United States Patent [191
`Failli et al.
`
`[54] RAPAMYCIN 42-SULFONATES AND
`42-(N-CARBOALKOXY) SULFAMATES
`USEFUL AS IMMUNOSUPPRESSIVE
`AGENTS
`
`[75]
`
`Inventors:
`
`Amedeo A. Failli, Princeton
`Junction, N.J.; Wenling Kao, Paoli;
`Robert J. Steffan, Langhorne, both
`of Pa.; Robert L. Vogel, Stratford,
`NJ.
`
`[73]
`
`Assignee:
`
`American Home Products
`Corporation, New York, NY.
`
`[21] Appl. No.: 846,637
`
`[22]
`[5 1]
`[52]
`[58]
`[56]
`
`Filed:
`
`Mar. 5, 1992
`
`Int. Cl.5 ................. .. A61K 31/395; CO7D 491/06
`US. Cl. .................................................. .. 540/456
`Field of ‘Search ....................... .. 546/456; 514/291
`
`References Cited
`U.S. PATENT DOCUMENTS
`
`3,929,992 12/1975 Seghal et a1. .... ............... .. 540/456
`5,091,389 2/1992 Ondekye et a1. .
`5,100,899 3/1992 Calne ................................. .. 540/456
`
`OTHER PUBLICATIONS
`Can. J. Physiol. Pharmacol. 55, 48(1977).
`
`Primary Examiner-Robert T. Bond
`Attorney, Agent, or Firm-—Walter Patton
`
`llllllllllllllIllIllllIllllllllllllllllllllllllIlllllllllllllllllllllllilll
`USOO5177203A
`[11] Patent Number:
`5,177,203
`[45] Date of Patent:
`Jan. 5, 1993
`
`ABSTRACT
`[57]
`A derivative of rapamycin of general formula (I)
`
`N
`
`°§ §o
`no
`
`0
`
`where R1 is alkyl, alkenyl, or alkynyl containing 1 to 6
`carbon atoms; or an aromatic moiety selected from the
`group consisting of phenyl and naphthyl or a heterocy
`clic moiety selected from the group consisting of thio
`phenyl and quinoliny] or NHCOzR2 wherein R2 is
`lower alkyl containing 1 to 6 carbon atoms or a pharma
`ceutically acceptable salt thereof, which by virtue of its
`immunosuppressive activity is useful in treating trans
`plantation rejection, host versus graft disease, autoim
`mune diseases, and diseases of in?ammation.
`
`10 Claims, No Drawings
`
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`1
`
`5,177,203
`
`RAPAMYCIN 42-SULFONATES AND
`42-(N-CARBOALKOXY) SULFAMATES USEFUL AS
`IMMUNOSUPPRESSIVE AGENTS
`
`20
`
`where R1 is alkyl, alkenyl, or alkynyl containing 1 to 6
`carbon atoms; or an aromatic moiety selected from the
`group consisting of phenyl and naphthyl or a heterocy
`clic moiety selected from the group consisting of thio
`phenyl and quinolinyl or NHCOZR2 wherein R2 is
`lower alkyl containing 1 to 6 carbon atoms or a pharma
`ceutically acceptable salt thereof.
`The rapamycin 42-sulfonates of this invention can be
`prepared by the standard literature procedure as out
`lined below.
`
`BACKGROUND OF THE INVENTION
`This invention relates to rapamycin 42-sulfonates and
`42-(N-carboalkoxy)sulfamates and a method for using
`them in the treatment of transplantation rejection, host
`versus graft disease, autoimmune diseases, diseases of
`in?ammation, and fungal infections.
`Rapamycin is a macrocyclic triene antibiotic pro
`duced by .S'treptomyces 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., J. Antibiot.
`28, 727 (1975); H. A. Baker et al., J. Antibiot. 31, 539
`(1978); U.S. Pat. Nos. 3,929,992; and U.S. Pat. No.
`3,993,749].
`..
`Rapamycin alone (U.S. Pat. No. 4,885,171) or in com
`bination with picibanil (U.S. Pat. No. 4,401,653) has
`been shown to have antitumor activity. R. Martel et al.
