United States Patent [19J
`Skotnicki et al.
`
`[54] CARBAMA1ES OF RAPAMYCIN
`
`[75]
`
`Inventors: Jerauld S. Skotnicki, Allentown,
`N.J.; Yvette L. Palmer, Newtown,
`Pa.; Wenling Kao, Paoli, Pa.; Magid
`A. Abou-Gharbia, Glen Mills, Pa.
`
`[73] Assignee: American Home Products
`Corporation, Madison, N.J.
`
`[21] Appl. No.: 259,701
`
`[22] Filed:
`
`Jun. 14, 1994
`
`Related U.S. Application Data
`[60] Continuation-in-part of Ser. No. 160,984, Dec. 1, 1993,
`abandoned, which is a division of Ser. No. 54,655, Apr.
`23, 1993, Pat. No. 5,302,584, which is a continuation-in(cid:173)
`part of Ser. No. 960,597, Oct. 13, 1992, abandoned.
`
`Int. Cl.6 ................... A61K 31/395; C07D 498/04
`[51]
`[52] U.S. CI •.................................... 514/291; 540/452;
`'
`540/456; 514/212; 514/218; 514/222.5;
`514/229.2; 514/233.2; 514/242; 514/241;
`514/253
`[58] Field of Search .................. 514/291, 63; 540/452,
`540/456
`
`[56]
`
`References Cited
`U.S. PATENT DOCUMENTS
`3,929,992 12/1975 Sehgal et al ........................ 424/122
`3,993,749 11/1976 Seugal et al . ............ : .......... 424/122
`4,316,885 2/1982 Rakhit ................................. 424/122
`4,375,464 3/1983 Sehgal et al ........................ 424/122
`4,401,653 8/1983 Eng ..................................... 424/114
`4,650,803 3/1987 Stella et al .......................... 514/291
`4,885,171 12/1989 Surendra et al .................... 424/122
`5,023,262 6/1991 Caufield et al ...................... 514/291
`5,023,263 6/1991 Von Burg ........................... 514/291
`5,023,264 6/1991 Caufield et al ...................... 514/291
`5,078,999 1/1992 Warner et al ....................... 424/122
`5,080,899 1/1992 Sturm et al ......................... 424/122
`5,091,389 2/1992 Ondeyka et al ..................... 514/291
`5,100,863 3/1992 Schiehser ............................ 514/183
`5,100,899 3/1992 Calne ................................... 5i4/291
`5,102,876 4/1992 Caufield .............................. 544/456
`5,118,677 6/1992 Caufield .............................. 540/456
`5,118,678 6/1992 Kao et al ............................. 540/456
`Bl 5,120,842 7/1993 Failli et al ........................... 540/456
`
`I lllll llllllll Ill lllll lllll lllll lllll lllll 111111111111111111111111111111111
`US005434260A
`[11] Patent Number:
`[45] Date of Patent:
`
`5,434,260
`Jul. 18, 1995
`
`5,120,842 6/1992 Failli et al ........................... 540/456
`5,130,307 7/1992 Failli et al ........................... 540/456
`5,138,051 8/1992 Hughes et al ....................... 540/456
`5,151,413 9/1992 Caufield et al ....................... 514/63
`5,169,851 12/1992 Hughes et al ....................... 540/456
`5,177,203 1/1993 Failli et al ........................... 540/456
`5,194,447 3/1993 Kao ..................................... 540/456
`5,221,670 6/1993 Caufield .............................. 514/183
`5,233,036 8/1993 Hughes ............................... 540/455
`5,260,300 11/1993 Hu ....................................... 540/456
`5,262,423 11/1993 Kao ..................................... 540/456
`5,286,730 2/1994 Caufield et al ...................... 514/291
`5,286, 731 2/1994 Caulfield et al. ................... 514/291
`5,302,584 4/1992 Kao et al ............................. 540/456
`
`FOREIGN PATENT DOCUMENTS
`507555 10/1992 European Pat. Off ............. 514/291
`W091/13899 9/1991 WIPO ................................. 540/456
`
`OTHER PUBLICATIONS
`Vezina et al. "J of Antibiotics", vol. 28, pp. 221-226
`(1975).
`Sehgal et al. "J. of Antibiotics", vol. 28, pp. 727-732
`(1975).
`Balcer et al. "J. of Antibiotics", vol. 31, pp. 539-545
`(1978.
`Stepkonuski "Transplantation Proceedings" vol. 23,
`No. 1 (1991) pp. 507-508.
`Galne et al: "The Lancet" Jun. 3, 1978 pp. 1183-1185.
`Morris et al. "Med. Sci Res." vol. 17 pp. 877-878 (1989).
`Martel et al, "Can J. Physiology and Pharmacology"
`vol. 55 pp. 48-51 (1977).
`Staruch et al. FASEB vol. 3, p. 3477 (1989).
`Staruch et al FASEB vol. 3, p. 5256 (1989).
`(Abstract) Fifth Int Conf Infamm. Res. Assoc. p. 121
`(1990).
`(Abstract) J. Heart and Lung Transplantation vol. 11 pt
`2 (1992).
`
`Primary Examiner-Robert T. Bond
`Attorney, Agent, or Firm-Arnold S. Milowsky
`
`[57]
`ABSTRACT
`A compound of the structure
`(Abstract continued on next page.)
