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`USOU581 1429A
`
`United States Patent
`
`1191
`
`|11| Patent Number:
`
`5,811,429
`
`Connell et al.
`
`|4s] Date of Patent:
`
`Sep. 22, 1998
`
`|54] BICYCLIC HlC'['lCR()CYC[.IC C()MP()UNl)S
`
`|5(i|
`
`|75]
`
`Invenlorsz Richard Connell, Trumbull, C-:1nn.;
`Siegfried Gnldmann; Ulrich Miiller,
`both of Wuppcrtal, Germany; Stefan
`Lohmer, Milan, Italy; Hilmar Bisehofl‘;
`Dirk Denzer, both of Wupperlal,
`Germany; Rudi Griitmlann, Solingen,
`Germany: Stefan Wohlfeil, Hildcn,
`Germany
`
`73] Assignee: Bayer Aktiengesellschaft, lJJVCl'kUSCl'],
`Germany
`
`21] App]. No.: 't'6l,92'l
`
`22]
`
`3l.)|
`
`Filed:
`
`Dec. 9, 1996
`
`Foreign Application Priority Data
`
`Dec. 15, 1995
`
`|DE|
`
`German)’ ...................... .. 195 46 913.5
`
`51]
`
`Int. Cl.” .................... ,. C{I':'D 239./72; C[|'1'D 413t00;
`A61K 311505; A61K 31535
`5144259; 514t234.5; 3344,1116;
`544284; 5441987; 544289
`
`52] U.S. CI.
`
`References Cited
`U.S. PATENT DOCUMENTS
`
`...................... .. 514-IISIJ
`5/1‘-J94 Whittaker et al.
`5,314,880
`4-E1995 Reilly et al.
`.......... ..
`5,409,926
`l"{)Rl:ll(_il\l PAl"l_’N'l' DOCUMljN'I‘S
`
`0 513 533 A2
`(J _''\6U [62 A1
`0 560 163 A1
`0 565 986 A2
`0 610 698 A2
`
`ll,-“I992
`‘J,-"1993
`9,-"1993
`10,-"1993
`8,-"1994
`
`Iiuropeau Pal. Off.
`European Pal. Off.
`European Pal. 011'.
`European Pal. Oil‘.
`European Paul. 013‘.
`
`.
`.
`.
`.
`.
`
`Pritttary I?.ratttim7r—l\’Iukund J. Shah
`A.ssi.str1rtt E.rrttttt'm’t‘—Bre1ida Coleni an
`
`Attorney, Agent, or I-Irtttfifiprung Kramer Schaefer &
`.l.ll'lS('.LI¢:C
`
`|S7|
`
`ABSTRACI"
`
`The hicyclic holcrc-cyclic compounds are prepared by reac-
`lion of correspondingly suhsliluled carboxylic acids wilh
`amines,
`in particular with phcnylglycinolaminc. The hicy—
`clic helerocyclic compounds according to the invention are
`suilahle as active compounds. in medicaments, in parlicular
`in mediearnenls having an antiatheroselerotie action.
`
`58] Field of Search ................................... .. 544t116, 284,
`5444287, 39; 514;234.5, 259
`
`6 Claims, No Drawings
`
`loft-ll]
`
`PENN EX. 2209
`
`CFAD V. UPENN
`lPR20l5-01836
`
`

`
`1
`BICYCLIC IIli:'[‘[i:I{()(_:Y(_:IJI(_: C()MI'()UNl)S
`
`2
`—continued
`
`5,811,429
`
`invention relates to bieyelie heterocyclic
`The present
`compounds, processes for their preparation and their use in
`medicaments,
`in particular as antiatherosclerotic medica-
`merits.
`
`It is known that increased blood levels of triglycerides
`(hype-rtriglyceridaemia)
`and
`cholesterol
`thypercholesterolaemia) are associated with the origin of
`atherosclerotic changes to the vascular wall and coronary
`heart disease.
`Moreover, a significantly increased risk of developing
`coronary heart disease exists if these two risk factors occur
`in combination, which in turn is accompanied by excessive
`production of apolipoprotein B—1U[I. There is therefore still
`a great need to provide active medicaments for combating
`atherosclerosis and coronary heart disease.
`Benzimiclazole derivatives having a PAF—antagonistic
`action furthermore are described in US. Pat. No. 5,314,880.
`The present
`invention relates to hicyelie heterocyclie
`compounds of the general formula {I}
`
`A
`
`in which
`
`7.
