JP2006225344A - 2-acylaminomethyl-3,4-dihydroxypyrrolidine type enzyme inhibitor - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To provide a new 2-acylaminomethyl-3,4-dihydroxypyrrolidine type compound useful as a tool or an agent for studying glycosidase inhibition, a synthesis intermediate necessary for simply obtaining the same in high purity, and its manufacturing method. <P>SOLUTION: The pyrrolidine compound is represented by formula (I) [wherein R is selected from the group consisting of hydrogen, a substituted or nonsubstituted alkyl group, a substituted or nonsubstituted aryl group, a substituted or nonsubstituted aralkyl group and a substituted or nonsubstituted alkylaryl group, and the configuration of the 2-position carbon is S or R, and those of the 3-position carbon and the 4-position carbon are selected from a combination of (3R, 4R) and (3S, 4S)]. <P>COPYRIGHT: (C)2006,JPO&NCIPI

Description

  The present invention relates to a pyrrolidine compound having enzyme inhibitory activity, a production method thereof, a synthetic intermediate in the production method, and an enzyme inhibitor containing the compound.

  Pyrrolidines, which are one of the basic skeletons of biologically active natural products, are important as functional materials including pharmaceuticals or research tools. In particular, hydroxypyrrolidine compounds having a structure similar to sugar are promising candidates for sugar-related enzyme inhibitors that are attracting attention as important biological activity regulators such as intercellular molecular recognition. Research is active. (For example, see Patent Documents 1 and 2 and Non-Patent Documents 1 and 2)

  Among them, compounds with a 3,4-dihydroxypyrrolidine skeleton are widely researched and developed because of their similarity to the substrate transition state of sugar hydrolase, and can be used as tools for further study of sugar hydrolase. Sex is shown. (For example, see Non-Patent Documents 3 and 4)

  As a glycosidase inhibitor with a higher degree of specificity, a modified compound with a functional group added to the 3,4-dihydroxypyrrolidine skeleton is considered promising, but the hydroxypyrrolidine necessary to synthesize such compounds Little progress has been made due to the lack of information on the various isomers of the class.

JP2000-95686 JP 9-509947 A Journal of Synthetic Organic Chemistry, 666-675 (2000) Phytochemistry, 56, 265-295 (2001) Chem. Biol. 8, 1061-1070 (2001) Bioorg. Med. Chem., 12, 3485-3495 (2004)

An object of the present invention is to provide novel 2-acylaminomethyl-3,4-dihydroxypyrrolidine-type enzyme inhibitors and methods for producing them.
The object of the present invention is to provide novel 2-acylaminomethyl-3,4-dihydroxypyrrolidine type compounds useful as glycosidase inhibition research tools or drugs, and synthetic intermediates necessary for obtaining them easily and with high purity. And provide its manufacturing method.

（Ｉ）
式中、Ｒは、水素、置換または非置換アルキル基、置換または非置換アリール基、置換または非置換アラルキル基、及び置換または非置換アルキルアリール基からなる群より選択され、２位炭素の立体配置はＳ体またはＲ体であり、３位炭素及び４位炭素の立体配置は（３Ｒ,４Ｒ）及び（３Ｓ,４Ｓ）の組合せから選択される。なお、各一般式において炭素原子横の＊のマークは、その炭素原子が不斉炭素であることを意味する。 The present invention provides a pyrrolidine compound having the following general formula (I).

(I)
Wherein R is selected from the group consisting of hydrogen, a substituted or unsubstituted alkyl group, a substituted or unsubstituted aryl group, a substituted or unsubstituted aralkyl group, and a substituted or unsubstituted alkylaryl group; Is S-form or R-form, and the configuration of the 3-position carbon and 4-position carbon is selected from the combination of (3R, 4R) and (3S, 4S). In each general formula, a mark * next to a carbon atom means that the carbon atom is an asymmetric carbon.

（ＩＩ）
式中、Ｒは、水素、置換または非置換アルキル基、置換または非置換アリール基、置換または非置換アラルキル基、及び置換または非置換アルキルアリール基からなる群より選択され、２位炭素の立体配置はＳ体またはＲ体であり、３位炭素及び４位炭素の立体配置は（３Ｒ,４Ｒ）及び（３Ｓ,４Ｓ）の組合せから選択され、Ｒ¹及びＲ²は、それぞれ独立に、水素、tert-ブチルジメチルシリル、メチルジ−tert-ブチルシリル、イソプロピルジメチルシリル、メチルジイソプロピルシリル、（トリフェニルメチル）ジメチルシリル、及びtert-ブチルジフェニルシリルからなる群より選択され、Ｒ³は水素またはアミノ保護基である。ただし、Ｒ¹とＲ²は同じであっても異なっていてもよく、及びＲ¹、Ｒ²及びＲ³は同時に水素を表さない。 The present invention also provides a compound having the following general formula (II):

(II)
Wherein R is selected from the group consisting of hydrogen, a substituted or unsubstituted alkyl group, a substituted or unsubstituted aryl group, a substituted or unsubstituted aralkyl group, and a substituted or unsubstituted alkylaryl group; Is the S-form or R-form, and the configuration of the 3-position carbon and 4-position carbon is selected from the combination of (3R, 4R) and (3S, 4S), and R ¹ and R ² are each independently hydrogen, selected from the group consisting of tert-butyldimethylsilyl, methyldi-tert-butylsilyl, isopropyldimethylsilyl, methyldiisopropylsilyl, (triphenylmethyl) dimethylsilyl, and tert-butyldiphenylsilyl, wherein R ³ is hydrogen or an amino protecting group is there. However, R ¹ and R ² may be the same or different, and R ¹ , R ² and R ³ do not represent hydrogen at the same time.