`[Can. J. Physiol. Pharmacol. 55, 48 (1976)] 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
`arthritis; and effectively inhibited the formation of IgE
`like antibodies.
`The immunosuppressive effects of rapamycin have
`been disclosed in FASEB 3, 3411 (1989). Rapamycin
`therefore is also useful in preventing transplant rejec
`tion [FASEB 3, 3411 (1989); FASEB 3, 5256 (1989); and
`R. Y. Calne et al., Lancet 1183 (1978)].
`Mono- and diacylated derivatives of rapamycin have
`been shown to be useful as antifungal agents (U .5. 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 esters described above would be at
`the 31- and 42-positions.
`DESCRIPTION OF THE INVENTION
`This invention relates to rapamycin 42-sulfonates and
`42-(N-carboalkoxy)sulfamates of general formula (I)
`
`The sulfonate formation between alcohol and sulfo
`nyl halide has been described [Jerry March, Advanced
`Organic Chemistry, 3rd edition, published in 1985, page
`444]. The speci?c reaction condition employed in this
`invention was developed by S. Rakhit of Ayerst Labo
`ratories and reported in U.S. Pat. No. 4,316,855 (Feb.
`23, 1982).
`The 42-(N-carboalkoxy)sulfamates of the present
`invention can also be prepared by reaction of rapamycin
`with an alkyl(carboxysulfamoyl)triethylammonium hy~
`droxide inner salt (Burgess Salts; see G. M. Atkins Jr.
`and E. M. Burgess, J. Am. Chem. Soc., 90, 4744, 1968;
`E. M. Burgess, H. R. Penton Jr. and E. A. Taylor, J.
`Org. Chem. 38, 26, 1978).
`
`45
`
`benzene, Li.
`
`a
`
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`

`
`3
`
`5,177,203
`
`-continued
`
`@ZOSOgNl-ICOgRZ
`
`""ocH;
`
`/ 0
`1|
`0
`
`N
`
`\
`0
`§ \0
`no
`
`0
`
`QCH;
`
`wherein R2 is as de?ned above.
`The pharmaceutically acceptable salts may be formed
`from inorganic cations such as sodium, potassium, and
`the like.
`The following examples illustrate the preparation of
`25
`representative compounds of this invention.
`
`30
`
`40
`
`EXAMPLE 1
`Rapamycin 42-ester with
`5-(dimethylamino)-l-naphthalenesulfonic acid
`A solution of 200 mg (0.22 mmol) of rapamycin in 2
`mL of pyridine was treated at 0° C. under anhydrous
`conditions with 840 mg (3.1 mmol) of dansyl chloride
`and stirred at room temperature for 24 hours. The reac
`35
`tion mixture was diluted at 0° C. with 30 mL of 2N HCl
`and extracted with ethyl acetate. The ethyl acetate
`extract was washed with brine, dried with MsSO4 and
`evaporated. The residue was chromatographed on silica
`gel. Elution with 25% ethyl acetate in benzene afforded
`150 mg of the title product as a light yellow powder,
`m.p. 101L104° c.
`IR: 3430 (OH), 1740 (sh), 1720 (both @O), 1650
`(amide C=O), 1450, 1355, 1170 (sulfonate), 1100, 985,
`960 cm-1.
`lH NMR (CDCI3, 400 MHz) 8 8.58 (d, 1H, H1), 8.32
`(d, 1H, H3), 8.25 (m, 1H, Hz), 7.53 (m, 2H, H5 and H6),
`7.19 (d, 1H, H4), 3.31, 3.13, 2.72 (all s, 3H, --O—
`
`45
`
`6H, -N
`
`CH3
`
`CH3
`
`CH3), 2.79 (s,
`
`50
`
`55
`
`65
`
`CH3
`
`CH3
`
`MS (neg. ion FAB) 1146 (M-), 912, 590, 250.