`
`R5
`
`/Lt\
`R3
`
`R"
`
`R1
`
`(U
`
`A represents a radical of the formula
`
`
`
`UT
`
`1U
`
`I5
`
`-
`
`30
`
`35
`
`4U
`
`45
`
`SU
`
`55
`
`6t]
`
`65
`
`wherein
`I, and M are identical or different and denote
`hydrogen, halogen,
`trifluoromethyl, carboxyl,
`cycloalkyl having 3 to 6 carbon atoms, hydroxyl,
`phenyl or straight—chain or branched alkyl, alkoXy—
`carbonyl or all-roxy having in each ease up to 6
`carbon atoms,
`Q denotes a nitrogen atom or the —(".l-I group,
`'1' denotes a group of the formula —SO._. or —CO or an
`oxygen or sulphur atom,
`V denotes an oxygen or sulphur atom,
`R5, R6, R7 and R3 are identical or different and denote
`hydrogen, straight-chain or branched alkyl having up to
`6 carbon atoms, benzyl or phenyl, which are option-
`ally substituted by halogen or by straight-chain or
`branched alkyl having up to 6 carbon atoms,
`R9 denotes lrilluoromethyl, benzyl or a 5- to 7—mcmbered,
`optionally benzo—fused hete-rocyclic radical having up
`to 3 heteroalorns from the series consisting of S, N
`andfor U, which is optionally substituted up to 3 times
`in an identical or ditferent manner by halogen, phenyl,
`hydroxy] or by straightachain or branched alkyl or
`alkoxy having in each ease up to 4 carbon atoms, or
`denotes
`
`a group of the formula —S(O),,—R1“,
`wherein
`a denotes the number 0, '1 or 2,
`R10 denotes straight-chain or branched alkyl or alkenyl
`having in each case up to 8 carbon atoms, which are
`optionally substituted by straight-chain or branched
`acyl having up to 6 carbon atoms or by aryl or aroyl
`having in each case up to 10 carbon atoms, which in
`turn can be substituted up to twice in an identical or
`ditferent manner by halogen,
`trifluoromethyl or by
`straight—chain or branched acyl having up to 5 carbon
`atoms, or denotes
`aryl having 6 to 10 carbon atoms, which is optionally
`substituted by halogen, hydroxyl, trilluoromethyl or
`straight—chain or branched alkyl or alkoxy having in
`each ease up to 5 carbon atoms,
`D and Isl are identical or dilferent and represent
`hydrogen, halogen,
`trifluoromethyl, hydroxyl or
`carboxyl, or represent straight—chain or branched
`alkyl, alkoxy or alkoxycarbonyl having in each ease
`up to 6 carbon atoms,
`7. represents an oxygen or sulphur atom,
`R‘ represents cycloall-zyl having 3 to '10 carbon atoms, or
`represents straight-chain or branched alkyl having 1
`to
`10 carbon atoms, or represents
`phenyl, which is optionally substituted up to twice in an
`identical or ditlerent manner by halogen, nitro,
`cyano, hydroxyl or straight-chain or branched alkyl
`or alkoxy having in c-ach case up to 4 carbon atoms,
`R2 represents hydrogen or straight—chain or branched
`alkyl having up to 3 carbon atoms,
`R3 represents hydrogen or straight—chain or branched
`alkyl having up to 5 carbon atoms, or represents
`
`20f4l]
`
`PENN EX. 2209
`
`CFAD V. UPENN
`lPR20l5-01836
`
`

`
`5,811,429
`
`3
`cycloalkyl having, 3 to 7 carbon atoms, or represents
`phenyl, or represents a 5- to 7-membered aromatic
`heterocyclic radical having up to 3 heleroatoms from
`the series consisting of S, N andfor 0, which are
`optionally substituted up to 3 times in an idcnticalor
`different manner by halogen, nitno, phenyl, hydroxyl
`or by straight—ehain or branched alkyl or alkoxy
`having up to 6 carbon atoms,
`R4 represents hydrogen, or represents a group of the 111
`formula —CH:—0H or C 30—C0—R“,
`wherein
`
`5
`
`R” denotes hydrogen, straight—chain or branched alkyl
`having up to 8 carbon atoms or phenyl, which is
`optionally substituted up to 3 times in an identical or
`different manner by halogen, hydroxyl, eyano or
`straight-chain or branched alkyl or alkoxy having in
`each ease up to 4 carbon atoms,
`and salts thereof.
`The bicyclic heterocyclic compounds according to the
`invention can also be in the form of their salts. Salts with
`
`'5
`
`-3n
`
`organic or inorganic bases or acids may be mentioned in
`general here.
`Physiologically acceptable salts are preferred in the oon- 15
`text of the present
`invention. Physiologically acceptable
`salts of the compounds according to the invention can be
`salts of the substances according to the invention with
`mineral acids, carboxylic acids or sulphonic acids. l’articu— 30
`larly preferred salts are, for example, those with hydrochlo-
`ric acid, hydrobromic acid, sulphuric acid, phosphoric acid,
`methanesulpltonic acid, ethanesulphonic acid,
`toluenesul—
`phonic acid, benzenesulphonic acid, naphthalenedisul—
`phonic acid, acetic acid, propionic acid, lactic acid, tartaric 35
`acid, citric acid, fumaric acid, maleic acid or benzoic acid.
`Physiologically acceptable salts can likewise be metal or
`ammonium salts of the compounds according to the inven—
`Iion which have a free carboxyl group. Particularly preferred
`salts are, for example, sodium, potassiurn, magnesium or 40
`calcium salts, as well as ammonium salts which are derived
`
`from ammonia or organic amines, such as, for example,
`ethylamine, di- or triethylamine, di- or triethanolamine,
`dicyclohexylaminc, dimcthylaminoethanol, arginine, lysine,
`mhylcncdiaminc or 2_phcnylc1hY111mi11c1
`Tlac compounds according to the invention can exist in
`1
`1
`_
`_
`1
`1
`_
`
`4,1
`-
`
`'
`'
`(diastereomers). The invention relates both to the enanti— so
`omers
`diastereomers and to the particular mixtures
`thereof
`lhese mixtures of the enantromers and diastere-
`omers can be separated into the stcreoisomerically uniform
`.
