(12; United States Patent
`Miiller et al.
`
`(10) Patent N0.:
`(45) Date of Patent:
`
`US 6,479,503 B2
`Nov. 12, 2002
`
`US0064-79503132
`
`(54) CYCL()/\I.KAN()-IN[)()I,l‘LAND
`-AZ/\lND0l.E DERIVATIVES
`
`(52) U.S. Cl.
`(58) Field of Search
`
`5141292; 546,185; 540787
`S46E85, 87; 514,292
`
`(75)
`
`Inventors: Ulrich Miiller, Wupperlal (DE);
`Richard Connell, Trumbull, Cl‘ (US);
`Siegfried Goldmann, Wuppcrlal (DE);
`Rudi Griitzlttann, Sulingen (DE);
`Martin Beuek, Nilford, Cl‘ (US);
`Hilmar Bisehoff, Wupperlal (DI-3); Dirk
`l)enzer, Wuppertal (DE); Anke
`Dtnndey-Bette, l-liickeswagen (DE);
`Stefan Wohlfeil, I-lilden (DE)
`
`(73) Assigneez Bayer Aktiengesellsehatt, Leverkusen
`(DE)
`
`[ *) Notice:
`
`Subject to any disclaimer, the term of this
`patent
`is extended or adjusted under 35
`U.S.C. 154[b] hy 0 days.
`
`(21) App]. No.: 09;s14,253
`
`(22)
`
`Filed:
`
`Mar. 21, 2001
`
`(65)
`
`Prior Publication Data
`
`Us 2[lU2,al'lll55f'135 A1 May 9, 20:12
`
`Related U.S. Application Data
`
`(:36)
`
`RI:l'I:I1.-noes Cited
`U.S. l’Al‘l:'.N'l‘ DOCUMl:LN'l‘S
`
`1_.-“I972 Maurer el :11.
`3,632,807 A
`10,-"I988 Gillard el :11.
`4,TI'5,o80 A
`SH996 Mijller el al.
`5,521,206 A
`5,684,014 A * l1;"l.0Fl’.-‘ Muller el al.
`5,?76,‘J{'l4 A
`?_.*”l9‘J.‘{ Mi.'Illcr el al.
`5,952,498 A
`9;’l9.‘J.<J
`I_.er|l'eI5 et al.
`FOREIGN PATENT DOCUMENTS
`
`S14.-‘"292
`
`EP
`LP
`El’
`EP
`EP
`EP
`
`0 234 T08 Al
`0 3LlU 626 A2
`1'} 310 1?") A2
`I’) 4.06 23? A2
`0 509 3:39
`0 61? 035 Al
`
`W198?
`U198‘)
`M1989
`7;'l‘)02
`l0;"l002
`9;'l9Q4
`
`OTHER PUl'3l.IC/\TIONS
`
`lletemcycles, vol. 22, No. '10, "1984 (pp. 2277-2279).
`
`* cited by examiner
`
`1’rr.'mar__v Ex(tminer—Alan L. Rotman
`A:~:.5'i5Irrru Ex(mtt'm:J'—Ri1a Desai
`(74) Attorney, Agent, or F.=Trm—Norris McLaughlin &
`Marcus
`
`(62) Division of application No. E19,-"3l3,(l35, Filed on May [7,
`1999, now Pat. No. 6,265,431, which is a division of
`application No. 085887381, filed on Jul. 3, 1997, which is a
`division of application No. (I8.*'535,6‘}8_. liled on Sep. 28.
`1995, now Pal. No. 5,684,014.
`
`F0l‘9i{.'.Il Appllctltlllfl Priority "1101
`[30)
`0“ 4 1994
`(DE)
`
`44,_§4.J,_,
`
`(57)
`
`ABS'I'RAC'l"
`
`Cycltlalcarttaitldiale ant] -21/.aim]ulc derivatives are I.JfCp2lft:Ll
`by reaction of appropriately substituted carboxylic acids
`with anmines. 'T'ne cycloahloanindole and —azaindole deriva-
`tives are suitable as active compounds for inedieatnents,
`preferably anliatherosclerolie rnedicaments.
`
`(51)
`
`Int. Cl.‘
`
`A61K 31.5437; C[I”z'D 4'/‘U06
`
`5 Claims, No Drawings
`
`1 of 70
`
`PENN EX. 2226
`CFAD V. UPENN
`lPR20l5-01836
`
`

`
`US 6,479,503 B2
`
`1
`CYCLOALKANO-INDOLE AND
`-AII.AINl)()I.lC [)It)RIVA'l‘lVES
`(‘ROSS—REFERENCE
`
`This application is a Divisional of application Ser. No.
`[)9;"3l3,(J35 now US. Pat. No. 6,265,431 filed May 17, 1999,
`which is a division of application -Ser. No. 08t"88?',781, filed
`Jul. 3, 1997, which is a division of application Ser. No.
`[)8f535,698, filed Sep. 28, 1995, now U.S. Pat. No. 5,684,
`014.
`
`The present invention relates to cycloalkano-indolc and
`-azcaindole derivatives, processes for their preparation and
`their use as medicaments, in particular as antiatheroselcrotic
`medicarnents.
