United States Patent [19]
`
`Fehr
`
`-
`
`[11] Patent Number:
`[45] Date of Patent:
`
`5,011,844
`Apr. 30, 1991
`
`[54] SUBSTITUTED
`4-AZATRICYCLO(22.3.1.04'9)0CI‘ACOS
`18-ENE DERIVATIVES, THEIR
`PREPARATION AND PHARMACEUTICAL
`COMPOSITIONS CONTAINING THEM
`
`[75] Inventor: Theodor Fehr, Dornach, Switzerland
`[73] Assignee: Sandoz Ltd., Basel, Switzerland
`[21] Appl. No.: 399,673
`[22] Filed:
`Aug. 25, 1989
`[30]
`Foreign Application Priority Data
`Aug. 26, 1988 [GB] United Kingdom ............... .. 8820348
`May 31, 1989 [GB] United Kingdom ............... .. 8912432
`
`[51] Int. Cl.5 ................ .. A61K 31/395; C07D 498/16;
`C12P 17/18
`[52] US. Cl. .................................. .. 514/291; 514/411;
`540/456; 435/898; 435/886; 435/118
`[58] Field of Search ................ .. 540/456; 514/63, 183,
`514/291, 411; 435/598, 886, 118
`
`[56]
`
`References Cited
`U.S. PATENT DOCUMENTS
`
`4,894,366 l/199O Okuhara et al. .................. .. 540/456
`
`OTHER PUBLICATIONS
`Jones et al., “J. Am. Chem. Soc.” (1989), vol. III, pp.
`1157-1159.
`
`Primary Examiner-Robert T. Bond
`Attorney, Agent, or Firm-Gerald D. Sharkin; Robert S.
`Honor; Thomas O. McGovern
`
`ABSTRACT
`[57]
`The compounds of formula I
`
`wherein
`either
`R1 is hydroxy,
`R2 is allyl or n-propyl and
`there is a single bond between the carbon atoms num
`bered 14 and 15
`
`or
`R1 is missing,
`R2 is allyl and
`there is a double bond between the carbon atoms
`numbered 14 and 15,
`have interesting immunosuppressant and anti-in?amma
`tory properties.
`I
`They are Obtained by fermentation or synthesis, e.g. by
`hydrogenation or dehydration.
`
`9 Claims, 4 Drawing Sheets
`
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`

`
`US. Patent
`
`Apr. 30, 1991
`
`Sheet 1 of 4
`
`5,011,844
`
`FIG. 1
`
`1200
`
`
`
`2000 1600
`
`3000
`
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`

`
`US. Patent-
`
`‘
`
`Apr. 30, 1991
`
`Sheet 2 of 4
`
`5,011,844
`
`FIG. 2
`
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`
`US. Patent
`
`Apr. 30, 1991
`
`Sheet 3 of 4
`
`5,011,844
`
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`

`
`US. Patent
`
`' Apr. 30, 1991
`
`Sheet 4 of 4
`
`5,011,844
`
`FIG. 1+
`
`m
`
`-m
`
`ppm
`
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`
`5,011,844
`
`SUBSTITUTED
`
`4-AZATRICYCLO(22.3.1.04'9)OCI‘ACOS-18-ENE
`
`DERIVATIVES, THEIR PREPARATION AND
`
`PHARMACEUTICAL COMPOSITIONS
`
`CONTAINING THEM
`
`10
`
`FIELD
`
`The invention relates to the ?eld of natural product
`
`15
`
`chemistry, in particular the chemistry of macrolides.
`
`The invention concerns a compound of formula I
`
`20
`
`OCH3 OCH3
`
`I
`
`CH3
`
`[2
`
`c‘:
`
`0
`
`R2
`
`CH3
`
`110
`
`H C
`3 \0
`
`0
`
`H3C
`
`H3C—O
`
`wherein
`
`either
`
`R1 is hydroxy,
`
`R2 is allyl or n-propyl and there is a single bond be
`
`tween the carbon atoms numbered 14 and 15
`
`or
`
`R‘ is missing,
`
`R2 is allyl and
`
`25
`
`30
`
`35
`
`50
`
`55
`
`there is a double bond between the carbon atoms
`
`numbered 14 and 15.