`_
`.
`Conmtuenui In a known manner"
`In the context of the invention, a heterocyclic radical, 55
`which is optionally benzo—fused,
`in general represents a
`saturated or unsaturated 5- to 7-mernbered, preferably 5- or
`6—membered, heterocyclic radical which can contain up to3
`heteroate-ms from the series consisting of S, N andfor O and, 6“
`
`4
`A represents a radical of the formula
`
`M
`’
`
`11
`0
`
`M
`
`.\‘
`/|J\
`
`X
`1
`
`Rs
`
`11
`
`M
`
`L
`
`T
`| l
`,
`I?
`
`O
`
`"l
`
`"‘*
`
`O
`
`M
`
`1'
`
`T
`
`>=V
`i
`_
`I\\
`
`O
`
`\./
`i
`_ AR,
`‘\
`
`'
`
`
`
`or
`M
`
`I1
`
`N\
`>_R9
`
`N
`
`\
`
`11111111111111
`
`_
`_
`_
`I‘ and M are ldcnllcal fir d1_frcmnl an_d dcnfflc
`hydrogen, [luor1rt1e,1cl:tlor1ne,brom1ne,trtIluoromethyl,
`ilfiblupmpyli LyLlUb.ul.y-1’ Ly.L10-pbmyli. L:.mlUliUEy1’
`ydroxyl, phenyl or straight-chain or branched alkyl,
`or
`up to
`car on atoms,
`Q 1111111111111 1 1111111111111 11111111 111 1111 _CH 1111111131
`T 1111111111111 11 gmup 111.1116 f.111111111a figofl 01 _CO 111 1111
`oxygen or Sulphur atom
`”
`V I
`_
`__ 1
`1_
`_ ‘I
`11
`( cnotcs an oxygen or su p ur atom,
`R5 R” R7 and R8 are identical or ditferent and denote
`Lydgugfln Sm]-,,h(1_chaiD or branch“, auwlshaving up [0
`5 L.a|.b_[;n 11m;S1 bl1_nzy1m_phmy11 which 1m,Up[iUn_
`any Substituted by fluorine‘ Chlorine, bromine or by
`5“-aj1gh[_Cha{n or branched _-,1ky1 having up to 5
`cflrljgn alums,’
`
`~
`=
`a
`b°1"3°[b]lhi°nY1= bcnzofblfuryls P}’ridY1= lhicnyls E‘-"Y1:
`l’)"m313"1a lhiamlyls Elxiimlyls imldiimlyls T"UTPh'3“")"] 07
`Dil3°l'idYlvQUi1'10lYlsfl-WY]: I3y|'i(lYl and lhicnyl 31": l'”'°f°“'°d- 65
`Preferred compounds of the general formula (1) are those
`in which
`
`1'1
`,
`l.l.tI]0
`,1mi azo
`, ur
`,
`rr
`, oxazo ' or
`ljliiiavdilylilwhicii are optionzilly substiifited up to 3 times
`in an identical or different manner by fluorine, chlorine,
`bromine, phenyl, hydroxyl or by straight—chain or
`branched alkyl or alkoxy having in each ease up to 3
`carbon atoms, or denotes
`
`3 BT40
`
`PENN EX. 2209
`CFAD V. UPENN
`lPR20l5-01836
`
`

`
`5
`a group of the formula %(0),—R1”,
`wherein
`
`a denotes the number 0 or 1,
`
`5,811,429
`
`6
`—continued
`
`R” denotes straight—chain or branched alkyl or alkenyl
`having in each ease up to 6 carbon atoms, which are
`optionally substituted by straight—ehain or branched
`acyl having up to 5 carbon atoms or by phenyl, benzoyl
`or naphthyl,
`\vhich in turn can be substituted up to
`twice in an identical or different manner by fluorine,
`chlorine, bromine, tritluoromethyl or by straightchain
`or branched acyl having up to 4 carbon atoms, or
`denotes naphthyl or phenyl, which are optionally sub-
`stituted by fluorine, chlorine, bromine, hydroxyl, trif-
`luoromethyl or straight-chain or branched alkyl or
`alkoxy having in each ease up to 4 carbon atoms,
`D and E are identical or different and represent
`hydrogen, fluorine, chlorine, bromine, trilluoromethyl,
`hydroxyl or straight—chain or branched alkyl or
`alkoxy having in cacli ease up to 4 carbon atoms,
`Z represents an oxygen or sulphur atom,
`R‘ represenLs cyclobutyl, cyclopenlyl, cyclohexl, cyclo-
`heptyl or cyclooctyl, or represents straight-chain or
`branched alkyl having up to 7 carbon atoms, or repre-
`ser1Ls
`
`phenyl, which is optionally substituted by fluorine,
`chlorine, bromine, nitro, cyano, hydroxyl or straight-
`chain or branched alkyl or alkoxy having in each
`case up to 3 carbon atoms,
`R2 denotes hydrogen or methyl,
`R3 represents hydrogen or straight—chain or branched
`alkyl having up to 4 carbon atoms, benzyl, cyclopropyl,
`cyclopentyl or cyclohexyl, or represents phenyl,
`pyridyl, thienyl or furyl, which are optionally substi-
`tuted up to twice in an identical or different manner by
`llLtorine, chlorine, bromine, phenyl, nitro, hydroxyl or