`
`West-Ward Exhibit 1067
`Skotnicki USP '260
`Page 001
`
`

`

`5,434,260
`
`Page 2
`
`Abstract-continued
`
`II
`
`-CN,
`-SR5,
`-ORS,
`halogen,
`-S03R5, -S02NR5R6, or Ar;
`R5 and R6 are each, independently, hydrogen, alkyl,
`alkenyl, alkynyl, or Ar;
`
`-S02Rs,
`
`XC=N
`I
`'
`I
`I
`
`/'
`' .... _ .....
`
`\
`
`~-c?<'<
`\
`I
`I
`\
`,,' , and
`,_,
`
`wherein R and R 1 are each, independently, hydrogen,
`
`0
`
`II -c,
`N-C=N,
`I
`I
`\
`R2
`I
`/
`\
`
`, '
`'-...-"'
`
`0
`II
`N
`-C
`'N-C-.:::' 'R3
`I
`'
`I
`I
`\
`/
`,
`'
`,_,
`
`, or
`
`R4
`0
`I
`II
`-c,N,.....c~N
`I
`I
`,
`\
`I ;
`'
`,_,
`
`R2 and R3 are each, independently, hydrogen, alkyl,
`-C02R5, -CQR5, -CN,
`alkenyl, alkynyl.
`-N02, -S02Rs, -S03Rs, -OR5, -SR5, or Ar;
`R4 is hydrogen, alkyl, alkenyl, alkynyl, -CF3,
`-NR5R6, -C02R5, -COR5, CQNR5R6, -N02,
`
`are each, independently, a 5-7 membered satu(cid:173)
`rated, unsaturated, or partially unsaturated hetero(cid:173)
`cyclic radical, that is optionally fused to a phenyl
`ring or a cycloalkane or cycloalkene ring, wherein
`the heterocyclic ring may optionally contain 0, S,
`or NR 8 in the heterocyclic ring, and may be option(cid:173)
`ally substituted by R7;
`R7 is alkyl of 1-6 carbon atoms, alkenyl, alkynyl,
`-CF3, -NR5R6, -C02Rs, -CQR5, CONR5R6,
`-N02, halogen, -ORS, -SR5, -CN, -S02Rs,
`-S03R5, -S02NR5R6, or Ar;
`R8 is hydrogen, alkyl, alkenyl, alkynyl, -CF3,
`-NR5R6, :._co2R5, -CQR5, CQNR5R6, -QR5,
`-SRs, -CN, -S02Rs, -S03R5, -S02NR5R6,
`or Ar;
`Ar is phenyl, naphthyl, or hetaryl, wherein the forego(cid:173)
`ing may be optionally substituted; with the proviso that
`R and RI are both not hydrogen, or a pharmaceutically
`acceptable salt thereof, which is useful as an immuno(cid:173)
`suppressive, antiinflammatory, antifungal, antiprolifera(cid:173)
`tive, and antitumor agent.
`
`1 Claim, No Drawings
`
`West-Ward Exhibit 1067
`Skotnicki USP '260
`Page 002
`
`

`

`1
`
`CARBAMATES OF RAPAMYCIN
`
`5,434,260
`
`2
`tory, antifungal, antiproliferative, and antitumor agents
`having the structure
`
`CROSS-REFERENCE TO RELATED
`APPLICATIONS
`This application is a continuation in part of Ser. No.
`08/160,984, filed Dec. 1, 1993 abandoned, which is a
`divisional of Ser. No. 08/054,655, filed Apr. 23, 1993
`(now U.S. Pat. No. 5,302,584), which is a continuation
`in part of Ser. No. 07 /960,597, filed Oct. 13, 1992 now
`abandoned.
`
`5
`
`10
`
`15
`
`20
`
`25
`
`BACKGROUND OF THE INVENTION
`This invention relates to carbamates of rapamycin
`and a method for using them for inducing immunosup(cid:173)
`pression, and in the treatment of transplantation rejec(cid:173)
`tion, graft vs. host disease, autoimmune diseases, dis(cid:173)
`eases of inflammation, solid tumors, fungal infections,
`and hyperproliferative vascular disorders.
`Rapamycin is a macrocyclic triene antibiotic pro(cid:173)
`duced by Streptomyces hygroscopicus, which was found
`to have antifungal activity, particularly against Candida
`albicans, both in vitro and in vivo [C. Vezina et al., J.
`Antibiot. 28, 721 (1975); S. N. Sehgal et al.~~- Antibiot.
`28, 727 (1975); H. A. Baker et al., J. Antib1ot. 31,539
`(1978); U.S. Pat. No. 3,929,992; and U.S. Pat. No.
`3,993,749].
`Rapamycin alone (U.S. Pat. No. 4,885,171 ) or in
`combination with picibanil (U.S. Pat. No. 4,401,653) has 30
`been shown to have antitumor activity. R. Martel et al.
`[Can. J. Physiol. Pharmacol. 55, 48 (1977)] disclosed
`that rapamycin is effective in the experimental allergic
`encephalomyelitis model, a model for multiple sclerosis;
`in the adjuvant arthritis model, a model for rheumatoid 35
`arthritis; and effectively inhibited the formation of IgE(cid:173)
`like antibodies.
