyl, 2, 5-or 2, 6-disubstituted benzofuranyl, 1, 4-or 1, 5-disubstituted indolyl, 3, 6-indazolyl or 3, 6-N-substituted-indazolyl, for example 3, 6-N-methyl-indazolyl.

The present invention also includes a process for preparing a compound of formula I, comprising:

a) for R in the formula₃Is Z-X₂，X₂A compound of formula I which is OH or a group of formula (a), removing the protecting group present in the compound of formula II:

x, R therein₁、R₂And R₄R 'as defined above'₃Is Z-X₂Wherein X is₂Is OH and R'₅Is an amino protecting group, or

b) For R in the formula₃Is Z-X₂，X₂Is a radical of formula (a) and wherein R₆And R₇A compound of formula I which is H, removing the protecting group present in the compound of formula III:

x, R therein₁、R₂、R₄And R'₅As defined above, and R "₃Is Z-X₂Wherein X is₂Is a residue of formula (a'):

wherein Z₁And R 'is as defined above'₆Or R'₇Is a hydrolyzable or hydrogenolyzable group, or R'₆And R'₇Together form a divalent bridging group optionally fused to a ring (e.g., a benzene ring),

and, if desired, converting the compound of formula I obtained in free form into the desired salt form and vice versa.

Process step a) can be carried out according to methods known in the art. Removal of the amino protecting group may conveniently be carried out according to methods known in the art, for example by hydrolysis, for example in an acidic medium such as using hydrochloric acid. Examples of amino protecting Groups are, for example, "Protective Groups in organic Synthesis" T.W.Greene, J.Wiley & Sons NY, 2 nd edition, Chapter 7, 1991 and those disclosed in the references therein, such as benzyl, p-methoxybenzyl, methoxymethyl, tetrahydropyranyl, trialkylsilyl, acyl, tert-butoxy-carbonyl, benzyloxycarbonyl, 9-fluorenylmethoxycarbonyl, trifluoroacetyl and the like.

In the residue of formula (a '), R'₆Or R'₇Can have the meaning of alkyl tert-butyl, phenyl or benzyl, for example, or together form a ring system, such as 1, 5-dihydro-2, 4, 3-benzodioxan *.

Process step b) may be carried out according to methods known in the art, for example by hydrolysis, for example when R'₆And R'₇Is hydrolyzed in basic medium, e.g. using a hydroxide such as barium hydroxide, or when R'₆And R'₇Is tert-butyl and is hydrolyzed in an acidic medium. It may also be carried out by hydrogenolysis, for example in the presence of a catalyst such as Pd/C, followed by hydrolysis, for example in an acidic medium such as HCl. The compounds of formulae V and VI and their salts used as starting materials are also novel and are part of the present invention.

The invention also includes a process for preparing a compound of formula II wherein X is O, comprising alkylating a compound of formula IV:

wherein R is₁、R’₃、R₄And R'₅As defined above, the above-mentioned,

to introduce the desired group-R by alkylation₂. Alkylation of the compound of formula IV may be carried out according to methods known in the art, for example by nucleophilic substitution, for example by reaction with an alkylating agent R₂-X₃Reaction of in which X₃Is methanesulfonate, toluenesulfonate, trifluoromethanesulfonate, p-nitrobenzenesulfonate or halogen, for example chlorine, bromine or iodine. Alkylation can also be achieved by using R according to the Mitsunobu scheme₂OH (e.g.as described in Hughes, Organic Preparation and Preparation International 28, 127- & 164, 1996 or D.L.Hughes, org.React.42, 335, 1992) in solution or according to solid phase support synthesis, for example by linking a compound of formula IV to a solid phase support synthesisOn a resin. Alternatively, it can be carried out using, for example, triphenylphosphine or diethyl azoformate bonded to a resin such as polystyrene.

Without specific description of the preparation of the starting materials, these compounds are known or can be prepared by methods analogous to those known in the art or as disclosed in the examples below.

The following examples are illustrative of the invention. Melting point (Mp) is uncorrected.

RT ═ room temperature

THF ═ tetrahydrofuran

AcOEt ═ ethyl acetate

DCM ═ dichloromethane

Example 1: (R) -2-amino-2-methyl-4- (6-pentoxynaphthalen-2-yl) butan-1-ol

N-Boc- (R) -2-amino-2-methyl-4- (6-pentoxynaphthalen-2-yl) butan-1-ol (21mg, 0.05mmol) was dissolved in a solution of hydrochloric acid in methanol and stirred at room temperature for 2 hours. The solvent was distilled off in vacuo and the residue was dried to give the title compound in the form of its hydrochloride salt. NMR (CDCl)₃/d₆DMSO＝2/1)δ7.65(d，J＝8.5Hz，1H)，7.63(d，J＝8.1Hz，1H)，7.57(s，1H)，7.32(d，J＝8.3Hz，1H)，7.07-7.17(m，2H)，4.06(t，J＝6.7Hz，2H)，3.71(s，2H)，2.78-2.88(m，2H)，2.0-2.1(m，2H)，1.8-1.9(m，2H)，1.38-1.53(m，4H)，1.41(s，3H)，0.95(t，J＝7Hz，3H)。ESI+MS：m/z＝316.5(MH)⁺。

N-Boc- (R) -2-amino-2-methyl-4- (6-pentoxynaphthalen-2-yl) butan-1-ol can be synthesized as follows:

protecting N-BocA mixture of (R) -2-amino-2-methyl-4- (6-hydroxynaphthalen-2-yl) butan-1-ol (485mg, 1.17mmol), potassium carbonate (300mg, 2.17mmol), 1-iodopentane (184. mu.l, 1.2mmol) and acetone (6ml) was heated under reflux overnight. The solvent was distilled off and the residue partitioned between water and AcOEt. The organic layer was dried over sodium sulfate and concentrated in vacuo. Silica gel chromatography (cyclohexane/AcOEt ═ 2/1) afforded the title compound as a colorless oil. NMR (CDCl)₃)δ7.65(d，J＝8.8Hz，1H)，7.64(d，J＝8.3Hz，1H)，7.55(s，1H)，7.29(dd，J＝1.6+8.3Hz，1H)，7.12(dd，J＝2.5+8.8Hz，1H)，7.09(d，J＝2.5Hz，1H)，4.65(s，1H)，4.06(t，J＝6.6Hz，2H)，3.74(d，J＝11.5Hz，1H)，3.67(d，J＝11.5Hz，1H)，2.67-2.86(m，2H)，2.08-2.18(m，1H)，1.9-2.01(m，1H)，1.8-1.89(m，2H)，1.36-1.55(m，4H)，1.43(s，9H)，1.26(s，3H)，0.95(t，J＝7.2Hz，3H)。¹H NMR(CDCl₃/d₆DMSO＝2/1)：7.65(d，J＝8.5Hz，1H)，7.63(d，J＝8.1Hz，1H)，7.57(s，1H)，7.32(d，J＝8.3Hz，1H)，7.07-7.17(m，2H)，4.06(t，J＝6.7Hz，2H)，3.71(s，2H)，2.78-2.88(m，2H)，2.0-2.1(m，2H)，1.8-1.9(m，2H)，1.38-1.53(m，4H)，1.41(s，3H)，0.95(t，J＝7Hz，3H)。

Examples 2 to 9: the examples shown in Table 1 were prepared as described in example 1.

Table 1:

		2H)，3.39-3.54(m，2H)，2.72(t，J＝8.7Hz，2H)，2.38-2.53(m，2H)，1.78-2.04(m，4H)，1.23(s，3H)
			9	-CH2-cyclohexyl radical	7.83(br.s.，3H)，7.73(d，J＝8.3Hz，1H)，7.72(d，J＝8.9Hz，1H)，7.59(s，1H)，7.31(dd，J＝8.4+1.6Hz，1H)，7.25(d，J＝2.4Hz，1H)，7.12(dd，J＝8.9+2.4Hz，1H)，5.51(t，J＝5Hz，1H)，3.86(d，J＝6.3Hz，2H)，3.38-3.54(m，2H)，2.71(t，J＝8.7Hz，2H)，1.62-1.94(m，8H)，1.02-1.32(m，5H)，1.22(s，3H)

Example 10: 2-amino-2- [2- (6-pentyloxy-naphthalen-2-yl) -ethyl ] -propane-1, 3-diol

2, 2-bis (hydroxymethyl) -3- (6-pentoxynaphthalen-2-yl) -propargylamine (53mg, 0.12mmol) was dissolved in AcOEt, palladium on carbon (10mg) and hydrogen were added and stirred under a hydrogen atmosphere for 5 hours. The mixture was filtered through celite and the solvent was evaporated to give the pure title compound.¹H NMR(d₆DMSO, free base): 7.70(d, J ═ 9.2Hz, 1H), 7.68(d, J ═ 9.9Hz, 1H), 7.33(s, 1H), 7.29(dd, J ═ 8.4+1.6Hz, 1H), 7.22(d, J ═ 2.4Hz, 1H), 7.08(dd, J ═ 8.9+2.5Hz, 1H), 4.44(t, J ═ 5Hz, 1H), 4.04(t, J ═ 6.6Hz, 2H), 3.28-3.35(m, 4H), 2.67-2.73(m, 2H), 1.7-1.8(m, 2H), 1.52-1.6(m, 2H), 1.3-1.48(m, 4H), 0.89(t, J ═ 7.1, 3H).

2, 2-bis (hydroxymethyl) -3- (6-pentoxynaphthalen-2-yl) -propargylamine can be synthesized as follows:

n-acetyl-2, 2-bis (acetoxymethyl) -3- (6-pentoxynaphthalen-2-yl) -propargylamine (33mg, 0.07mmol) was dissolved in methanol and treated with excess 1N NaOH. The mixture was heated to reflux for 5 hours and then acidified with 1N HCl. The precipitated hydrochloride salt was collected by filtration, washed with water and obtained in pure form. The free base can be obtained by treatment with base and extraction with AcOEt. Mp (HCl)199 deg.C; mp (free base) 144-147 ℃.

N-acetyl-2, 2-bis (acetoxymethyl) -3- (6-pentoxynaphthalen-2-yl) -propargylamine can be synthesized as follows:

2-bromo-6-pentoxynaphthalene (84mg, 0.3mmol), N-acetyl-2, 2-bis (acetoxymethyl) -propynylamine (69mg, 0.3mmol), tetrakis-triphenylphosphine palladium (17mg, 0.01mmol), cuprous iodide (5mg, 0.03mmol) and triethylamine (0.12. mu.l) were mixed in dry DMF (1ml) and heated under argon overnight at 100 ℃. After cooling, the mixture was partitioned between water and AcOEt and the organic layer was washed with brine, dried over magnesium sulfate and evaporated. Purification by silica gel chromatography (cyclohexane/AcOEt ═ 2/3) gave the pure title compound, mp 106-.