`
`wherein
`R1 is hydroxy or protected hydroxy,
`R2 is hydrogen, hydroxy or protected hydroxy,
`R3 is methyl, ethyl, propyl or allyl,
`n is an integer of 1 or 2 and the symbol of a line and
`dotted line is a single bond or a double bond,
`and salts thereof.
`As is evident from the above formula, there are many
`asymmetry centers and therefore, a large number of
`possible stereoisomers exist for any given meaning of
`the substituents.
`On the other hand, although on page 4 in EP 184 162
`it is mentioned that there may be one or more conior
`mer(s) or stereoisomeric pairs such as optical and geo
`metrical isomers due to asymmetric carbon atom(s) and
`double bond(s), for none of the compounds speci?cally
`disclosed in EP 184 162 is there any indication of the
`exact stereochemical con?guration.
`This is so in particular for the compound named FK
`900506 (FK 506), which is the object of Examples 1 to
`3 therein, its derivative hydrogenated at the allyl group
`to an n-propyl group, which is the object of Example 21
`therein, and its derivative dehydrated between positions
`45
`, 14 and 15, which is disclosed in Example 17 therein.
`From the formula and the names indicated on page 32,
`95 and 98 of EP 184 162 it is not apparent what con?u
`ration FK 506 and these two derivatives have.
`The con?guration of FK 506 has however been pub
`lished in the scienti?c literature, e.g. in H. Tanaka et al.,
`J. Am. Chem. Soc. 109 (1987) 5031-5033, T. Kino et al.,
`J. Antibiotics 40 (1987) 1249-1255 and T. Taga et al.,
`Acta Cryst. C43 (1987) 751—753.
`It appears therefrom that FK 506 and, by implication,
`the two derivatives thereof mentioned above, have the
`con?guration indicated above for formula I of the pres
`ent invention, except that at the carbon atom numbered
`17 the con?guration is reversed, ie it is the R con?gu
`ration, whereas in formula 1 above the S con?guration
`is shown.
`
`Formula I is meant to cover the compounds in free form
`
`and, where such forms may exist, in salt form.
`
`BACKGROUND ART
`
`Fujisawa E? 184162 discloses a group of compounds
`
`represented by formula A
`
`65
`
`SUMMARY
`It has now been found that, surprisingly, the com
`pounds of formula I, which are novel and are the stereo
`isomers of FK 506, its dihydrogenated derivative and its
`dehydrated derivative, but with the opposite con?gura
`tion at the carbon atom in position 17, have an excellent
`
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`
`5,011,844
`4
`3
`immunosuppressant and antiinflammatory, e.g. antip
`(b) for the preparation of the compound of formula I
`soriatic activity.
`wherein
`R1 is missing and R2 is allyl,
`dehydrating the corresponding compound of formula I
`wherein
`R1 is hydroxy or
`(c) for the preparation of the compound of formula I
`wherein
`R1 is hydroxy and R2 is n-propyl,
`hydrogenating the corresponding compound of formula
`I wherein
`R1 is allyl.
`The invention also concerns a pharmaceutical com
`position containing a compound of formula I as de?ned
`above together with a pharmaceutically acceptable
`carrier or diluent.
`It also concerns a compound of formula I as de?ned
`above for use as a pharmaceutical.
`It also concerns the use of a compound of formula I as
`de?ned above in the preparation of a pharmaceutical
`composition, comprising mixing a compound of for
`mula I with a pharmaceutically acceptable carrier or
`diluent.
`It further concerns a process for the preparation of a
`pharmaceutical composition comprising mixing a com
`pound of formula I as de?ned above with a pharmaceu
`tically acceptable carrier or diluent.