`by straight-chain or branched alliyl or alkoxy having up
`to 4 carbon atoms,
`
`R4 represents hydrogen, or represents a group of the
`formula —CH2—OH or —(.‘H2O—CO—Rl',
`wherein
`
`R” denotes hydrogen, straight—chain or branched alkyl
`having up to 6 carbon atoms or phcnyl, which is
`optionally substituted up to twice in an identical or
`different manner by fluorine, chlorine, bromine, cyano,
`hydroxyl or straightchain or branched alkyl or alkoxy
`having in each case up to 3 carbon atoms,
`and salts thereof,
`
`1U
`
`I5
`
`-
`
`30
`
`35
`
`4U
`
`45
`
`SU
`
`Particularly preferred compounds of the general formula
`(I) are those in which
`
`55
`
`
`
`or
`
`M
`
`L
`
`wherein
`
`N
`
`‘HN
`\
`
`L and M are identical or ditferent and denote hydrogen,
`fluorine, chlorine, bromine, hydroxyl, phenyl or
`straight-chain or branched alkyl or alkoxy having in
`each ease up to 4 carbon atoms,
`0 denotes a nitrogen atom or the —CH group,
`'1' denotes a group of the formula —S[),; or —CO or an
`oxygen or sulphur atom,
`V denotes an oxygen or sulphur atom,
`R5, R", R7 and R” are identical or different and denote
`hydrogen, straight-chain or branched Ell.li}«'l having, up to
`4 carbon atoms, benzyl or phenyl, which are option-
`ally substituted by fluorine, chlorine, bromine or by
`straight-chain or branched alkyl having up to 4
`carbon atoms,
`R” denotes trifluorornethyl, benzyl, benzothienyl, thienyl,
`pyridyl,
`irnidazolyl,
`fury] or
`thiazolyl, which are
`optionally substituted up to 3 times in an identical or
`different manner by iluorine, chlorine, bromine,
`phenyl, hydroxyl or by straight—ehain or branched alkyl
`or alkoxy having in each ease up to 3 carbon atoms, or
`denotes
`
`A represents a radical of the formula
`M
`
`M
`
`L
`
`N
`T AR,’
`
`I
`
`L
`
`T
`
`N>:v’
`
`\
`
`a group of the formula —S[O)fl—Rm,
`wherein
`a denotes the number 0 or 1,
`R1” denotes straight-chain or branched alkyl or alkenyl
`having in each case up to 5 carbon atoms, which are
`optionally substituted by straight-chain or branched
`acyl having up to 4 carbon atoms or by phenyl, benzoyl
`or naphthyl, which in turn can be substituted up to
`twice in an identical or different manner by fluorine,
`chlorine, bromine, triiluoromcthyl or by straight—chain
`or branched acyl having up to 3 carbon atoms, or
`denotes
`
`till
`
`65
`
`4-of4l]
`
`PENN EX. 2209
`
`CFAD V. UPENN
`lPR20l5-01836
`
`

`
`5,811,429
`
`7
`naphthyl or phenyl, which are optionally substituted by
`fluorine, chlorine, bromine, hydroxyl,
`tri['luorom-
`ethyl or straight-chain or branched alkyl or alkoxy
`having in each ease up to 3 carbon atoms,
`D and E are identical or different and represent hydrogen,
`fluorine, chlorine, bromine or triliuoromethyl,
`Z represents oxygen,
`
`5
`
`R1 represents cyclopentyl, cyclohexyl, cyclolieptyl or
`cyclooctyl, or represents straight—chain or branched 1”
`alkyl having up to 6 carbon atoms,
`R2 represents hydrogen or methyl,
`R3 represents phenyl and
`
`R4 represents the group —Cll,_.—0ll,
`
`'5
`
`8
`_oom{m1od
`
`_
`h
`N /ll\CHs
`
`D
`
`O
`
`_
`5
`,\/$_/ 0”
`Ill
`
`Characlcrlzcd in that
`acids of tho gcncfal fom-H113 (11)
`
`I
`
`UH
`
`R]
`
`E
`
`(H,
`
`30
`
`A_CH,
`
`I)
`
`in which
`
`A, D, E, Z and R‘ have the meaning given,
`am roaolod Wm-, Compounds of [he general formula (III)
`
`R3
`/J\
`H.\'R*
`
`R‘
`
`1,, which
`
`(:11;
`
`R?‘ R3 and R4 have the meaning given‘
`
`in inert solvents and in the presence of bases andfor auxil—
`iaries.
`
`The process according to the invention can be illustrated
`by way of example by the following equation:
`
`Suitable solvents here are inert organic solvents which do
`and Salts thereof
`not change under the reaction conditions. These include
`.
`,
`.
`.
`.
`A
`_ _
`,
`_
`.