Example 11: 2-amino-2- [2- (4-heptyloxy-naphthalen-1-yl) -ethyl ] -propane-1, 3-diol

The title compound was prepared according to the procedure described for example 10 using the appropriate starting materials.¹HNMR(CDCl₃Free base): 8.33(d, J ═ 8.2Hz, 1H), 7.96(d, J ═ 8.5Hz, 1H), 7.43-7.56(m, 2H), 7.21(d, J ═ 7.8Hz, 1H), 6.71(d, J ═ 7.8Hz, 1H), 4.1(t, J ═ 6.4Hz, 2H), 3.69(d, J ═ 10.6Hz, 2H), 3.6(d, J ═ 10.6Hz, 2H), 2.98-3.07(m, 2H), 1.78-1.96(m, 4H), 1.5-1.62(m, 2H), 1.35-1.46(m, 6H)，0.9(t，J＝7Hz，3H)。

Example 12: phosphoric acid mono- [ (R) -2-amino-2-methyl-4- (6-pentoxynaphthalen-2-yl) but-1-yl ] ester

N-Boc protected (R) -2-amino-2-methyl-4- (6-pentoxynaphthalen-2-yl) butan-1-ol (60mg, 0.14mmol) was dissolved in dry THF and treated with tetrazole (30mg, 0.42mmol) and di-tert-butyl-N, N-diisopropylphosphoramide (78mg, 0.28mmol) at room temperature. The mixture was stirred under argon overnight, then aqueous hydrogen peroxide (160 μ l of 30% aqueous solution, 1.4mmol) was added. After 1 hour, the mixture was treated with excess aqueous sodium thiosulfate and then extracted with AcOEt. The combined organic layers were washed with water and brine, dried over sodium sulfate and concentrated in vacuo. Silica gel chromatography (cyclohexane/AcOEt ═ 2/1) gave pure di-tert-butyl phosphate (R) -2-amino-2-methyl-4- (6-pentoxynaphthalen-2-yl) but-1-yl ester: NMR (CDCl)₃) δ 7.58(d, J ═ 8.7Hz, 1H), 7.56(d, J ═ 8.4Hz, 1H), 7.47(s, 1H), 7.22(dd, J ═ 8.4+1.7Hz, 1H), 6.99-7.06(m, 2H), 4.79(br.s, 1H), 4.02(dd, J ═ 10+5.5Hz, 1H), 3.99(t, J ═ 6.6Hz, 2H), 3.82(dd, J ═ 10+5.5Hz, 1H), 2.6-2.72(m, 2H), 2.04-2.17(m, 1H), 1.88-1.98(m, 1H), 1.72-1.83(m, 2H), 1.43(s, 18H), 1.37(s, 9.9, 1H), 1.88-1.31 (t, 1H), 1.31.31H, 1H), 1.31.31 (t, 1H). The intermediate (54mg, 0.089mmol) was stirred with saturated methanol hydrochloride solution at room temperature overnight. The solvent was distilled off, and the residue was purified by HPLC to give mono- [ (R) -2-amino-2-methyl-4- (6-pentoxynaphthalen-2-yl) but-1-yl phosphate as a colorless powder]And (3) an ester. Mp 266-.¹HNMR(CD₃OD/DCl＝10/1)：7.7(d，J＝9Hz，1H)，7.68(d，J＝9.3Hz，1H)，7.62(s，1H)，7.34(dd，J＝8.4+1.8Hz，1H)，7.17(d，J＝2.5Hz，1H)，7.1(dd，J＝9+2.5Hz，1H)，4.15(dd，J＝11.1+5.1Hz，1H)，4.11(t，J＝6.5Hz，2H)，4.04(dd，J＝11.1+4.6Hz，1H)，2.78-2.9(m，2H)，1.98-2.16(m，2H)，1.8-1.87(m，2H)，1.39-1.55(m，4H)，1.45(s，3H)，0.96(t，J＝7.2Hz，3H)。

Examples 13 and 14: the examples shown in Table 2 were prepared according to the method described in example 12.

Table 2:

example 15: (R) -2-amino-2-methyl-4- (2-pentyl-benzoxazol-5-yl) -butan-1-ol

The method A comprises the following steps:

reacting [ (R) -1-hydroxymethyl-1-methyl-3- (2-pentyl-benzoxazol-5-yl) -propyl]A solution of tert-butyl carbamate (31mg, 0.079mmol) in diethyl ether (2ml) was treated with a solution of 2M HCl in diethyl ether and stirred at room temperature for 2 hours. The reaction was stopped by the addition of 28% aqueous ammonium hydroxide (2ml), methanol (2ml) and DCM (1 ml). After evaporation of the solvent in vacuo, the residue is purified by chromatography on silica gel (eluent: CH)₂Cl₂/MeOH/28％NH₄OH: 9/1/0.1). Crystallization from n-pentane gave the title compound as a colorless solid.¹H-NMR(DMSO-d₆)δ：7.52(d，J＝7.9Hz；1H)，7.45(s；1H)，7.15(d，J＝7.9Hz；1H)，3.30(bs；2H)，2.88(bs；2H)，2.77-2.55(m；2H)，1.78(bs；2H)，1.63(bs；2H)，1.32(bs；4H)，1.03(s；3H)，0.85(bs；3H)。

Preparation of [ (R) -1-hydroxymethyl-1-methyl-3- (2-pentyl-benzoxazol-5-yl) -propyl ] -carbamic acid tert-butyl ester

A solution of [ (R) -3- (3-amino-4-hydroxy-phenyl) -1-hydroxymethyl-1-methyl-propyl ] -carbamic acid tert-butyl ester (500mg, 1.61mmol) and ethyl hexanoimidate hydrochloride (318mg, 1.77mmol) in DCM (6ml) was stirred at room temperature for 16 h. The solvent was evaporated in vacuo and then chromatographed on silica gel (eluent: DCM/methanol 40/1). The product containing fractions were combined and then evaporated in vacuo. Crystallization from pentane gave the title compound as a colorless solid. mp: 92.7/92.6 ℃.

Preparation of [ (R) -3- (3-amino-4-hydroxy-phenyl) -1-hydroxymethyl-1-methyl-propyl ] -carbamic acid tert-butyl ester

An argon-purified solution of [ (R) -1-hydroxymethyl-3- (4-hydroxy-3-nitro-phenyl) -1-methyl-propyl ] -carbamic acid tert-butyl ester (3.02g, 8.87mmol) in ethanol (40ml) was treated with 10% Pd on charcoal (0.5 g). The argon was replaced with hydrogen and the reaction was carried out at normal pressure for 2 hours. The reaction suspension was filtered and the filtrate was evaporated to dryness in vacuo. Chromatography on silica gel (eluent: DCM/methanol 20/1 to 10/1) followed by crystallization from diethyl ether afforded the title compound as colorless crystals. mp 122.1/123.0 ℃.

Preparation of [ (R) -1-hydroxymethyl-3- (4-hydroxy-3-nitro-phenyl) -1-methyl-propyl ] -carbamic acid tert-butyl ester

Reacting [ (R) -1-hydroxymethyl-3- (4-hydroxy-phenyl) -1-methyl-propyl]-solution of tert-butyl carbamate (5.298g, 17.936mmol) in dry ethanol (27ml) with Fe (NO)₃)₃·9H₂O (5.797g, 14.348mmol) and stirring at 40 ℃ for 2 hours changed the color from dark blue to reddish brown. After cooling to room temperature, the reaction mixture was partitioned between 1N HCl (200ml) and DCM (200 ml). The aqueous layer was washed twice with DCM and the combined organic layers were MgSO₄Dried and concentrated in vacuo. Flash chromatography (eluent: DCM/methanol 40/1) afforded the title compound as a light yellow amorphous solid.¹H-NMR(CDCl₃)δ：10.45(s；1H)，7.92(d，J＝2.2Hz；1H)，7.45(dd，J＝2.2，8.6Hz；1H)，7.09(d，J＝8.6Hz；1H)，3.68(m；2H)，2.73-2.52(m；2H)，2.10-1.8(m；2H)，1.45(s；9H)，1.23(s；3H)。

The method B comprises the following steps:

sodium borohydride (262mg, 6.925mmol) was suspended in dry ethanol (7ml), cooled to-10 ℃ with CaCl₂(284mg，3.4625mmol) and stirred for 45 minutes while warming to 0 ℃. A solution of (R) -2-amino-2-methyl-4- (2-pentyl-benzoxazol-5-yl) -butyric acid methyl ester (147mg, 0.462mmol) in dry ethanol (2ml) was added to the reaction slurry and stirred at 8-10 ℃ for 2 hours. The precipitate was collected through a sintered funnel and washed with ethanol. The filtrate and the ethanol washings were combined and then concentrated in vacuo. The residue was partitioned between 1N aqueous NaOH and chloroform. The organic layer was washed with MgSO₄Dried and concentrated in vacuo. Chromatography on silica gel (eluent: CH)₂Cl₂/MeOH/28％NH₄OH: 9/1/0.1) gave the title compound as an amorphous colorless solid.