`EXAMPLE 2
`Rapamycin 42-ester with 4-methylbenzenesulfonic acid
`A solution of 6.0 g (31.6 mmol) p-toluenesulfonyl
`chloride in 25 rnL pyridine was added to a solution of
`10.0 g (10.9 mmol) rapamycin at 0° C. and the resulting
`solution was stirred at 20° C. for 22 hours. Cold 2N HC]
`(240 mL) was added and the product was extracted into
`ethyl acetate, washed with brine, dried over MgSO4
`and evaporated to a yellow solid. Chromatography on
`silica gel eluted with 20% ethyl acetate in methylene
`chloride afforded 5.3 g product as a white solid, mp.
`108°-1l6° C.
`IR(KBr): 3410, 2910, 1710, 1640, 1440, 1160 and 655
`cm-1.
`‘ NMR (CD013, 400 MHz): 6 7.80 (d, 2H, aromatic),
`732 (d, 2H, aromatic), 3.33 (s, 3H), 3.14 (s, 3H), 3.13 (s,
`3H), 2.44 (s, 3H).
`MS (neg. ion FAB): 1067 (M-), 590, 171, 155.
`EXAMPLE 3
`Rapamycin 42-ester with 2-thiophenesulfonic acid
`A solution of 0.18 g (0.2 mmol) rapamycin and 0.13 g
`(0.71 mmol) 2-thiophenesulfonyl chloride in 2 mL pyri
`dine was heated at 55° C. for 4 hours, then cooled to 20°
`C. and treated with 40 mL 1N HCl. The product was
`extracted into ethyl acetate, washed with brine, dried
`over MgSO4 and stripped of solvent. Chromatography
`on silica gel eluted with 20% ethyl acetate in methylene
`chloride afforded 40 mg title compound as a white solid,
`m.p. ll4°~119° C.
`,
`IR (KBr): 3420, 2915, 1712, 1644, 1440, 1365, 1170
`and 660 cm"1.
`~
`NMR (CDCI3, 400 MHz): 8 7.67 (1H, aromatic), 7.62
`(1H, aromatic), 7.07 (1H, aromatic), 3.29 (s, 3H, OCH3),
`3.14 (s, 3H, OCHg), 3.09 (s, 3H, OCT-I3).
`MS (neg. ion FAB): 1059 (M--), 912, 590, 163.
`EXAMPLE 4
`Rapamycin 42’ester with
`4-[[4-(dimethylamino)phenyl]aza]benzenesulfonic acid
`Dabsyl chloride (0.83 g, 2.57 mmol) was added to a
`solution of 0.54 g (0.59 mmol) rapamycin in 30 mL dry
`pyridine and the solution heated at 65°—70° C. for 24
`hours. Upon cooling, the reaction mixture was parti
`tioned between 200 mL 2N HCl and 50 mL ethyl ace
`
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`
`5
`tate. The product was dried over MgSO4, stripped of
`solvent and chromatographed on silica gel eluted with
`30% ethyl acetate in methylene chloride, to afford the
`title compound as a bright red solid, mp. 118°-133°
`IR (KBr): 3430, 2930, 1720, 1600, 1360, 1142, 683 and
`602 cm-1.
`NMR (CDCI3, 400 MHz): 8 8.00 (2H, aromatic), 7.93
`(4H, aromatic), 6.76 (2H, aromatic), 3.33 (s, 3H, OCHg),
`3.135 (s, 3H, OCH3), 3.126 (s, 3H, OCH3).
`MS (pos. FAB): 1223 (MNa+), 1169, 1137, 918, 306.
`EXAMPLE 5
`Rapamycin 42-ester with l-naphthalene sulfonic acid
`l-Naphthalenesulfonyl chloride (0.48 g, 0.2.1 mmol)
`was added to a solution of (0.54 g, 0.59 mmol) rapamy
`cin in 11 mlL pyridine and the resulting solution was
`stirred at 20° C. for 44 hours. Cold 2N HCl (75 mL) was
`added and the product was extracted into ethyl acetate,
`washed with brine, dried over MgSO4 and evaporated
`to a tan solid. Chromatography on slica gel eluted with
`20% ethyl acetate in methylene choride yielded 30 mg
`product as a white solid, mp. l10°-13l° C.