`
`increased blood levels of triglycerides
`is known that
`It
`[hypertriglyeeridaemia)
`and
`cholesterol
`(hypercholesterolaemiai are associated with the genesis of
`atherosclerotic vessel wall changes and coronary heart dis-
`eases.
`
`5
`
`1U
`
`I5
`
`Adistinctly increased risk of the development of coronary .
`heart disease is moreover present if these two risk factors
`occur in combination, which is accompanied, in turn, with
`an overproduction of apolipoprotein B-100. There is
`therefore, as before, a great need to make available effective
`medicarnents for the control of atherosclerosis and coronary
`heart diseases.
`
`The present invention relates to eycloalkano—indo]e and
`—azaindole derivatives of the general formula (1)
`
`30
`
`(U
`
`R3
`
`R‘
`
`R l
`
`R3
`
`l
`
`l
`
`N
`
`1&5
`
`l
`\
`I-'.—I
`cu; Y R?
`D
`
`R°
`
`in which
`
`R' and R2, including the double bond connecting tltetn,
`together form a phenyl or pyridyl ring or a ring of the
`formula
`
`wherein
`
`R3 denotes hydrogen or straight-chain or branched alkyl
`having up to 4 carbon atoms,
`R3 and R4, including the double bond connecting them,
`together form a phenyl
`ring or a 4-
`to 8-membered
`cycloaukene or oxocycloalkene radical,
`all ring systems mentioned under RJFRS and R3,-‘R4 option-
`ally being substituted up to 3 times by identical or
`different halogen,
`triflnnrnmethyl, earhoxyl or
`hydroxyl substituents, by straight-chain or branched
`alkoxy or alkoxycarbonyl each having up to 6 carbon
`atoms or by straiht—chain or branched alkyl having up
`to 6 carbon atoms, which, for its part, can be substituted
`by hydroxyl or by straight-chain or branched alkoxy
`having up to 4 carbon atoms,
`
`2
`D represents hydrogen, cycloalkyl having 4 to 12 carbon
`atoms or straight-chain or branched alkyl having up to
`12 carbon atoms,
`ll. represents the —(I[)— or —(TS— group,
`
`L represents an ogygen or sulphur atom or a group of the
`formula —NR ,
`wherein
`
`R9 denotes hydrogen or straight-chain or branched alkyl
`having up to 6 catbon atoms, which is optionally substituted
`by hsydroxyl or phenyl,
`R represents phenyl or a 5- to 7-membered sated or
`unsaturated heterocycle having up to 3 heteroatoms from the
`series consisting of S, N andfior O, the cycles optionally
`being substituted up to 3 li111es by identical or different nitro,
`carboxyl, halogen or cyano substituents or by straight-chain
`or branched alkenyl or alkoxycarbonyl each having up to 6
`carbon atoms or by staight—chain or branched alkyl having
`up to 6 carbon atoms, which is optionally substituted by
`hydroxyl, carboxyl or by straight-chain or branched alkoxy
`or alkoxycarbonyl each having up to 6 carbon atoms, and.-“or
`the cycles optionally being substituted by a group of the
`formula —DR'" or —NRl lRl2,
`wherein
`
`R10 denotes hydrogen or straight-chain or branched alkyl
`or alkenyl each having up to 6 carbon atoms,
`R” and R” are identical or different and denote phenyl,
`hydrogen or straight-chain or branched alkyl having up to 6
`carbon atoms or straight-chain or branched acyl having up to
`8 carbon atoms, which is optionally substituted by a group
`of the fon'nula NRHRH,
`wherein
`
`Rl 3 and R” are identical or different and denote hydrogen
`or straight-chain or branched aeyl having up to 8 carbon
`atoms,
`R6 represents hydrogen, carboxyl or straight—chain or
`branched alkoxycarbonyl having up to 5 carbon atoms, or
`represents straight—ehain or branched alkyl having 11p to 6
`carbon atoms, which is optionally substituted by hydroxyl or
`by a group of the formula —0—C0—R”,
`wherein
`
`R15 denotes phenyl which is optionally substituted up to
`3 times by identical or dillierent halogen or hydroxyl sub-
`slituents or by straight-chain or branched alkyl having up to
`5 carbon atoms, or straight—chain or branched al.kyl or
`alkenyl each having up to 22 carbon atoms, each of which
`is optionally substituted by a group of the formula —0R",
`wherein
`
`the group of the formula
`
`R1“ is hydrogen, benzyl, triphenylmethyl or straight-chain
`or branched acyl having up to 6 carbon atoms,
`R7 represents hydrogen or
`R6 and R7 together represent
`=(),
`if appropriate in an isomeric form and their salts.
`The cycloal.kan—indole and —azaindole derivatives accord-
`ing to the invention can also be present in the form of their
`salts. In general, salts with organic or inorganic bases or
`acids may be mentioned here.