The compound represented by the general formula (II) is a synthetic intermediate useful in the production of the compound of the general formula (I).
In the general formula (II), R ¹ and R ² are preferably hydrogen. R ³ is preferably a benzyl group, a tert-butoxycarbonyl group or a benzyloxycarbonyl group.

  The present invention also provides a method for producing a pyrrolidine compound represented by the above general formula (I) by deprotecting the hydroxy protecting group and / or amino protecting group of the compound of the above general formula (II).

（III）
式中、２位炭素の立体配置はＳ体またはＲ体であり、３位炭素及び４位炭素の立体配置は（３Ｒ,４Ｒ）及び（３Ｓ,４Ｓ）の組合せから選択され、Ｒ¹及びＲ²は、それぞれ独立に、水素、tert-ブチルジメチルシリル、メチルジ−tert-ブチルシリル、イソプロピルジメチルシリル、メチルジイソプロピルシリル、（トリフェニルメチル）ジメチルシリル、及びtert-ブチルジフェニルシリルからなる群より選択され、Ｒ³は水素またはアミノ保護基である。ただし、Ｒ¹とＲ²は同じであっても異なっていてもよく、及びＲ¹、Ｒ²及びＲ³は同時に水素を表さない。 The present invention also provides a method, wherein the compound represented by the general formula (II) is further produced by acylating a compound represented by the following general formula (III). .

(III)
In the formula, the configuration at the 2-position carbon is S-form or R-form, and the configuration at 3-position carbon and 4-position carbon is selected from the combination of (3R, 4R) and (3S, 4S), and R ¹ and R ² are each independently selected from the group consisting of hydrogen, tert-butyldimethylsilyl, methyldi-tert-butylsilyl, isopropyldimethylsilyl, methyldiisopropylsilyl, (triphenylmethyl) dimethylsilyl, and tert-butyldiphenylsilyl; R ³ is hydrogen or an amino protecting group. However, R ¹ and R ² may be the same or different, and R ¹ , R ² and R ³ do not represent hydrogen at the same time.

  The present invention also provides an enzyme inhibitor comprising a pyrrolidine compound represented by the general formula (I) as an active compound.

  According to the present invention, a novel pyrrolidine compound having high optical purity can be obtained. In addition, the pyrrolidine compound of the present invention exhibits a specific enzyme inhibitory action due to the influence of the functional group provided. The present invention also provides an easily purified synthetic intermediate in which the amino group and / or hydroxyl group of the hydroxypyrrolidine derivative is protected.

（Ｉ） The pyrrolidine compound showing the enzyme inhibitory action of the present invention will be described. The pyrrolidine compound showing the enzyme inhibitory action of the present invention is represented by the general formula (I). Hereinafter, the term “pyrrolidine compound of the present invention” refers to a pyrrolidine compound which is any stereoisomer represented by the general formula (I), unless otherwise specified.

(I)

In the formula, the configuration at the 2-position carbon is S-form or R-form, and the configuration at 3-position carbon and 4-position carbon is selected from a combination of (3R, 4R) and (3S, 4S).
Wherein R is selected from the group consisting of hydrogen, a substituted or unsubstituted alkyl group, a substituted or unsubstituted aryl group, a substituted or unsubstituted aralkyl group, and a substituted or unsubstituted alkylaryl group; Is S-form or R-form, and the configuration of the 3-position carbon and 4-position carbon is selected from the combination of (3R, 4R) and (3S, 4S).

  In the above description, the alkyl group means a linear, branched or cyclic aliphatic hydrocarbon group, preferably an alkyl group having 1 to 10 carbon atoms, more preferably an alkyl group having 1 to 3 carbon atoms. The alkyl group may be substituted, and examples of the substituent on the alkyl group include a halogen, a hydroxyl group, an amino group, an alkyl group having 1 to 10 carbon atoms, and an alkoxy group having 1 to 10 carbon atoms.

  Aryl groups also include heterocyclic compounds containing atoms other than carbon. The aryl group is preferably an aryl group having 6 to 10 carbon atoms, preferably a phenyl group or a naphthyl group. Examples of the substituent on the aryl group include a halogen, a hydroxyl group, an amino group, an alkyl group having 1 to 10 carbon atoms, and an alkoxy group having 1 to 10 carbon atoms.

  The alkyl group and aryl group in the aralkyl group and alkylaryl group have the same meaning as defined above. The substituents in the aralkyl group and the alkylaryl group are also synonymous with the substituents described for the alkyl group and the aryl group.