`IR (KBr): 3440, 2925, 1720, 1645, 1450, 1175 and 765
`cm—1.
`NMR (CDCI3, 400 MHz): 6 8.65 (1H), 8.26 (1H), 8.10
`(2H), 7.93 (1H), 7.70 (1H), 7.62-7.53 (complex, 2H),
`3.32 (s, 3H, OCH3), 3.13 (s, 3H, OCI-I3), 2.64 (s, 3H,
`OCH3).
`MS (neg. FAB): 1103 (M-), 912, 590.
`EXAMPLE 6
`Rapamycin 42-ester with S-quinolinesulfonic acid
`A solution of (0.30 g, 0.33 mmol) rapamycin and (0.29
`g, 1.28 mmol) S-quinolinesulfonyl chloride in 5 mL
`pyridine was stirred at 20° C. for 24 hours. The reaction
`mixture was partitioned between 2N HCl (10 mL) and
`ethyl acetate.
`The organic layer was washed with brine, dried over
`MgSO4, stripped of solvent and chromatographed on
`silica gel eluted with 30% ethyl acetate in methylene
`chloride, affording 130 mg of title compound as a white
`solid, mp l20°—165" C.
`IR (KBr): 3430, 2925, 1715, 1640, 1170, 985 and 785
`cm-1.
`NMR (CDCI3, 400 MHz): 6 9.18 (1H), 8.49 (1H), 8.25
`(1H), 8.09 (1H), 7.65 (1H), 7.55 (1H), 3.32 (s, 3H,
`OCH3), 3.13 (s, 3H, OCH3), 2.60 (s, 3H, OCT-13).
`MS (neg. FAB): 1104 (M_), 912, 590, 208.
`EXAMPLE 7
`Rapamycin 42-methanesulfonate, hemiethylacetate,
`hemihydrate
`Under an atmosphere of nitrogen, an ice cold solution
`of rapamycin (0.46 g, 0.5 mmol) and triethyl amine (0.14
`mL, 1.0 mmol) in 5 mL of dry CHiClz was treated
`dropwise with methanesulfonyl chloride (0.943 mL,
`0.55 mmol). The ice bath was removed and the solution
`stirred at ambient temperature for one hour. The reac
`tion mixture was diluted with CHgClz and washed suc
`cessively with H20 and brine. After drying (Na2SO4),
`the solvent was removed in vacuo to give a yellow
`foam. Puri?cation by ?ash chromatography (silica
`Merck 60, ethyl acetate-hexane 1:1) afforded the title
`compound (0.37 g, 75% white solid). '
`-'
`NMR (400 MHz, CDCl3): 8 1.65 (3H, CH3C=C ,
`1.74+1.75 (2s, 3H, CHgH), 3.06 (s, 3H, CH3SO2),
`
`50
`
`55
`
`65
`
`5,177,203
`6
`3.l3 (s, 3H, OCH3), 3.34 (s, 3H, OCH3), 3.4003 (5, 3H,
`OCH3).
`MS (neg. ion FAB, m/z): 991 (M)—, 590, 399.
`Anal. calc’d for C5ZH31NO15S+05 1120-1-05
`C4H3O2: C, 62.05; H, 8.29; N, 1.34; Found: C, 61.63; H,
`8.34; N, 1.49.
`
`5
`
`30
`
`45
`
`EXAMPLE 8
`Rapamycin 42-(2,2,2-tri?uoroethane sulfonate),
`dihydrate
`Under an atmosphere of nitrogen, a solution of rapa
`mycin (0.46 g, 0.5 mmol) and triethylamine (0.15 mL,
`1.1 mmol) in 2 mL of dry CHgClz was treated in one
`portion with 2,2,2-tri?uoroethane sulfonyl chloride
`(0.06 mL, 0.55 mmol). The solution was stirred over
`night at ambient temperature. The solvent was evapo
`rated in vacuo to give a yellow foam. The crude prod
`uct mixture was puri?ed by MPLC (silica Lichrosorb
`60, Merck 440‘37 mm, ethyl acetate-hexane 1:2, flow
`rate 20 mL/min) to give the title compound.