`In the context of the present invention, physiologically
`acceptable salts are preferred. 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. Particu-
`larly preferred salts are, for example, those with hydrochlo-
`ric acid, hydrobrornic acid, sulphuric acid, phosphoric acid,
`melhanesulphonic acid, elhanesulphonic acid,
`toluenesul-
`phonic acid, benzenesulphonie acid, naphthalenedisul—
`phonic acid, acetic acid, propionic acid, lactic acid, tartaric
`acid, citric acid, fumaric acid, maleic acid or benzoic acid.
`
`35
`
`4U
`
`45
`
`SU
`
`55
`
`fit]
`
`65
`
`20f7l]
`
`PENN EX. 2226
`
`CFAD V. UPENN
`lPR20l5-01836
`
`

`
`US 6,479,503 B2
`
`4
`L represents an oxygen or sulphur atom or represents a
`group of the formula —NR9,
`wherein
`
`3
`Physiologically acceptable salts can also be metal or
`ammonium salts of the compounds according to thc inven-
`tion which have a free carboxyl group. Particularly preferred
`salts are, for example, sodium, potassium, magnesium or
`R9 denotes hydrogen or staight—chain or branched alkyl
`calcium salts, and also ammonium salts which are derived
`5 having up to 5 carbon atoms, which is optionally substituted
`.
`_
`.
`_
`.
`_
`_
`.
`_
`_
`_
`_
`‘
`H
`b h dmx lm hm I
`tron: ammonia, or organic amines, such as, for example
`_
`_
`_
`_
`y Sy
`y
`P
`y ‘
`etliylamine, di- or triethylamine, di- or triethanolatnitie,
`R 1'°P1'°5‘_5m-‘5 _Ph°"Y_1~ P}’1'1dYl-fufyls‘h1C"}’10T1m1FlaZ°lYl»
`dicyclohexylamine, dimethylaminoethanol, arginine, lysine,
`each of which is optionally substituted up to 2 times by
`cthytemdiaminc or 3_phgny1e1[-,y1amfnc_
`identical or dillierent nitro, carboxyl,
`fluorine, chlorine,
`11].»;
`5]1'uQ[u[¢-_-,,
`Including the double bond of parent
`eyeloalkene radical (R3t'R") in the context of the invention in 1” bromine or cyano subslitucnls. by straight-chain or branched
`general represents a 4 —to 8—membere(| hydrocarbon radical,
`alkcnyl Or alkoxy carbonyl each having up to 4 carbon atoms
`preferably a 5- to 8-memhered hydrocarbon radical,
`for
`orby straight-chain orbranched alkyl having up to5 carbon
`example 3 (;y(;1o[)u[cnc‘ cy(;]0pemcnc, cyclohexenc, cycI0—
`atoms, which is optionally substituted by hyClt'0Xyl, carboxyl
`hcptcnc or cyctooctcnc tactic-a1_ ‘the cyclopcmcnc, H or by staight—chain or branched alkoxy or alkoxycarbonyl
`cyclohcxcnc, cyclooctenc or cycloheptcnc radicals are prc— '
`each having up to 5 carbon atoms, andfor the cycles are
`fa;-1-cd
`optionally substituted by a group of the formula —OR10 or
`Hctcrocycle [R5] in the context of the invention in general —NR“Rl2.
`represenLs a saturated or unsaturated 5-
`to 7'-memhered
`wherein
`h°‘°1'0'3)’C1'-‘ta Prcfcrably 3 5' ‘0 5'm‘''mb‘'r‘'°d h°‘°"°"-'Y'3l°s
`.,n R” denotes hydrogen or straight-chain or branched alkyl
`which can contain up to 3 heteroatoms from the series ”
`or alkcnyt cach having up to 4 carbon men-13_
`conslsnng 0t S’ N‘
`‘md’lOr‘ 0' Eflxamplcs which may be
`R” and R12 are identical or ditferent and denote phenyl,
`mcntmnccl all“: pyrldyl’ ‘h'°“§’1s furylf pyfmlyl’ “W17”lyl’
`hydrogen or straight-chain or branched alkyl having up to 5
`m(_amly1'1m1d“‘£°1y1’ mmpholmyl or pIpc"dyl' Pyndyl ‘ind
`carbon atoms or denote straight—chain or branched acyl
`Ihlfnyl are preferred‘
`,
`,
`_
`_
`_
`35 having up to 6 carbon atoms, which is optionally substituted
`Ihe compounds according to the invention can exist in
`by a group 01- [ha ronnula _NR1fiR1nl’
`stereoisomeric forms which either behave as image and
`whcrcm
`_
`_
`_
`mirror image (enantiomers), or do which do not behave as
`,3
`ifreldenucalm ‘J1m’“’““‘_“d denote hydrogen
`image and mirror image (diastereomers). The invention
`R i_"-“JR
`relates both to the eiiantiomeis and tliastereoiners and their m "T 'ii'”“'1gh1'd-'a'" "T branchul acyl havmg up [0 6 Carhm
`respective mixtures. These mixtures of the enantiomers and '
`“1”m_S'
`_
`_
`diastereomers can be separated in a known manner into the
`R0 WPWSCHIS h)'dT"’geT1 Carbilxyl
`"T -‘ifialghbchiim '5"
`Stcrcoisomcrically uniform ,_-_0n5timem_.;_
`branched alkoxycarbonyl having up to 4 carbon atoms, or
`Preferred compounds of the general formula (1) are those
`1‘‘-'*IJ1‘°3°"l-‘5 511‘?t11t’,l1l'‘-'l1**i11 01' b1'a“‘v'h°‘l alkyl ha‘-“i118 UP 10 5
`in which
`carbon atoms, which is optionally substituted by hydroxyl or
`R‘ and R2, including the double bond connecting them, 35 by 3 Qpm‘-‘I’ 01' H19 fmmula —O—C0—Rl5r
`together form a phenyl or pyridyl ring or a ring of the
`wherein
`formula
`R13 denotes phenyl which is optionally substituted up to
`3 times by identical or different lluorine, chlorine, bromine
`40 or hydroxyl substitucnts or by straight—chain or branched
`alkyl having up to 4 carbon atoms, or straight-chain or
`branclied alkyl or alkenyl each having up to 20 catboti
`atoms, each of which is optionally substituted by a group of
`the formula —OR'fi,
`.