`It further concerns a method for the prevention or
`treatment of conditions requiring immunosuppression
`or of in?ammatory conditions, comprising administer
`ing a therapeutically effective amount of a compound of
`formula I as de?ned above together with a pharmaceu
`tically acceptable carrier or diluent to a subject in need
`of such treatment, e.g. a method of treatment of im- '
`mune-mediated conditions of the eye comprising topi
`callyladministering to the eye surface a therapeutically
`effective amount of a compound of formula I as de?ned
`above in a pharmaceutically acceptable ophthalmic
`vehicle. DR
`
`20
`
`DETAILED DESCRIPTION
`The compounds of formula I are novel. They may be
`prepared in accordance with standard procedures.
`The compounds of formula I wherein R1 is hydroxy
`and R2 is allyl (Compound No. l; “l7-epi-FK506”) or
`wherein R2 is missing and R2 is allyl (Compound No. 3;
`“dehydro-l7-epi-FK506”) may be isolated in known
`manner from e.g. Streptomyces tsukubaensis No. 9993
`using the general procedures described in EP 184 162
`and in the Examples hereafter. Thus, an appropriate
`Streptomyces strain such as Streptomyces tsukubaensis
`No. 9993 may be cultivated in an appropriate culture
`medium and the above two compounds isolated from
`the resultant culture. Cultivation is effected by incuba
`tion, e.g. as described in EP 184 162 or in Example 1
`hereunder. The pH is kept between about 6 and about 8,
`preferably at about 6.8. The temperature may vary
`between about 18° C. and about 35° C., it preierably is
`kept at around 27'’ C.
`The compound of formula I wherein R1 is hydroxy
`and R1 is n-propyl (Compound No. 2; “dihydro-l7-epi
`FKSOG”) may e.g. be prepared in known manner by
`hydrogenation of Compound No. 1, e.g. by catalytic
`reduction using palladium on charcoal as a catalyst. The
`temperature may e.g. vary from about 5° C. to about 30
`° C., preferably about room temperature is used. The
`reaction is preferably effected in the presence of.an inert
`organic solvent such as an alcohol, e.g. ethanol.
`Compound No. 3 may e. g. also be prepared in known
`manner by dehydration of Compound No. l, e.g. by
`catalytic dehydration in an acidic solution. Preferably
`an inert organic solvent such as an ester, e.g. acetic acid
`ethyl ester, is used. The temperature may vary between
`about 5° C. and about 30° C., the reaction preferably is
`effected at about room temperature.
`'
`The compounds of the invention may be isolated and
`40
`puri?ed from the reaction or isolation mixture in known
`manner.
`The producing strain, Streptomyces tsukubaensis No.
`9993, is disclosed in Fujisawa EP 184162. Samples are
`available from the Fermentation Research Institute,
`Tsukuba, Ibaraki 305, Japan under the provisions of the
`Budapest Treaty, under deposit No. FERM BP-927.
`This strain has been redeposited on April 27, 1989 with
`the Agricultural Research Culture Collection Intema
`tIonaI Depository, Peoria, Ill. 61604, USA under the
`provisions of the Budapest Treaty, under deposit No.
`NRRL l8488.
`Compound No. 1 may e.g. also be produced by total
`synthesis according to the procedure published for the
`total synthesis of FK 506 (T. K. Jones et al., J. Am.
`Chem. Soc. 111 [198911157-1159) using corresponding
`epimeric starting materials.
`The invention thus concerns the compounds of for
`mula I as de?ned above.
`It also concerns a process for the preparation of a
`compound of formula I as de?ned above which com
`prises
`(a) for the preparation of the compounds of formula I
`wherein
`R1 is hydroxy or missing and R2 is allyl,
`cultivating an appropriate Streptomyces strain such as
`Streptomyces tsukubaensis No. 9993 and isolating the
`compounds from the resultant mixture,
`
`45
`
`65
`
`EXPLANATION OF THE FIGURES
`FIG. 1: IR-spectrum of Compound No. 1.