`NMR (400 MHz, CDC13): 8 1.65 (s, 3H, CH3C=C),
`1.749+1.752 (25, 31-1, CH3C=C), 3.14 (s, 3H, OCH3),
`3.34 (s, 3H, OCH3), 3.37 (s, 3H, OCH3).
`MS (neg. ion FAB, m/z): 1059 (M)-, 590, 560, 427,
`163.
`Anal. calc’d f0!‘ C52H31NO15S+2 H20: C, 58.02; H,
`7.72; N, 1.28; Found: C, 57.94; H, 7.96; N, 1.22.
`EXAMPLE 9
`42-O-[[(Methoxycarbonyl)]amino]sulfonyl]rapamycin
`Under anhydrous conditions, a solution of rapamycin
`(0.5 g, 0.55 mmol) and methyl(carboxysulfamoyl)trie
`thylammonium inner salt (0.25 g, 1.2 mmol, prepared as
`described by Burgess et al., J. Org. Chem. 38, 26, 1978)
`in 5 mL of benzene was stirred at ambient temperature
`overnight. The reaction mixture was then diluted with
`EtOAc (50 mL) and the solution was washed with
`water and brine and dried (Na2SO4). Removal of the
`solvent in vacuo yielded an off-white solid which was
`further puri?ed by MPLC (silica Merck 6O Lichroprep,
`440‘37 mm, ethyl acetate-hexane 2:1->methanol) to
`provide the title product as a yellow solid (0.247 g,
`43%).
`_ 1H NMR (CDC13, 400 MHz): 6 1.65 (s, 3H,
`CH3C=C), 1.78 (s, 3H, CH3C=C , 3.13 (m, 3H,
`CH3O), 3.39 (m, 6H, CHgO), 3.71 (s, 3H, CO2CH3).
`MS (neg. ion FAB, m/z): 1050 (M—H)—.
`The comitogen-induced thymocyte proliferation pro
`cedure (LAF) was used as an in vitro measure of the
`immunosuppressive effects of representative com
`pounds. Brie?y, cells from > the thymus of normal
`BALB/c mice were 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 vari
`ous concentrations of rapamycin, cyclosporin A, or test
`compound. Cells are harvested and incorporated; radio
`activity is determined. Inhibition of lymphoprolifera
`tion is assessed in percent change in counts per minute
`from non-drug treated controls. The results are ex
`pressed by the following ratio:
`
`3l-I-control thmus cells — PP-ramycin-treated thmus cells
`3l-l-control thymus cells - H3-test compound-treated cells
`
`A mixed lymphocyte reaction (MLR) occurs when
`lymphoid cells from genetically distinct animals are
`
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`5,177,203
`7
`8
`combined in tissue culture. Each stimulates the other to
`suppressive agent, the substantial increase in survival
`undergo blast transformation which results in increased
`time of the skin grant when treated with the compounds
`DNA synthesis that can be quanti?ed by the incorpora
`of the present invention further demonstrate their utility
`tion of tritiated thymidine. Since stimulating a MLR is a
`as immunosuppressive agents.
`function of disparity at Major Histocompatibility anti
`Based on the results of these standard pharmacologi
`gens, an in vivo popliteal lymph node (PLN) test proce
`cal test procedures, the compounds of this invention are
`dure closely correlates to host vs. graft disease. Brie?y,
`useful in the prevention and treatment of transplant
`irradiated spleen cells from BALB/c donors are in
`rejection such as heart, kidney, liver, bone marrow, and
`jected into the right hind foot pad of recipient C3H
`skin transplants; graft versus host disease; autoimmune
`mice. The drug is given daily, p.o. from Day 0 to Day
`and proliferative diseases such as, systemic lupus ery
`4. On Day 3 and Day 4, tritiated thymidine is given i.p.,
`thematosus, rheumatoid arthritis, type 1 diabetes, multi
`b.i.d. On Day 5, the hind popliteal lymph nodes are
`ple sclerosis, glomerular nephritis, Hashimoto’s thy
`removed and dissolved, and radioactivity counted. The
`roiditis, myastenia gravis, uveitis and psoriasis; diseases
`corresponding left PLN serves as the control for the
`of in?ammation such as dermatitis, eczema,iseborrhea
`PLN from the injected hind foot. Percent suppression is
`and in?ammatory bowel disease; and fungal infections.