`_
`_
`_
`_
`process has lurtherrnore been found ior the preparation
`f the com mmds of the metal formula (I) accmdin m the Sn etheis, such as dtethyl ether oi tetraliydrofutan, halogenated
`9
`,
`P
`3
`3
`hydrocarbons, such as methylene Cl]lOI'1ClC, chloroform, car-
`‘"V°““°“=
`bon tetrachloride, 1,2—diehloroethane,
`trichloroethane,
`tetrachloroethane, 1,2—dichloroethar1e or trichloroethylene,
`hydrocarbons, such as benzene, xylene,
`toluene, hexane,
`cyelohexane or petroleum fractions, nitromethane,
`1
`-5 dtmethylformamlde, acetone, acetonttrtle or hexameth-
`ylphosphoric acid triamide. It
`is also possible to employ
`mixtures of the solvents. Methylene eliloride,
`tetrahydroluran, toluene or dlmethylformamlde are particu-
`larly preferred.
`Inorganic or organic bases can in general be employed as
`bases for the process according to the invention.
`'l'hese
`include, preferably, alkali metal hydroxides, such as, for
`example, sodium hydroxide or potassitim hydroxide, alka-
`line earth metal hydroxides, such as, for example, barium
`35 hydroxide, alkali metal carbonates, such as sodium carbon-
`ate or potassium carbonate, alkaline earth metal carbonates,
`such as calcium carbonate, or alkali metal or alkaline earth
`metal alcoholates, such as sodium or potassium methanolate,
`sodium or potassium ethanolate or potassium tert—butylatc,
`40 or organic amines [trialkyl(C,—Cfi)amines), such as
`triethylamine, or heterocyclic compounds, such as 1,4-
`diazabicyclo[2.2.2]octane (DABCO),
`'l,8—diazabicyclo
`[5.4.0]undec-7-ene (DBU), pyridine, diaminopyridine,
`rnethylpiperidine or morpholine,
`It
`is also possible to
`45 employ alkali metals, such as sodium, and hydrtdcs thereof
`such as sodium hydride, as bases. Sodium carbonate, potas-
`siutu carbonate and trielltylatuinc are preferred.
`The base is employed in an amount of 1 mol to 5 mol,
`p,-ofo.-3h]y 1 mot to 3 mo], per molo oftho compound of tho
`5o gm-low] formula (11),
`Dehydrating reagents are also suitable auxiliaries. These
`include,
`for cxample, carbodiimide-s,
`such as
`diisopropylcarbodiimide, dicyclohexylcarbodiimide or
`N—(3—dimethylaminopropyl)—N'—ethylcarbo(Iiimide
`55 hydrochloride, or carbonyl compounds, such as
`carbonyldiimidazole, or 1,2—oxazolium compounds, such as
`2-ethyl-5-phenyl-I,2-oxazolium 3-sulphonate, or propane-
`phosphoric anhydride or
`iso—butyl chloroform ate or
`benzotriazolyloxy-tris-(dimethylamino)phosphonium
`an hexalluorophosphate or phosphonic acid diphenyl ester-
`amide or methanesulphonyl chloride, if appropriate in the
`presence of bases, such as
`triethylamine or
`N-ethylmorpholine or N-methylpiperidine or dicyclohexyl-
`carbodiimide and N-hydroxysuccinimide.
`in a temperature
`The reaction is in general carried out
`range from 0° C.
`to 150° (7., preferably from +2U° C.
`to
`+11U° C.
`
`7
`1‘
`A
`
`(-"H:
`
`U
`
`N
`
`(R}.ph._-ny1g1yg1n01__Hon'1‘
`1'JCC|, Fla
`
`
`“"15-"-0'0-—’ RT
`
`0
`
`O”
`
`65
`
`5 BT40
`
`PENN Ex. 2209
`CFAD V. UPENN
`lPR20l5-01836
`
`

`
`(IVJ
`
`(Vi
`
`'5
`
`i<13—CiI_s
`
`
`
`5,811,429
`
`9
`The reaction can be carried out under normal, increased or
`reduced pressure (for example 0.5 to 5 bar). It is in general
`carried out under normal pressure.
`The compounds of the general formula (II) are new in
`mom cases and can be prepared’ fgr example’ by a process
`in which Compounds of the general formula (IV)
`
`5
`
`10
`Suitable bases are the customary inorganic bases. These
`include, preferably, alkali metal hydroxides or alkaline earth
`tnetal hydroxides, such as, for example, sodium hydroxide,
`potassium hydroxide or barium hydroxide, or alkali metal
`carbonates, such as sodium carbonate or potassium carbon-
`ate or sodium bicarbonate. Sodium hydroxide or potassium
`hydroxide are particularly preferably employed.
`Suitable solvents are water or the organic solvents cus-
`A—H
`.
`\. 1
`tomary for hydrolysis. These include, preferably, alcohols,
`in whfl
`A has the meaning given Elm rmcmd with compounds 0f m such as methanol,cthanol,propanol, isopropanol orbutanol,
`’
`‘
`‘
`or ethers, such as tetrahydrofuraii or dioxanc, or diinetliyl—
`lb” 35°”°“11 f°”““1" (V)
`formamide or dimethyl sulphoxide. Alcohols, such as
`methanol, ethanol, propanol or isopropanol, are particularly
`preferably used. It is also possible to employ mixtures of the
`solvents mentioned.