`45 wherein
`R‘ 6 is hydrogen, benzyl, triphenylmethyl or straight-c|:iain
`_
`or Egdmhul diglhhivlng up 10 5 Ldrbon Numb’
`Whfigaiirentttes hydrogen or stright-chain or branched alkyl
`I
`f
`I
`Re rcpclrcgtirn H yl mgen or
`1
`E
`having up to 3 carbon atoms,
`R3 and R4, including the double bond connecting them, 50 :0 an
`toga Er lclimscm he group 0 H6 0mm a
`.
`together form a phenyl ring or a cyclopentene, cyclohexene,
`.
`'
`.
`._
`.
`.
`_
`_
`_
`_
`_
`_
`_
`_
`if appropriate in an isomeric form, and their salts.
`_
`eyeloheptene, eyelooctene, oxoeyelopentene,
`,
`_
`,
`_
`oxocyclohcxgm’ oxocycloheplene or Umcyclooclenc
`Particularly preferred compounds ot the general tormula
`radical,
`(I) are [how
`all ring systems mentioned under RJFR3 and l{3fR" option— 55 in which
`ally being substituted up to 2 times by identical or different
`R‘ and R2, including the double bond connecting them,
`fluorine, chlorine, broiiiiiic,
`trilluoroiiietliyl, carboxyl or
`togetlier form a phenyl or pyridyl ring or a ring of the
`hydroxyl substitucnts, by straight—chain or branched alkoxy
`formula
`or alkoxycarbonyl each having up to 4 carbon atoms or by
`straight-chain or branched alkyl having up to 4 carbon 6“
`atoms, which, in turn, can be substituted by hydroxl or by
`
`H
`
`I‘-Rs
`'
`
`O
`
`straight—chai.n or branched alkoxy having up to 3 carbon
`atoms.
`
`NR3?
`
`D represents hydrogen, cyclobutyl, eyclopentyl,
`cyclohexyl, cycloheptyl, cyclooctyl or straight-chain or 65
`braiiclicd alkyl having up to 10 carbon atoms,
`IL represents the —CO— or —(.‘S— group,
`
`O
`
`3 of 7"
`
`PENN EX. 2226
`CFAD V. UPENN
`lPR20l5-01836
`
`

`
`US 6,479,503 B2
`
`wherein
`
`R8 denotes hydrogen or methyl,
`
`R3 and R“, including the double bond connecting them,
`together form a phenyl ring or a cyclopentene, cyclohexene,
`eycloheptene, cyclooctene, oxocyclopentene,
`oxocyclohexene, oxocycloheptene or oxocyclooctene
`radical,
`
`all ring systems mentioned under RWR2 and l{3fR" option-
`ally being substituted up to 2 times by identical or different
`fluorine, chlorine, bromine,
`trifluoromethyl, carboxyl or
`hydroxyl substituents, by straight—chain or branched alkoxy
`or alkoxycarbonyl each having up to 3 carbon aton1s or by
`straight-chain or branched alkyl having up to 3 carbon
`atoms, which, for its part, can be substituted by hydroxyl,
`methoxy or ethoxy,
`
`D represents hydrogen, cyclopentyl, cyclohexyl,
`eyeloheptyl, cyelooctyl or straight—chain or branched
`alkyl having up to 6 carbon atoms,
`
`1:" represents the —(T()— or —(ES— group,
`I, represents an oxygen or sulphur atom or represents a
`group of the fonnula —_\lR9,
`
`wherein
`
`R9 denotes hydrogen or straight—chain or branched alkyl
`having up to 4 carbon atoms, which is optionally substituted
`by hydroxyl or phenyl,
`
`R5 represents phenyl, pyridyl or thienyl, each ofwhich is
`optionally substituted up to 2 times by identical or ditfercnt
`nitro, carboxyl,
`fluorine, chlorine, bromine or cyano
`subslituents, by straight-chain or branched alkenyl or
`alkoxycarbonyl each having up to 3 carbon atoms or by
`straight-chain or branched alkyl having up to 4 carbon
`atoms, which is optionally substituted by hydroxyl, carboxyl
`or by straight—ehain or branched alkoxy or alkoxycarbonyl
`each having up to 4 carbon atoms, andfor the cycles are
`optionally substituted by a group of the formula —OR"-’ or
`_NR11R12‘
`
`wherein
`
`Rm denotes hydrogen or straight—chain or branched alkyl
`or alkenyl each having up to 3 carbon atoms,
`
`R” and R” are identical or ditferent and denote phcnyl,
`hydrogen or straight —chain or branched alkyl having up to 4
`carbon atoms or denote straight—chain or branched acyl
`having up to 5 carbon atoms, which is optionally substituted
`by a group of the formula —NR‘3R“,
`
`wherein
`