`calculated using the non-drug treated animals as allo
`The compounds may be administered neat or with a
`genic control. Rapamycin at a dose of 6 mg/kg, p.o.
`pharmaceutical carrier to a mammal in need thereof.
`gave 86% suppression, whereas cyclosporin A at the
`The pharmaceutical carrier may be solid or liquid.
`same dose gave 43% suppression. Results are expressed
`A solid carrier can include one or more substances
`by the following ratio:
`which may also act as ?avoring agents, lubricants, solu
`bilizers, suspending agents, ?llers, glidants, compression
`aids, binders or tablet-disintegrating agents; it can also
`be an encapsulating material. In powders, the carrier is
`a ?nely divided solid which is in admixture with the
`?nely 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
`lose, sodium carboxymethyl cellulose, polyvinylpyr
`rolidine, low melting waxes and ion exchange resins.
`Liquid carriers are used in preparing solutions, sus
`pensions, emulsions, syrups, elixirs and pressurized
`compositions. The active ingredient can be dissolved or
`suspended in a pharmaceutically acceptable liquid car
`rier such as water, an organic solvent, a mixture of both
`or pharmaceutically acceptable oils or fats. The liquid
`carrier can contain other suitable pharmaceutical addi
`tives such as solubilizers, emulsi?ers, buffers, preserva
`tives, sweeteners, ?avoring agents, suspending agents,
`thickening agents, colors, viscosity regulators, stabiliz
`ers or osmo-regulators. Suitable examples of liquid car
`riers for oral and parenteral administration include
`water (partially containing additives as above, e.g. cel
`lulose derivatives, preferably sodium carboxymethyl
`cellulose solution), alcohols (including monohydric
`alcohols and polyhydric alcohols, e. g. glycols) and their
`derivatives, and oils (e.g. fractionated coconut oil and
`arachis oil). For parenteral administration, the carrier
`can also be an oily ester such as ethyl oleate and isopro
`pyl myristate. Sterile liquid carriers are useful in sterile
`liquid form compositions for parenteral administration.
`The liquid carrier for pressurized compositions can be
`halogenated hydrocarbon or other. pharmaceutically
`acceptable propellent.
`Liquid pharmaceutical compositions which are sterile
`solutions or suspensions can be utilized by, for example,
`. intramuscular intraperitoneal or subcutaneous injection.
`Sterile solutions can also be administered intravenously.
`The compound can also be administered orally either in
`liquid or solid composition form.
`Preferably, the pharmaceutical composition is in unit
`dosage form, e.g., as tablets or capsules. In such form,
`the composition is sub-divided in unit dose containing
`
`The second in vivo test procedure is designed to
`determine the survival time of pinch skin graft from
`male DBA/Z donors transplanted to male BALB/c
`recipients. The method is adapted from Billingham R.
`30
`E. and Medawar P. B., J. Exp. Biol. 28:385-402 (1951).
`Brie?y, a pinch skin graft from the donor is grafted on
`the dorsum of the recipient as a homograft, and an
`autograft is used as control in the same region. The
`recipients are treated with either varying concentra
`35
`tions of cyclosporin A as test control or the test com
`pound, intraperitoneally. Untreated recipients serve as
`rejection control. The graft is monitored daily and ob
`servations are recorded until the graft becomes dry and
`forms a blackened scab. This is considered as the rejec
`tion day. The mean graft survival time (number of days
`iSD.) of the drug treatment group is compared with
`the control group.
`
`BIOLOGICAL DATA
`The following table summarizes the results of repre
`sentative compounds of this invention in these three
`standard test procedures.