`The ester hydrolysis is in general carried out in a tem-
`perature range from 0° C. to +l00° (7., preferably from +20°
`C. to +80° C.
`The ester liydrolysis is in general carried out under normal
`_
`_
`pressure. However,
`it
`is also possible to carry out
`the
`1“ whlch
`an hydrolysis under reduced pressiire or under increased pres-
`D: 11- i-'5 and K1 have the meaning ghfcns
`sure (for cxamplc from 05 to 5 bar),
`R13 represents halogen, preferably chlorine or bromine,
`In carrying out the ester hydrolysis, he base is in general
`iiflll
`employed in an amount of t
`to 3 mol, preferably 1
`to 1.5
`R” represents straight-chain or branched alkyl having up
`mol, per mole of the ester. Molar amounts of the reactants
`to 6 carbon atoms, in inert solvents and in the presence
`of bases andtor auxiliaries, and, finally, the esters are 15 are particularly preferably used.
`hydrolysed by customary methods.
`The compounds of the general formula (III) are known
`Customary organic solvents which do not change under
`per se.
`the reaction conditions are suitable lor the reaction. These
`The compounds of the general formula (I) according to
`include, preferably. ethers, such as diethyl ether, dioxane,
`the invention have a pharmacological action spectrum which
`tetrahydrofuran or glycol dimethyl ether, or hydrocarbons,
`so cannot bi: fort,-,_s,~,.,-,t-,1-1_
`such as benriene, toluene, xylene, liexatie, cyclohexane or
`They can be used as active compounds in medicaments
`petroleum fractions, or halogenated hydrocarbons, such as
`for reducing changes to vascular walls and for treatment of
`methylene chloride, chloroform, carbon tetrachloride,
`coronary heart disease, cardiac insufficiency, disturbances in
`dicliloroetliylene,
`trieliloroetliyleiie or chlorobenzene, or
`cerebral performance,
`iscliaeiiiic cerebral diseases,
`ethyl acetate, triethylamine, pyridine, dimethyl sulphoxide, 35 apoplexy, circulatory disturbances, disturbances in micro-
`dimethylforrnamide, hexamethylphosphoric acid triamide,
`circulation and thromboses.
`acetonitrile, acetone or nitromethane. It is also possible to
`The proliferation of smooth iiiuscle cells, furtheriiiore,
`use mixtu res of
`the
`solvents mentioned.
`plays a decisive role in the occlusion of vessels. The
`Dimethylformamide, tetrahydrofnran and dimelhyl sulphox-
`compounds according to the invention suitable for inhibiting
`ide are preferred.
`40 this proliferation and therefore for preventing atheroscle-
`Bases which can be employed are in general the above—
`rotic processes.
`mentioned bases, but in particular the alkali metal hydride!-3,
`The compounds according to the invention are (listin-
`such as sodium hydride.
`guished by a reduction in ApoB—l00—associated lipoproteins
`The base is in general employed in an amount of U.05 mol
`(VII) and its breakdown products, such as, for example,
`to '10 mol, preferably 1 mol
`to 2 mol, per mole of the 45 I.I)I.], Ap0B—l(l(l, triglycerides and cholesterol.Thcy there-
`compound of the formula (IV).
`fore have valuable pharmacological properties which are
`The auxiliary is in general employed in an amount of0.0'l
`superior to the prior art.
`mol to 100 mol, preferably 0.1 mol to 10 mol, per mole of
`Suprisingly, the action of the compounds according to the
`the compound of the general formula (IV).
`invention initially comprises a reduction in or complete
`The reaction is in general carried out in a temperature 5U inhibition of the formation andlor release of ApoB—1[t(t—
`range from —30° C. to +10U° C., preferably —10° C. to +60°
`associated lipoproleins from hepatic cells, which results in a
`C.
`lowering of the plasma \r'I.l)I.level. This lovvering in V[.[)[.
`must be accompanied by a lowering of the plasma level of
`The reaction is in general carried out under normal
`ApoB—100, LDL, triglycerides and cholesterol; several of
`pressure. However,
`it
`is also possible to carry out
`the
`processes under increased pressure or under reduced pres- 55 the abovementioned riskffactors which contribute to
`sure [for example in a range from 0.5 to 5 bar).
`changes to the vascular wall are thus reduced simulta-
`The above mentioncd dehydrating reagents are suitable
`neously.
`auxiliaries.
`The compounds according to the invention can therefore
`The various aminefamidc derivatives are also prepared
`be employed for prevention and treatmentof atherosclerosis,
`using a large number of known methods, such as,
`for on obesity, pancreatitis and constipation.
`example, coupling of acid derivatives from peptide chcm—
`1. Inhibition of the release of/\poB-100-associated lipopro-
`istry [in this context, cf. Bodanszky, The Practice of Peptide
`teins
`Synthesis: Springer Verlag, Volume 21, 1984 and Organic
`The test for detection of the inhibition of the release of
`Chemistry; Functional Group Transformations: Academic
`Aptill-lilll-associated lipoproteius from hepatic cells was
`Press, Volume 1241].