`FIG. 2: NMR-spectrum of Compound No. 1.
`FIG. 3: IR-spectrum of Compound No. 2.
`FIG. 4: NMR-spectrum of Compound No. 2.
`The following Examples illustrate the invention and
`are not limitative.
`
`EXAMPLE 1
`Fermentation
`
`' process variant (a), cultivation
`(A) Starting culture on agar
`An agar culture of strain Streptomyces tsukubaensis
`No. 9993 is grown for 14 days at 27° C. on the following
`medium:
`
`Yeast extract (Bacto)
`Malt extract (Bacto)
`Dextrose (Bacto)
`Agar (Bacto)
`dcmineralised water ad
`
`4.0 g
`10.0 g
`4.0 g
`20.0 g
`1000 ml
`
`The pH value is set to 6.6 with NaOH/I-I2SO4 prior to
`sterilization. Sterilization is effected for 20 minutes at
`120° C.
`
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`
`5
`(B) Preculture
`The spores and mycelium from 6 starting cultures are
`suspended in 90 m1 of a 0.9 % solution of sodium chlo
`ride. erlenmeyer ?asks containing each 1 liter of precul- 5
`ture medium are inoculated with 7 ml of this suspension.
`The preculture medium has the following composition:
`
`Glycerine
`Starch
`Glucose
`Cotton seed extract
`(Pharmamedia)
`Yeast extract (Gistex)
`CaCO3
`demineralised water ad
`
`10.0 g
`10.0 g
`5.0 g
`10.0 g
`
`5.0 g
`2.0 g
`1000 ml
`
`10
`
`15
`
`5,011,844
`6
`ration with thrice 70 l of methanol/ water 9:1 and thrice
`70 l of hexane.The methanol/water phase is then evap
`orated to dryness under reduced pressure and the resi~
`due is chromatographed on a column containing 25 kg
`Sephadex LH20 in methanol and then on a column
`containing 20 kg silicagel Merck (0.04 to 0.063 mm)
`using tert-butylmethylether as an eluent. After 50 l of
`elution, fractions of 6.2 l are collected. Fractions 11 to
`13 contain mainly FK506. Fractions 14 to 16 are col
`lected and brought to crystallization by dissolution in
`150 ml of ether and addition of 100 ml of hexane. The
`product is recrystallized from acetonitrile. The title
`compound (Compound No. l) is obtained. It has the
`following characteristics:
`M.P. 180°-l84° C. (dec.) (from methanol, ether or
`acetonitrile),
`_
`colorless crystals,
`[a]1)22= -4.0° (0:072 in methanol),
`elementary analysis:
`found C 65.6, H 8.7, N 1.8, 0 24.0%;
`calc. C 65.7, H 8.7, N 1.7, 0 23.9%.
`elementary formula: C44H69NO12 (804.0),
`mass spectrum FAB 804.5=(MH+),
`
`The pH value is set to 6.8 prior to sterilization, which
`takes place for 20 minutes at 120° C.
`The propagation of this preculture is effected for 96 20
`hours at 27° C. at 200 rpm on an agitator with an excen
`tricity of 50 mm.
`(C) Intermediate culture
`25
`Two 500 l aliquots of preculture medium are inocu
`lated in a 750 1 steel fermentor with 5 liters each of
`preculture and incubated for 48 hours at 27° C. Rotation
`speed is 100 rpm and aeration is 0.5 1 per minute per
`liter of medium.
`
`30
`
`768.5 (MI-I3°-36),
`576.3 (MI-I30 —228,
`100%
`UV-spectrum in methanoI: Amx=end absorption
`(MeOH),
`IR-spectrum in KBr: see FIG. 1,
`1I-I-NMR-spectrum in CDCL3, 360 MHz with tet
`ramethylsilane as internal standard: see FIG. 2.