`The immunosuppressive effects of rapamycin have
`been disclosed in FASEB 3, 3411 (1989). Cyclosporin A
`and FK-506, other macrocyclic molecules, also have 40
`been shown to be effective as immunosuppressive
`agents, therefore useful in preventing transplant rejec(cid:173)
`tion [FASEB 3, 3411 (1989); FASEB 3, 5256 (1989); R.
`Y. Calne et al., Lancet 1183 (1978); and U.S. Pat. No.
`5,100,899].
`Rapamycin has also been shown to be useful in pre(cid:173)
`venting or treating systemic lupus erythematosus [U.S.
`Pat. No. 5,078,999], pulmonary inflammation [U.S. Pat.
`No. 5,080,899], insulin dependent diabetes mellitus
`[Fifth Int. Conf. Inflamm. Res. Assoc. 121 (Abstract), 50
`(1990)], and smooth muscle cell proliferation and inti(cid:173)
`mal thickening following vascular injury [Morris, R. J.
`Heart Lung Transplant 11 (pt. 2): 197 (1992)].
`Mono- and diacylated derivatives of rapamycin (es(cid:173)
`terified at the 28 and 43 positions) have been shown to 55
`be useful as antifungal agents (U.S. Pat. No. 4,316,885)
`and used to make water soluble prodrugs of rapamycin
`(U.S. Pat. No. 4,650,803). Recently, the numbering
`convention for rapamycin has been changed; therefore
`according to Chemical Abstracts nomenclature, the 60
`esters described above would be at the 31- and 42- posi(cid:173)
`tions. U.S. Pat. No. 5,118,678 discloses carbamates of
`rapamycin that are useful as immunosuppressive, anti(cid:173)
`inflammatory, antifungal, and antitumor agents.
`
`45
`
`DESCRIPTION OF THE INVENTION
`This invention provides derivatives of rapamycin
`which are useful as immunosuppressive, antiinflamma-
`
`65
`
`.
`.
`2
`1
`wherem R and R are each, mdependently, hydrogen,
`-CONH-[(CR3R4)m(-A-(CR5R_6)n)p]q-B;
`,.--
`\
`'
`-co-N
`, __
`i.
`"'
`I
`'
`
`R9
`
`/
`, or
`-co-N
`'-.RIO
`
`\
`
`R3, R4, RS, R6, and Bare each, independently, hydro(cid:173)
`gen, alkyl of 1-6 carbon atoms, alkenyl of 2-7 car(cid:173)
`bon atoms, alkynyl of 2-7 carbon atoms, hydroxy(cid:173)
`alkyl of 1-6 carbon atoms, alkoxyalkyl of 2-12
`carbon atoms, alkylthioalkyl of2-12 carbon atoms,
`alkylaminoalkyl of2-12 carbon atoms, dialkylami(cid:173)
`noalkyl of 3-12 carbon atoms, arylalkyl of 7-10
`carbon atoms, cycloalkyl of 3-8 carbon atoms,
`-OR7, -SR7, halogen, -CN, -N02, -CF3,
`-COR7, -C02R7, -CONHR7, -S02R7, -O-
`S03R7, -NR7R8, -NHCOR7, -NHS02R7, or
`Ar;
`R 7 and R 8 am each, independently, hydrogen, alkyl of
`1-6 carbon atoms, arylalkyl of7-10 carbon atoms,
`alkenyl of2-7 carbon atoms, alkynyl of2-7 carbon
`atoms, hydroxyalkyl of 1-6 carbon atoms, alkoxy(cid:173)
`alkyl of 2-12 carbon atoms, alkylthioalkyl of 2-12
`carbon atoms, alkylaminoalkyl of 2-12 carbon
`atoms, dialkylaminoalkyl of 3-12 carbon atoms,
`cycloalkyl of 3-8 carbon atoms, or Ar;
`R9 and R 10 are each, independently, alkyl of 1-6 car(cid:173)
`bon atoms, alkenyl of2-7 carbon atoms, alkynyl of
`2-7 carbon atoms, hydroxyalkyl of 1-6 carbon
`atoms, alkoxyalkyl of 2-12 carbon atoms, alkylthi(cid:173)
`oalkyl of 2-12 carbon atoms, alkylaminoalkyl of
`2-12 carbon atoms, dialkylaminoalkyl of 3-12 car(cid:173)
`bon atoms, arylalkyl of7-10 carbon atoms, cycloal(cid:173)
`kyl of 3-8 carbon atoms, -CF3, -COR7,
`-C02R7, -CONHR7, -S02R7, or Ar;
`A is -CH2-, -NR7-, -0-, -S-, -SO-,
`-S02-, -PR7-, -CO-, -NHCO-, -NH(cid:173)
`SO-, or -P(O)(R7)-;
`Ar is phenyl, naphthyl, pyridyl, quinolyl, isoquinolyl,
`quinoxalyl, thienyl, thionaphthyl, fury!, benzofu(cid:173)
`ryl, benzodioxyl, benzoxazolyl, benzoisoxazolyl,
`indolyl, thiazolyl, isoxazolyl, pyrimidinyl, pyrazi(cid:173)
`nyl, imidazolyl, benzopyranyl, benz[b]thiopheno-
`
`West-Ward Exhibit 1067
`Skotnicki USP '260
`Page 003
`
`

`

`5,434,260
`
`4
`pyrrolidinyl, or imidazolyl group that may be option(cid:173)
`ally substituted as described above.
