United States Patent [191
`K20 et a1.
`
`[54] CARBAMATES OF RAPAMYCIN
`
`[75] Inventors: Wenling Kao, Paoli. Pa.; Robert L.
`Vogel, Stratford. N..l.; John H.
`Musser, Alameda, Calif.
`
`[73] Assignee:
`
`American Home Products
`Corporation, New York, N.Y.
`
`[211 Appl. No.: 686,728
`
`[22] Filed:
`
`Apr. 17, 1991
`
`[51] Int. Cl.5 ................. .. A61K 31/395; C07D 491/06
`[52] US. Cl. .................................. .. 514/183; 514/321;
`514/63: 540/452: 540/456
`[58] Field of Search .............. .. 540/456. 452; 514/183.
`514/321, 63
`
`[56]
`
`References Cited
`U.S. PATENT DOCUMENTS
`
`3.929.992 12/1975 Seghal et a1. ..................... .. 424/122
`3.993.749 11/1976 Seghal et a1. ......... ..
`424/122
`4.316.885 2/1982 Rakhit ........ ..
`424/122
`4.401.653 8/1983 Eng ........ ..
`424/114
`4.650.803 3/1987 Stella et a1.
`546/90
`4.885.171 12/1989 Surendra et a1. ................. .. 424/122
`
`OTHER PUBLICATIONS
`
`J. Antibiot. 28. 721~726 (1975).
`J. Antibiot. 28, 727—732 (1975).
`J: Antibiot. 31. 539~545 (1978).
`Can. J. Physiol. Pharmacol. 55.48 (1977).
`FASEB 3.3411 (1989).
`FASEB 3.5256 (1989).
`Lancet 1183 (1978).
`Med. Sci. Res. 17-. 877 (1989).
`
`Primary Exam1'ner-—Robert T. Bond
`Attorney, Agent, or Firm—Arnold S. Milowsky
`
`ABSTRACT
`[5 7]
`A compound of the structure
`
`llllllllllllllllllllllIllllllllllllllllllllllllllllllllllllllllllllllllllll
`5,118,678
`Jun. 2, 1992
`
`USOO5118678A
`Patent Number:
`[11]
`[45] Date of Patent:
`
`\ 0M6
`
`0
`
`(j
`
`o
`
`Y
`O§ E0
`HO
`
`0
`
`wherein
`R1 and R2 are each. independently, hydrogen or
`—CONH(CR3R4)H—X:
`R3 and R4 are each. independently. hydrogen. alkyl,
`aralkyl, cycloalkyl. halogen. or trifluoromethyl;
`X ix hydrogen. lower alkyl. cycloalkyl. tri?uoro
`methyl. nitro, alkoxy, carboalkoxy. aralkyl. halo,
`dialkylamino. thioalkyl. or Y;
`Y is a phenyl group which may be optionally mono-.
`di-, or tri- substituted with a group selected from
`alkyl. aralkyl, alkoxy, cyano. halo. nitro, carbalk
`oxy. tri?uoromethyl, dialkylamino, or alkylthio:
`
`with the proviso that R1 and R2 are not both hydro
`gen and when n=0, X is lower alkyl, cycloalkyl,
`aralkyl. or Y;
`or a pharmaceutically acceptable salt thereof when X is
`dialkylamino, which by virtue ofits immunosuppressive
`activity is useful in treating transplantation rejection,
`host vs. graft disease, autoimmune diseases and diseases
`of in?ammation; by virtue of its antitumor activity is
`useful in treating solid tumors; and by virtue of its anti
`fungal activity is useful in treating fungal infections.
`
`23 Claims, No Drawings
`
`NOVARTIS EXHIBIT 2130
`Par v Novartis, IPR 2016-00084
`Page 1 of 7
`
`

`
`BACKGROUND OF THE INVENTION
`This invention relates to carbamates of rapamycin 5
`and a method for using them in the treatment of trans
`plantation rejection, host vs. graft disease. autoimmune
`diseases, diseases of in?ammation. solid tumors, and
`fungal infections.
