(19)
`
`J
`
`
`
`(12)
`
`Europaisches Patentamt
`European Patent Office
`Office europeen des brevets
`
`(11)
`
`E P 0 9 4 9 2 6 0 A 1
`
`EUROPEAN PATENT APPLICATION
`published in accordance with Art. 158(3) EPC
`
`(43) Date of publication:
`13.10.1999 Bulletin 1999/41
`
`(21 ) Application number: 97949235.2
`
`(22) Date of filing: 25.12.1997
`
`(84) Designated Contracting States:
`AT BE CH DE DK ES Fl FR GB GR IE IT LI LU MC
`NL PT SE
`
`(30) Priority: 26.12.1996 JP 34785196
`26.12.1996 JP 34785396
`26.12.1996 JP 34789596
`
`(71) Applicants:
`• Ube Industries, Ltd.
`Ube-shi, Yamaguchi-ken 755-0052 (JP)
`• TANABE SEIYAKU CO., LTD.
`Chuo-ku, Osaka-shi, Osaka 541-8505 (JP)
`
`(72) Inventors:
`• KITA, Jun-ichiro
`Ube Res. Laboratory
`Yamaguchi 755 (JP)
`
`(51) int. CI.6: C07D 4 0 1 / 1 2
`// (A61 K31/445, C07M7:00)
`
`(86) |nternationa| app|ication number:
`PCT/JP97/04826
`
`(87) International publication number:
`WO 98/29409 (09.07.1998 Gazette 1998/27)
`
`• FUJIWARA, Hiroshi
`Ube Res. Lab. Ube Ind.
`Yamaguchi 755 (JP)
`• TAKAMURA, Shinji
`Ube Res. Laboratory,
`Yamaguchi 755 (JP)
`• YOSHIOKA, Ryuzo
`Shimamotocho, Mishima-gun Osaka 618 (JP)
`. OZAKI, Yauhiko
`Osaka 572 (JP)
`. YAMADA Shin-ichi
`Hyogo 665 (JP)
`
`(74) Representative: HOFFMANN - EITLE
`Patent- und Rechtsanwalte
`Arabellastrasse 4
`81925 Munchen (DE)
`
`(54) ACID-ADDITION SALTS OF OPTICALLY ACTIVE PIPERIDINE COMPOUND AND PROCESS
`FOR PRODUCING THE SAME
`
`The present invention is to provide a benzenesulfonic acid salt and a benzoic acid salt of (S)-4-[4-[(4-chloroph-
`(57)
`enyl)(2-pyridyl)methoxy]piperidino]butanoic acid represented by the formula (I):
`
`( I )
`
`wherein * represents an asymmetric carbon, which are excellent in antihistaminic activity and anti-allergic activ-
`ity, and a process for producing the same.
`
`Printed by Xerox (UK) Business Services
`2.16.7/3.6
`
`O
`CO
`CM
`<7>
`<7>
`O
`Q_
`LU
`
`MYLAN Ex. 1017, Page 1
`
`

`
`EP 0 949 260 A1
`
`Description
`
`Technical field
`
`[0001] This invention relates to benzenesulfonic acid salt or benzoic acid salt of (S)-4-[4-[(4-chlorophenyl)(2-pyri-
`dyl)methoxy]piperidino]butanoic acid which is excellent in antihistaminic activity and antiallergic activity, a process for
`preparing the same, and an optically resolving method of 4-[(4-chlorophenyl)(2-pyridyl)methoxy]piperidine which is
`important as a racemic intermediate thereof. The acid addition salt has little hygroscopicity and excellent in physico-
`chemical stability so that it is particularly suitable compound as a medicine. Also, the present invention relates to a med-
`ical composition containing the compound as an effective ingredient.