`This invention also discloses preferred arnidino car(cid:173)
`bamates having the structure
`
`3
`lyl, benzimidazolyl, benzthiazolyl, benzodioxolyl,
`piperidinyl, morpholinyl, piperazinyl, tetrahydro(cid:173)
`furanyl, or pyrrolidinyl; wherein the Ar group may
`be optionally mono-, di-, or trisubstituted with a
`group selected from alkyl of 1-6 carbon atoms, 5
`arylalkyl of 7-10 carbon atoms, alkoxy of 1-6 car(cid:173)
`bon atoms, cyano, halo, hydroxy, nitro, carbalkoxy
`of 2-7 carbon atoms, trifluoromethyl, amino, dial(cid:173)
`kylamino of 1-6 carbon atoms per alkyl group,
`dialkylaminoalkyl of 3-12 carbon atoms, hydroxy- 10
`alkyl of 1-6 carbon atoms, alkoxyalkyl of 2-12
`carbon atoms, alkylthio of 1-6 carbon atoms,
`-S03H, -P03H, and -C02H;
`
`II
`
`wherein R and R 1 are each, independently, hydrogen,
`
`0
`II
`-c,
`N-C=N
`I
`I
`\
`2
`I
`I
`R
`I
`I,
`
`'
`,_ ...
`
`/
`
`0
`II
`-C
`'N-v '-~3
`.... __ ..,
`
`I
`,
`
`I
`/
`
`, or
`
`N
`
`R4
`0
`I
`II
`-c, ,,....c::,,..
`N ~N
`,
`j.
`I
`\
`,_ ....
`
`R2 and R3 are each, independently, hydrogen, alkyl of
`1-6 carbon atoms, alkenyl of 2-7 carbon atoms,
`alkynyl of 2-7 carbon atoms, -C02R5, -CORS,
`-CN, -N02, -S02Rs, -S03Rs, -ORS, -SRS,
`or Ar;
`R4is hydrogen, alkyl of 1-6 carbon atoms, alkenyl of
`2-7 carbon atoms, alkynyl of 2-7 carbon atoms,
`-CF3, -NR5R6, -C02Rs, -COR5, CONR5R6,
`-N02, halogen, -ORS, -SR5, -CN, -S02Rs,
`-S03R5, -S02NR5R6, or Ar;
`R 5 and R 6 are each, independently, hydrogen, alkyl of
`1-6 carbon atoms, alkenyl of 2-7 carbon atoms,
`alkynyl of 2-7 carbon atoms, or Ar;
`
`XN-C~
`,
`\
`:
`I
`, ___ ..,
`\
`I
`
`,
`
`and
`
`+
`
`XN/C.:::::,N
`I
`I
`I
`I
`
`,
`'
`,_ ....
`
`is a nitrogen containing hetcrocycle that may be
`saturated, unsaturated, or partially unsaturated,
`and may be optionally mono-, di-, or tri- substituted
`with a group selected from alkyl of 1-6 carbon
`atoms, arylalkyl of 7-10 carbon atoms, alkoxy of 25
`1-6 carbon atoms, cyano, halo, hydroxy, nitro,
`carbalkoxy of 2-7 carbon atoms, trifluoromethyl,
`amino, dialkylamino of 1-6 carbon atoms per alkyl
`group, dialkylaminoalkyl of 3-12 carbon atoms,
`hydroxyalkyl of 1-6 carbon atoms, alkoxyalkyl of 30
`2-12 carbon atoms, alkylthio of 1-6 carbon atoms,
`-S03H, -P03H, and -C02H;
`with the proviso that R 1 and R 2 are not both hydrogen;
`m=0-6;
`n=0-6;
`p=0-1;
`q=0-1;
`or a pharmaceutically acceptable salt thereof.
`The pharmaceutically acceptable salts are those de(cid:173)
`rived from such inorganic cations such as sodium, po- 40
`tassium, and the like; organic bases such as: mono-, di-,
`and trialkyl amines of 1-6 carbon atoms, per alkyl group
`and mono-, di-, and trihydroxyalkyl amines of 1-6 car(cid:173)
`bon atoms per alkyl group, and the like; and organic and
`inorganic acids as: acetic, lactic, citric, tartaric, sue- 45
`cinic, maleic, malonic, gluconic, hydrochloric, hydro(cid:173)
`bromic, phosphoric, nitric, sulfuric, methanesulfonic,
`and similarly known acceptable acids.
`It is preferred that the aryl portion of the arylalkyl
`substituent is a phenyl, piperazinyl, piperidinyl, or pyri- 50
`dyl group that is optionally mono-, di-, or tri-substituted
`with a group selected from .alkyl of 1-6 carbon atoms,
`arylalkyl of 7-10 carbon atoms, alkoxy of 1-6 carbon
`atoms, cyano, halo, nitro, carbalkoxy of 2-7 carbon
`atoms, trifluoromethyl, amino, dialkylamino of 1-6 car- 55
`bon atoms per alkyl group, alkylthio of 1-6 carbon
`atoms, -S03H, -P03H, and -C02H. The term alkyl
`includes both straight chain and branched alkyl groups.