（ＩＩ）
式中、Ｒは、水素、置換または非置換アルキル基、置換または非置換アリール基、置換または非置換アラルキル基、及び置換または非置換アルキルアリール基からなる群より選択され、２位炭素の立体配置はＳ体またはＲ体であり、３位炭素及び４位炭素の立体配置は（３Ｒ,４Ｒ）及び（３Ｓ,４Ｓ）の組合せから選択され、Ｒ¹及びＲ²は、それぞれ独立に、水素、tert-ブチルジメチルシリル、メチルジ−tert-ブチルシリル、イソプロピルジメチルシリル、メチルジイソプロピルシリル、（トリフェニルメチル）ジメチルシリル、及びtert-ブチルジフェニルシリルからなる群より選択され、Ｒ³は水素またはアミノ保護基である。ただし、Ｒ¹とＲ²は同じであっても異なっていてもよく、及びＲ¹、Ｒ²及びＲ³は同時に水素を表さない。 Examples of intermediates useful in synthesizing the pyrrolidine compounds of the present invention include compounds having the general formula (II).

(II)
Wherein R is selected from the group consisting of hydrogen, a substituted or unsubstituted alkyl group, a substituted or unsubstituted aryl group, a substituted or unsubstituted aralkyl group, and a substituted or unsubstituted alkylaryl group; Is the S-form or R-form, and the configuration of the 3-position carbon and 4-position carbon is selected from the combination of (3R, 4R) and (3S, 4S), and R ¹ and R ² are each independently hydrogen, selected from the group consisting of tert-butyldimethylsilyl, methyldi-tert-butylsilyl, isopropyldimethylsilyl, methyldiisopropylsilyl, (triphenylmethyl) dimethylsilyl, and tert-butyldiphenylsilyl, wherein R ³ is hydrogen or an amino protecting group is there. However, R ¹ and R ² may be the same or different, and R ¹ , R ² and R ³ do not represent hydrogen at the same time.

  In the above definition, the definition of the alkyl group, aryl group, aralkyl group, alkylaryl group, and substituent on each group is as described for formula (I).

Depending on the synthesis method, R ¹ and R ^{2 in} the formula (II) may pass through a hydroxy protecting group other than the protecting group. The hydroxy protecting group in this case is an ether type protecting group, an ester type protecting group, a carbonate type protecting group, etc., which are usually used for protecting a hydroxyl group, such as “Protective Groups in Organic Synthesis” TW Green, John Wiley & Sons. The protecting group for hydroxy group described in, can be used as appropriate. R ² may be the same protecting group as R ¹ .

With respect to R ³ , the amino protecting group may be any protecting group commonly used to protect secondary amino groups. Specifically, amino acids described in tert-butyloxycarbonyl group (Boc group), benzyloxycarbonyl group (Cbz group), benzyl group, or “Protective Groups in Organic Synthesis” TW Green, John Wiley & Sons. A protecting group or the like can be appropriately used.

The method for producing the pyrrolidine compound of the present invention will be described below.
The enzyme inhibitor of the present invention, 2-acylaminomethyl-3,4-dihydroxypyrrolidine, is a synthetic intermediate, N-protected-2-aminomethyl-3,4-dihydroxypyrrolidine (Non-patent Document Chem. Pharm. Bull., 1987, 35, 3995-3999 .; Patent Document: Japanese Patent Application No. 2004-052693) can be synthesized by selectively modifying the 2-position amino group. In that case, the configuration of the product depends on the synthetic intermediate used.

Scheme 1 outlines one embodiment of a method for producing the pyrrolidine compound of the present invention. Optical functional cyclic nitrone (III) 1- and 2-position functional group conversion is performed, aminomethyl group is present at the 2-position, the 3,4-position is an optional hydroxy protecting group, and the nitrogen atom on the ring is substituted. The pyrrolidine compound (II) protected with any amino protecting group is obtained. The pyrrolidine compound (I) of the present invention can be produced by acylating the aminomethyl group at the 2-position and removing all protecting groups for this (II).

スキーム１ 2-アシルアミノメチル-3,4-ジヒドロキシピロリジン体の合成（例）

Scheme 1 Synthesis of 2-acylaminomethyl-3,4-dihydroxypyrrolidine (example)

  More specifically, a method for producing the pyrrolidine compound of the present invention using a specific protecting group will be described according to Scheme 2. Scheme 2 shows a route for the synthesis of (2R, 3S, 4S) 2-acylaminomethylpyrrolidine compounds.