`
`Background art
`
`[0002] A piperidine compound (II) represented by the formula (II):
`
`( I D
`
`where in A represents a lower alkyl group, hydroxyl group, a lower alkoxy group, amino group, a lower alkylamino
`group, phenyl group or a lower alkyl -substituted phenyl group,
`
`5
`
`10
`
`15
`
`20
`
`25
`
`30
`
`or a salt thereof described in Japanese Provisional Patent Publication No. 25465/1 990 has characteristics that a sec-
`35 ondary effect such as stimulation or suppression on the central nerves, which often appears in the conventional antihis-
`taminic compound, can be reduced as little as possible, and is expected to be a medicine for therapeutic treatment of
`allergic skin diseases such as a nettle rash, eczema, dermatitis and the like, allergic rhinitis, sneeze, mucus, cough due
`to respiratory inflammation such as cold and the like, and bronchial asthma.
`[0003] For producing the piperidine compound (II) effectively as a more preferred optical isomer for a medicine, it is
`40 desired to use the optically resolved product as a starting material by optically resolving an intermediate. However, this
`piperidine compound (II) has one asymmetric carbon atom but the method of isolating its optically active isomer from
`the racemic mixture has not been known as of today.
`It has been generally known that optical isomers show different pharmacological activity or safety and there
`[0004]
`are also differences in the metabolic rates and the protein binding ratios therebetween (Pharmacia, 25 (4), pp. 31 1 -336,
`45 1989). Accordingly, for providing a medicine, a pharmaceutical^ preferable optical isomer with high optical purity is
`required. Also, in order to secure high quality of said optical isomer as a medicine, it is desirable that the isomer has
`superior properties in physicochemical stability.
`[0005] The present inventors have studied intensively to solve the above problems. As the result, they have found that
`a benzenesulfonic acid salt or a benzoic acid salt of optically active (S)-4-[4-[(4-chlorophenyl)(2-pyridyl)methoxy]-pipe-
`ridino]butanoic acid represented by the following formula (I) has excellent stability which is preferred as a medicine
`whereby accomplished the present invention.
`
`50
`
`Disclosure of the invention
`
`55
`
`[0006] The first invention relates to a benzenesulfonic acid salt or a benzoic acid salt of an optically active piperidine
`compound (I) represented by the formula (I):
`
`2
`
`MYLAN Ex. 1017, Page 2
`
`

`
`EP 0 949 260 A1
`
`( I )
`
`10
`
`15
`
`wherein * represents an asymmetric carbon, which has an absolute configuration of (S).
`[0007] The second invention relates to a process for preparing a benzenesulfonic acid salt or a benzoic acid salt of
`an optically active piperidine compound by reacting the optically active piperidine compound represented by the above
`formula (I) with an absolute configuration of (S) with benzenesulfonic acid or benzoic acid to form a salt.
`[0008] The third invention relates to a medical composition which comprises a benzenesulfonic acid salt of (S)-4-[4-
`[(4-chlorophenyl)(2-pyridyl)methoxy]piperidino]butanoic acid or a benzoic acid salt of the same as an effective ingredi-
`ent.
`[0009] The invention further relates to a process for preparing a benzenesulfonic acid salt or a benzoic acid salt of the
`optically active piperidine compound (I) represented by the above formula (I) which comprises reacting (+)-4-[(4-chlo-
`rophenyl)(2-pyridyl)methoxy]piperidine with the optically active propionic acid compound (VII) represented by the fol-
`lowing formula (VII) or the optically active N-acyl-amino acid; separating and collecting less soluble diastereomeric salt
`25 by utilizing the difference in solubilities of the formed two kinds of diastereomeric salts; decomposing the resulting salt;
`reacting an ester represented by the formula (V):
`
`20
`
`30
`
`35
`
`40
`
`45
`
`50
`
`55
`
`o
`
`O R
`
`(V)
`
`wherein R represents a lower alkyl group such as methyl group, ethyl group, etc., and W represents a leaving
`group such as a halogen atom or a reactive ester group such as methanesulfonyloxy group, p-toluenesulfonyloxy group,
`etc.,
`with the resulting (S)-4-[(4-chlorophenyl)(2-pyridyl)-methoxy]piperidine to obtain (S)-4- [(4-chlorophenyl)(2-pyri-
`dyl)methoxy]piperidine butanoic acid ester represented by the formula (VI):
`
`( V I )
`
`CI
`
`wherein R and * have the same meaning as defined above, hydrolyzing the resulting compound; and reacting
`the hydrolyzed compound with benzenesulfonic acid or benzoic acid to form a salt.