`65 carried out in vitro with cultured hepatic cells, preferably
`The ester hydrolysis is carried out by customary methods,
`with cells of the human line HepG2. These cells are cultured
`by treating the esters with customary bases in inert solvents.
`under standard conditions in medium for culture of eukary-
`
`6 BT40
`
`PENN Ex. 2209
`CFAD V. UPENN
`lPR20l5-01836
`
`

`
`5,811,429
`
`5
`
`11
`otie cells, preferably in RPMI 1640 with 10% foetal calf
`serum. HepG2 cells synthesize and secrete into the culture
`supernatant ApoB-100-associated lipoprotein particles,
`.
`.
`.
`.
`.
`.
`.
`which are in principle biiilt tip similarly to the VIDI. and
`LDL particles which are to be found in plasma.
`These particles can be detected with an immunoassay for
`human LDL. This immunoassay is carried out with ai:itibod—
`ies which have been induced against human LDL in rabbits
`under standard conditions. The anti-[.[)l. antibodies (rab-
`anti—I.l)I.—Ab) were purified by alfinity chromatography on 1”
`an iminunosorberit with human LDL. These puritied rab—
`ariti-LDL-Ab are adsorbed onto the surface of plastic. This
`adsorption is expediently carried out on the plastic surface of
`niicrotiter plates with 96 wells, preferably on MaxiSorp
`plates.
`ll" Apol}-llltt-associated particles are present in the '5
`supernatant of Hep—G2 cells, these can bond to the inso|u—
`bilized rab-anti-LDL-Ab, and an immune complex bonded
`_
`.
`_
`.
`._
`.
`.
`_
`to the plastic surface is lormed. Non-bonded proteins are
`removed by washing. The immune complex on the plastic
`surface is detected with monoclonal antibodies, which have 3“
`been induced against human LDL and purified under stan—
`dard conditions. These antibodies were conjugated with the
`enzyme peroxidase. Peroxidase converts the colourless sub-
`strate TMB into a colourert product in the presence of H302.
`After acidilication of the reaction mixture with [I2S()4__ the 35
`specific absorption of light is determined at 450 nm, this
`being a measure of the amount of ApoB—100—associated
`particles which had been secreted by the Ilep(i2 cells into
`the culture supernatant.
`Surprisingly, the compounds according to the invention
`inhibit
`the release of ApoB-100-associated particles.
`'lhe
`“:50 Value indlcaicl thc.C0.nCe.m.raIl0n (if Substance at whicll
`the absorption ot light is inhibited by 5U% compared with
`.
`the control (solvent control without the substance).
`
`12
`—continued
`
`_
`Flxatiiple ho.
`
`‘“1‘° B
`ICi-,0 [oh-I]
`
`4:3
`
`4;
`44
`47
`:2
`5;]
`51
`51;
`
`5?
`53
`50
`60
`61
`02
`
`E‘
`-,0
`“ti
`73
`
`3-,
`T8
`
`31
`
`69.0
`
`11.3
`15-‘:
`
`fig
`5.?
`57 7-9
`162:?
`19:;
`5.0
`351-0
`29.2
`9?
`35:9
`122.9
`3:--"
`
`35:5
`2).
`333-9
`
`14:;
`2?9.3
`
`35
`
`1-jxamph; N0_
`
`mm B
`[cm [net]
`
`1,
`3
`4
`5_
`E;
`5
`0
`1“
`li
`13
`14
`la
`“’
`2“
`1
`24
`25
`
`53
`29
`ii}
`',i_
`33
`.14
`-‘5_
`_l[’;
`37
`33
`asi
`
`E3‘?
`11:5
`298.5
`
`15'“:3
`43
`is
`
`1,6
`14_3
`130.2
`93“-2
`-‘*“"-"
`2483
`1411
`15.3
`13
`
`319
`9.3
`111‘:
`
`_.—,_3
`26.6
`“-3
`is
`353
`sir
`
`2. Determination of VLDI. secretion in vivo on the hamster
`,
`_
`_
`_
`_
`_
`.
`.
`The effect of the test substances on VLDL secretion in
`.
`.
`.
`.
`_
`.
`.
`35 vivo is investigated on the hamster. lior this, after premedi-
`cation with atropine (83 mgfkg s.e.), golden hamsters are
`aiiaosthotized with Ketavet (83 mg/kg s.c.) and Ncmbutal
`()0 mgfkg i.p.). When the animals have become reflex-tree,
`the v._i1igtilaris is exposed and eanniilated. 0.25 mllkg of a
`40 20% strength solution of Triton WR—1339 in physiological
`saline solution is then administered. This detergent inhibits
`lipoprotein lipase and thus leads to an increase in the
`triglyceride level because of an absence of catabolism of
`secreted VIDI. particles. This increase in triglycerides can
`45 be used as a measure for the rate of VLDL secretion. Blood
`was taken [mm the animals by puncture of the retroorbital
`venous plexus before and one and two hours after admin-
`istration of the detergent. The blood is incubated at room
`temperature for 2 hours and then at 4° C. overnight in order
`50 to eoiiclude coagulation coiiipletely. Ttiereafter, it is centri-
`fuged at
`l(],{l[)t) g for 5 minutes. The triglyceride concen-
`1-1‘
`‘mm th ~ bi‘ —ri‘~ii-i-
`'-d ‘mm--‘d
`ra ion in
`serum us o -alrtl.