TBS＝ｔ−ブチルジメチルシリル、Boc=t-ブチルオキシカルボニル
スキーム２ （２Ｒ,３Ｓ,４Ｓ）2-アシルアミノメチルピロリジン化合物の合成のルート

TBS = t-butyldimethylsilyl, Boc = t-butyloxycarbonyl Scheme 2 Route of synthesis of (2R, 3S, 4S) 2-acylaminomethylpyrrolidine compounds

The compound (5) is synthesized via a known compound (4) derived from the compound (1) (Japanese Patent Laid-Open No. 2002-88059). In introducing the tert-butoxycarbonyl group into the nitrogen atom of compound (4), a commercially available or prepared protecting group introduction reagent can be used. Boc ₂ O can be used in excess, but for ease of isolation and purification, the amount used is preferably 1 to 10 equivalents, more preferably 1.5 to 3 equivalents per 1 equivalent of amino nitrile. is there. As the base to be coexisted, an organic base such as tertiary amine represented by triethylamine or pyridine and an inorganic base represented by potassium carbonate can be used. Examples of the reaction solvent include methanol, ethanol and other alcohols, water, ethyl acetate, chloroform, methylene chloride, tetrahydrofuran, diethyl ether, 1,2-dimethoxyethane, 1,4-dioxane, and mixed solutions thereof. Can be used. The reaction temperature is usually preferably about -20 to 50 ° C, particularly preferably 0 to 20 ° C. The reaction time is usually preferably about 5 minutes to 2 days. The TBDMS group of the N-Boc aminonitrile derivative thus obtained can be removed by the action of fluoride ions. For this reaction, a method using commercially available TBAF (tetrahydrofuran solution) can be applied. The amount of TBAF (tetrahydrofuran solution) used is 2 to 10 equivalents, preferably 1.5 to 3 equivalents, with respect to 1 equivalent of the N-Boc aminonitrile derivative. As the reaction solvent, tetrahydrofuran or the like can be usually used. The reaction temperature is usually preferably about -10 to 50 ° C, particularly preferably 0 to 20 ° C. The reaction time is usually preferably about 5 minutes to 2 days. The diastereomeric excess of 1-tert-butoxycarbonyl-2-cyano-3,4-dihydroxypyrrolidine produced in this reaction is similar to that of the starting material, but this compound can be purified by recrystallization. It is. As the recrystallization solvent, methanol, ethanol and other alcohols, acetone, ethyl acetate and the like can be used. The cyano group of 1-tert-butoxycarbonyl-2-cyano-3,4-dihydroxypyrrolidine undergoes addition of hydrogen in the presence of a heterogeneous catalyst typified by palladium-activated carbon to the corresponding aminomethyl group. Converted. Although palladium-activated carbon can be used in excess, it is desirable to limit the amount used to avoid loss due to physical adsorption of the product. The amount used is preferably 1 to 200% equivalent, more preferably 10 to 50% equivalent to 1 equivalent of raw material. The reducing agent used in this reaction is preferably hydrogen or a gas containing hydrogen, and a known method can be applied as the supply method. For example, a gas containing hydrogen may be passed through the reaction solution, the reaction solution may be stirred in a hydrogen gas atmosphere, or hydrogen may be transferred from cyclohexadiene or the like in the reaction system. As reaction solvents, alcohols such as methanol and ethanol, water, ethyl acetate, acetic acid, formic acid, tetrahydrofuran, diethyl ether, 1,2-dimethoxyethane, 1,4-dioxane, and mixed solutions thereof are usually used. it can. The reaction temperature is usually preferably about -20 to 50 ° C, particularly preferably 0 to 20 ° C. The reaction time is usually preferably about 5 minutes to 2 days.

  The terminal amino group of compound (5) gives the corresponding amide form (acylated amino form) by reaction with carboxylic acids. In the construction of the amide bond, a general chemical that appropriately activates the carboxylic acid can be used, and a known peptide synthesis method can also be used as appropriate. That is, the carboxylic acid used is selected and introduced from acid anhydrides, acid halides, active esters and the like. When carboxylic acid is used, it is reacted as an active ester using N-hydroxysuccinimide or the like and N- (3-dimethylaminopropyl) -N′-ethylcarbodiimide hydrochloride (EDCI) or dicyclohexylcarbodiimide (DCC). However, it is particularly effective to use O-benzotriazol-1-yl-N, N, N ′, N′-tetramethyluronium hexafluoroborate (HBTU) as an additive type condensing agent. Although these chemicals can be used in excess, for ease of isolation and purification, the amount used is preferably 0.1 to 10 equivalents, more preferably 0 to 1 equivalent of aminomethyl compound (5). 0.5-3 equivalents. As the base to be coexisted, an organic base such as tertiary amine represented by triethylamine or pyridine and an inorganic base represented by potassium carbonate can be used. Examples of the reaction solvent include methanol, ethanol and other alcohols, water, ethyl acetate, chloroform, methylene chloride, tetrahydrofuran, diethyl ether, 1,2-dimethoxyethane, 1,4-dioxane, and mixed solutions thereof. Can be used. The reaction temperature is usually preferably about -20 to 50 ° C, particularly preferably 0 to 20 ° C. The reaction time is usually preferably about 5 minutes to 2 days. The N-Boc group of the resulting condensate can be removed by the action of trifluoroacetic acid (TFA). In this reaction, TFA can be used as a solvent, but a mixed solution with water may be used. The amount of TFA used is 0.1 to 10 ml, preferably 1 to 3 ml, based on 1 mmol of the N-Boc amide derivative, and may be a mixed solution with half to the same amount of water. The reaction temperature is usually preferably about −10 to 80 ° C., particularly preferably 10 to 30 ° C. The reaction time is usually preferably about 30 minutes to 2 days.