Preparation of (R) -2-amino-2-methyl-4- (2-pentyl-benzoxazol-5-yl) -butyric acid methyl ester

A suspension of (R) -2-amino-4- (3-hexanoylamino-4-hydroxy-phenyl) -2-methyl-butyric acid methyl ester trifluoroacetate salt (460mg, 1.02mmol) in dry toluene is heated at 200 ℃ for 10 min under pressure (microwave Emrys optizer). After cooling to room temperature, the reaction mixture was taken up in AcOEt and saturated NaHCO₃The aqueous solution was partitioned. Chromatography on silica gel (eluent: CH)₂Cl₂MeOH 10/1) gave the title compound as an amorphous colorless solid.¹H-NMR(DMSO-d₆)：7.51(d，J＝8.3Hz；1H)，7.42(d，J＝1.2Hz；1H)，7.12(dd，J＝8.3，1.2Hz；1H)，3.60(s；3H)，2.88(t，J＝7.4Hz；2H)，2.71-2.63/2.58-2.50(m；2H)，1.92(bs；2H)，1.90-1.82/1.80-1.73(m；2H)，1.43-1.28(m；4H)，1.22(s；3H)，0.85(m；3H)。

Preparation of (R) -2-amino-4- (3-hexanoylamino-4-hydroxy-phenyl) -2-methyl-butyric acid methyl ester trifluoroacetate salt

A solution of (S) -2-isopropyl-3, 6-dimethoxy-5-methyl-2, 5-dihydro-pyrazine (652.5. mu.l, 3.291mmol) in dry THF (7ml) at-70 ℃ was treated with a solution of n-butyllithium in n-hexane (2.1ml of a 1.6M solution, 3.29mmol) under argon. After stirring for 10 min, a solution of 5- (2-iodo-ethyl) -2-pentyl-benzoxazole (753mg, 2.19mmol) in dry THF (5ml) was added and the reaction was mixedThe mixture was stirred at-65 ℃ for 90 minutes and then at 0-5 ℃ for 2 hours. The reaction was partitioned between AcOEt and saturated aqueous ammonium chloride. The organic layer was washed with MgSO₄Dried and concentrated in vacuo. Flash chromatography on silica gel (pentane/diethyl ether 4/1) afforded the crude product 5- [2- ((2R, 5S) -5-isopropyl-3, 6-dimethoxy-2-methyl-2, 5-dihydro-pyrazin-2-yl) -ethyl]-2-pentyl-benzoxazole. The crude product was dissolved in acetonitrile (21ml), treated with water (21ml) and trifluoroacetic acid (450 μ l) and stirred at room temperature for 3 days. The solvent was removed in vacuo and reverse phase chromatography (Waters, C-18, eluent: water/acetonitrile gradient with 0.1% trifluoroacetic acid) was performed to afford the title compound as an amorphous solid.¹H-NMR(CDCl₃)δ：8.50(s；1H)，6.98(s；1H)；8.86/6.83(2br s；2H)，3.71(s；3H)，2.75-2.35(m；4H)，2.20(br s；2H)，1.70(br s；2H)，1.61(s；3H)，1.40-1.20(m；4H)，0.90(m；3H)。

Preparation of 5- (2-iodo-ethyl) -2-pentyl-benzoxazole

A0 ℃ solution of 2- (2-pentyl-benzooxazol-5-yl) -ethanol (1.63g, 6.99mmol) in dry THF (49ml) was treated with triphenylphosphine (2.018b, 7.693mmol) and imidazole (1.05g, 15.38mmol) under argon. After complete dissolution, iodine (1.95g, 7.69mmol) was added in small portions so that the reaction temperature did not exceed 4 ℃. After stirring at 0 ℃ for 3 hours, the reaction mixture was partitioned between diethyl ether and saturated aqueous ammonium chloride. The organic layer was evaporated in vacuo to form a suspension in diethyl ether, filtered and the filtrate was concentrated in vacuo. Chromatography on silica gel (eluent: pentane/diethyl ether 4/1) followed by crystallization from pentane gives the title compound as colorless crystals. mp: 47.7/48.7 ℃.

Preparation of 2- (2-pentyl-benzoxazol-5-yl) -ethanol

A solution of (2-pentyl-benzoxazol-5-yl) -acetic acid (2.11g, 8.53mmol) in triethyl orthoacetate (4.67ml, 25.6mmol) was heated under pressure (in three batches in a microwave Emrys optizer) at 180 ℃ for 8 min. After cooling to room temperature, the reaction mixture was partitioned between AcOEt and saturated aqueous sodium bicarbonate. Will haveThe organic layer was washed twice with water and MgSO₄Dried and concentrated in vacuo. 1.88g of crude (2-pentyl-benzoxazol-5-yl) -acetic acid ethyl ester (2.31g, 99%) was dissolved in dry ethanol (43ml) and added to Ca (BH)₄)₂Prepared from a suspension of sodium borohydride (3.87g, 102.4mmol) in dry ethanol (130ml), cooled to-10 ℃ with CaCl₂(5.68g, 51.2mmol) and stirred for 45 minutes while warming to 0 deg.C). After stirring at 8 ℃ for 2 hours, the reaction mixture was filtered and the precipitate was washed with ethanol. The filtrate was combined with the washings and then concentrated in vacuo, treated with MeOH and concentrated again. Chromatography on silica gel (eluent: DCM/methanol 10/1) gave the title compound as a colorless solid.¹H-NMR(DMSO-d₆)δ：7.51(d，J＝8.1Hz；1H)，7.49(s；1H)，7.16(dd，J＝8.3，1.6Hz；1H)，4.60(t，J＝5.2Hz；1H)，3.61(m；2H)，2.88(t，J＝7.4Hz；2H)，2.80(t，J＝7.0Hz；2H)，1.76(m；2H)，1.30(m；4H)，0.85(m；3H)。

Preparation of (2-pentyl-benzoxazol-5-yl) -acetic acid

A solution of (3-amino-4-hydroxy-phenyl) -acetic acid (4.65g, 27.8mmol) in methanol (79ml) was treated with ethyl hexamethyleneimidate hydrochloride (5.00g, 27.8mmol) and stirred at room temperature for 14 h. The solvent was evaporated in vacuo and the residue was purified by chromatography on silica gel (eluent: DCM/methanol 10/1). Crystallization from cyclohexane/n-pentane gave the title compound as colorless crystals. mp: 58.2/57.1 ℃.

Example 16: (R) -2-amino-2-methyl-4- (2-phenyl-benzooxazol-5-yl) -butan-1-ol

The procedure described in example 15 (method A) was followed using [ (R) -1-hydroxymethyl-1-methyl-3- (2-phenyl-benzooxazol-5-yl) -propyl]Tert-butyl carbamate gives the title compound as a colorless solid. mp.127.9-128.1 deg.C.¹H-NMR(DMSO-d₆)δ：8.18(m；2H)，7.66(d，J＝8.4Hz；1H)，7.60(m；3H)，7.52(d，J＝8.4Hz；1H)，4.57(s；1H)，3.15(s；2H)，2.72(m；2H)，1.58(m；2H)，0.97(s；3H)。

Preparation of [ (R) -1-hydroxymethyl-1-methyl-3- (2-phenyl-benzooxazol-5-yl) -propyl ] -carbamic acid tert-butyl ester

The title compound was prepared according to the procedure described for example 15 using ethyl benzimidate hydrochloride. mp: 205.8/204.7 ℃.

Example 17: (R) -2-amino-2-methyl-4- [2- (4-hydroxyphenyl) -benzooxazol-5-yl ] -butan-1-ol

The title compound was obtained as a colorless solid by the method described in example 15 (method a) using the appropriate starting materials. mp 229.3-231.2 ℃.¹H-NMR(DMSO-d₆)δ：8.00(d，J＝8.8Hz；2H)，7.62(d，J＝8.4Hz；1H)，7.55(s；1H)，7.18(d，J＝8.4Hz；1H)，6.97(d，J＝8.8Hz；2H)，5.51(s；1H)，3.47(m；2H)，2.73(m；2H)，1.85(m；2H)，1.22(s；3H)。

Preparation of tert-butyl { (R) -1-hydroxymethyl-1-methyl-3- [2- (4-hydroxyphenyl) -benzooxazol-5-yl ] } -3-carbamate

The title compound was prepared according to the procedure described for the preparation of example 15 using ethyl 4-hydroxy-benzimidinate hydrochloride. mp 136.5/133.2 ℃.

Example 18: (R) -2-amino-4- [2- (3-chloro-4-methyl-phenyl) -benzooxazol-5-yl ] -2-methyl-butan-1-ol

The title compound was obtained as a colorless solid by the method described in example 15 (method a) using the appropriate starting materials.¹H-NMR(DMSO-d₆)δ：8.13(s；1H)，8.03(d，J＝8.0Hz；1H)，7.64(d，J＝8.4Hz；1H)，7.58(m；2H)，7.26(d，J＝8.4Hz；1H)，4.56(s；1H)，3.15(s；2H)，2.72(m；2H)，2.42(s；1H)，1.57(m；2H)，0.97(s；3H)。

Preparation of { (R) -3- [2- (3-chloro-4-methyl-phenyl) -benzooxazol-5-yl ] -1-hydroxymethyl-1-methyl-propyl } -carbamic acid tert-butyl ester

The title compound was prepared according to the procedure described for example 15 using ethyl 3-chloro-4-methyl-benzimidinate hydrochloride.¹H-NMR(DMSO-d₆)δ：8.13(d，J＝1.7Hz；1H)，8.02(dd，J＝1.7，7.9Hz；1H)，7.65(d，J＝8.4Hz；1H)，7.59(d，J＝8.2Hz；1H)，7.59(d，J＝8.2Hz；1H)，7.56(s；1H)，7.24(dd，J＝1.7，8.4Hz；1H)，6.25(s；1H)，4.71(m；1H)，3.40(m；2H)，2.62(m；2H)，2.42(s；3H)，1.95(m；1H)，1.82(m；1H)，1.39(s；9H)，1.18(s；3H)。

Example 19: (R) -2-amino-4- [2- (4-butoxy-phenyl) -benzooxazol-5-yl ] -2-methyl-butan-1-ol

The title compound was obtained as a colorless solid by the method described in example 15 (method a) using the appropriate starting materials.¹H-NMR(DMSO-d₆)δ：8.09(d，J＝8.8Hz；2H)，7.60(d，J＝8.3Hz；1H)，7.52(s；1H)，7.18(d，J＝8.3Hz；1H)，7.12(d，J＝8.8Hz；2H)，4.56(s；1H)，4.07(t；J＝6.5Hz；2H)，3.15(s；2H)，2.70(m；2H)，1.72(m；2H)，1.56(m；2H)，1.42(m；2H)，0.95(s；3H)，0.92(t，J＝6.5Hz；3H)。

Preparation of { (R) -3- [2- (4-butoxy-phenyl) -benzooxazol-5-yl 1-1-hydroxymethyl-1-methyl-propyl } -carbamic acid tert-butyl ester

The title compound was prepared according to the procedure described for example 15 using ethyl 4-butoxy-benzimidate hydrochloride. mp 152.1/152.5 ℃.