`The structure of this compound has also been ana
`5 lyzed by X-ray diffraction analysis and compared with
`that for FK 506. The structure was re?ned to an R
`factor of 0.046 using 3200 observed reflections. The
`main insight gained thereby is that the conformation of
`the 2l-membered ring is stabilised by an intramolecular
`hydrogen bond (010 . . . 022) and is signi?cantly differ
`ent from the ring conformation found in the published
`crystal structure of FK 506.
`
`40
`
`(D) Main culture
`6000 l of main culture medium are inoculated in two
`4500 1 steel fermentors with 600 l of intermediate cul
`ture. The main culture medium has the following com
`position:
`
`Soluble starch
`Corn steep (Roquette)
`Yeast extract (Gisten)
`CaCO3
`demineralised water ad
`
`45.0 g
`10.0 g
`10.0 g
`1.0 g
`1000 ml
`
`The pH is set to 6.8 with NaOI-I prior to sterilization.
`The corn steep is presterilized for 20 minutes at 120° C
`Sterilization of the whole medium is effected at 120° C
`for 20 minutes.
`Incubation is effected for 96 hours at 27° C, 50 rpm,
`0.5 bar and an aeration rate of 0.5 1 per minute per liter
`of medium. Foam formation is reduced using a silicone
`antifoam agent.
`
`EXAMPLE 2
`
`50
`
`EXAMPLE 3
`18,14 a-Dihydroxy-l2-[2'-(4"(R)-hydroxy-341
`_ (R)-methoxycyclohex- l "(R)-yl)-1,-methyl-trans~vinyl]
`23a,25B-dimethoxy-l3a,19,2la27B-tetramethyl
`17Bl7B-propyl-l1,28-dioxa-4-azatricyclo[22.3.1.04"
`9]octacos- l 8-trans-ene-2,3, 10, l 6-tetraone
`[=C0mpound No. 2; “dihydro-l7-epi-FK506”]
`[Formula I: R1=OH; R2=n-propyl; single bond in
`14,15-position]
`[process variant c), hydrogenation]
`1.6 g of the Compound No. 1 is dissolved in 80 ml of
`ethanol, mixed with 80 mg of 10 % palladium on char
`coal and hydrogenated for 10 minutes at normal pres
`sure and room temperature. The catalyst is then filtered
`off, the filtrate evaporated to dryness, and the residue
`chromatographed with tert-butylmethylether on 180 g
`silicagel. The fractions are checked by high pressure
`liquid chromatography and the fractions containing the
`hydrogenation product are collected and crystallized
`from diethylether/hexane. The title compound (Com
`pound No. 2) is obtained. It has the following character
`istics:
`M.P 154-156° C. (dec.):
`[a]D2Z: — 19.1“ (c: 1.10 in methanol,
`
`55
`
`vinyl]-23a25B-dimethoxy-130.,19,2la,27B-tetrame-
`thyll 1,28-dioxa-4-azatricyclo
`[22.3.1.04'9]octacos-18-trans-ene-2,3,10,16-tetraone
`[32 Compound No. 1; “17-epi-FK506”
`[Formula I: R1=OH; R2=allyl; single bond in
`14,15-position
`[process variant (a), isolation ]
`6200 1 of fermentation medium are stirred for 6 hours
`at room temperature with 6000 1 of ethyl acetate and 65
`thereafter the two phases are separated in a separator.
`The ethyl acetate phase is evaporated to dryness under
`reduced pressure. The extract is then defatted by sepa
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`

`
`5,011,844
`8
`7
`As regards immunosuppressant activity, in the mixed
`Elementary analysis: found: C 65., H 9.0, N 1.8, 0 24.0
`lymphocyte reaction [T. Meo, Immunological Methods,
`%; calc. C 65.6, H 8.9, N 1.7, 0 23.8 %;
`Elementary formula: C44H7iNO12 (806.0),
`L. Lefkovits and B. Permis, Eds., Academic Press, NY.