  In Scheme 2 described above, the diastereomeric excess of 2-acylaminomethyl-3,4-dihydroxypyrrolidine to be finally produced is similar to that of 2-aminomethyl, but each step described in each step Purification with is possible. Therefore, 2-acylaminomethyl-3,4-dihydroxypyrrolidine having high purity can be easily obtained.

  2-Acylaminomethyl-3,4-dihydroxypyrrolidine can be purified using ion exchange resins such as DOWEX50WX-8 (manufactured by Dow Chemical Co.), and after purification, concentrated with hydrochloric acid and dried. Thus, the hydrochloride is obtained.

  In general, glycosidase is a general term for endoglycosidases such as chitinase and lysozyme, and exoglycosidases such as glucosidase, but the substrate recognition method is different. As an inhibitor for glycosidase that shows strict substrate recognition, a higher molecular design is required, but the pyrrolidine compound of the present invention exhibits a glycosidase inhibitory action, and the effect varies depending on the functional group to which it is bound. This fact is an index for estimating the enzyme-inhibitor affinity outside the active site of the enzyme.

In particular, the 2-acylaminomethyl compounds of the present invention are notable for specific enzyme inhibitory activity or relatively enzyme selective. In particular, α-glucosidase, β-hexosaminidase, β-glucosidase, and specific β-glucuronidase have an inhibitory effect, but other enzymes have almost no inhibitory effect. Such enzyme specificity was not found in the conventionally known pyrrolidine compounds. A compound having such enzyme-selective inhibitory activity is very useful because it has various uses as a therapeutic agent, a medical experimental tool, etc. for a disease related to the enzyme or a sugar chain having a similar binding mode. .
For example, α-glucosidase inhibition controls blood glucose levels in the treatment of non-insulin dependent diabetes, β-hexosaminidase inhibition and α-mannosidase inhibition suppresses glycoprotein sugar chain processing in cancer cells, and β-glucosidase This suggests that inhibition may help to suppress abnormal substrate accumulation in lysosomal storage diseases.
Next, the present invention will be described more specifically with reference to examples.

Production method of compound (5) (reference example)
(2R, 3S, 4S) 3,4-bis- (tert-butyldimethylsilyloxy) -2-cyanopyrrolidine (4) (2.39 g, obtained from L-tartaric acid by a known method (Japanese Patent Laid-Open No. 2002-88059) To a solution of 6.70 mmol) in methanol (120 ml), Boc ₂ O (2.19 g, 10.1 mmol) and triethylamine (2.18 ml, 20.1 mmol) were added and stirred at room temperature for 12 hours. After completion of the reaction, the reaction solution was concentrated and the residue was purified by silica gel flash column chromatography (n-hexane: ethyl acetate = 19: 1) and (2R, 3S, 4S) 3,4-bis- (tert-butyl Dimethylsilyloxy) -1-tert-butyloxycarbonyl-2-cyanopyrrolidine (2.8 g, 92%) was obtained. This compound is a mixture with the 2-position epimer (de = 76 to 88%), which is equivalent to the purity of the raw material (4). In addition, it is difficult to separate each of the 2-position epimers with a silica gel column. To a solution of this (2R, 3S, 4S) 3,4-bis- (tert-butyldimethylsilyloxy) -1-tert-butyloxycarbonyl-2-cyanopyrrolidine (2.41 g, 5.27 mmol) in THF (48 ml) , Tetra n-butylammonium fluoride (1M in THF, 13.2 ml, 13.2 mmol) was added, and the mixture was stirred at room temperature for 12 hours. The reaction solution was concentrated to remove most of THF, and then diluted with ethyl acetate, washed with saturated brine, dried and concentrated to quantitatively obtain the desilylated product as crude crystals. Crude crystals containing the 2-position epimer (de = 76-88%) were purified by recrystallization from ethyl acetate to obtain pure (2R, 3S, 4S) 1 -tert-butyloxycarbonyl-2-cyano-3, 4-Dihydroxypyrrolidine is isolated in 50-70% yield. Pure (2R, 3S, 4S) 1-tert-butyloxycarbonyl-2-cyano-3,4-dihydroxypyrrolidine (13.5 mg, 0.191 mmol) purified by recrystallization was dissolved in ethanol (6 ml), and 20 % Palladium hydroxide-activated carbon (6.8 mg) and acetic acid (0.75 ml) were added, and the mixture was stirred at room temperature for 30 minutes in a hydrogen gas atmosphere. After completion of the reaction, the solution was filtered through Celite and concentrated to quantitatively obtain the corresponding N-Boc aminomethyl derivative (5). Furthermore, it is isolated as salt-free by ion exchange column (eluted with 1.3N ammonia water) with DOWEX50WX-8 (manufactured by Dow Chemical).