Example 20: (R) -2-amino-4- [ ((E) -2-but-2-enyl) -benzooxazol-5-yl ] -2-methyl-but-1-ol

The title compound was obtained as a colorless solid by the method described in example 15 (method a) using the appropriate starting materials.¹H-NMR(DMSO-d₆)δ：7.51(d，J＝8.3Hz；1H)，7.43(s；1H)，7.15(d，J＝8.3Hz；1H)，5.66(m；2H)，4.57(s；1H)，3.63(m；2H)，3.16(s；2H)，2.68(m；2H)，1.68(s；3H)，1.54(m；2H)，0.96(s；3H)。

Preparation of { (R) -3- [ ((E) -2-but-2-enyl) -benzooxazol-5-yl ] -1-hydroxymethyl-1-methyl-propyl } -carbamic acid tert-butyl ester

The title compound was prepared according to the procedure described for the preparation of example 15 using ethyl (E) -pent-3-enimidoate hydrochloride.¹H-NMR(DMSO-d₆)δ：7.52(d，J＝8.3Hz；1H)，7.42(s；1H)，7.14(d，J＝8.3Hz；1H)，6.23(s；1H)，5.16(m；2H)，4.69(s；1H)，3.62(m；2H)，3.38(m；2H)，2.58(m；2H)，1.92/1.78(m；2H)，1.66(s；3H)，1.66(m；2H)，1.38(s；9H)，1.15(s；3H)。

Example 21: (R) -2-amino-2-methyl-4- (2-pent-4-ynyl-benzoxazol-5-yl) -butan-1-ol

The title compound was obtained as a colorless solid by the method described in example 15 (method a) using the appropriate starting materials.¹H-NMR(DMSO-d₆)δ：7.52(d，J＝8.3Hz；1H)，7.45(s；1H)，7.16(d，J＝8.3Hz；1H)，4.54(s；1H)，3.14(s；2H)，2.98(t，J＝7.3Hz；2H)，2.81(s；1H)，2.67(m；2H)，2.30(m；2H)，1.95(m；2H)，1.53(m；2H)，0.95(s；3H)。

Preparation of [ (R) -1-hydroxymethyl-1-methyl-3- (2-pent-4-ynyl-benzoxazol-5-yl) -propyl ] -carbamic acid tert-butyl ester

The title compound was prepared according to the procedure described for the preparation of example 15 using ethyl hex-5-ynimidate hydrochloride.¹H-NMR(DMSO-d₆)δ：7.52(d，J＝8.3Hz；1H)，7.43(s；1H)，7.14(d，J＝8.3Hz；1H)，6.23(s；1H)，4.69(s；1H)，3.38(s；2H)，2.98(t，J＝7.3Hz；2H)，2.80(s；1H)，2.58(m；2H)，2.30(m；2H)，1.95(m；2H)，1.92/1.78(m；2H)，1.37(s；9H)，1.16(s；3H)。

Example 22: (R) -2-amino-4- (2-cyclopropylmethyl-benzooxazol-5-yl) -2-methyl-butan-1-ol

The title compound was obtained as a colorless solid by the method described in example 15 (method a) using the appropriate starting materials.¹H-NMR(DMSO-d₆)δ：7.52(d，J＝8.3Hz；1H)，7.45(s；1H)，7.15(d，J＝8.3Hz；1H)，4.56(s；1H)，3.14(s；2H)，2.82(d，J＝7.0Hz；2H)，2.67(m；2H)，1.54(m；2H)，1.14(m；1H)；0.95(s；3H)，0.54(m；2H)，0.27(m；2H)。

Preparation of [ (R) -3- (2-cyclopropylmethyl-benzooxazol-5-yl) -1-hydroxymethyl-1-methyl-propyl ] -carbamic acid tert-butyl ester

The title compound was prepared according to the procedure described for the preparation of example 15 using ethyl 2-cyclopropyl-acetimidate hydrochloride.¹H-NMR(DMSO-d₆)δ：7.52(d，J＝8.3Hz；1H)，7.43(s；1H)，7.13(d，J＝8.3Hz；1H)，6.23(s；1H)，4.70(s；1H)，3.38(s；2H)，2.82(d，J＝7.0Hz；2H)，2.58(m；2H)，1.93/1.80(m；2H)，1.38(s；9H)，1.16(m；4H)；0.52(s；3H)，0.27(m；2H)。

Example 23: ((R) -2-amino-2-methyl-4- [2- (4-trifluoromethyl-phenyl) -benzooxazol-5-yl ] -butan-1-ol

The title compound was obtained as a colorless solid by the method described in example 15 (method a) using the appropriate starting materials.¹H-NMR(DMSO-d₆)δ：8.37(d，J＝8.2Hz；2H)，7.96(d，J＝8.2Hz；2H)，7.69(d，J＝8.3Hz；1H)，7.64(s；3H)，7.30(d，J＝8.3Hz；1H)，4.58(s；1H)，3.16(s；2H)，2.72(m；2H)，1.57(m；2H)；0.96(s；3H)。

{ (R) -1-hydroxymethyl-1-methyl-3- [2- (4-trifluoromethylphenyl) -benzooxazol-5-yl ] } -carbamic acid tert-butyl ester

The title compound was prepared according to the procedure described for the preparation of example 15 using ethyl 4-trifluoromethylbenzimidinate hydrochloride.¹H-NMR(DMSO-d₆)δ：8.37(d，J＝8.2Hz；2H)，7.96(d，J＝8.2Hz；2H)，7.71(d，J＝8.3Hz；1H)，7.62(s；3H)，7.29(d，J＝8.3Hz；1H)，6.26(s；1H)，4.72(s；1H)，3.40(s；2H)，2.63(m；2H)，1.95/1.83(m；2H)；1.39(s；9H)，1.18(s；3H)。

Example 24: (R) -2-amino-4- (2-biphenyl-4-yl-benzooxazol-5-yl) -2-methyl-butan-1-ol

The title compound was obtained as a colorless solid by the method described in example 15 (method a) using the appropriate starting materials.¹H-NMR(DMSO-d₆)δ：8.23(d，J＝8.2Hz；2H)，7.83(d，J＝8.2Hz；2H)，7.71(d，J＝7.4Hz；2H)，7.58(d，J＝8.0Hz；1H)，7.55(s；1H)，7.46(m；1H)，7.38(m；1H)，7.22(d；J＝8.4Hz；1H)，3.22(m；2H)，2.72(m；2H)，1.64(m；2H)，1.02(s；3H)。

Preparation of [ (R) -3- (2-biphenyl-4-yl-benzooxazol-5-yl) -1-hydroxymethyl-1-methyl-propyl ] carbamic acid tert-butyl ester:

the title compound was prepared according to the procedure described for the preparation of example 15 using ethyl biphenyl-4-carboximidic acid hydrochloride.¹H-NMR(DMSO-d₆)δ：8.25(d，J＝8.3Hz；2H)，7.92(d，J＝8.3Hz；2H)，7.77(d，J＝7.4Hz；2H)，7.68(d，J＝8.0Hz；1H)，7.58(s；1H)，7.51(m；1H)，7.43(m；1H)，7.23(d；J＝8.4Hz；1H)，6.25(s；1H)，4.72(s；1H)，3.40(m；2H)，2.64(m；2H)，1.96/1.83(m；2H)，1.89(s；9H)，1.18(s；3H)。

Example 25: (R) -2-amino-4- (2-benzo [1, 3] dioxol-5-yl-benzooxazol-5-yl) -2-methyl-butan-1-ol

The title compound was obtained as a colorless solid by the method described in example 15 (method a) using the appropriate starting materials.¹H-NMR(DMSO-d₆)δ：7.73(dd，J＝8.2，1.7Hz；1H)，7.60(s；1H)，7.59(d，J＝8.3Hz；1H)，7.54(s；1H)，7.20(d，J＝8.3Hz；1H)，7.12(d，J＝8.2Hz；1H)，6.15(s；2H)，4.55(s；1H)，3.15(s；2H)，2.70(m；2H)，1.57(m；2H)，0.95(s；3H)。

Preparation of [ (R) -3- (2-benzo [1, 3] dioxol-5-yl-benzooxazol-5-yl) -1-hydroxymethyl-1-methyl-propyl ] -carbamic acid tert-butyl ester:

the title compound was synthesized according to the procedure described for example 15 using 2-benzo [1, 3]]And dioxole-5-ethyl formimidate hydrochloride.¹H-NMR(DMSO-d₆)δ：7.73(dd，J＝8.2Hz，1.7Hz；1H)，7.61(s；1H)，7.59(d，J＝8.3Hz；1H)，7.50(s；1H)，7.18(d，J＝8.3Hz；1H)，7.12(d，J＝8.2Hz；1H)，6.24(s；1H)，6.15(s；2H)，4.71(s；1H)，3.39(m；2H)，2.62(m；2H)，1.95/1.81(2m；2H)，1.40(s；9H)，1.18(s；3H)。

Example 26: (R) -2-amino-4- [2- (3-ethoxy-phenyl) -benzooxazol-5-yl ] -2-methyl-butan-1-ol

The title compound was obtained as a colorless solid by the method described in example 15 (method a) using the appropriate starting materials.¹H-NMR(DMSO-d₆)δ：7.74(d，J＝7.8Hz；1H)，7.64(d，J＝8.3Hz；1H)，7.63/7.57(2s；2H)，7.49(t，J＝8.0Hz；1H)，7.23(d，J＝8.3Hz；1H)，7.16(m；1H)，4.56(s；1H)，4.13(q，J＝7.0Hz；2H)，3.16(s；2H)，2.71(m；2H)，1.57(m；2H)，1.36(t，J＝7.0Hz；3H)，0.96(s；3H)。

Preparation of { (R) -3- [2- (3-ethoxy-phenyl) -benzooxazol-5-yl ] -1-hydroxymethyl-1-methyl-propyl } -carbamic acid tert-butyl ester:

the title compound was prepared according to the procedure described for example 15 using ethyl 3-ethoxy-benzimidinate hydrochloride.¹H-NMR(DMSO-d₆)δ：7.74(d，J＝7.8Hz；1H)，7.65(d，J＝8.3Hz；1H)，7.64/7.55(2s；2H)，7.49(t，J＝8.0Hz；1H)，7.23(d，J＝8.3Hz；1H)，7.16(m；1H)，6.24(s；1H)，4.72(m；1H)，4.13(q，J＝7.0Hz；2H)，3.40(m；2H)，2.63(m；2H)，1.96/1.82(m；2H)，1.40(s；9H)，1.36(t，J＝7.0Hz；3H)，1.18(s；3H)。

Example 27: (R) -2-amino-2-methyl-4- [2- (4-phenoxy-phenyl) -benzooxazol-5-yl ] -butan-1-ol

Method (method) according to example 15Method A) the title compound is obtained as a colorless solid using the appropriate starting materials.¹H-NMR(DMSO-d₆)δ：8.16(d，J＝8.6Hz；2H)，7.62(d，J＝8.3Hz；1H)，7.55(s；1H)，7.46(m；2H)，7.23(m；2H)，7.14(m；4H)，4.57(s；1H)，3.15(s；2H)，2.71(m；2H)，1.57(m；2H)，0.96(s；3H)。

Preparation of { (R) -1-hydroxymethyl-1-methyl-3- [2- (4-phenoxy-phenyl) -benzooxazol-5-yl ] -propyl } -carbamic acid tert-butyl ester

The title compound was prepared according to the procedure described for the preparation of example 15 using ethyl 4-phenoxy-benzimidate hydrochloride.¹H-NMR(DMSO-d₆)δ：8.17(d，J＝8.6Hz；2H)，7.63(d，J＝8.3Hz；1H)，7.535(s；1H)，7.46(m；2H)，7.25(m；1H)，7.20(m；1H)，7.14(m；4H)，6.24(s；1H)，4.71(s；1H)，3.39(m；2H)，2.62(m；2H)，1.95/1.82(2m；2H)，1.39(s；9H)，1.17(s；3H)。