`Rapamycin is a macrocyclic triene antibiotic pro
`duced by Streptomyces hygroscopicus, which was found
`to have antifungal activity, particularly against Candida
`albt'cans, 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. No. 3,929,992; and US. Pat. No.
`3,993,749].
`Rapamycin alone (U.S. Pat. No. 4,885,171) or in com
`bination with picibanil (US. Pat. No. 4,401,653) has
`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
`arthritis; and effectively inhibited the formation of IgE
`like antibodies.
`The immunosuppressive effects of rapamycin have
`been disclosed in FASEB 3, 34l1 (1989). Cyclosporin A
`and FK-506, other macrocyclic molecules. also have
`been shown to be effective as immunosuppressive
`agents, therefore 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 (es
`teri?ed at the 28 and 43 positions) have been shown to
`be useful as antifungal agents (U.S. Pat. No. 4,316.885)
`and used to make water soluble prodrugs of rapamycin
`(US. 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; posi
`tions.
`
`25
`
`with the proviso that R1 and R2 are not both hydro
`gen and when n=0, X is lower alkyl of 1-6 carbon
`atoms, cycloalkyl of 3-8 carbon atoms, aralkyl of
`7-10 carbon atoms, or Y;
`or a pharmaceutically acceptable salt thereof when X is
`dialkylamino of 1-6 carbon atoms per alkyl group.
`The pharmaceutically acceptable salts are those de
`rived from such organic and inorganic acids as: acetic,
`lactic, citric, tartaric, succinic, maleic, malonic, glu
`conic, hydrochloric. hydrobromic, phosphoric, nitric,
`sulfuric, methanesulfonic, and similarly known accept
`able acids.
`Of these compounds, preferred members are those in
`which R2 is hydrogen; those in which R1 is hydrogen;
`those in which n is O and X is Y; those in which R2 is
`hydrogen. n is O. and X is Y; and those in which n is 0,
`X is Y, and Y is phenyl, 4-fluorophenyl, 2,4
`difluorophenyl. 4-nitrophenyl, or 4-methylphenyl.
`The compounds of this invention carbamylated at the
`42-position or at both the 31- and 42~positions can be
`prepared by reacting rapamycin with an isocyanate
`having the general structure
`
`1
`
`CARBAMATES or RAPAMYCIN
`
`2
`
`5,118,678
`wherein
`R1 and R2 are each. independently. hydrogen or
`-CONH(CR3R4),,—X;
`R3 and R4 are each. independently. hydrogen. alkyl of
`l-6 carbon atoms. aralkyl of 7-10 carbon atoms.
`cycloalkyl of 3-8 carbon atoms. halogen, or tri?uo
`romethyl:
`X is hydrogen, lower alkyl of 1-6 carbon atoms,
`cycloalkyl of 3-8 carbon atoms, trifluoromethyl,
`nitro, alkoxy of l-6 carbon atoms, carboalkoxy of
`2-7 carbon atoms, aralkyl of 7-10 carbon atoms,
`halo, dialkylamino of 1-6 carbon atoms per alkyl
`group. thioalkyl of l-6 carbon atoms, or Y;
`Y is a phenyl group which may be optionally mono-,
`di-, or tri-substituted with a group selected from
`alkyl of 1-6 carbon atoms, aralkyl of 7-10 carbon
`atoms, alkoxy of 1-6 carbon atoms, cyano, halo,
`nitro, carbalkoxy of 2-7 carbon atoms, trifluoro
`methyl, dialkylamino of 1-6 carbon atoms per alkyl
`group, or alkylthio of 1-6 carbon atoms;
`
`20
`
`DESCRIPTION OF THE INVENTION
`This invention provides derivatives of rapamycin
`which are useful as immunosuppressive. anti-in?amma
`tory, antifungal, and antitumor agents having the struc
`ture
`
`(,3
`§ YO
`
`HO
`
`in the presence of a base, such as pyridine.