Physical property values of compound (5) (reference example)
Molecular formula: C ₁₀ H ₂₀ N ₂ O ₄
Molecular weight: 232.28
¹ H NMR (CD ₃ OD) δ 4.36-3.95, 3.65-3.25 and 3.20-3.10 (each m, 7H), 1.50 and 1.48 (each s, 9H, t-Bu). (Due to the Boc group in the structure Observed as a rotamer mixture)
¹³ C NMR (CD ₃ OD) δ 179.11, 158.44, 82.01, 77.81, 74.92, 58.90, 58.31, 54.10, 40.94, 28.77, 28.73, 28.64, 23.29, 18.38. (Observed as a rotamer mixture due to the Boc group in the structure Was)

General production method of compound (6) -A-
N-Boc aminomethyl compound (5) was dissolved in methanol (20 times the weight of compound (5)), and triethylamine (3 equivalents of compound (5)) and acid anhydride (3 equivalents of compound (5)) Alternatively, acid chloride (3 equivalents of compound (5)) was added and stirred at room temperature for 2 hours. After completion of the reaction, the condensate obtained by concentrating the reaction solution was purified by acetylation, silica gel column, and deacetylation as necessary. To the collected condensate, TFA (10 times the weight of compound (5)) was added and stirred for 1 hour. After completion of the reaction, the reaction solution is concentrated and the resulting residue is subjected to ion exchange column (eluted with 1.3N aqueous ammonia) using DOWEX50WX-8 (manufactured by Dow Chemical Co.), concentrated with 1M aqueous hydrochloric acid, and lyophilized. This gave pure hydrochloride.

General production method of compound (6) -B-
Carboxylic acid was dissolved in methylene chloride (20 times the weight of compound (5)), and N-hydroxysuccinimide (1.2 equivalents of compound (5)) and N- (3-dimethylaminopropyl) -N′-ethylcarbodiimide Hydrochloric acid salt (1.2 equivalent of compound (5)) was added and stirred at room temperature for 1 hour, and then N-Boc aminomethyl compound (5) was added and stirred for 10 hours. After completion of the reaction, the condensate obtained by concentrating the reaction solution was purified by acetylation, silica gel column, and deacetylation as necessary. To the collected condensate, TFA (10 times the weight of compound (5)) was added and stirred for 1 hour. After completion of the reaction, the reaction solution is concentrated and the resulting residue is subjected to ion exchange column (eluted with 1.3N aqueous ammonia) using DOWEX50WX-8 (manufactured by Dow Chemical Co.), concentrated with 1M aqueous hydrochloric acid and freeze-dried. This gave pure hydrochloride.

General process for preparing compound (6) -C-
N-Boc aminomethyl compound (5) is dissolved in ethanol (20 times the weight of compound (5)) and O-benzotriazol-1-yl-N, N, N ′, N′-tetramethyl at 0 ° C. Uronium hexafluoroborate (HBTU) (3 equivalents of compound (5)) was added and stirred at room temperature for 3 hours. After completion of the reaction, the condensate obtained by concentrating the reaction solution was purified by acetylation, silica gel column, and deacetylation as necessary. To the collected condensate, TFA (10 times the weight of compound (5)) was added and stirred for 1 hour. After completion of the reaction, the reaction solution is concentrated and the residue obtained is ion exchange column (eluted with 1.3N ammonia water) using DOWEX50WX-8 (manufactured by Dow Chemical Co.), concentrated with 1M hydrochloric acid and lyophilized. This gave pure hydrochloride.

Physical property value of compound (6) Acetic acid conjugate hydrochloride synthesized by method A (6a)
Molecular formula: C ₇ H ₁₅ N ₂ O ₃ Cl
Molecular weight: 210.66
Specific rotation: -15.3 ° (c 1.0, H ₂ O)
¹ H NMR (D ₂ O) δ 4.18 (d, 1H, J = 4.8 Hz, H-4), 4.06 (br s, 1H, H-3), 3.69 (ddd, 1H, J = 3.3, 8.6, 8.6 Hz, H-2), 3.49 (dd, 1H, J = 4.4, 13.0 Hz, H-5a), 3.44-3.41 (m, 2H, H-2 '), 3.06 (d, 1H, J = 13.0 Hz, H-5b), 1.81 (s, 3H, Ac).
¹³ C NMR (D ₂ O) δ 175.10, 74.42, 73.85, 61.42, 51.00, 36.04, 21.67.

Benzoic acid conjugate hydrochloride synthesized by Method A (6b)
Molecular formula: C ₁₂ H ₁₇ N ₂ O ₃ Cl
Molecular weight: 272.73
Specific rotation: -1.8 ° (c 1.3, H ₂ O)
¹ H NMR (D ₂ O) δ 7.59 (d, 2H, J = 0.0 Hz, aromatic), 7.46-7.32 (m, 3H, aromatic), 4.24 (d, 1H, J = 4.4 Hz, H-4), 4.15 (br s, 1H, H-3), 3.86 (ddd, 1H, J = 3.2, 5.9, 7.4 Hz, H-2), 3.68 (dd, 1H, J = 7.4, 14.4 Hz, H-2'a ), 3.63 (dd, 1H, J = 5.9, 14.4 Hz, H-2'b), 3.57 (dd, 1H, J = 4.4, 13.0 Hz, H-5a), 3.12 (d, 1H, J = 13.0 Hz) , H-5b).
¹³ C NMR (D ₂ O) δ 171.49, 132.78, 132.45, 128.78, 127.12, 74.60, 73.89, 61.59, 51.13, 36.61.