`
`Best mode for carrying out the invention
`
`[001 0] A benzenesulfonic acid salt or a benzoic acid salt of (S)-piperidine compound (I) can be produced by the
`method represented by the following reaction scheme (1):
`
`3
`
`MYLAN Ex. 1017, Page 3
`
`

`
`EP 0 949 260 A1
`
`R e a c t i o n s c h e m e
`
`(1)
`
`wherein HX represents benzenesulfonic acid or benzoic acid, and * has the same meaning as defined above,
`(hereinafter referred to as a salt-forming reaction).
`[001 1 ]
`In the salt-forming reaction, benzenesulfonic acid or benzoic acid can be used in an amount of 0.8 to 2.5-fold
`mole, preferably 0.9 to 1 .2-fold mole based on 1 mole of the (S)-piperidine compound (I).
`[001 2] As a solvent to be used in the salt-forming reaction, it is not particularly limited so long as it does not participate
`in the reaction, and there may be mentioned, for example, nitriles such as acetonitrile and propionitrile; esters such as
`methyl acetate and ethyl acetate; alcohols such as methanol, ethanol, 1-propanol, 2-propanol, etc.; acetone, dimethyl-
`formamide, etc., and preferably ethanol, 2-propanol, acetonitrile and ethyl acetate. The solvent to be used in the present
`invention may be used alone or may be in admixture of the above-mentioned two or more kinds of optional solvents.
`[0013] An amount of the solvent to be used in the salt-forming reaction is usually 0.5 to 30 liters, preferably 0.8 to 20
`liters, more preferably 1 to 10 liters per mole of the (S)-piperidine compound (I).
`[001 4] A temperature of the salt-forming reaction is, for example, 5 to 50 °C, preferably 1 0 to 35 °C, and a temperature
`at the time of salt precipitation is, for example, -30 °C to 30 °C, preferably -10 °C to 15 °C. Also, a method of addition is
`not particularly limited, but, for example, there may be mentioned a method in which benzenesulfonic acid or benzoic
`acid dissolved in a solvent is added to a mixed solution of the (S)-piperidine compound (I) and a solvent.
`[001 5] The formed salt of the (S)-piperidine compound (I) can be easily obtained in accordance with the conventional
`method in this field of technology by, for example, collecting after separation with filtration, centrifugation, etc., washing
`and then drying.
`[0016] Next, a process for preparing an (S)-piperidine compound (I) of the present invention will be explained.
`[001 7] The (S)-piperidine compound (I) of the present invention can be prepared by the method shown in the following
`reaction scheme (2):
`
`R e a c t i o n s c h e m e
`
`(2)
`
`CI
`
`(VI)
`
`CI
`
`( i )
`
`wherein W represents a leaving group, including a halogen atom such as chlorine atom, bromine atom, iodine
`atom, etc.; or a reactive ester group such as methanesulfonyloxy group, p-toluenesulfonyloxy group, etc., and R repre-
`
`MYLAN Ex. 1017, Page 4
`
`

`
`5
`
`10
`
`15
`
`20
`
`25
`
`30
`
`35
`
`40
`
`45
`
`so
`
`EP 0 949 260 A1
`sents a lower alkyl group such as methyl group, ethyl group, etc., and * has the same meaning as defined above.