`The 31-carbamylated compounds of this invention
`can be prepared by protecting the 42-alcohol of rapa
`mycin with a protecting group, such as with a tert-butyl
`dimethylsilyl group. followed by carbamylation of the
`31-position with an isocyanate with the general struc
`ture shown above. Removal of the protecting group
`provides the 31-carbamy1ated compounds. In the case
`of the tert-butyl dimethylsilyl protecting group. depro
`tection can be accomplished under mildly acidic condi
`tions.
`Having the 3l-position carbamylated and the 42-posi
`tion deprotected, the 42-position can be reacted with a
`different isocyanate than was reacted with the 31
`alcohol, to give compounds having different carba
`mates at the 31- and 42-positions. Alternatively, the
`42-carbamylated compounds, prepared as described
`above, can be reacted with a different isocyanate to
`provide compounds having different carbamates at the
`31- and 42-positions.
`
`55
`
`60
`
`65
`
`NOVARTIS EXHIBIT 2130
`Par v Novartis, IPR 2016-00084
`Page 2 of 7
`
`

`
`5,118,678
`4
`3
`rejection control. The graft is monitored daily and ob
`The isocyanates used to prepare the compounds of
`servations are recorded until the graft becomes dry and
`the invention are commercially available or can be pre
`pared by methods that are disclosed in the literature.
`forms a blackened scab. This is considered as the rejec
`lmmunosuppressive activity was evaluated in an in
`tion day. The mean graft survival time (number of days
`vitro standard pharmacological test procedure to mea
`15B.) of the drug treatment group is compared with
`sure lymphocyte proliferation (LAF) and in two in vivo
`the control group.
`standard pharmacological test procedures. The ?rst in
`The following table summarizes the results of repre
`vivo procedure was a popliteal lymph node (PLN) test
`sentative compounds of this invention in these three
`procedure which measured the effect of compounds of
`standard test procedures.
`this invention on a mixed lymphocyte reaction and the
`TABLE 1
`second in vivo procedure evaluated the survival time of
`a pinch skin graft.
`LAF
`PLN
`lCgMnM)
`(ratio)
`The comitogen-induced thymocyte proliferation pro
`cedure (LAF) was used as an in vitro measure of the
`immunosuppressive effects of representative com
`pounds. Briefly, cells from the thymus of normal
`BALB/c mice are cultured for 72 hours with FHA and
`lL-l and pulsed with tritiated thymidine during the last
`six hours. Cells are cultured with and without various
`concentrations of rapamycin. cyclosporin A, or test
`compound. Cells are harvested and incorporated radio
`activity is determined. Inhibition of lymphoprolifera
`tion is assessed as percent change in counts per minute
`from non-drug treated controls. The results are ex
`pressed as an IC50.
`
`10
`
`25
`
`Compound
`
`Example I
`Example 2
`Example 3
`Example 4
`Example 5
`Example 6
`Example 7
`Example 8
`Example 9
`Example 10
`Example ll
`Example 12
`Example 14
`Example 15
`Rapamycin
`
`8.6
`3.9
`> 1000
`57.9
`3% * ‘
`10.4
`+
`7.4
`8.7
`>l0O0
`> 1000
`> l000
`>l0OO
`>l0OO
`3.2-9.4
`
`0.71
`0.47
`+
`0.90
`+
`0.25
`-,>
`+
`0.72
`-
`4-
`#-
`-»
`-—
`1.00
`
`Skin Graft
`(days + SD)
`
`8.5 i 1.6
`8.5 t 1.2
`4
`+
`+
`8.3 i 1.0
`+
`8.8 : .3
`8.0 + l 7
`
`-
`-
`+
`-'
`17.0 : 1.7
`
`3l-l-control thymus cells — H3-rapamvcin-treated thvmus cells
`3H-control thymus cells — Hiltest compound~treated cells
`
`- Not evaluated.
`’ ‘ Percent inhibition at 100 nM.