Pivalinamide synthesized by Method B (6c)
Molecular formula: C ₁₀ H ₂₁ N ₂ O ₃ Cl
Molecular weight: 252.74
Specific rotation: -10.9 ° (c 0.7, H ₂ O)
¹ H NMR (D ₂ O) δ 4.19 (d, 1H, J = 4.4 Hz, H-4), 4.04 (br s, 1H, H-3), 3.69 (ddd, 1H, J = 3.0, 6.6, 7.2 Hz, H-2), 3.50 (dd, 1H, J = 4.4, 13.0 Hz, H-5a), 3.47 (dd, 1H, J = 7.2, 14.7 Hz, H-2'a), 3.41 (dd, 1H , J = 6.6, 14.7 Hz, H-2'b), 3.12 (d, 1H, J = 13.0 Hz, H-5b), 0.99 (s, 9H, t-Bu).
¹³ C NMR (D ₂ O) δ 183.50, 74.43, 73.76, 61.48, 51.08, 38.38, 36.21, 26.36.

Hexanoic acid conjugate hydrochloride synthesized by Method A (6d)
Molecular formula: C ₁₁ H ₂₃ N ₂ O ₃ Cl
Molecular weight: 266.76
¹ H NMR (D ₂ O) δ 4.19 (d, 1H, J = 4.3 Hz, H-4), 4.05 (br s, 1H, H-3), 3.70 (ddd, 1H, J = 3.0, 6.8, 7.0 Hz, H-2), 3.50 (dd, 1H, J = 4.3, 13.0 Hz, H-5a), 3.46 (dd, 1H, J = 7.0, 14.4 Hz, H-2'a), 3.41 (dd, 1H , J = 6.8, 14.4 Hz, H-2'b), 3.08 (d, 1H, J = 13.0 Hz, H-5b), 2.08 (t, 2H, J = 7.3 Hz, C (= O) CH _2- C ₄ H ₇ ), 1.39 (quint, 2H, J = 7.2 Hz, C (= O) CH ₂ -CH ₂ -C ₃ H ₅ ), 1.15-1.05 (m, 4H, C (= O) C ₂ H ₄ -C ₂ H ₄ -CH ₃ ), 0.66 (t, 3H, J = 7.2 Hz, C (= O) C ₄ H ₈ -CH ₃ ).
¹³ C NMR (D ₂ O) δ 178.28, 74.33, 73.80, 61.30, 51.03, 35.78, 35.45, 30.44, 24.80, 21.58, 13.12.

L-Phenylalanine conjugate hydrochloride synthesized by method C (6e)
Molecular formula: C ₁₄ H ₂₂ N ₃ O ₃ Cl
Molecular weight: 315.80
Specific rotation: + 29.3 ° (c 1.1, H ₂ O)
¹ H NMR (D ₂ O) δ 7.27-7.12 (m, 5H, Ph), 4.17 (d, 1H, J = 4.6 Hz, H-4), 4.08 (t, 1H, J = 7.5 Hz, C (= O) CH <), 3.78 (br s, 1H, H-3), 3.52 (ddd, 1H, J = 3.0, 6.6, 7.2 Hz, H-2), 3.48 (dd, 1H, J = 4.6, 13.0 Hz , H-5a), 3.45 (dd, 1H, J = 6.6, 14.3 Hz, H-2'a), 3.39 (dd, 1H, J = 7.2, 14.3 Hz, H-2'b), 3.08 (d, 1H, J = 13.0 Hz, H-5b), 3.05 (dd, 1H, J = 7.5, 13.9 Hz, -CH ₂ -Ph), 3.01 (dd, 1H, J = 7.5, 13.9 Hz, -CH ₂ -Ph ).
¹³ C NMR (D ₂ O) δ 169.72, 133.78, 129.29, 129.21, 128.04, 73.99, 73.85, 60.54, 54.39, 51.07, 40.34, 36.74, 35.52.

L-valine conjugate hydrochloride synthesized by method C (6f)
Molecular formula: C ₁₀ H ₂₂ N ₃ O ₃ Cl
Molecular weight: 267.75
Specific rotation: 0 ° (c 0.6, H ₂ O)
¹ H NMR (D ₂ O) δ 4.21 (d, 1H, J = 4.0 Hz, H-4), 4.08 (br s, 1H, H-3), 3.76 (ddd, 1H, J = 3.3, 7.3, 7.3 Hz, H-2), 3.66 (d, 1H, J = 5.9 Hz, C (= O) CH <), 3.58-3.49 (m, 3H, H-5a, 2'a, 2'b), 3.11 ( d, 1H, J = 12.8 Hz, H-5b), 2.05 (oct, 1H, J = 6.6 Hz, CH (CH ₃ ) ₂ ), 0.84 and 0.83 (each d, 6H, J = 6.6 Hz, CH (CH ₃ ) ₂ ).
¹³ C NMR (D ₂ O) δ 170.10, 74.31, 73.84, 60.52, 58.56, 51.16, 35.90, 29.71, 17.63, 16.74.