`R13 and R” are identical or different and denote hydrogen
`or straight—ehain or branched acyl having up to 5 carbon
`atoms,
`
`R“ represents hydrogen, earboxyl or straight—chain or
`branched alkoxycarbonyl having up to 3 carbon atoms, or
`represents straight-chain or branched alkyl having up to 4
`
`1U
`
`I5
`
`30
`
`35
`
`4U
`
`45
`
`SU
`
`55
`
`oil
`
`65
`
`6
`carbon atoms. which is optionally substituted. by hydroxyl
`or by a group of the fonnula —()—C()—R15,
`wherein
`
`R15 denotes phenyl which is optionally substituted up to
`3 times by identical or difi'crcnt straight-chain or branched
`alkyl having up to 3 carbon atoms, or denotes straight—cha1n
`or branched alkyl or alkenyl each having up to 19 carbon
`atoms, each of which is optionally substituted by a group of
`the formula —OR'fi,
`
`wherein
`
`triphenylmethyl or
`R” denotes hydrogen, benzyl,
`straight—chain or branched acyl having up to 4 carbon atoms,
`
`R7 represents hydrogen or
`
`R6 and R7 together represent the group of the formula
`:0,
`
`if appropriate in an isomeric form, and their salts.
`
`A process for the preparation of the compounds of the
`general formula {]) according to the invention has addition-
`ally been found, characterized in that carboylic acids of the
`general formula (11)
`
`([1)
`
`R3
`
`R‘
`
`R1
`
`R3
`
`N
`
`/ \.[/CTCJEH,
`
`D
`
`in which
`
`R‘, R2, R3, R4 and D have the meaning indicated, are
`amidated using compounds of the general formula (Ill)
`
`nil)
`
`in which
`
`R5 has the meaning indicated
`
`and
`
`R” has the indicated meanig of R°, but (lees not represent
`carboxyl,
`
`in an inert solvent and in the presence of bases andfor
`auxiliaries,
`
`functional groups are varied by
`if appropriate,
`and,
`hydrolysis, esterification or reduction.
`
`The process according to the invention can be illustrated
`by the following reaction scheme:
`
`4-of?!)
`
`PENN EX. 2226
`
`CFAD V. UPENN
`lPR20l5-01836
`
`

`
`US 6,479,503 B2
`
`
`
`co»_.n
`
`IW/\/OH
`
`T}ichIoromeLltaitejtrietltyla mine
`l-|tyr.lt'ox_v-IH-13eItzott'i:izolc and
`N'-(3—dimcth}.'lantLt1oprcpylJ-
`N-elhylca rhorliimidc hydrochlo ride
`
`
`
`Separation of
`d iastcreoni c rs
`
`
`
`Suitable solvents for the amidation arc in this case inert
`organic solvents which do not change under the reaction
`conditions. These include ethers, such as dielhyl ether or
`tetrahydrofuran halogenohydrcarbons such as 55
`dichloromethane,
`trichioromcthane,
`tetrachloromcthane,
`trichloroethane,
`tetrachloroelhane,
`l
`,2-dichloroelhane or
`trichlorocthylene, hydrocarbons such as hcmicnc, xylene,
`toluene, hexane, cyclohexane or petroleum fractions,
`nitrornethane, dimelhylformamide, acetone, acetonitrile or on
`hexamethylphosphorarnide.
`It
`is also possible to employ
`mixtures of the solvents. Dichloromethanc, tetrahydrofuran,
`acetone and dimethylformamide are particularly preferred.
`Bases which can be employed for the process according
`to the invention are in general inorganic or organic bases. 65
`These preferably include alkali metal hydroxides such as, for
`example, sodium hydroxide or potassium hydroxide, alka-
`
`line earth metal hydroxides, such as, for examnple, barium
`hydroxide, alkali metal carbonates such as sodium carbonate
`or potassium carbonate, alkaline earth metal carbonates such
`as calcium carbonate, or alkali metal alkoxides such as
`sodium or potassium mcthoxide, sodium or potassium
`ethoxide or potassium tert-butoxide, or organic amines
`(trialkyl(C,—C,_)amines) such as triethylaminc, or hetero-
`cycles such as 1,4diazabicyclo[2.2.2]octane (DABCO), 1,8-
`dia7.ahicyclo[5.4.0]undec-7-ene (DBU), pyridine,
`diaminopyridine, methylpiperidine or morpholine. It is also
`possible to employ alkali metals such as sodium and their
`hydridcs such as sodium hydride as bases. Sodium and
`potassium carbonate and triethylamine are preferred.