`(1979) p. 227-239], they elicit suppression of mixed
`788.9
`Mass spectrum: FAB 806.9=(MI-I+)'
`lymphocytes at a dosage of from about 0.15 nM to about
`(MH+ — 18), 770.9 (MI-1+ — 36), 578.6 (MI-1+ —228),
`10 nM. They are further active at a concentration of
`100 %.
`UV-spectrum in methanol: Amax=end absorption
`from about 0.5 nM to about 10 nM in the test of the
`primary humoral immune response on sheep red blood
`(MeOH).
`cells in vitro (R.I. Mishell and R. W. Dutton, Science
`IR-spectrum in KBr: see FIG. 3.
`153 [l966]l004-l006; R. I. Mishell and R. W. Dutton, J.
`lI-l-NMR-spectrum in CDCl3, 360 MHz with tet
`Exp. Med. 126 [l967]423—442).
`ramethylsilane as internal standard: see FIG. 4.
`As regards anti-in?ammatory activity, in the oxazo
`lone allergy test (mouse) (described in EP 315978) the
`compounds elicit an activity between 20% and 70%
`upon topical administration at a concentration of 0.01
`%.
`The compounds of formula I are therefore useful as
`immunosuppressant and antiinflammatory agents in the
`prevention and treatment of conditions requiring immu
`nosuppression and of inflammatory conditions, such as
`(a) the prevention and treatment of
`resistance in situations of organ or tissue transplanta
`tion, e.g. of heart, kidney, liver, bone marrow and
`skin,
`graft-versus-host disease, such as foIIowin bone mar
`row grafts,
`autoimmune diseases such as rheumatoid arthritis,
`systemic Lupus erythematosus, Hashimoto’s
`thyroidis, multiple sclerosis, Myasthenia gravis,
`diabetes type I and uveitis,
`cutaneous manifestations of immunologically
`mediated illnesses, such as Alopecia areata, and
`(b) treatment of in?ammatory and hyperproliferative
`skin diseases, such as psoriasis, atopical dermatitis,
`contact dermatitis and further eczematous dermatit
`ises, seborrhoeic dermatitis, Lichen planus, Pemphi~
`gus, bullous Pemphigoid, Epidermolysis bullosa, urti
`caria, angioedemas, vasculitides, erythemas, cutane
`ous eosinophilias, Lupus erythematosus and acne.
`The compounds may be administered systemically or
`topically.
`For these indications the appropriate dosage will, of
`course, vary depending upon, for example, the host, the
`mode of administration and the nature and severity of
`the condition being treated. However, in general, satis
`factory results are indicated to be obtained systemically
`at daily dosages of from about 0.15 mg/kg to about 1.5
`mg/kg animal body weight. For the larger mammal an
`indicated daily dosage is in the range from about 0.01
`mg to about 100 mg of a compound of formula I, conve
`niently administered, for example, in divided doses up
`to four times a day.
`For topical use satisfactory results are obtained with
`local administration of a 1~3 % concentration of active
`substance several times daily, e.g. 2 to 5 times daily.
`Examples of indicated galenical forms are lotions, gels
`and creams.
`The compound of the invention may be administered
`by any conventional route, in particular enterally, e.g.
`orally, e.g. in the form of tabIets or capsules, or topi
`cally, e. g. in the form of lotions, gels or creams.
`Pharmaceutical compositions comprising a com
`pound of formula I as de?ned above in association with
`at least one pharmaceutical acceptable carrier or diluent
`may be manufactured in conventional manner by mix
`ing with a pharmaceutically acceptable carrier or dilu
`ent. Unit dosage forms contain, for example, from about
`0.0025 mg to about 50 mg of a compound of formula I.