Enzyme inhibitory activity of 2-acylaminomethyl-3,4-dihydroxypyrrolidine type compounds;
Inhibition experiments on common glycosidases on the market were performed as follows. To each of the synthesized pyrrolidine compounds and substrates (see Table 1 below) adjusted to about 5 mM, each enzyme (see Table 1 below for the concentration) was added, and the resulting p-nitrophenol was quantified from the absorbance at 400 nm. Thus, the decomposition rate of the substrate was determined, and the enzyme reaction inhibition rate of each compound was determined. The results are shown in Tables 2-5. In the table, compound (6) is hydrochloride.

Table 1

  The enzyme inhibitor of the present invention has various uses as a therapeutic agent for a disease related to an enzyme or a sugar chain having a similar binding mode, a medical experimental tool, and the like.

Claims (5)

A pyrrolidine compound having the following general formula (I):

(I)
Wherein R is selected from the group consisting of hydrogen, a substituted or unsubstituted alkyl group, a substituted or unsubstituted aryl group, a substituted or unsubstituted aralkyl group, and a substituted or unsubstituted alkylaryl group; Is S-form or R-form, and the configuration of the 3-position carbon and 4-position carbon is selected from the combination of (3R, 4R) and (3S, 4S). In each general formula, a mark * next to a carbon atom means that the carbon atom is an asymmetric carbon. A pyrrolidine compound having the following general formula (II):

(II)
Wherein R is selected from the group consisting of hydrogen, a substituted or unsubstituted alkyl group, a substituted or unsubstituted aryl group, a substituted or unsubstituted aralkyl group, and a substituted or unsubstituted alkylaryl group; Is the S-form or R-form, and the configuration of the 3-position carbon and 4-position carbon is selected from the combination of (3R, 4R) and (3S, 4S), and R ¹ and R ² are each independently hydrogen, selected from the group consisting of tert-butyldimethylsilyl, methyldi-tert-butylsilyl, isopropyldimethylsilyl, methyldiisopropylsilyl, (triphenylmethyl) dimethylsilyl, and tert-butyldiphenylsilyl, wherein R ³ is hydrogen or an amino protecting group is there. However, R ¹ and R ² may be the same or different, and R ¹ , R ² and R ³ do not represent hydrogen at the same time. A method for producing a pyrrolidine compound represented by the general formula (I) by deprotecting the hydroxy protecting group and / or the amino protecting group of the compound of the general formula (II).

(II)
Wherein R is selected from the group consisting of hydrogen, a substituted or unsubstituted alkyl group, a substituted or unsubstituted aryl group, a substituted or unsubstituted aralkyl group, and a substituted or unsubstituted alkylaryl group; Is the S-form or R-form, and the configuration of the 3-position carbon and 4-position carbon is selected from the combination of (3R, 4R) and (3S, 4S), and R ¹ and R ² are each independently hydrogen, selected from the group consisting of tert-butyldimethylsilyl, methyldi-tert-butylsilyl, isopropyldimethylsilyl, methyldiisopropylsilyl, (triphenylmethyl) dimethylsilyl, and tert-butyldiphenylsilyl, wherein R ³ is hydrogen or an amino protecting group is there. However, R ¹ and R ² may be the same or different, and R ¹ , R ² and R ³ do not represent hydrogen at the same time.

(I)
Wherein R is selected from the group consisting of hydrogen, a substituted or unsubstituted alkyl group, a substituted or unsubstituted aryl group, a substituted or unsubstituted aralkyl group, and a substituted or unsubstituted alkylaryl group; Is S-form or R-form, and the configuration of the 3-position carbon and 4-position carbon is selected from the combination of (3R, 4R) and (3S, 4S). In each general formula, a mark * next to a carbon atom means that the carbon atom is an asymmetric carbon. The method of Claim 3 including producing the compound represented by general formula (II) by acylating the compound represented by general formula (III).

(III)
In the formula, the configuration at the 2-position carbon is S-form or R-form, and the configuration at 3-position carbon and 4-position carbon is selected from the combination of (3R, 4R) and (3S, 4S), and R ¹ and R ² are each independently selected from the group consisting of hydrogen, tert-butyldimethylsilyl, methyldi-tert-butylsilyl, isopropyldimethylsilyl, methyldiisopropylsilyl, (triphenylmethyl) dimethylsilyl, and tert-butyldiphenylsilyl; R ³ is hydrogen or an amino protecting group. However, R ¹ and R ² may be the same or different, and R ¹ , R ² and R ³ do not represent hydrogen at the same time. An enzyme inhibitor comprising the pyrrolidine compound represented by the general formula (I) according to claim 1 as an active compound.