`The base is employed in an amount from 1 mol to 5 mol,
`preferably from 1 me] to 3 mol, relative to 1 mol of the
`compound of the general formula (II).
`
`50f7l]
`
`PENN EX. 2226
`
`CFAD V. UPENN
`lPR20l5-01836
`
`

`
`US 6,479,503 B2
`
`9
`The reaction is in general carried out in a temperature
`range from 0° C.
`to 150° C., preferably from +20° C. to
`+110° C.
`The reaction can be carried out at normal, increased or
`reduced pressure (e.g. 0.5 to 5 bar). In general, the reaction
`is carried out at normal pressure.
`The amidation can optionally proceed via the activated
`stage of the acid halides, which can be prepared from the
`corresponding acids by reaction with thionyl chloride, phos-
`phorus trichloride. phosphorus pentachloride, phosphorus
`tribromide or oxalyl chloride.
`The abovcmentioncd bases can optionally also be
`employed for the amidation as acid—binding auxiliaries.
`Suitable auxiliaries are also dehydrating reagents. These
`include,
`for cxarnple, carbodiimides such as
`diisopropylcarbodiimide, dieyclohexylcarbodiimide and
`N—(3—dimethylaminopropyl)N'—ethy|carbodiimirle hydro-
`chloride or carbonyl compounds such as carbonyldiidazole
`or 1,2-oxazolium compounds such as 2-ethyl-5-phenyl-l
`,2—oxazolium—3—sulphonate or propancphosphonic anhy-
`dride or isobutyl chloroformate or benzotria7olyloxy—tris—
`dimethylamino)phosphonium hexafluorophosphate or
`diphenyl phosphoramidate 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.
`The acid—binding agents and dehydrating reagents are in
`general employed in an amount from 0.5 to 3 mol, preferably
`from 1
`to l.5 mol, relative to 1 mol of the corresponding
`carboxylic acids.
`for example
`funetiotial groups,
`The variation of
`hydrolysis, esteritication and reduction, and also separation
`of isomers and salt formation is carried out by customary
`methods.
`The carboxylic acids of the general formula (II) are new
`and can be prepared by reacting compounds of the general
`formula (I V)
`
`1U
`
`I5
`
`30
`
`35
`
`(W)
`
`4U
`
`min “E
`
`l
`
`/\|/D
`
`in which
`
`I) has the meaning indicated,
`for example
`T represents a typical
`leaving group,
`chlorine, bromine, iodine, tosylate or mesylate, prefer-
`ably bromine,
`
`anrl
`
`Rm represents ((I,—(I,,)-alkyl, with compounds of the
`general formula
`
`(V:
`
`R“
`
`it‘
`
`R ‘
`
`R3,
`
`H
`
`in which
`
`R‘, R3, R3 and R" have the meaning indicated, in inert
`solvents, if appropriate in the presence of a base.
`
`45
`
`SU
`
`55
`
`fit]
`
`65
`
`10
`the process are the customary
`Suitable solvents for
`organic solvents which do change under the reaction con-
`ditions. These preferably include ethets such as dietliyl
`ether, dioxane, tetrahydrofuran, glycol dimethyl ether, or
`hydrocarbons such as benzene,
`toluene, xylene, hexane,
`cyclohexane or petroleum fractions, or halogenohydrocar-
`bons such as dichloromethane,
`trichloro methane,
`tetrachloromethane, dichloroethylene,
`trichloroethylenc or
`chlorobenzene, or ethyl acetate,
`triethylamine, pyridine,
`dimethyl
`sulphoxidc, dimethylformamide,
`hexaniethylphosplioramide, acetotiitrile, acetone or
`nitromcthane.
`It
`is also possible to use mixtures of the
`solvents mentioned. Dimethylformamirle and tetrahydrofu-
`ran are preferred.
`The bases employed for the process according to the
`invention can in general be inorganic or organic bases. These
`preferably include alkali metal hydroxides, for example,
`sodium hydroxide or potassium hydroxide, alkaline earth
`metal
`ltydroxides,
`for example, barium hydroxide, alkali
`metal carbonates such as sodium carbonate or potassium
`carbonate, alkaline earth metal carbonates such as calcium
`carbonate, or alkali metal alkoxides such as sodium or
`potassium methoxide, sodium or potassium ethoxide or
`potassium tert-butoxide, or organic amines (trialkyl(C,—C,;)
`amines) such as triethylamine, or heterocycles such as
`"l,4diazabicyclo[2.2.2]octane [D/\BCO), 1,8-diaxabicyclo
`[5.4.0]undec-7-ene (DBU), pyridine, diamninopyridine,
`methylpiperidine or morpholine.
`It
`is also possible to
`employ alkali metals such as sodium or their hydrides such
`as sodium hydride as bases. Sodium hydride, potassium
`carbonate,
`tricthylamine, pyridine and potassium tert-
`butoxide, DBU or DABCO are preferred.
`In general, the base is employed in an amount from 0.05
`mol to "[0 mol, preferably from '1 mol to 2 mol, relative to
`1 mol of the compound of the formula (IV).