`
`EXAMPLE 4
`l7B-Allyl- IB-hydroxy-12-[2'-(4”(R)-hydroxy-3"(R)
`methoxycyclohex- l "(R)-yl)- l '-methyl-trans-vinyl]
`230.,25/3-dimethoxy- 1 3a,
`19,2 1 a,27B-tetramethyl-1 l,28-dioxa~4-azatricyclo
`[22.3.1.04'9]soctacos-l4-trans,l 8-trans-diene-2,3,l0,l6
`tetraone
`Compound No. 3; “dehydro-l7-epi-FK506”]
`[Formula I: R1 missing; R2=allyl; double bond in
`l4,15-position]
`(a) synthetically [process variant (1)), dehydration] :
`1 g Compound No. l is dissolved in l l of ethyl ace
`tate and 10 ml IN HCl are added. Agitation is main
`tained for 5 days. Then the reaction mixture is neutral
`ized with 10 ml of IN NaOH and washed with 500 ml of
`water. The organic phase is dried over sodium sulfate
`and evaporated to dryness. The residue is subjected to
`chromatographic separation over silicagel H using
`methyl tert-butylether as an eluent. The fractions are
`checked by HPLC. The product is recrystallized from
`ether. The title compound (Compound No. 3) is ob
`tained. It has the following characteristics:
`M.P. l89°—191° C. (from ether).
`colourless crystals.
`[a]D22= 131.9” (c=0.84 in CHC13).
`elementary formula: C44H$7NO11 (786.0).
`UV-spectrum in methanol: km,‘ 230 log e’=l.2l15;
`323 log e'=0.2l38.
`retention time upon high pressure liquid chromatog
`raphy (HPLC)
`in gradient 1 (in 20 min from 50:50 to 10:90): 16.64
`mm.
`in gradient 2 (in 20 min from 90:10 to 10:90): 22.48
`min.
`HPLC system: column: Lichrosorb RP18 Merck
`(250X4 mm); ?ow rate: 2 ml/min; detection UV
`50
`220 nm/0.1;
`solvents: buffer triethylamine-phosphate pH 3.5 0.05
`M 10 % acetonitrile / acetonitrile
`(b) By fermentation (process variant a), isolation]
`After crystallization of FK506 from fractions 11 to 13
`(see Example 2) the supernatant is chromatographed
`over silicagel using hexane/methyl tert-butylether/me
`thanol 5:4:1 as an eluent. The fractions are checked by
`HPLC and the fraction having a retention time of 17.25
`min is rechromatographed over silicagel H with methyl
`tert-butyIether. Upon recrystallization from ether the
`title compound is obtained (M.P. l89°—l93° C.).
`The compounds of the invention possess pharmaco
`logical activity. They are, therefore, useful as pharma
`ceuticals.
`In particular, they possess immunosuppressant and
`anti-inflammatory activity.
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`5,011,844
`Topical administration is e.g. to the skin. A further
`form of topical administration is to the eye, for the
`treatment of immune-mediated conditions of the eye,
`such as: auto-immune diseases, e.g. uveitis, keratoplasty
`and chronic keratitis; allergic conditions, e.g. vernal 5
`conjunctivitis; in?ammatory conditions and corneal
`transplants, by the topical administration to the eye
`surface of a compound of formula I as de?ned above in
`a pharmaceutically acceptable ophthalmic vehicle.
`The ophthalmic vehicle is such that the compound of 10
`formula I is maintained in contact with the ocular sur
`face for a sufficient time period to allow the compound
`to penetrate the corneal and internal regions of the eye,
`e.g. the anterior chamber, posterior chamber, vitreous
`body, aqueous humor, vitreous humor, cornea, iris/cili
`ary, lens, choroid/retina and sclera.
`The pharmaceutically acceptable ophthalmic vehicle
`may be e.g. an ointment, vegetable oil, or an encapsulat
`ing material.
`Compound No. l is preferred for the above systemic 20
`and topical indications.