`is eerrriine wi
`e .11
`of a modified eoniniercially obtainable eiizynie test
`(Merekotest® Triflyceride No. 14354). 100 at of test
`55 reagent are added to "100 Iiitl of serum in 96-well plates and
`the plates are irieubated at room temperature for 10 minutes.
`The optical density is then determined at a wavelength of
`492 nm in an automatic plate reader [SI.T—Speetra). Serum
`samples having a triglyceride concentration which is too
`at! high are diluted with physiological saline solution. The
`triglyceride concentration contained in the samples is deter-
`mined with the aid ofa standard curve measured in parallel.
`In this model, test substances are administered either intra-
`.
`.
`.
`.
`.
`.
`venously immediately belore administration of the detergent
`65 or orally or subcutaneously before initiation of the anaes-
`ttiesia.
`3. Inhibition of intestinal tiglycende absorption in vivo (rats)
`
`7 BT40
`
`PENN EX. 2209
`CFAD V. UPENN
`lPR20l5-01836
`
`

`
`5,811,429
`
`lu
`
`l"’“““t3"-‘
`6
`3
`9
`'0
`
`39
`30
`33
`33
`
`:29
`40
`
`45
`46
`43
`5:
`
`5-,:
`90
`
`79
`7'4
`77
`
`14
`—continued
`
`Inhibitiot‘. of intestinal absorption
`Oi ”ltél&'°¢'ld°$ in \"l\"° (HHS!
`<5 mgkg
`,5 0,91%
`2-1 nag,-‘kg
`-‘_ msfks
`":3 mg’?
`In
`,_,r=_z mgrkg
`;-;-‘l[) mgfkg
`3-5 ntsfke
`3 ”’§r:l‘S
`
`E mzrkg
`:->6 mg.:'l-cg
`“E “Fig
`>3 :3‘:
`.,.,g mggkg
`3 mg.-‘kg
`”“: “‘%il‘_t5
`E
`3 mggkg
`0 ntsfks
`”lg mgfig
`:,3 $31‘:
`»2 mgrkg
`3 01838
`
`13
`The substances which are to be investigated in vivo for
`their inhibiting action on triglyceride absorption are admin-
`istered orally to male Wistar rats having a body weight of
`between 170 and 230 g. For this purpose, the animals are
`divided into groups of 6 animals 18 hours before adminis— 5
`tration of the substance, and their food is then withdrawn.
`Drinking water is available to the animals ad libituin. The
`animals of the control groups are given an aqueous traga—
`carith suspension or a tragacanth suspension which con1-
`prises olive oil. The lragacanth-olive oil suspension is pre-
`pared with an Ultra—'l‘urr:tx. The substances to be
`investigated are suspended in a corresponding tragaeanth—
`olive oil suspension, likewise with an Ultra—'l'urrax, directly
`before administration of the substance.
`Before administration by at stomach tube, blood is taken
`from each rat by puncture ol‘ the retroorbital venous plexus '5
`for determination of the basal serum triglyceride content.
`The tragacaiitli suspeiisioii, the tra_gacaiitli—iilive oil sLI.spciI—
`sions without
`a substance [control animals) or
`the
`substances, suspended in a corresponding tragaeanth olive
`oil suspension, are then administered to the fasting animals 3“
`using a stomach tube. Further blood for determination of the
`postprandial serum triglyceride increase is as a rtile taken 1,
`2 and 3 hours after the administration by stomach tube.
`The blood samples are centrifuged and, after isolation of
`the serum, the triglycerides are determined photometrically 15
`with an
`]:Lt’OS—Analyzer 5060 (tippendorf
`(jeriitcbau,
`Netheler & l-Iinz GmbH, Hamburg. The triglycerides are
`determined completely enzymatically with a commercially
`m
`available UV test.
`4. Inhibition of VI .l)]. secretion in vivo (rats)
`The postprandial increase in serum triglycerides is deter— '
`The action of the test substances on VLDL secretion is
`mined by subtraction of the triglyceride prevalue of each
`also investigated on the rat. For this, 500 mgflrg body weight
`animal from its corresponding postprandial triglyceride con-
`(2.5 mgfkg] of Triton WR—1339, dissOlve(| in physiological
`ceritrations (1, 2 and 3 hours aliter administration).
`Thc aw-,1-age islakcn Ofthc (tilfel-enccs (in mmobq) al cach as saline solution, is administered lntravenoiislyinro the tail
`point in time ('1, 2 and 3 hours) in the groups, and the means " V91" 0f 7315- T719“ WR'l?39 11'1l'l1l‘1l5 llpflprmcln l1l335‘3 and
`of the lncrcasg in Serum triglycerides (Al-G) Gl‘ lhe animals
`thus leads to an lllt-Jftiii.‘-96' in the triglyceride and cholesterol
`lrcaml wllh lhe Substance are wmpared will] [ha animals
`level due to inhibition or Vl.l)[.catabolism. lhese increases
`whicli received only the tr2tgacantli—oil suspension.
`can be l"i*§d as a m