`[0018] The step A is an N-alkylation reaction of (S)-piperidine intermediate (IV), and the reaction can proceed by
`using 1 to 3-fold mole, preferably 1 to 1 .5-fold mole of the ester (V) based on 1 mole of the (S)-piperidine intermediate
`(IV). The above reaction can be carried out in an inert solvent. As a suitable solvent, there may be mentioned, for exam-
`pie, water; lower alcohols such as methanol, ethanol, propanol, butanol, etc.; nitriles such as acetonitrile, propionitrile,
`etc.; aromatic hydrocarbons such as benzene, toluene, xylene, etc.; ethers such as 1,4-dioxane, tetrahydrofuran, etc.;
`ketones such as acetone, methyl ethyl ketone, methyl isobutyl ketone, etc.; amides such as N,N-dimethylformamide,
`etc.; and preferably water, acetonitrile, acetone, and N,N-dimethylformamide. These solvents may be used alone or
`may be used in admixture of two or more kinds with a suitable mixing ratio.
`[001 9] The reaction is preferably carried out in the presence of a base, and as the preferred base, there may be men-
`tioned, for example, alkali metal hydroxides such as sodium hydroxide, etc.; alkaline earth metal hydroxides such as cal-
`cium hydroxide, etc.; alkali metal carbonates such as potassium carbonate, etc.; alkaline earth metal carbonates such
`as calcium carbonate, etc.; alkali metal acidic carbonates such as sodium hydrogen carbonate, etc.; alkali metal
`hydrides such as sodium hydride, etc.; alkaline earth metal hydrides such as calcium hydride, etc.; alkali metal alkox-
`ides such as sodium methoxide, etc.; trialkylamines such as triethylamine, etc., and a pyridine compound, etc., and
`preferably sodium carbonate, potassium carbonate, sodium hydrogen carbonate or potassium hydrogen carbonate.
`These bases are each used in an amount of 1 to 3-fold moles, preferably 1 to 1 .5-fold moles based on 1 mole of the
`(S)-piperidine intermediate (IV) when the base is monovalent. When the base is divalent, it is used in an amount of 0.5
`to 1 .5-fold mole, preferably 0.6 to 1 -fold mole based on the same.
`[0020] Also, as a reaction accelerator, a small amount of a metal iodide such as, for example, sodium iodide or potas-
`sium iodide may be added. The reaction can be carried out at a reflux temperature of the reaction mixture, for example,
`5 to 150 °C, preferably 20 to 100 °C. The reaction time is 2 to 24 hours.
`[0021] The step B is a hydrolysis reaction of an (S)-ester (VI). The reaction can be carried out in an aqueous alcohol
`such as aqueous methanol, aqueous ethanol, etc., and by using an inorganic base such as sodium hydroxide, potas-
`sium hydroxide, etc. in an amount of 1 to 5-fold mole, preferably 1 to 3-fold mole per mole of the (S)-ester (VI). A reac-
`tion temperature is, for example, 5 to 90 °C, preferably 1 5 to 70 °C. A reaction time is generally 1 to 1 0 hours. After
`completion of the reaction, the reaction mixture is subjected to neutralization treatment by using a mineral acid such as
`hydrochloric acid, sulfuric acid, etc. or an organic acid such as acetic acid, oxalic acid, etc. to produce an (S)-piperidine
`compound (I).
`[0022] To obtain an optical isomer in general, methods such as an asymmetric synthesis, optical resolution by frac-
`tional crystallization or by an enzyme such as lipase, fractionation by an optical resolution column, and the like have
`been known. For preparing an optically active (S)-piperidine compound (I) efficiently in the present invention, as shown
`in the following reaction scheme (3):
`
`R e a c t i o n scheme
`
`(3)
`
`wherein * represents an asymmetric carbon, (+)-4-[(4-chlorophenyl)(2-pyridyl)methoxy]piperidine which is a
`starting compound and represented by the formula (III) is previously optically resolved and the resulting optically active
`(S)-4-[(4-chlorophenyl)(2-pyridyl)methoxy]piperidine represented by the formula (IV) is used as a synthetic intermedi-
`ate.