Example 28: (R) -2-amino-4- [2- (3, 5-bis-trifluoromethyl-phenyl) -benzooxazol-5-yl ] -2-methyl-butan-1-ol

The title compound was obtained as a colorless solid by the method described in example 15 (method a) using the appropriate starting materials.¹H-NMR(DMSO-d₆)δ：8.65(s；2H)，8.40(s；1H)，7.72(d；J＝8.3Hz；1H)，7.64(s；1H)，7.33(s，J＝8.3Hz；1H)，4.58(s；1H)，3.18(s；2H)，2.73(m；2H)，1.58(m；2H)，0.95(s；3H)。

Preparation of { (R) -3- [2- (3, 5-bis-trifluoromethyl-phenyl) -benzooxazol-5-yl ] -1-hydroxymethyl-1-methyl-propyl } -carbamic acid tert-butyl ester:

the title compound was prepared according to the procedure described for example 15 using ethyl 3, 5-bis-trifluoromethyl-benzimidinate hydrochloride.¹H-NMR(DMSO-d₆)δ：8.68(s；2H)，8.40(s；1H)，7.73(d；J＝8.3Hz；1H)，7.64(s；1H)，7.32(s，J＝8.3Hz；1H)，6.27(s；1H)，4.72(s；1H)，3.40(m；2H)，2.64(m；2H)，1.96/1.83(2m；2H)，1.40(s；9H)，1.18(s；3H)。

Example 29: (R) -2-amino-4- (2-furan-2-yl-benzooxazol-5-yl) -2-methyl-butan-1-ol

The title compound was obtained as a colorless solid by the method described in example 15 (method a) using the appropriate starting materials.¹H-NMR(DMSO-d₆)δ：8.59(s；1H)，7.92(s；1H)，7.58(d，J＝8.3Hz；1H)，7.51(s；1H)，7.31(d，J＝8.3Hz；1H)，7.05(s；1H)，4.58(s；1H)，3.16(s；2H)，2.70(m；2H)，1.58(m；2H)，0.95(s；3H)。

Preparation of [ (R) -3- (2-furan-2-yl-benzooxazol-5-yl) -1-hydroxymethyl-1-methyl-propyl ] -carbamic acid tert-butyl ester:

the title compound was prepared according to the procedure described for the preparation of example 15 using ethyl furan-2-carboximidic acid hydrochloride.¹H-NMR(DMSO-d₆)δ：8.60(s；1H)，7.92(s；1H)，7.60(d，J＝8.3Hz；1H)，7.50(s；1H)，7.20(d，J＝8.3Hz；1H)，7.05(s；1H)，6.23(s；1H)，4.70(s；1H)，3.40(s；2H)，2.61(m；2H)，1.94/1.82(2m；2H)，1.40(s；9H)，1.18(s；3H)。

Example 30: (R) -2-amino-4- {2- [ (E) -2- (3, 4-dimethoxy-phenyl) -vinyl ] -benzooxazol-5-yl } -2-methyl-butan-1-ol

The title compound was obtained as a colorless solid by the method described in example 15 (method a) using the appropriate starting materials.¹H-NMR(DMSO-d₆)δ：7.70(d；1H)，7.54(d；1H)，7.49(d；1H)，7.44(d；1H)，7.29(dd；1H)，7.21(d；1H)，7.19(d；1H)，7.00(d；1H)，3.83(s；3H)，3.80(s；3H)，3.15(bs；2H)，2.69(m；2H)，1.56(m；2H)，1.40(bs；2H)，0.95(s；3H)。

((R) -3- {2- [ (E) -2- (3, 4-dimethoxy-phenyl) -vinyl ] -benzooxazol-5-yl } -1-hydroxy-methyl-1-methyl-propyl) -carbamic acid tert-butyl ester:

the title compound was prepared according to the procedure described for the preparation of example 15 using (E) -3- (3, 4-dimethoxy-phenyl) -acryloyl imidoic acid ethyl ester hydrochloride. Characterised by¹H-NMR Signal (CDCl)₃)δ：3.94/3.96(2s；6H)，1.44(s；9H)，1.25(s；3H)。

Example 31: (R) -2-amino-4- [2- (2, 2-difluoro-benzo [1, 3] dioxol-5-yl) -benzooxazol-5-yl ] -2-methyl-butan-1-ol

The title compound was obtained as a colorless solid by the method described in example 15 (method a) using the appropriate starting materials.¹H-NMR(DMSO-d₆)δ：8.14(s；1H)，8.04(dd；1H)，7.64(t；2H)；7.59(s；1H)，7.26(d；1H)，4.72(bs；1H)，3.22(m；2H)，2.73(m；2H)，1.62(m；2H)，1.00(s；3H)。

Preparation of ((R) -3- [2- (2, 2-difluoro-benzo [1, 3] dioxol-5-yl) -benzooxazol-5-yl ] -1-hydroxy-methyl-1-methyl-propyl } -carbamic acid tert-butyl ester:

the title compound was synthesized according to the procedure described for example 15 using 2, 2-difluoro-benzo [1, 3]]And dioxole-5-ethyl formimidate hydrochloride. Characterised by¹H-NMR Signal (CDCl)₃)δ：8.15(d；1H)，8.06(dd，1H)，7.65/7.63(2d；2H)，7.56(d；1H)，7.24(dd；1H)，1.40(s；9H)，1.18(s；3H)。

Example 32: (R) -2-amino-2-methyl-4- [2- (3-phenoxy-phenyl) -benzooxazol-5-yl ] -butan-1-ol

The title compound was obtained as a colorless solid by the method described in example 15 (method a) using the appropriate starting materials.¹H-NMR(DMSO-d₆)δ：7.90(d；1H)，7.68-7.52(m；4H)，7.45(m；2H)，7.25(m；3H)，7.12(d；2H)，4.63(bs；1H)，3.18(s；2H)，2.70(m；2H)，1.57(m；2H)，0.97(s；3H)。

Preparation of { (R) -1-hydroxymethyl-1-methyl-3- [2- (3-phenoxy-phenyl) -benzooxazol-5-yl ] -propyl } -carbamic acid tert-butyl ester:

the title compound was prepared according to the procedure described for the preparation of example 15 using ethyl 3-phenoxy-benzimidinate hydrochloride. Characterised by¹H-NMR Signal (CDCl)₃) δ: 3.23/3.17 (AB-system; 2H), 1.44 (s; 9H), 1.25 (s; 3H).

Example 33: (R) -2-amino-4- [2- (4-cyclopentyloxy-3-methoxy-phenyl) -benzooxazol-5-yl ] -2-methyl-butan-1-ol

The title compound was obtained as a colorless solid by the method described in example 15 (method a) using the appropriate starting materials.¹H-NMR(DMSO-d₆)δ：7.73(dd；1H)，7.62(d；1H)，7.60(d；1H)，7.53(s；1H)，7.19(d；1H)，7.14(d；1H)，4.90(bs；1H)，4.60(bs；1H)，3.83(s；3H)，3.18(s；2H)，3.70(m；2H)，1.92(m；2H)，1.74(m；2H)，1.60(m；2H)，0.96(s；3H)。

Preparation of { (R) -3- [2- (4-cyclopentyloxy-3-methoxy-phenyl) -benzooxazol-5-yl ] -1-hydroxy-methyl-1-methyl-propyl } -carbamic acid tert-butyl ester:

the title compound was prepared according to the procedure described for the preparation of example 15 using ethyl 4-cyclopentyloxy-3-methoxy-benzimidinate hydrochloride. Characterised by¹H-NMR Signal (DMSO-d)₆)δ：3.83(s；3H)，1.39(s；9H)，1.18(s；3H)。

Example 34: (R) -2-amino-4- [2- (2, 3-dimethoxy-phenyl) -benzooxazol-5-yl ] -2-methyl-butan-1-ol

The title compound was obtained as a colorless solid by the method described in example 15 (method a) using the appropriate starting materials.¹H-NMR(DMSO-d₆)δ：7.60(bs；3H)，7.28(bs；3H)，3.90(bs；6H)，3.20(bs；2H)，2.75(bs；2H)，1.60(bs；1H)，0.98(bs；3H)。

Preparation of { (R) -3- [2- (2, 3-dimethoxy-phenyl) -benzooxazol-5-yl ] -1-hydroxymethyl-1-methyl-propyl } -carbamic acid tert-butyl ester:

the title compound was prepared according to the procedure described for the preparation of example 15 using ethyl 2, 3-dimethoxy-benzimidinate hydrochloride. Characterised by¹H-NMR Signal (DMSO-d)₆)δ：3.85/3.87(2s；6H)，1.39(s；9H)，1.19(s；3H)。

Example 35: (R) -2-amino-4- [2- (2, 5-dimethoxy-phenyl) -benzooxazol-5-yl ] -2-methyl-butan-1-ol

The title compound was obtained as a colorless solid by the method described in example 15 (method a) using the appropriate starting materials.¹H-NMR(DMSO-d₆)δ：7.64(d；1H)，7.57(s；1H)，7.27(d；2H)，7.24(dd；1H)，6.63(t；1H)，4.60(bs；1H)，3.34(s；6H)，3.17(s；2H)，2.70(m；2H)，1.58(m；2H)，0.95(s；3H)。

Preparation of { (R) -3- [2- (2, 5-dimethoxy-phenyl) -benzooxazol-5-yl ] -1-hydroxymethyl-1-methyl-propyl } -carbamic acid tert-butyl ester:

the title compound was prepared according to the procedure described for the preparation of example 15 using ethyl 2, 5-dimethoxy-benzimidinate hydrochloride. Characterised by¹H-NMR Signal (CDCl)₃)δ：3.90(s；6H)，1.44(s；9H)，1.26(s；3H)。

Example 36: (R) -2-amino-2-methyl-4- [2- (4-phenyl-5-trifluoromethyl-thiophen-2-yl) -benzooxazol-5-yl ] -butan-1-ol

The title compound was obtained as a colorless solid by the method described in example 15 (method a) using the appropriate starting materials.¹H-NMR(DMSO-d₆)δ：8.03(s；1H)，7.67(d；1H)，7.61(s；1H)，7.56-7.45(m；5H)，7.30(dd；1H)，4.59(bs；1H)，3.17(s；2H)，2.73(m；2H)，1.58(m；2H)，0.97(s；3H)。

Example 37: (R) -2-amino-2-methyl-4- (2-phenyl-benzofuran-5-yl) -butan-1-ol

The title compound was synthesized according to the method described in Synthesis (2003), (11), 1667-1670.¹H-NMR(DMSO-d₆)：7.88(d，J＝7.39Hz，2H)，7.48(t，J＝6.12Hz，2H)，7.42(s，1H)，7.40(m，2H)，7.34(s，1H)，7.12(d，J＝7.68Hz，1H)，4.57(br，s，1H，OH)，3.17(s，2H)，2，68(m，J＝8.65Hz，2H)，1.58(t，J＝8.8Hz，2H)，1.45(br，s，2H；-NH₂)，0.98(s，3H)。MS(ESI+)：296.3[M+H]⁺。

Preparation of 5- (2-iodo-ethyl) -2-phenyl-benzofuran

2- (2-phenyl-benzofuran-5-yl) -ethanol (11.18 g; 46.9mmol) was dissolved in DCM (170ml) and N-iodosuccinimide (NIS; 11.31 g; 50.27mmol) and triphenylphosphine (15.39 g; 52.67mmol) were added with stirring. The reaction mixture was allowed to stand at room temperature overnight, and then the reaction solution was treated with 6% NaHCO₃Extracting with aqueous solution twice, and extracting the organic layer with Na₂SO₄And (5) drying. Filtration and removal of the solvent gave the pure title compound after crystallization from methanol/DCM/cyclohexane. The mother liquor was collected and purified on silica gel (cyclohexane/ethyl acetate 95/5 as mobile phase).