`It is preferred that
`
`15
`
`20
`
`35
`
`60
`
`65
`
`is a pyridyl, pyrazinyl, piperidinyl, morpholinyl, pipera(cid:173)
`zinyl, pyrrolidinyl, thiazolyl, pyrimidinyl, isoxazolyl,
`
`are each, independently, a 5-7 membered satu(cid:173)
`rated, unsaturated, or partially unsaturated hetero-
`
`West-Ward Exhibit 1067
`Skotnicki USP '260
`Page 004
`
`

`

`10
`
`15
`
`20
`
`5
`cyclic radical, that is optionally fused to a phenyl
`ring or a cycloalkane or cycloalkene ring of 5-7
`carbon atoms, wherein the heterocyclic ring may
`optionally contain 0, S, or NR 8 in the heterocyclic
`ring, and may be optionally substituted by R7;
`R 7 is alkyl of 1-6 carbon atoms, alkenyl of 2-7 carbon
`atoms, alkynyl of 2-7 carbon atoms, -CF3,
`-NRSR6, -C02Rs, -CORs, CONR5R6, -N02,
`-ORS,
`-SRS,
`-CN,
`-S02RS,
`halogen,
`-S03Rs, -S02NR5R6, or Ar;
`RS is hydrogen, alkyl of 1-6 carbon atoms, alkenyl of
`2-7 carbon atoms, alkynyl of 2-7 carbon atoms,
`-CF3, -NR5R6, -C02Rs, -COR5, CONRSR6,
`-ORS,
`-SRS, -CN,
`-S02Rs,
`-S03R5,
`-S02NR5R6, or Ar;
`Ar is phenyl, naphthyl, or hetaryl, wherein the fore(cid:173)
`going may be optionally mono-, di-, or tri-sub(cid:173)
`stituted with a group selected from alkyl of 1-6
`carbon atoms, alkenyl of 2-7 carbon atoms, alkynyl
`of 2-7 carbon atoms, arylalkyl of 7-10 carbon
`atoms, alkoxy of 1-6 carbon atoms, cyano, halo, hy(cid:173)
`droxy, nitro, carbalkoxy of2-7 carbon atoms, trifluoro(cid:173)
`methyl, trifluoromethoxy, amino, dialkylamino of 1-6
`carbon atoms per alkyl group, dialkylaminoalkyl of
`3-12 carbon atoms, hydroxyalkyl of 1-6 carbon atoms, 25
`alkoxyalkyl of2-12 carbon atoms, alkylthio of 1-6 car(cid:173)
`bon atoms, -S03H, and -C02H;
`with the proviso that R and RI are both not hydrogen,
`or a pharmaceutically acceptable salt thereof.
`For the compounds having structure II (immediately 30
`above), the terms alkyl of 1-6 carbon atoms, alkenyl of
`2-7 carbon atoms, and alkynyl of 2-7 carbon atoms,
`include both straight chain as well as branched carbon
`chains. When any of the generic terms (i.e., R5) are
`contained more than once in a given compound, each 35
`may be the same or different. The pharmaceutically
`acceptable salts of the compounds having structure II
`are the same as was defined following the compounds of
`structure I.
`For the compounds having structure II, hetaryl is 40
`defined as an unsaturated or partially saturated hetero(cid:173)
`cyclic radical of 5-12 atoms having 1 ring or 2 fused
`rings. Preferred heterocyclic radicals include unsatu(cid:173)
`rated heterocyclic radicals such as furanyl, thiophenyl,
`pyrrolyl, pyrazolyl, imidazolyl, 1,2,3-triazolyl, 1,2,4- 45
`triazolyl, 1,2-dithiolyl, 1,3-dithiolyl, 1,2,3-oxathiolyl,
`isoxazolyl, oxazolyl,
`thiazolyl,
`isothiazolyl, 1,2,3-
`oxadiazolyl,
`1,2,5-oxadiazolyl,
`1,3,4-oxadiazolyl,
`1,2,3,4-oxatriazolyl, 1,2,3,5-oxatriazolyl, 1,2,3-dioxazo(cid:173)
`lyl, 1,2,4-dioxazolyl, 1,3,2-dioxazolyl, 1,3,4-dioxazolyl, 50
`1,2,5-oxathiazolyl, 1,3-oxathiolyl, 1,2-pyranyl, 1,4-pyra(cid:173)
`nyl, pyridinyl, pyridazinyl, pyrimidinyl, pyrazinyl,
`1,3,5-triazinyl, 1,2,4-triazinyl, 1,2,3-triazinyl, 1,2,4-
`oxazinyl, 1,3,2-oxazinyl, 1,2,6-oxazinyl, 1,4-oxazinyl,
`isoxazinyl, 1,2,5-oxathiazinyl, 1,4-oxazinyl, o-isoxazinyl, 55
`p-isoxazinyl,
`1,2,5-oxathiazinyl,
`1,2,6-oxathiazinyl,
`1,3,5,2-oxadiazinyl, azepinyl, oxepinyl, thiepinyl, 1,2,4-
`diazepinyl, benzofuranyl, isobenzofuranyl, thionaphthe(cid:173)
`nyl, indolyl, indolenyl, 2-isobenzazolyl, 1,5-pyrindinyl,
`pyrano[3,4-b)pyrrolyl, benzpyrazolyl, benzisoxazolyl, 60
`benzoxazolyl, anthranilyl, 1,2-benzopyranyl, quinolinyl,
`isoquinolinyl, cinnolinyl, quinazolinyl, naphthyridinyl,
`pyrido[3,4-b ]pyridinyl, pyrido[ 4,3-b ]pyridinyl, pyri(cid:173)
`do[2,3-b )pyridinyl, 1,3,2-benzozazinyl, 1,4,2-benzoxazi(cid:173)
`nyl, 2,3,1-benzoxazinyl, 3,1,4-benzoxazinyl, 1,2-ben- 65
`zisoxazinyl, 1,4-benzisoxazinyl, carbazolyl, purinyl, and
`partially saturated heterocyclic radicals selected from
`the list above. All of the preferred heterocyclic radicals
`
`5,434,260
`
`6
`contain at least one double bond. When the heterocyclic
`radical is partially saturated, one or more of the olefms
`in the unsaturated ring system is saturated; the partially
`saturated heterocyclic radical still contains at least one
`5 double bond. It is more preferred that hetaryl is pyridi(cid:173)
`nyl.