`[0023] The said optical resolution can be effectively carried out by the following procedure. That is, by reacting a
`racemic piperidine compound represented by the formula (III):
`
`5
`
`MYLAN Ex. 1017, Page 5
`
`

`
`EP 0 949 260 A1
`
`NH
`
`( I I I )
`
`CI
`
`with an optically active propionic acid compound (VII) represented by the formula (VII):
`
`Z
`
`Y-
`
`S
`
`( V I I )
`
`N02 C O O H
`
`wherein Y represents a hydrogen atom or a halogen atom; Z represents a lower alkoxy group; and * represents
`an asymmetric carbon,
`or an optically active N-acyl-amino acid, separating and collecting a less soluble diastereomeric salt by utilizing the dif-
`ference in solubilities of the formed two kinds of diastereomeric salts; and decomposing the resulting salt to give an opti-
`cally active piperidine intermediate (IV) represented by the formula (IV):
`
`( I V )
`
`CI
`
`wherein * has the same meanings as defined above.
`[0024] As specific examples of the optically active propionic acid compound (VI I) used as an optically resolving agent,
`there may be mentioned a compound in which, in the formula (VII), Y is hydrogen atom or chlorine atom, and Z is meth-
`oxy group. Among these, as preferred examples, there may be mentioned (2R,3R)-2-hydroxy-3-(4-methoxyphenyl)-3-
`(2-nitro-5-chlorophenylthio)propionic acid and (2R,3R)-2-hydroxy-3-(4-methoxyphenyl)-3-(2-nitrophenylthio)-propionic
`acid, and among these, (2R,3R)-2-hydroxy-3-(4-methoxyphenyl)-3-(2-nitro-5-chlorophenylthio)propionic acid is partic-
`ularly preferred.
`[0025] Also, as an acyl group of the optically active N-acyl-amino acid which is used as an optically resolving agent,
`there may be mentioned an aliphatic acyl group such as acetyl group, propionyl group, etc. ; an aromatic acyl group such
`as tosyl group, etc.; and an aralkyloxycarbonyl group such as benzyloxycarbonyl group, etc. The optically active N-acyl-
`amino acid can be prepared by the well-known acylation of various kinds of neutral, acidic and basic L-amino acids
`which are constitutional ingredient of protein or non-natural type D-amino acids. As the amino acid, there may be pref-
`erably mentioned L-phenylalanine, L-leucine, L-glutamic acid, L-methionine, L-valine, L-threonine and D-phenylglycine.
`[0026] As the preferred specific examples of the optically active N-acyl-amino acids, there may be mentioned N-
`acetyl-L-phenylalanine, N-acetyl-L-leucine, N-benzyloxycarbonyl-L-phenylalanine, N-benzyloxycarbonyl-L-valine, N-
`
`MYLAN Ex. 1017, Page 6
`
`

`
`EP 0 949 260 A1
`
`5
`
`15
`
`benzyloxycarbonyl-L-threonine and N-benzyloxycarbonyl-L-serine, and more preferably, there may be mentioned N-
`acetyl-L-phenylalanine.
`[0027] An amount of the optically active propionic acid compound (VII) of the formula (VII) or the optically active N-
`acyl-amino acid to be used as the optically resolving agent is not particularly limited but basically 0.5 to 1 .5-fold mole,
`preferably 0.6 to 1.1 -fold mole based on 1 mole of the racemic piperidine intermediate (III) of the formula (III).
`[0028] As the racemic piperidine intermediate (III) of the formula (III) to be used as the starting material in the present
`invention, there may be used an equimolar mixture of the (S)-isomer and the (R)-isomer, but the mixing ratio may not
`necessarily be even and there may be used as a mixture in which either one of the isomers is excessively contained.
`[0029] The racemic piperidine intermediate (III) of the formula (III) may be used as an acid addition salt such as a
`10 hydrochloride. In that case, for example, when a suitable alkali (e.g., sodium hydroxide) is added to the reaction system,
`a free piperidine compound is produced by causing salt-exchange. Also, the optical isomer of the optically active propi-
`onic acid compound (VII) of the formula (VII) or the optically active N-acyl-amino acid may be used as a salt with a base.