Preparation of 2- (2-phenyl-benzofuran-5-yl) -ethanol

Ethyl 2-phenyl-benzofuran-5-yl) -acetate (Ota et al; journal of the chemical society, Perkin Transactions 1: organic and Bio-Organic Chemistry (1972-1999) (1988), (11), 3029-35; 13.14 g; 46.87mmol) was dissolved in dry THF. Adding LiAIH₄(2.85 g; 74.99mmol) and then the reaction mixture was left at 80 ℃ for 1 hour. After cooling to room temperature, the reaction mixture was slowly poured into saturated Na₂SO₄Aqueous solution (150ml) and then filtered through Hyflo. The solvent was removed under reduced pressure, the residue was dissolved in AcOEt and extracted twice with water. The organic layer was washed with Na₂SO₄And (5) drying. Removal of the solvent gave the pure title compound.

Example 38: (R) -2-amino-4- [2- (4-fluoro-phenyl) -benzofuran-5-yl ] -2-methyl-butan-1-ol

The title compound was synthesized according to the procedure described for the preparation of example 30, using 4-fluoro-phenacyl chloride as the synthesis of [2- (4-fluoro-phenyl) -benzofuran-5-yl]-starting material of ethyl acetate.¹H-NMR(DMSO-d₆)δ：7.94(d，J＝8.78Hz，1H)，7.92(d，J＝8.72Hz，1H)，7.48(d，J＝8.33Hz，1H)，7.42(s，1H)，7.35-7.29(m，3H)，7.12(d，J＝8.34Hz，1H)，4.58(br，s，1H；-OH)，3.17(s，2H)，2，68(m，J＝8.65Hz，2H)，1.58(t，J＝8.33Hz，2H)，1.23(br，s，2H；-NH₂)，0.98(s，3H)。MS(ESI+)：[M+H]⁺。

Example 39: 2-amino-2- [2- (2-phenyl-benzofuran-5-yl) -ethyl ] -propane-1, 3-diol

The title compound was synthesized according to the procedure described in Journal of Medicinal Chemistry (2000 Jul 27), 43(15), 2946-61) using 5- (2-iodo-ethyl) -2-phenyl-benzofuran as the alkylating agent in scheme 6 (step g).¹H-NMR(DMSO-d₆)δ：7.88(d，J＝7.58Hz，2H)，7.55-7.30(m，6H)，7.12(d，J＝8.34Hz，1H)，4.43(br，s，2H；-OH)，3.26(m，4H)，2，68(m，2H)，1.56(m，2H)，1.28(s，2H；-NH₂)。MS(ESI+)：312.4[M+H]⁺。

Example 40: 2-amino-2- [2- (3-ethyl-1-methyl-1H-indazol-5-yl) -ethyl ] -propane-1, 3-diol

The title compound was synthesized according to the method described in Tetrahedron Letters (2002), 43(45), 8095-8097, using 5- (2-iodo-ethyl) -2-phenyl-benzofuran as alkylating agent in the Schollkopf reaction.¹H-NMR(DMSO-d₆)：7.57(d，J＝8.21Hz，1H)，7.27(s，1H)，6.92(d，J＝8.27Hz，1H)，4.45(br，s，2H；-OH)，3.90(s，3H；-NCH₃)，3.25(q，J＝8.08Hz，4H)，2.85(q，J＝7.58Hz，2H)，2.72(m，2H)，1.57(m，2H)，1.28(t，J＝7.52Hz，3H)。MS(ESI+)：312.4[M+H]⁺。

Example 41: 2-amino-2- [2- (3-heptyl-1-methyl-1H-indazol-5-yl) -ethyl ] -propane-1, 3-diol

The title compound was synthesized according to the method described in Tetrahedron Letters (2002), 43(45), 8095-8097; 5- (2-iodo-heptyl) -2-phenyl-benzofuran (prepared in analogy to the preparation of 5- (2-iodo-ethyl) -2-phenyl-benzofuran) was used as alkylating agent in the Schollkopf reaction.¹H-NMR(DMSO-d₆)δ：7.55(d，J＝8.22Hz，1H)，7.27(s，1H)，6.91(d，J＝8.27Hz，1H)，4.99(br，s，3H；-NH₂；-OH)，3.90(s，3H；-NCH₃)，3.25(m，4H)，2.82(t，J＝7.51Hz，2H)，2.71(m，2H)，1.69(m，2H)，1.61(m，2H)，1.35-1.15(m，8H)，0.85(t，J＝6.37Hz，3H)。MS(ESI+)：348.5[M+H]⁺

Example 42: phosphoric acid mono- [ (R) -2-amino-2-methyl-4- (2-pentyl-benzoxazol-5-yl) -butyl ] ester

Reacting [ (R) -1-methyl-3- (2-pentyl-benzoxazol-5-yl) -1-phosphonooxymethyl-propyl]A solution of ammonium tert-butyl carbamate (18mg, 0.0369mmol) in diethyl ether (2ml) was treated with 2M HCl in diethyl ether and stirred at room temperature for 2 hours. The reaction was stopped by the addition of 28% aqueous ammonium hydroxide (2ml), methanol (2ml) and DCM (1 ml). After evaporation of the solvent in vacuo, the residue is chromatographed using preparative reverse phase chromatography (X-Terra, C-18, eluent: 10mM NH)₄HCO₃Water/acetonitrile gradient). The title compound was obtained as an amorphous solid. MS (MH-) 269.3.

Preparation of [ (R) -1-methyl-3- (2-pentyl-benzoxazol-5-yl) -1-phosphonooxymethyl-propyl ] -carbamic acid tert-butyl ester ammonium salt

10% Palladium on carbon (50mg) was added to [ (R) -1-methyl-1- (3-oxo-1, 5-dihydro-benzo [ e ] under argon atmosphere][1，3，2]Dioxaphospha * -3-yloxymethyl) -3- (2-pentyl-benzoxazol-5-yl) -propyl]-tert-butyl carbamate (168.5mg, 0.2942mmol) in ethanol (20 ml). The reaction was allowed to proceed for 8 hours with hydrogen instead of argon. 28% aqueous ammonium hydroxide solution was added to alkalinity and the solvent was evaporated in vacuo. The residue was purified by preparative reverse phase chromatography (X-Terra, C-18, eluent: 10mM NH)₄HCO₃Gradient of water/acetonitrile solution of (a). The title compound was obtained as an amorphous solid. MS (MH-) 469.3.

Preparation of [ (R) -1-methyl-1- (3-oxo-1, 5-dihydro-benzo [ e ] [1, 3, 2] dioxaphospha * -3-yloxymethyl) -3- (2-pentyl-benzoxazol-5-yl) -propyl ] -carbamic acid tert-butyl ester

(R) -1-hydroxymethyl-1-methyl-3- (2-pentyl-benzoxazol-5-yl) -propyl under argon]A solution of tert-butyl carbamate (151mg, 0.387mmol) and tetrazole (85mg, 1.21mmol) in dry THF (3ml) was treated with N, N-diethyl-1, 5-dihydro-2, 4, 3-benzo-dioxaphospha * -3-amine (185mg, 0.77mmol) and stirred at room temperature for 1 hour. After the reaction mixture was cooled to 0 ℃, aqueous hydrogen peroxide (0.4ml of a 30 w/w% solution) was added and the reaction solution was stirred at room temperature for 1 hour. The reaction mixture was washed with 10% Na₂S₂O₃The partition was between aqueous solution and AcOEt. The organic layer was washed with MgSO₄Dried, concentrated in vacuo and purified over silica gel (eluent: DCM/methanol 30/1). The title compound was obtained as a colorless amorphous solid.¹H-NMR(DMSO-d₆) δ: 7.52(d, J ═ 8.3 Hz; 1H), 7.50-7.40 (m; 5H), 7.15(dd, J ═ 8.3, 1.5 Hz; 1H), 6.8 (bs; 1H), 5.32-5.26/5.16-5.04 (m; 4H), 4.17/4.06 (AB-system, J ═ 9.5, 4.5 Hz; 2H), 2.88(t, J ═ 7.5 Hz; 2H), 2.71-2.58 (m; 2H), 2.14-2.08 (m; 1H), 1.81-1.68 (m; 3H), 1.38 (s; 9H); 1.28 (s; 3H), 1.38-1.28 (m; 4H), 0.85 (m; 3H).