`I claim:
`1. A compound of formula I
`
`15
`
`or
`R1 is missing, R2 is allyl and
`there is a double bond between the carbon atoms
`numbered 14 and 15.
`4. A process for the preparation of the compound of
`formula I wherein
`R1 is hydroxy, R2 is n-propyl and
`there is a single bond between the carbon atoms num
`bered l4 and 15
`which comprises
`hydrogenating the corresponding compound of for
`mula I wherein R2 is allyl.
`5. A pharmaceutical composition containing a com
`pound according to claim 1 together with a pharmaceu
`tically acceptable carrier or diluent.
`6. A method for the prevention or treatment of condi
`tions requiring immunosuppression or of in?ammatory
`conditions, such as
`(a) the prevention and treatment of
`resistance in situations oi organ or tissue transplanta
`tion e.g. of heart, kidney, liver, bone marrow and
`skin,
`graft-versus-host disease, such as following bone mar
`row grafts,
`autoimmune diseases such as rheumatoid arthritis,
`systemic Lupus erythematosus, Hashimoto’s
`thyroidis, multiple sclerosis, Myasthenia gravis,
`diabetes type I and uveitis,
`cutaneous manifestations of immunologically
`mediated illnesses, such as Alopecia areata, and
`(b) the treatment of in?ammatory and hyperprolifera
`tive skin diseases, such as psoriasis, atopical dermati
`tis, contact dermatitis and further eczematous derma
`titises, seborrhoeic dermatitis, Lichen planus, Pem
`phigus, bullous Pemphigoid, Epidermolysis bullosa,
`urticaria, angioedemas, vasculitides, erythemas, cuta
`neous eosinophilias, Lupus erythematosus and acne,
`comprising administering, a therapeutically effective
`amount of a compound according to claim 1 together
`with a pharmaceutically acceptable carrier or diluent
`to a subject in need of such treatment.
`7. A method of treatment of immune-mediated condi
`tions of the eye, such as: auto-immune diseases, e.g.
`uveitis, keratoplasty or chronic keratitis; allergic condi
`tions, e. g. vernal conjunctivitis; in?ammatory condi
`45
`' tions or corneal transplants, which comprises topically
`administering to the eye surface a therapeutically effec
`tive amount of a compound according to claim 1 in a
`pharmaceutically acceptable ophthalmic vehicle.
`8. A process for the preparation of a compound of
`claim 1 wherein R1 is hydroxy or is missing and R2 is
`allyl, which comprises cultivating an appropriate Strep
`tomyces strain such as Streptomyces tsukubaensis No.
`9993 until a sufficient amount of the compound is pro
`duced, and isolating the compound from the resulting
`culture.
`9. A process for the preparation of a compound of
`claim 1 wherein R1 is missing and R2 is allyl, which
`comprises dehydrating the corresponding compound of
`claim 1 in which R1 is hydroxy and isolating the com
`pound.
`
`25
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`so
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`35
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`4O
`
`HO
`
`H C
`3 \o
`
`CH3
`
`‘:2
`5
`
`I
`
`0
`
`R2
`
`O
`
`I'I3C
`
`H3C-O
`
`wherein
`either
`R1 is hydroxy,
`R2 is allyl or n-propyl and
`there is a single bond between the carbon atoms num
`bered l4 and 15
`
`50
`
`R1 is missing,
`R2 is allyl and
`there is a double bond between the carbon atoms
`numbered 14 and 15.
`2. The compound according to claim 1 wherein R1 is
`hydroxy, R2 is allyl and there is a single bond between
`the carbon atoms numbered 14 and 15.
`3. The compounds according to claim 1 wherein
`either
`R1 is hydroxy, R1 is n-propyl and
`there is a single bond between the carbon atoms num
`bered l4 and 15
`
`*
`
`it
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`* t
`
`‘I
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`65
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`NOVARTIS EXHIBIT 2017
`Par v Novartis, IPR 2016-00084
`Page 10 of 10