`In that case, when an acid such as hydrochloric acid is added to the reaction system, a free optically active propionic
`acid compound (VII) or a free optically active N-acyl-amino acid is produced, respectively.
`[0030] As the solvent to be used for the optical resolution of a racemic piperidine intermediate (III), there may be men-
`tioned, for example, alcohols such as methanol, ethanol, propanol, etc.; ketones such as acetone, methyl ethyl ketone,
`etc.; esters of a carboxylic acid such as methyl acetate, ethyl acetate, etc.; nitriles such as acetonitrile, propionitrile, etc.;
`ethers such as dioxane, tetrahydrofuran, etc.; amides such as dimethylformamide, etc.; water and the like. More pre-
`ferred are esters, nitriles, alcohols or water, and particularly preferred are alcohols or water. These solvents may be
`20 used alone but may be used in admixture of two or more kinds with a suitable mixing ratio depending on the necessity,
`particularly a mixed solvent of alcohols and water is preferred. An amount of the solvent to be used is not particularly
`limited, but it may be used in an amount of, for example, 2 to 50 parts by weight, preferably 5 to 50 parts by weight based
`on 1 part by weight of the racemic piperidine intermediate (III).
`In the method of optical resolution, the difference in solubilities between the resulting two kinds of diastereo-
`[0031]
`25 meric salts is sufficiently large so that a less soluble diastereomeric salt can be easily precipitated from a reaction mix-
`ture by allowing the mixture to stand or with stirring without any further treatment for crystallization.
`[0032] With regard to the conditions of dissolving the racemic piperidine intermediate (III) of the formula (III) and the
`optically active propionic acid compound (VII) of the formula (VII) or the optically active N-acyl-amino acid in a solvent
`and subsequent precipitation of a less soluble diastereomeric salt, there is no specific limitation. However, dissolution
`30 of the both compounds in a solvent can be carried out, for example, by slightly heating or under heating, and the sub-
`sequent precipitation of a less soluble diastereomeric salt can be carried out, for example, under cooling or slightly heat-
`ing.
`[0033] For precipitating the less soluble diastereomeric salt from the reaction mixture, it is generally not necessary to
`add seed crystals. However, in order to make the precipitation easier, the same kind of crystals of the desired diaster-
`eomeric salt may be added as seed crystals.
`[0034] Also, after the less soluble diastereomeric salt is separated, a mother liquor is concentrated to separate and
`collect the other diastereomeric salt which is the more soluble diastereomeric salt, and then the salt is decomposed. Or
`else, the mother liquor after separating the less soluble diastereomeric salt may be extracted with a suitable organic sol-
`vent to recover the remaining optically active piperidine intermediate (IV) which is an enantiomer.
`[0035] The purity of the separated and collected diastereomeric salt can be improved by recrystallization, depending
`on the necessity.
`[0036] A salt is removed from the diastereomeric salt thus collected by the conventionally known salt decomposition
`method whereby the desired optically active piperidine intermediate (IV) can be obtained. For example, an optically
`active piperidine intermediate (IV) can be obtained by dissolving the salt in a suitable solvent (e.g., a mixed solvent of
`45 water-dimethylformamide, etc.), treating with a suitable alkali (e.g., sodium hydroxide, potassium hydroxide, etc.),
`extracting with a suitable extraction solvent (e.g., diethyl ether, ethyl acetate, chloroform, methylene chloride, toluene,
`etc.), and evaporating the extraction solvent.
`[0037] Further, an aqueous layer after extraction is treated by a suitable mineral acid (e.g., hydrochloric acid, sulfuric
`acid, etc.) and the aqueous layer is extracted with a suitable solvent (e.g., diethyl ether, ethyl acetate, chloroform, meth-
`50 ylene chloride, toluene, etc.), an optically active propionic acid compound (VII) or optically active N-acyl-amino acid,
`which is an optically resolving agent, can be recovered.