Example 43: phosphoric acid mono- { (R) -2-amino-4- [2- (3-ethoxy-phenyl) -benzooxazol-5-yl ] -2-methyl-butyl } ester hydrochloride

Reacting [ (R) -3- (3-amino-4-hydroxy-phenyl) -1- (di-tert-butoxy-phosphoryloxymethyl) -1-methyl-propyl]A solution of tert-butyl carbamate (157.5mg, 0.313mmol) and 3-ethoxy-benzimidate ethyl ester hydrochloride in dry methanol (2ml) was stirred in a closed vial (microwave reactor) at 120 ℃ for 20 min. After cooling to room temperature, the precipitate was filtered off, washed twice with MeOH, three times with water and three times with diethyl ether. After drying in vacuo, the precipitate was dissolved in a mixture of diethyl ether/HCl (4ml, 2M) and methanol (1 ml). The solvent was evaporated to give the title compound as colorless crystals.¹H-NMR(CD₃OD)：δ＝7.76(d；1H)，7.71(s；1H)，7.61(bs；2H)，7.47(m；1H)，7.37(m；1H)，7.14(d；1H)，4.24-4.00(m；4H)，2.89(m；2H)，2.10(m；2H)，1.47(s；6H)；MS(ESI+)：421.4[M+H]⁺。

Preparation of [ (R) -3- (3-amino-4-hydroxy-phenyl) -1- (di-tert-butoxy-phosphoryloxymethyl) -1-methyl-propyl ] -carbamic acid tert-butyl ester:

(R) -3- (4-benzyloxy-3-nitro-phenyl) -1- (di-tert-butoxy-phosphoryloxymethyl) -1-methyl-propyl under argon]A solution of tert-butyl carbamate (5.36g, 8.6mmol) in ethanol (250ml) was treated with 10% palladium on carbon (1.22 g). The argon gas was replaced with hydrogen gas, and the reaction mixture was stirred at room temperature for 16 hours. After filtration and removal of the solvent in vacuo, the residue was purified by silica gel column chromatography (dichloromethane/methanol 10/1) to give the title compound as a colorless solid.¹H-NMR(CD₃OD)：δ＝6.61-6.57(m；2H)，6.41(dd；1H)，6.22(s；1H)，4.55(s；1H)，4.41/3.93(2m；2H)，2.43(m；1H)，2.05(m；1H)，1.70(m；1H)，1.48(s；18H)，1.44(s；9H)，1.28(s；3H)；MS(ESI+)：503.4[M+H]⁺。

Preparation of [ (R) -3- (4-benzyloxy-3-nitro-phenyl) -1- (di-tert-butoxy-phosphoryloxymethyl) -1-methyl-propyl ] -carbamic acid tert-butyl ester

(R) -3- (4-benzyloxy-3-nitro-phenyl) -1-hydroxymethyl-1-methyl-propyl under argon]A solution of tert-butyl carbamate (3.71g, 8.62mmol) in tetrahydrofuran (71ml) was treated with tetrazole (1.89g, 27.0mmol) and di-tert-butyl diethylaminophosphite (4.8ml, 17.24 mmol). After stirring for 100 minutes at room temperature, hydrogen peroxide (9ml of a 30% aqueous solution) was carefully added (exothermic). After stirring at room temperature for a further 60 minutes, the reaction mixture is taken up in Na₂S₂O₃The partitioning was performed between solution (10% in water) and AcOEt. The organic layer was dried over magnesium sulfate and evaporated in vacuo (colorless oil). MS (ESI +): 623.4[ M + H]⁺。

Preparation of [ (R) -3- (4-benzyloxy-3-nitro-phenyl) -1-hydroxymethyl-1-methyl-propyl ] -carbamic acid tert-butyl ester:

reacting [ (R) -3- (3-amino-4-hydroxy-phenyl) -1-hydroxymethyl-1-methyl-propyl]A solution of tert-butyl carbamate (3.27g, 9.60mmol) in dimethylformamide (64ml) was treated with potassium carbonate (2.65g, 19.2mmol) and benzyl bromide (1.15ml, 9.6mmol) and then stirred at 80 ℃ for 20 h. After filtration, the filtrate was evaporated in vacuo, redissolved in dichloromethane and extracted with 1N aqueous HCl, saturated aqueous sodium bicarbonate and brine. The organic phase is dried over magnesium sulfate and the solvent is removed in vacuo, then the title compound is crystallized from pentane to give the title compound as pale yellow crystals.¹H-NMR(DMSO-d₆)：δ7.70(d；1H)，7.55-7.29(m；7H)，6.27(bs；1H)，5.75(s；2H)，4.69(t；1H)，3.37(m；2H)，2.51(m；2H)，1.87(m；1H)，1.72(m；1H)，1.36(s；9H)，1.15(s；3H)；MS(ESI+)：431.3[M+H]⁺。

Examples 44 to 48: the examples shown in Table 3 were prepared as described in example 43.

Table 3:

example 49: phosphoric acid mono- [ (R) -2-amino-2-methyl-4- (2-phenyl-benzofuran-5-yl) -butyl ] ester

The title compound was synthesized according to the method described in Synthesis (2003), (11), 1667-1670.¹H-NMR(MeOD+DCl)δ：7.86(d，J＝7.96，2H)，7.50-7.30(m，5H)，7.19(d，J＝8.44Hz，1H)，7.13(s，1H)，4.09(m，2H)，2.82(m，2H)，2.09(m，2H)，1.47(s，3H)。MS(ESI+)：376.4[M+H]⁺。

Example 50: phosphoric acid mono- { (R) -2-amino-4- [2- (4-fluoro-phenyl) -benzofuran-5-yl ] -2-methyl-butyl } ester

The title compound was synthesized according to the method described in Synthesis (2003), (11), 1667-1670.¹H-NMR(MeOD+DCl)δ：7.91(d，J＝8.65，2H)，7.90(d，J＝8.65，2H)，7.49(s，1H)，7.44(d，J＝8.21，1H)，7.20(m，2H)，7.09(s，1H)，4.09(m，2H)，2.82(m，2H)，2.08(m，2H)，1.46(s，3H)。³¹p-NMR(MeOD+DCl)δ：-0.45；MS(ESI-)：392[M-H]^-。

The compounds of formula I in free form or in pharmaceutically acceptable salt form exhibit valuable pharmacological properties, such as properties modulating lymphocyte recirculation, as demonstrated, for example, in vitro and in vivo tests, and are therefore suitable for use in therapy.

A. In vitro

The compounds of formula I have binding affinity for the human S1P receptor as determined in the following assay:

sphingosine-1-phosphate (S1P) receptor performance test

EDG-1 (S1P) using the human S1P receptor₁)、EDG-3(S1P₃)、EDG-5(S1P₂)、EDG-6(S1P₄) And EDG-8 (S1P)₅) The compounds were tested for activator activity. GTP [ gamma- ] induced by compounds³⁵S]Functional receptor activation was assessed by quantifying the ability to bind to membrane proteins prepared from transfected CHO or RH7777 cells stably expressing the appropriate human S1P receptor. The assay technique used was SPA (scintillation proximity based assay). Briefly, compounds dissolved in DMSO were serially diluted and incubated in 50mM Hepes, 100mM NaCl, 10mM MgCl₂10 μ M GDP, 0.1% fat-free BSA and 0.2nM GTP [ γ -³⁵S](1200Ci/mmol) was added to the membrane protein expressing S1P receptor immobilized on SPA-beads (Amersham-Pharmacia) (10-20. mu.g/well). After incubation in 96-well microtiter plates for 120 min at RT, unbound GTP [ gamma-³⁵S]. Membrane bound GTP [ gamma- ] cells were read using a TOPcount plate reader (Packard)³⁵S]The luminescence of the initiated SPA beads was quantified. Calculation of EC Using Standard Curve fitting software₅₀. In this assay, the compound of formula I is paired with S1P₁Receptors have binding affinities < 50 nM.

Compounds of the examples	S1P1EC50[nM]	S1P3EC50[nM]	S1P4EC50[nM]	S1P5EC50[nM]
					12	0.3 agonists	8.4 agonists	0.4 agonists	0.1 agonists
13	0.06 agonists	5.7 agonists	1.2 agonists	0.4 agonists
					14	2.8 agonists	47 inverse agonist	1.0 agonists	0.8 agonists
42	17 agonists	> 1000 agonists	9 agonists	5 agonists
					49	1.6 agonists	1180 agonists	0.7 agonists	0.4 agonists

B. In vivo: lymphopenia of blood

The compound of formula I or excipient was administered orally to rats by gavage. Hematology analyses were performed on rat tail blood samples taken on day 1 and 2, 6, 24, 48 and 72 hours post-administration, and baseline values were obtained for individuals from the day 1 blood samples. In this assay, the compounds of formula I reduce peripheral blood lymphocytes when administered at a dose of 0.03 to 3 mg/kg. For example, the following results were obtained: peripheral blood lymphocyte reduction by more than 50%

Example 1: after 6 hours of oral administration of 0.02mg/kg, 24 hours of oral administration of 0.3mg/kg, 48 hours of oral administration of > 1 mg/kg.

Example 6: 0.07mg/kg for 6 hours after oral administration.

Example 25: after 6 hours of oral administration of 0.06mg/kg, 48 hours of oral administration of 0.3 mg/kg.

Example 26: after 0.03mg/kg was orally administered for 6 hours, and after 0.2mg/kg was orally administered for 48 hours.

Example 37: 0.40mg/kg for 6 hours after oral administration.

Thus, the compounds of formula I are useful in the treatment and/or prevention of diseases or disorders mediated by lymphocyte interactions, for example in transplantation, such as acute or chronic rejection of cell, tissue or organ allografts or xenografts or delayed graft function (delayed graft function), graft-versus-host disease, autoimmune diseases such as rheumatoid arthritis, systemic lupus erythematosus, hashimoto's thyroiditis, multiple sclerosis, myasthenia gravis, type I or II diabetes and disorders related thereto, vasculitis, pernicious anemia, sjogren's syndrome, uveitis, psoriasis, Graves 'ophthalmopathy (Graves' ophthalmopathy), alopecia areata and the like, allergic diseases such as allergic asthma, atopic dermatitis, allergic rhinitis/conjunctivitis, allergic contact dermatitis, inflammatory diseases optionally accompanied by abnormal reactions such as inflammatory bowel disease, Crohn's disease or ulcerative colitis, intrinsic asthma, inflammatory lung injury, inflammatory liver injury, inflammatory glomerular injury, atherosclerosis, osteoarthritis, irritant contact dermatitis and other eczematous dermatitis, seborrheic dermatitis, skin symptoms of immune-mediated conditions, inflammatory eye disease, keratoconjunctivitis, myocarditis or hepatitis, ischemia/reperfusion injury such as myocardial infarction, stroke, gut ischemia, renal failure or hemorrhagic shock, traumatic shock, angiogenesis, alzheimer's disease, cancer such as breast cancer, T-cell lymphoma or T-cell leukemia, infectious diseases such as toxic shock (e.g. superantigen-induced), septic shock, adult respiratory distress syndrome or viral infections such as AIDS, viral hepatitis, chronic bacterial infections or senile dementia. Examples of cell, tissue or solid organ transplants include, for example, pancreatic islets, stem cells, bone marrow, corneal tissue, neuronal tissue, heart, lung, combined heart-lung, kidney, liver, intestine, pancreas, trachea or esophagus. For the above uses, the required dosage will, of course, vary with the mode of administration, the particular condition being treated and the effect desired.

In general, satisfactory results are obtained with systemic administration at daily doses of about 0.03 to 2.5mg/kg body weight. In larger mammals, such as humans, a suitable daily dosage is from about 0.5mg to about 100mg, which may conveniently be administered, for example, in divided doses up to 4 times per day or in sustained release form. Suitable unit dosage forms for oral administration contain from about 0.1 to 50mg of the active ingredient.