`For the amidino carbamates having structure II, it is
`preferred that the
`
`and
`
`+
`xN....-c~N
`I
`I
`.... _ ...
`\
`I
`'
`
`/
`
`moieties are pyrrolyl, 3,4-dihydropyrrolyl, pyrolidinyl,
`pyrazolyl,
`imidazolyl,
`4,5-dihydroimidazolyl,
`2-
`pyrazolinyl, 1,2,4-triazolyl, isoxazolyl, oxazolyl, thia(cid:173)
`zolyl, isothiazolyl, pyridinyl, 1,2,3,6-tetrahydropyridi(cid:173)
`nyl, piperidinyl, pyridazinyl, pyrimidinyl, 1,4,5,6-tet(cid:173)
`rahydropyrimidinyl, pyrazinyl, piperazinyl, 1,3,5-triazi(cid:173)
`nyl, hexahydro-1,3,5-triazinyl, 1,2,4-triazinyl, 1,3,2-
`oxazinyl, 1,4-oxazinyl, morpholinyl, azepinyl, indolyl,
`indolinyl, indolenyl, benzoxazolyl, quinolinyl, 1,2,3,4-
`tetrahydroquinolinyl,
`5,6, 7 ,8-tetrahydroquinolinyl,
`isoquinolinyl, 3,4-dihydroisoquinolinyl, 1,2,3,4-tetrahy(cid:173)
`droisoquinolinyl, and, 5,6, 7,8-tetrahydroisoquinolinyl
`radicals that are optionally substituted by RB. The
`fourth ring structure, above, is the more preferred of the
`four ring systems, and within this ring system it is more
`preferred that the radical is 4,5-dihydroimidazolyl or
`1,4,5,6-tetrahydropyrimidinyl. It is also preferred that
`the R7 substituent, when present, is phenyl.
`The compounds of this invention carbamylated at the
`42-position or at both the 31- and 42-positions can be
`prepared by converting the 42- and/or 31-alcohols of
`rapamycin to a carbonate (see Example 1 ) followed by
`reaction with an appropriately substituted amine to
`provide the desired carbamate. The following scheme
`illustrates the preparation of the compound of Example
`4.
`
`C)-0
`
`H
`
`West-Ward Exhibit 1067
`Skotnicki USP '260
`Page 005
`
`

`

`5,434,260
`
`7
`-continued
`
`Example4
`
`8
`concentrations of rapamycin, cyclosporin A, or test
`compound. Cells are harvested and incorporated radio(cid:173)
`activity is determined. Inhibition of lymphoprolifera(cid:173)
`tion is assessed as percent change in counts per minute
`S from non-drug treated controls. For each compound
`evaluated, rapamycin was also evaluated for the pur(cid:173)
`pose of comparison. An ICso was obtained for each test
`compound as well as for rapamycin. When evaluated as
`a comparator for the representative compounds of this
`10 invention, rapamycin had an IC50 ranging from 0.3-0.9
`nM. The results obtained are provided as an IC50 and as
`the percent inhibition of T-cell proliferation at 0.1 µM.
`The results obtained for the representative compounds
`of this invention were also expressed as a ratio com(cid:173)
`pared with rapamycin. A positive ratio indicates immu-
`nosuppressive activity. A ratio of greater than 1 indi(cid:173)
`cates that the test compound inhibited thymocyte pro(cid:173)
`liferation to a greater extent than rapamycin. Calcula(cid:173)
`tion of the ratio is shown below.