`The process according to the invention is in general
`carried out in a temperature range from —3[]° C. to +100“ (T,
`preferably from —1(J° C. to +6U° C.
`The process according to the invention is in general
`carried out at normal pressure. However, it is also possilnle
`to carry out the process at elevated pressure or at reduced
`pressure (e.g. in a range from 0.5 to 5 bar).
`The compounds of the general formula (III) are known
`per se.
`The compounds of the general formula (IV) are known or
`can be prepared in analogy to known methods.
`The compounds of the general formula (V) are known or
`can be prepared in analogy to known methods.
`The compounds of the general formula (I) according to
`the invention have an unforeseeable spectrum of pharma-
`cological action.
`They can be used as active compounds in medicaments
`for the reduction of changes to vessel walls and for the
`treatment of coronary heart disorders, cardiac iiisufliciency,
`brain power disorders, ischacrnic brain disorders, apoplexy,
`circulatory disorders, disorders of the microcirculation and
`thromboses.
`
`the proliferation of smooth muscle cells
`Furthermore,
`plays a decisive part
`in the occlusion of vessels. The
`compounds according to the invention are suitable for
`inhibiting this proliferation and thus preventing atheroscle-
`rotic processes.
`The compounds according to the invention are distin-
`guished by a lowering of the ApoB—1U0—associated lipopro—
`teins (VI.|)I. and its degradation products, e.g. I.lJI.), of
`ApoB—100, of triglycerides and of cholesterol. They thus
`have useful, superior pharmacological properties in com-
`parison with the prior art.
`
`6of7l]
`
`PENN EX. 2226
`
`CFAD V. UPENN
`lPR20l5-01836
`
`

`
`US 6,479,503 B2
`
`E, V
`\' ' 0'
`2
`5
`?
`
`ED
`
`fk
`'"” [mg’ *2] p'0'
`10-15
`_-L5
`10-2”
`
`12
`11
`Surprisingly, the act ion of the compounds according to the
`hamsters are anaesthetized with Ketaset (83 mgfkg s.c.) and
`Nembutal (50 mgfkg i.p.) after premedication with atropine
`invention consists first in a decrease or complete inhibition
`(83 mgfkg s.c.). When the animals have become reflex-free,
`of the formation andfor the release of Apofl-"I00-associated
`the jugular vein is exposed and cannulated. 0.25 ml,-‘kg of a
`lipoproteins from liver cells, which results in a lowering of
`5 20% strength solution of Triton WR—l339 in physiological
`the Vl.Di_ plasma level. This lowering of VLDL must be
`saline solution is then administered. This detergent inhibits
`accompanied by a lowering of the plasma level of ApoB1UU,
`1|“; ]iI'}(]p]'()[t;if1 ]jpa_s-,3 and [hug lgadg [.3 a (1,.-,3 in the trigtyc.
`l.Di., triglycerides and cholesterol, a number of the above-
`eride level as a result of a lack of catabolism of secreted
`m°mi01'1'-3d Ti-5k f«'lCl0F5 which 3“? li1"0lVC(l
`in VCSSCI Wall
`VLDL particles. This triglyceride rise can be used as a
`Ch'fm3‘35 31'“ lb“-S -‘5iml1lla“‘WU5lY dc°:r"'3-“*‘3‘l-
`The compounds according to the invention can therefore lu measure of mg VLDL Sacrefion mtg
`bf‘ °mPl°Y“_‘l R” _ll“‘ P“""”ll}°" “"1 ”_“‘*l'_1“”‘l “I
`Blood is taken from the animals before and also one and
`alhcr0sl:k."r.OSlS’ chewy‘ pancrcmllls and O0nSnl.')an0n'.
`two hours after administration of the detergent by puncture
`l.lnhll-mm" of [he mlease 0fAp0H'l(l(lAfiwclalcd ldmp‘
`ofthe retroorbital venous plexus. The blood is incubated for
`m°.mF’
`.
`.
`.
`.
`.
`two hours at room temperature and then overnight at 4“ C.
`The test for detecting the inhibition of the release of
`.
`_
`.
`_
`’
`.‘
`.
`_’
`A oli-100-associated 1i )0 roteins from liver cells was car-
`'5 In Md“ 10 and domng wmplelelif It 13 than Cenmfuged ‘H
`v
`-
`v
`I p
`-
`-
`-
`"I0 000
`for 5 minutes. The tri
`lvceride concentration in the
`-p
`ried out in vitro using cultured liver cells, preferably using
`’
`3
`.
`,
`.g *
`,
`.
`.
`..