The compounds of the formula I can be administered by any conventional route, in particular enterally, for example orally, for example in the form of tablets or capsules, or parenterally, for example in the form of injectable solutions or suspensions, topically, for example in the form of lotions, gels, ointments or creams, or topically in the form of nasal or suppository. Pharmaceutical compositions comprising a compound of formula I in free form or in pharmaceutically acceptable salt form in association with at least one pharmaceutically acceptable carrier or diluent may be prepared in accordance with conventional procedures by mixing with a pharmaceutically acceptable carrier or diluent.

The compounds of formula I may be administered in free form or in pharmaceutically acceptable salt form, for example as described above. Such salts can be prepared by conventional methods and have activities of the same order of magnitude as the free compounds.

According to the above, the present invention also provides:

1.1 a method of preventing or treating a disorder or disease mediated by lymphocytes, such as those described above, in a subject in need of such treatment, which comprises administering to said subject an effective amount of a compound of formula I, or a pharmaceutically acceptable salt thereof;

1.2 a method of preventing or treating acute or chronic transplant rejection or T cell mediated inflammatory or autoimmune diseases, such as those described above, in a subject in need of such treatment, which comprises administering to said subject an effective amount of a compound of formula I or a pharmaceutically acceptable salt thereof;

2. a compound of formula I in free form or in pharmaceutically acceptable salt form for use as a medicament, e.g. in any of the methods described under 1.1 or 1.2 above.

3. A pharmaceutical composition comprising a compound of formula I, in free form or in pharmaceutically acceptable salt form, in association with a suitable pharmaceutically acceptable diluent or carrier, for use, for example, in any of the methods described under 1.1 or 1.2 above.

4. A compound of formula I or a pharmaceutically acceptable salt thereof for use in the preparation of a pharmaceutical composition for use in any of the methods described under l.1 or 1.2 above.

The compounds of formula I may be administered as the sole active ingredient or, for example, as an adjuvant in combination with other drugs, for example, immunosuppressants or immunomodulators or other anti-inflammatory agents, for example, for the treatment or prophylaxis of acute or chronic rejection of allografts or xenografts or inflammatory or autoimmune diseasesThe above-mentioned drugs for autoimmune disorders, or chemotherapeutic agents such as malignant cell antiproliferative agents. For example, the compounds of formula I may be used in combination with: calcineurin inhibitors, e.g. cyclosporin A, FK 506 or ISA_TX₂47; mTOR inhibitors, such as rapamycin, 40-O- (2-hydroxyethyl) -rapamycin, CCI779, ABT578, AP23573, AP23464, AP23675, AP23841, TAFA-93, biolipimus (biolimus)7, or bimimous 9; ascomycins with immunosuppressive properties, such as ABT-281, ASM981, and the like; S1P receptor agonists, such as FTY720 or analogs thereof; a corticosteroid; cyclophosphamide; azathioprine; methotrexate; leflunom; mizoribine; mycophenolic acid; mycophenolate mofetil; 15-deoxyspandex or an immunosuppressive homolog, analog or derivative thereof; immunosuppressive monoclonal antibodies, e.g., monoclonal antibodies against leukocyte receptors such as MHC, CD2, CD3, CD4, CD7, CD8, CD25, CD28, CD40, CD45, CD58, CD80, CD86 or their ligands; other immunomodulatory compounds, such as recombinant binding molecules having at least a portion of the extracellular domain of CTLA4 or a mutant thereof, e.g., having at least the extracellular portion of CTLA4 or a mutant thereof bound to a non-CTLA 4 protein sequence, e.g., CTLA4Ig (e.g., designated ATCC 68629) or a mutant thereof, e.g., LEA 29Y; adhesion molecule inhibitors, such as LFA-1 antagonists, ICAM-1 or-3 antagonists, VCAM-4 antagonists or VLA-4 antagonists; or chemotherapeutic agents, such as paclitaxel, gemcitabine, cisplatin, doxorubicin, or 5-fluorouracil; or an anti-infective agent.

When a compound of formula I is administered in combination with other immunosuppressive/immunomodulatory, anti-inflammatory, chemotherapeutic or anti-infective therapies, the dosage of the co-administered immunosuppressive, immunomodulatory, anti-inflammatory, chemotherapeutic or anti-infective compounds will, of course, vary with the type of drug co-administered, e.g., whether it is a steroid or calcineurin inhibitor, the particular drug employed, the condition being treated, etc. In accordance with the foregoing, in another aspect, the present invention provides:

5. a method as described above, which method comprises co-administration, e.g. concurrently or sequentially, of a therapeutically effective and non-toxic amount of a compound of formula I and at least one second drug substance, e.g. an immunosuppressive, immunomodulatory, anti-inflammatory or chemotherapeutic drug, e.g. as described above.

6.A pharmaceutical combination, e.g. a pharmaceutical pack, comprising a) a first active agent which is a compound of formula I as disclosed herein in free form or in pharmaceutically acceptable salt form, and b) at least one co-active agent, e.g. an immunosuppressive, immunomodulatory, anti-inflammatory, chemotherapeutic or anti-infective drug. The pharmaceutical pack may include instructions for its administration.

The terms "co-administration" or "co-administration" and the like as used herein are meant to encompass the administration of a selected therapeutic agent to a single patient and are intended to encompass treatment regimens in which the active agents need not be administered by the same route of administration or simultaneously.

The term "pharmaceutical combination" as used herein refers to a product obtained by mixing or combining more than one active ingredient, and includes both fixed and non-fixed combinations of active ingredients. The term "fixed combination" means that the active ingredients, e.g. a compound of formula I and a co-agent, are administered to a patient simultaneously in a single entity or dosage form. The term "non-fixed combination" means that the active ingredients, e.g. a compound of formula I and a co-agent, are administered to a patient as separate entities either simultaneously, concurrently or sequentially with no specific time limits, wherein the administration provides therapeutically effective levels of the 2 compounds in the patient. The latter is also applicable to cocktail therapy, e.g. administering 3 or more active ingredients.

Claims (10)

1. A compound of formula I, in free or salt form:

wherein:

R₁is C_1-6An alkyl group; by hydroxy, C_1-2Alkoxy or C substituted by 1 to 6 fluorine atoms_1-6An alkyl group; c_2-6An alkenyl group; or C_2-6An alkynyl group;

x is O, CH₂C ═ O or straightConnecting a bond;

R₂is optionally substituted C_1-7Alkyl, optionally substituted C_1-7Alkenyl, optionally substituted C_1-7Alkynyl, optionally substituted C_3-6Cycloalkyl, optionally substituted phenyl or optionally substituted heteroaryl, wherein C is substituted_1-7Alkyl radical, C_1-7Alkenyl radical, C_1-7Alkynyl or C_3-6Cycloalkyl has 1 to 5 substituents selected from the group consisting of: hydroxy, halogen, C_1-4Alkyl radical, C_1-4Alkoxy, C substituted by 1 to 5 halogen atoms_1-4Alkoxy radical, C_3-6Cycloalkyl radical, C_3-6Cycloalkoxy, C_3-6Cycloalkyl radical C_1-5Alkyl radical, C_3-6Cycloalkoxy group C_1-5Alkyl, cyano, optionally substituted phenyl, optionally substituted phenyloxy, optionally substituted heteroaryl, optionally substituted heteroaryloxy, optionally substituted heterocyclyl optionally linked through oxygen;

and wherein the phenyl, phenyloxy, heteroaryl, heteroaryloxy, heterocyclyl optionally linked through oxygen may be substituted independently of each other by 1 to 5 substituents selected from the group consisting of: hydroxy, halogen, C_1-4Alkyl, C substituted by 1 to 5 fluorine atoms_1-4Alkyl radical, C_1-4Alkoxy, C substituted by 1 to 5 fluorine atoms_1-4Alkoxy, cyano, phenyl and phenyl substituted with 1 to 5 substituents selected from the group consisting of: hydroxy, halogen, C_1-4Alkyl, C substituted by 1 to 5 fluorine atoms_1-4Alkyl radical, C_1-4Alkoxy, C substituted by 1 to 5 fluorine atoms_1-4Alkoxy and cyano; or phenyl, phenyloxy, heteroaryl, heteroaryloxy, heterocyclyl which are optionally linked via oxygen, may be fused independently of one another with a heterocyclyl;

R₃is Z-X₂Wherein Z is CH₂CHF or CF₂And X₂Is OH or a group of the formula (a)

Wherein Z₁Is a direct bond、CH₂、CHF、CF₂Or O, and R₆And R₇Independently of one another, H, C optionally substituted by 1, 2 or 3 halogen atoms_1-4Alkyl or benzyl;

and R is₄And R₅Independently of one another, H, C optionally substituted by 1, 2 or 3 halogen atoms_1-4An alkyl or acyl group;

the fused rings a and b are independently of each other C_5-6Cycloalkyl, aryl, heterocyclyl or heteroaryl.

2. The compound of claim 1, in free form or in salt form, wherein X is O or a direct bond.

3. A compound according to claim 1, in free form or in salt form, wherein a and b together form 2, 6-or 2, 7-disubstituted naphthyl, 2, 5-or 2, 6-disubstituted benzoxazolyl, 2, 5-or 2, 6-disubstituted benzothienyl, 2, 5-or 2, 6-disubstituted benzofuranyl, 1, 4-or 1, 5-disubstituted indolyl, 3, 6-indazolyl or 3, 6-N-substituted-indazolyl.

4. A compound according to claim 1, wherein R is₁Is CH₃Or CH₂-OH。

5. A compound according to claim 1, wherein R is₂Is C_1-7Alkyl, substituted C_1-7Alkyl, substituted phenyl, phenylalkyl or substituted phenylalkyl, preferably C_3-7Alkyl or C substituted by 1 to 5 fluorine atoms_3-7An alkyl group.

6.A compound according to claim 1, wherein R is₄And R₅Is hydrogen.

7. A compound according to any one of claims 1 to 6, or a pharmaceutically acceptable salt thereof, for use as a medicament and for the manufacture of a medicament.

8. A pharmaceutical composition comprising a compound according to any one of claims 1 to 6, or a pharmaceutically acceptable salt thereof, in association with a pharmaceutically acceptable diluent or carrier.

9. A pharmaceutical composition comprising a compound according to any one of claims 1 to 6 in free form or in pharmaceutically acceptable salt form and at least one active agent in combination.

10. A method for preventing or treating a lymphocyte-mediated condition or disease, for preventing or treating acute or chronic transplant rejection or a T cell-mediated inflammatory or autoimmune disease in an individual, comprising administering to the individual in need thereof an effective amount of a compound according to any one of claims 1 to 6, or a pharmaceutically acceptable salt thereof.