`
`20
`
`Alternatively, the compounds of this invention car(cid:173)
`bamylated at the 42-position or at both the 31- and
`42-positions can be prepared by reacting rapamycin 15
`with an appropriately substituted isocyanate under neu(cid:173)
`tral conditions or in the presence of a base, such as
`pyridine. Preparation of carbamates of rapamycin using
`this method was disclosed in U.S. Pat. No. 5,118,678,
`which is hereby incorporated by reference. The 31-car-
`bamylated compounds of this invention can be prepared
`by protecting the 42-alcohol of rapamycin with a pro(cid:173)
`tecting group, such as with a tertbutyl dimethylsilyl
`group, followed by carbamylation of the 31-position by
`the procedures described above. Removal of the pro(cid:173)
`Representative compounds of this invention were
`tecting group provides the 31-carbamylated com- 25
`also evaluated in an in vivo test procedure designed to
`pounds. In the case of the tert-butyl dimethylsilyl pro(cid:173)
`determine the survival time of pinch skin graft from
`tecting group, deprotection can be accomplished under
`male BAB/c donors transplanted to male C3H(H-2K)
`mildly acidic conditions. Having the 31-position car(cid:173)
`recipients. The method is adapted from Billingham R.
`bamylated and the 42-position deprotected, the 42-posi(cid:173)
`E. and Medawar P. B., J. Exp. Biol. 28:385-402, (1951).
`tion can be carbamylated using a different amine (via 30
`Briefly, a pinch skin graft from the donor was grafted
`the carbonate) or isocyanate than was reacted with the
`on the dorsum of the recipient as a allograft, and an
`31-alcohol, to give compounds having different carba(cid:173)
`isograft was used as control in the same region. The
`mates at the 31- and 42- positions. Alternatively, the
`recipients were treated with either varying concentra(cid:173)
`42-carbamylated compounds, prepared as described
`tions of test compounds intraperitoneally or orally.
`above, can be reacted with a different amine (via the 35
`Rapamycin was used as a test control. Untreated recipi(cid:173)
`carbonate) or isocyanate to provide compounds having
`ents serve as rejection control. The graft was monitored
`different carbamates at the 31- and 42-positions.
`daily and observations were recorded until the graft
`The amines and isocyanates used to prepare the com(cid:173)
`became dry and formed a blackened scab. This was
`pounds of the invention are commercially available or
`considered as the rejection day. The mean graft survival
`can be prepared by methods that are disclosed in the 40
`time (number of days±S.D.) of the drug treatment
`literature.
`group was compared with the control group. The fol(cid:173)
`This invention also covers analogous carbamates of
`lowing table shows the results that were obtained. Re(cid:173)
`other rapamycins such as, but not limited to, 29-deme(cid:173)
`sults are expressed as the mean survival time in days.
`thoxyrapamycin, [U.S. Pat. No. 4,375,464, 32-deme(cid:173)
`Untreated (control) pinch skin grafts are usually re(cid:173)
`thoxyrapamycin under C.A. nomenclature]; rapamycin 45
`jected within 6-7 days. The results shown in Table 1 are
`derivatives in which the double bonds in the 1-, 3-,
`based on a dose of 4 mg/kg of test compound. A sur(cid:173)
`and/or 5-positions have been reduced [U.S. Pat. No.
`5,023,262]; 42-oxorapamycin [U.S. Pat. No. 5,023,262];
`vival time of 12.0±1.7 days was obtained for rapamycin
`29-desmethylrapamycin [U.S. Pat. No. 5,093,339, 32-
`at 4 mg/kg.
`desmethylrapamycin under C.A. nomenclature]; 7,29- SO
`The following table summarizes the results of repre(cid:173)
`bisdesmethylrapamycin [U.S. Pat. No. 5,093,338, 7,32-
`sentative compounds of this invention in these two
`desmethylrapamycin under C.A. nomenclature]; and
`standard test procedures.
`15-hydroxy- and 15,27-bishydroxy- rapamycin [U.S.
`TABLE 1
`Pat. No. 5,102,876]. The disclosures in the above cited
`U.S. Pat. Nos. are hereby incorporated by reference.
`EVALUATION OF IMMUNOSUPPRESSIVE ACTIVITY*
`Immunosuppressive activity for representative com(cid:173)
`LAF
`Skin Graft
`(days± SD)
`pounds of this invention was evaluated in an ill vitro
`Compound
`(ratio)
`ICso (nM)
`standard pharmacological test procedure to measure
`0.54
`Example 2
`0.63
`10.8 ± 1.0
`Example 3
`0.20
`1.70
`9.8 ± 1.5
`lymphocyte proliferation (LAF) and in an ill vivo stan(cid:173)
`Example4
`0.78
`1.21
`dard pharmacological test procedure which evaluated 60 - - - - - - - - - - - - - - - - - - - - (cid:173)
`the survival time of a pinch skin graft.
`•Calculation of the ratio was described supra.
`The comitogen-induced thymocyte proliferation pro(cid:173)
`cedure (LAF) was used as an in vitro measure of the
`immunosuppressive effects of representative com(cid:173)
`pounds. Briefly, cells from the thymus of normal 65
`BALB/c mice are cultured for 72 hours with PHA and
`IL-1 and pulsed with tritiated thymidine during the last
`six hours. Cells are cultured with and without various
`
`ICso of Rapamycin
`IC50 of Test Compound
`
`55
`
`The results of these standard pharmacological test
`procedures demonstrate immunosuppressive activity
`both in vitro and in vivo for the compounds of this
`invention. Positive ratios in the LAF test procedures
`indicates suppression of T-cell proliferation, thereby
`demonstrating the immunosuppressive activity of the
`
`West-Ward Exhibit 1067
`Skotnicki USP '260
`Page 006
`
`

`

`25
`
`9
`compounds of this invention. As transplanted pinch skin
`grafts are typically rejected within 6-7 days without the
`use of an immunosuppressive agent, the increased sur(cid:173)
`vival time of the skin graft when treated with the com(cid:173)
`pounds of this invention further demonstrates their 5
`utility as immunosuppressive agents. Additionally, the
`results obtained in the skin graft test procedure further
`demonstrates the ability of the compounds of this inven(cid:173)
`tion to treat or inhibit transplantation rejection.