`[0038] A specific example of the optically resolving process using (2R,3R)-2-hydroxy-3-(4-methoxyphenyl)-3-(2-nitro-
`5-chlorophenylthio)propionic acid as an optically resolving agent is described below.
`
`35
`
`40
`
`55
`
`7
`
`MYLAN Ex. 1017, Page 7
`
`

`
`EP 0 949 260 A1
`
`That is, by acting (2R,3R)-2-hydroxy-3-(4-methoxy-phenyl)-3-(2-nitro-5-chlorophenylthio)propionic acid (an
`[0039]
`optically resolving agent) on (+)-4-[(4-chlorophenyl)(2-pyridyl)methoxy]piperidine, separating and collecting a salt of
`(S)-4-[(4-chlorophenyl)(2-pyridyl)methoxy]piperidine precipitating as a less soluble diastereomeric salt and (2R,3R)-2-
`hydroxy-3-(4-methoxyphenyl)-3-(2-nitro-5-chlorophenylthio)propionic acid, and then decomposing the said salt, (S)-4-
`[(4-chlorophenyl)(2-pyridyl)methoxy]-piperidine can be obtained.
`In the present specification, the less soluble diastereomeric salt means a diastereomeric salt having less sol-
`[0040]
`ubility in a solvent than the other between a pair of diastereomeric salts.
`[0041] The racemic piperidine intermediate (III) of the formula (III), which is a starting material, is described in Japa-
`nese Provisional Patent Publication No. 25465/1990. The optically active propionic acid compound (VII) of the formula
`(VII), which is used as an optically resolving agent, can be prepared according to the method as described in, for exam-
`ple, Japanese Patent Publication No. 13994/1988.
`
`(Pharmacological test)
`
`[0042] By using an (S)-ester and (R)-ester of the following optically active piperidine ester compound, difference in
`pharmacological effects between optical isomers were studied.
`
`8
`
`MYLAN Ex. 1017, Page 8
`
`

`
`EP 0 949 260 A1
`
`(S)-ester: ethyl (S)-4-[4-[(4-chlorophenyl)(2-pyridyl)methoxy]piperidino]butanoate fumaric acid salt (prepared in Refer-
`ence example 3)
`
`(R)-ester: ethyl (R)-4-[4-[(4-chlorophenyl)(2-pyridyl)methoxy]piperidino]butanoate fumaric acid salt (prepared in Refer-
`ence example 4)
`
`5
`
`Protective effects on histamine-induced death
`
`[0043] By using Hartley male guinea pigs with a body weight of 250 to 550 g, protective effects on histamine-induced
`10 death were tested according to the method of Lands et al. (Journal of Pharmacological Experimental Therapy, vol. 95,
`p. 45, 1949 (A.M. Lands, J.O. Hoppe, O.H. Siegmund, and F.F. Luduena, J. Pharmacol. Exp. Ther., 95, 45 (1949))). Test
`animals were fasted overnight (about 14 hours), and then 5 ml/kg of a test substance was orally administrated. Two
`hours after administration of the test substance, 1 .25 mg/kg of histamine hydrochloride was intravenously administrated
`to induce histamine shock. After induction, symptom of the test animals was observed and the histamine-shock appear-
`ing time was measured, and termination of respiration or restoration was also observed. The test results are shown in
`Table 1 .
`
`15
`
`(S)-ester
`
`(R)-ester
`
`Table 1
`Protective effects on histamine-induced death
`Test substance
`Administrated amount
`Survival ratio (%)
`n
`(mg/kg, p.o.)