`Cons of the human Hm, HCPGZ T-hcsc cells are Cultured
`serum thusobtainedisdetermined withthe aid ota modihed
`under standard conditions in medium for
`the culture of
`cummerclally avallablfl mzyme msl (Mer‘5l"“l"{Sl® mgl3""'"
`eukaryotic cells, preferably in RPMI 1640 With 10% foetal
`ends N0- '1 43_54)- 100rd of serum are treated with 100 #1 of
`calf SCI-um_ Hcpfig cells Symhesim and SOC}-cm mm the -30
`test reagent in 9whole plates and incubated at room tem-
`culture supernatant ApoB—l(l0—associated lipoprotein par—
`I‘°"3“""5 fm "lo mlm"35"- Thc nllllcfll ‘l9-1"-‘illy l-5 llm" ‘l°lC"'
`tieles which in principle are built up in a similar manner to
`mi“°d 3‘ 3 Wflwlfinglh of 493 “M in 51“
`“11l0T1'1*1li‘3 P1310‘
`the VI.[)I_ and I.lJI. particles which are to be found in the
`Wading 3PP3T3l“-*5 [SIT Spectral S3“-1"“ Samples l'"i"’ll"g 3“
`plagma
`excessively high triglyceride concentration are diluted with
`Thase panictcs can he dammed using an immun0a,.-_.;ay for 35 physiological saline solution. The triglyceride concentration
`human I.[)I_. This immunoassay is carried out using anti-
`‘-Tlnlalnfid in fl“? 53mPl°-*5
`dflermlned Witl} lhe aid "ll 3
`bodies which haw been induced against human LDL in
`standard curve measured in parallel. In this model,
`test
`rabbits under standard conditions. The aiiti—LDL antibodies
`5"b5l““"'“5 are adminislered in”‘“'°“°‘-‘S13’ ciiht” imm°‘li'
`(rabbit 31-,1j_[_[)[_ Ab) wgww purified by afijily chmmal(,g_
`ately before administration of the detergent or orally or
`mph}; on an inununosm-hem using human {Dru Thcsc 3n subcutaneously before initiation of anaesthesia.
`purified rabbit anti—LDL Ab are adsorbed on the surface of
`plastic. Expediently,
`this adsorption is carried out on the
`plastic surface of microtitre plates having 96 wells, pref'er—
`ably on Maxisorp plates. If ApoB—10U—associated particles
`are present in the supernatant of l[e(i2 cells, they can be 35
`bound to the insolubilized rabbit anti—i_DI. Ab, and an
`immune complex results which is bound to the plastic
`surface. Unbound proteins are removed by washing. The
`3. inhibition oflntcstinal Triglyceride Absorption in vivo
`immune complex located on the plastic surface is detected
`(Rats)
`using monoclonal antibodies which have been induced =ttJ
`The substances which are to be investigated for their
`against human I_DI. and purified according to standard
`triglyceride absorption-inhibiting action in vivo are admin-
`conditions. These antibodies were conjugated with the
`istered orally to male Wistar rats having a body weight of
`enzyme peroxidase. Peroxidase converts the colourless sub—
`between I70 and 230 g. For this purpose, the animals are
`strate TMB into a coloured product in the presence oflI2O2.
`After acidification of the reaction mixture with H2804, the 45 divided into groups of 6 animals 18 hours before substance
`specific light absorption at -150 rim is detemined, which is a
`administration and food is then withdrawn from them.
`measure of the amount of ApoB—1(J0assoeiated particles
`Drking water is available to the animals ad libitum The
`which have been secreted into the culture supernatant by the
`animals of the control groups receive an aqueous tragacanth
`lIepG2 cells.
`suspension or a tragacanth suspension which contains olive
`Surprisingly, the compounds according to the invention 50 oil. 'lhc tragacantl1—olive oil suspension is prepared using an
`inhibit the release of the /\poRl()U-associated particles. The
`Ultra-Turrax. The substances to be investigated are sus-
`l(f50 value indicates at which substance concentration the
`pended in an appropriate tragacanth—olive oil suspension
`light absorption is inhibited by 50% in comparison with the
`likewise using the Ultra—'l‘urrax, directly before substance
`control (solvent control without substance).
`administration.
`To determine the basal serum triglyceride content, blood
`is taken from each rat by puncture of the retroorbital venous
`pleirus before stomach tube application. The tragacanth
`suspension,
`the tragacanth—olive oil suspensions without
`substance (control arimals) or the substances suspended in
`an an appropriate tragacanth-olive oil suspension are then
`administered to the fasting animals using a stomach tube.
`Further taking of blood to determine the postprandial serum
`triglyceride rise is carried out, as a rule, 1, 2 and 3 hours after
`stomach tube application.
`The blood samples are centrifuged and, after recovering
`the serum, the triglycerides are determined photometrically
`using an ILPOS analyzer 5060 (lippendorf Geratebau,
`
`55
`
`"""-N3“
`1
`5
`-:1
`‘
`
`1(i5fJl"ll_R “mill
`33
`1.1
`1:2
`“'
`
`2. Detenriination of the Vl.l)I. Secretion in vivo in the
`llamster
`The effect of the test substances on VLDL secretion in
`vivo is investigated in the hamster. To do this, golden
`
`65
`
`7 of 70
`
`PENN Ex. 2226
`CFAD V. UPENN
`lPR20l5-01836
`
`

`
`US 6,479,503 B2
`
`'
`
`14
`13
`enzyme test (Boehring Mannheim®, No. 1442350} at a
`Netheler & Hinz GmbH, Hamburg). The determination of
`the triglycerides is carried out completely enzymatically
`wavelength of 546 nm. Samples with triglyceride or cho-
`using a standard commercial UV test.
`lesterol conce