`Based on the results of these standard pharmacologi- 10
`cal test procedures, the compounds are useful in the
`treatment or inhibition of transplantation rejection such
`as kidney, heart, liver, lung, bone marrow, pancreas
`(islet cells), cornea, small bowel, and skin allografts, and
`heart valve xenografts; in the treatment or inhibition of 15
`graft vs. host disease; in the treatment or inhibition of
`autoimmune diseases such as lupus, rheumatoid arthri(cid:173)
`tis, diabetes mellitus, myasthenia gravis, and multiple
`sclerosis; and diseases of inflammation such as psoriasis,
`dermatitis, eczema, seborrhea, inflammatory bowel 20
`disease, pulmonary inflammation (including asthma,
`chronic obstructive pulmonary disease, emphysema,
`acute respiratory distress syndrome, bronchitis, and the
`like), and eye uveitis.
`Because of the activity profile obtained, the com(cid:173)
`pounds of this invention also are considered to have
`antitumor, antifungal activities, and antiproliferative
`activities. The compounds of this invention therefore
`also useful in treating solid tumors, adult T-cell leuke- 30
`mia/lymphoma, fungal infections, and hyperprolifera(cid:173)
`tive vascular diseases such as restenosis and atheroscler(cid:173)
`osis. When used for restenosis, it is preferred that the
`compounds of this invention are used to treat restenosis
`that occurs following an angioplasty procedure. When 35
`used for this purpose, the compounds of this invention
`can be administered prior to the procedure, during the
`procedure, subsequent to the procedure, or any combi(cid:173)
`nation of the above.
`When administered for the treatment or inhibition of 40
`the above disease states, the compounds of this inven(cid:173)
`tion can be administered to a mammal orally, parenter(cid:173)
`ally, intranasally, intrabronchially, transdermally, topi(cid:173)
`cally, intravaginally, or rectally.
`It is contemplated that when the compounds of this 45
`invention are used as an immunosuppressive, antirejec(cid:173)
`tion, or antiinflammatory agent, they can be adminis(cid:173)
`tered in conjunction with one or more other im(cid:173)
`munoregulatory agents. Such other immunoregulatory
`agents include, but are not limited to azathioprine, corti- 50
`costeroids, such as prednisone and methylprednisolone,
`cyclophosphamide, rapamycin, cyclosporin A, FK-506,
`OKT-3, and ATG. By combining the compounds of this
`invention with such other drugs or agents for inducing
`immunosuppression or treating inflammatory condi- 55
`tions, lesser amounts of each of the agents are required
`to achieve the desired effect. The basis for such combi(cid:173)
`nation therapy was established by Stepkowski whose
`results showed that the use of a combination of rapamy(cid:173)
`cin and cyclosporin A at subtherapeutic doses signifi- 60
`candy prolonged heart allograft survival time. [Trans(cid:173)
`plantation Proc. 23:507 (1991)].
`The compounds of this invention can be formulated
`neat or with a pharmaceutical carder to a mammal in
`need thereof. The pharmaceutical carder may be solid 65
`or liquid. When formulated orally, it has been found
`that 0.01 % Tween 80 in PHOSAL PG-50 (phospho(cid:173)
`lipid concentrate with 1,2-propylene glycol, A. Natter-
`
`5,434,260
`
`10
`mann & Cie. GmbH) provides an acceptable oral formu(cid:173)
`lation.
`A solid carrier can include one or more substances
`which may also act as flavoring agents, lubricants, solu(cid:173)
`bilizers, suspending agents, fillers, glidants, compression
`aids, binders or tablet-disintegrating agents; it can also
`be an encapsulating material. In powders, the carder is
`a finely divided solid which is in admixture with the
`finely divided active ingredient. In tablets, the active
`ingredient is mixed with a carrier having the necessary
`compression properties in suitable proportions and
`compacted in the shape and size desired. The powders
`and tablets preferably contain up to 99% of the active
`ingredient. Suitable solid carriers include, for example,
`calcium phosphate, magnesium stearate, talc, sugars,
`lactose, dextrin, starch, gelatin, cellulose, methyl cellu(cid:173)
`lose, sodium carboxymethyl cellulose, polyvinylpyr(cid:173)
`rolidine, low melting waxes and ion exchange resins.
`Liquid carriers are used in preparing solutions, sus(cid:173)
`pensions, emulsions, syrups, elixirs and pressurized
`compositions. The active ingredient can be dissolved or
`suspended in a pharmaceutically acceptable liquid car(cid:173)
`rier such as water, an organic solvent, a mixture of both
`or pharma