`TABLE 1
`Biological Activity
`LAF Assay
`PLN
`(R/A ratio)
`(R/A ratio)
`0.26
`-—
`0.21
`—
`0.23
`1.23 (i.p.)
`0.03
`-—
`0.19
`0.92 (i.p.)
`1.32
`0.08 (i.p.)
`1.70
`0.36 (i.p.)
`0.85
`0.83 (i.p.)
`0.01
`0.93 (i.p.)
`
`Skin Graft Model
`(days + SD)
`8.0 i 0.9
`8.7 :L 1.2
`9.3 i 0.8
`—
`9.5 i 0.3
`10.7 i 2.1
`9.83 i 0.98
`10.0 i 1.4
`10.33 i 0.24
`
`The results of these standard pharmacological test
`procedures demonstrate high immunosuppressive activ
`ity both in vitro and in vivo for the compounds of the
`present invention. A positive ratio in the LAF and PLN
`65
`test procedures indicates suppression of T-cell prolifera
`tion. As transplanted pinch skin grafts are typically
`rejected within 6-7 days without the use of an immuno
`
`3H-PLN cells control C311 mouse —
`3H-PLN cells rapamvcin-treated C311’ mouse
`H-PLN cells control (3311 mouse —
`3H-PLN cells test compound-treated C3H mouse
`
`20
`
`25
`
`Example
`1
`2
`3
`4
`5
`6
`7
`8
`9
`
`45
`
`50
`
`55
`
`60
`
`NOVARTIS EXHIBIT 2119
`Par v Novartis, IPR 2016-00084
`Page 5 of 6
`
`

`
`5,177,203
`10
`where R1 is alkyl, alkenyl, or alkynyl containing 1 to 6
`appropriate quantities of the active ingredient; the unit
`carbon atoms; or an aromatic moiety selected from the
`dosage forms can be packaged compositions, for exam
`group consisting of phenyl and naphthyl or a heterocy
`ple, packeted powders, vials, ampoules, pre?lled syrin
`clic moiety selected from the group consisting of thio
`ges or sachets containing liquids. The unit dosage form
`phenyl and quinolinyl or NHCO1R2 wherein R2 is
`can be, for example, a capsule or tablet itself, or it can be
`lower alkyl containing 1 to 6 carbon atoms or a pharma
`the appropriate number of any such compositions in
`ceutically acceptable salt thereof.
`package form. The dosage to be used in the treatment
`2. A compound of claim 1 wherein R1 is S-(dime
`must be subjectively determined by the attending physi
`thylamino)-l-naphthyl or a pharmaceutically accept
`able salt thereof.
`3. A compound of claim 1 wherein R1 is 4-methylphe
`nyl or a pharmaceutically acceptable salt thereof.
`4. A compound of claim 1 wherein R1 is Z-thiophenyl
`or a pharmaceutically acceptable salt thereof.
`5. A compound of claim 1 ‘wherein R1 is 4-[[4-(dime
`thylamino)phenyl]aza]phenyl or a pharmaceutically
`acceptable salt thereof.
`6. A compound of claim 1 wherein R1 is l-naphthyl
`or a pharmaceutically acceptable salt thereof.
`7. A compound of claim 1 wherein R1 is 8-quinolinyl
`or a pharmaceutically acceptable salt thereof.
`8. A compound of claim 1 wherein R1 is methyl or a
`pharmaceutically acceptable salt thereof.
`9. A compound of claim 1 wherein R1 is 2,2,2-tri
`fluoroethyl or a pharmaceutically acceptable salt
`thereof.
`10. A compound of claim 1 wherein R1 is [methox
`ycarbonyl1amino or a pharmaceutically acceptable salt
`thereof.
`
`cian.
`
`.
`
`What is claimed is:
`1. A compound of formula (I)
`
`N
`
`\
`
`0% \o
`no
`
`0
`
`0502-111
`
`(1)
`
`15
`
`20
`
`25
`
`* t i t t
`
`30
`
`35
`
`45
`
`50
`
`55
`
`65
`
`NOVARTIS EXHIBIT 2119
`Par v Novartis, IPR 2016-00084
`Page 6 of 6