`0.01
`0.02
`0.03
`0.06
`0.1
`0.3
`0.6
`1.0
`3.0
`10.0
`n: number of used test animals
`
`8
`7
`8
`8
`8
`8
`8
`8
`8
`8
`
`0
`42.9
`62.5
`100
`100
`0
`0
`50.0
`62.5
`100
`
`40
`
`45
`
`so
`
`55
`
`Inhibitory effect on 7 days homologous PCA reaction
`
`[0044] By using Hartley male guinea pigs with a body weight of 250 to 550 g, inhibitory effect on PCA reaction was
`tested according to the method of Levine et al. (Journal of Immunology, vol. 106, p. 29, 1971 (B.B. Levine, H. Chang,
`Jr., and N.M. Vaz, J. Immunol. 106, 29 (1971))). 0.05 ml of guinea pig anti-BPO • BGG-lgE serum diluted 32-fold with a
`physiological saline was hypodermically administrated to the guinea pig through two points of right and left sandwiching
`the median line of the back which had been shaved one day before.
`[0045] After 7 days, 1ml of a 1 % Evans Blue physiological saline solution containing 500 ng of benzylpenicilloyl
`bovine serum albumin (BPO • BSA) was intravenously administrated to induce a PCA reaction. Thirty minutes later,
`exsanguination was carried out, and the skin was peeled off and the amount of the leaked dye was measured according
`to the method of Katayama et al. (Microbiological Immunology, vol. 22, p. 89, 1978 (S. Katayama, H. Shinoya and S.
`Ohtake, Microbiol. Immunol., 22, 89 (1978))). The test animals were fasted overnight (about 16 hours) and the test sub-
`stances were orally administrated 2 hours before the administration of the antigen. The test results are shown in Table
`2.
`
`9
`
`MYLAN Ex. 1017, Page 9
`
`

`
`EP 0 949 260 A1
`
`(S)-ester
`
`(R)-ester
`
`Table 2
`Inhibitory effect on 7 days homologous PCA reaction
`Test substance
`Administrated amount
`Inhibitory ratio (%)
`n
`(mg/kg, p.o.)
`0.01
`0.02
`0.03
`0.06
`0.1
`0.3
`1.0
`3.0
`10.0
`n: number of used test animals
`
`10
`8
`9
`8
`8
`8
`8
`8
`8
`
`37.3
`46.3
`56.9
`63.4
`58.8
`-3.1
`13.6
`45.8
`59.5
`
`30
`
`[0046] From the test results shown in Table 1 , both of the (S)-ester and the (R)-ester showed inhibitory activities dose-
`25 dependency, and ED50 values of the (S)-ester and the (R)-ester obtained from the dose-response curve are 0.023
`mg/kg and 1 .0 mg/kg, respectively, which means that the (S)-ester showed about 43-folds higher potency than the (R)-
`ester. Also, in the inhibitory effect on PCA reaction shown in Table 2, both of the (S)- and (R)-esters showed inhibitory
`activities dose-dependently. It can be estimated that the maximum inhibitory ratio of this test is expected to be about 70
`% or so, and when the activities are compared with a dose which inhibits 50 % of the maximum value (i.e., 35 %), the
`(S)-ester showed about 100-fold or more potency than the (R)-ester. These results show the clear difference in phar-
`macological effects between optical isomers and the (S)-ester is confirmed to be superior to the (R)-ester.
`[0047] The above-mentioned (S)-ester is, however, hygroscopic as shown in the stability test results (Table 4) below.
`Though the (S)-piperidine compound of the formula (I), which is a metabolite of the (S)-ester, substantially shows the
`same pharmacological effects as the (S)-ester, the (S)-piperidine compound (I) itself is quite unlikely to crystallize and
`35 usually obtained as an amber syrup. Therefore, both the (S)-ester and (S)-piperidine compound (I) are difficult to secure
`and maintain high quality as a medicinal product.
`[0048] Thus, with regard to various acid addition salts of the (S)-piperidine compounds of the formula (I), crystalliza-
`tion was investigated by the following methods.
`
`40
`
`(Experiment 1)
`
`(S)-Piperidine compound of the formula (I) was dissolved in an organic solvent