`
`30
`
`A mixed lymphocyte reaction (MLR) occurs when
`lymphoid cells from genetically distinct animals are
`combined in tissue culture. Each stimulates the other to
`undergo blast transformation which results in increased
`DNA synthesis that can be quanti?ed by the incorpora
`tion oftritiated thymidine. Since stimulating a MLR is a
`function of disparity at Major Histocompatibility anti
`gens. an in vivo popliteal lymph node (PLN) test proce
`dure closely correlates to host vs. graft disease. Brie?y,
`irradiated spleen cells from BALB/c donors are in
`40
`jected into the right hind foot pad of recipient C3H
`mice. The drug is given daily. p.o. from Day 0 to Day
`4. On Day 3 and Day 4, tritiated thymidine is given i.p.,
`b.i.d. On Day 5, the hind popliteal lymph nodes are
`removed and dissolved, and radioactivity counted. The
`corresponding left PLN serves as the control for the
`45
`PLN from the injected hind foot. Percent suppression is
`calculated using the non-drug treated animals as allo
`genic control. Rapamycin at a dose of 6 mg/kg. p.o.
`gave 86% suppression, whereas cyclosporin A at the
`same dose gave 43% suppression. Results are expressed
`by the following ratio:
`
`50
`
`3H-PLN cells control C3l-l mouse —
`3H-PLN cells rapamycin-treated C3H mouse
`3H-PLN cells control C3H mouse —
`3H-PLN cells test compound-treated C3H mouse
`
`55
`
`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
`60
`recipients. The method is adapted from Billingham R.
`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
`65
`recipients are treated with either varying concentra
`tions of cyclosporin A as test control or the test com
`pound, intraperitoneally. Untreated recipients serve as
`
`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 and PLN test
`procedures indicate suppression of T cell proliferation.
`As a transplanted pinch skin grafts are typically re
`jected within 6—7 days without the use of an immuno
`suppressive agent, the increased survival time of the
`skin graft when treated with the compounds of this
`invention further demonstrates their utility as immuno
`suppressive agents.
`Because the compounds ofthis invention are structur
`ally similar to rapamycin and have a similar activity
`pro?le to rapamycin, the compounds of this invention
`also are considered to have antitumor and antifungal
`activities.
`Based on the results of these standard pharmacologi
`cal test procedures. the compounds are useful in the
`treatment of transplantation rejection such as, heart,
`kidney, liver. bone marrow, and skin transplants; auto
`immune diseases such as, lupus, rheumatoid arthritis,
`diabetes mellitus, myasthenia gravis, and multiple scle
`rosis; and diseases of in?ammation such as, psoriasis,
`dermatitis, eczema. seborrhea, in?ammatory bowel
`disease, and eye uveitis; solid tumors; and fungal infec
`tions.
`The compounds may be administered neat or with a
`pharmaceutical carrier to a mammal in need thereof.
`The pharmaceutical carrier may be solid or liquid.
`A solid carrier can include one or more substances
`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
`
`NOVARTIS EXHIBIT 2130
`Par v Novartis, IPR 2016-00084
`Page 3 of 7
`
`

`
`5,118,678
`6
`Elution with 30% ethyl acetate in benzene afforded 15
`mg of the title compound as a white solid, mp 99°—l03°.
`IR=KBr max 3440 (OH). 1740 (C20), 1625 (amide
`C—_—0), 1460, 1855, 1200, 1070, 990 cm—1. 1H NMR
`(CDCl3, 400 MHz). 57.38 (t, 2H. aromatic protons,
`ortho to ?uorine), 7.00 (t, 2H. aromatic protons meta to
`?uorine), 3.39 (s. 3H. CH3O). 3.33 (s. 3H. C1330); 3.14
`(s, 3H CH3O). MS (neg. ion FAB) 1050 (M—), 590,458,
`167.
`
`0
`
`5
`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 injec
`tion. Sterile solutions can also be administered intrave
`nously. 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
`appropriate quantities of the active ingredient; the unit
`dosage forms can be packaged compositions, for exam
`ple, packeted powders. vials, ampoules, pre?lled syrin
`ges or sachets containing liquids. The unit dosage form
`can be, for example, a capsule or tablet itself, or it can be
`the appropriate number of any such compositions in
`package form. The dosage to be used in the treatment
`must be subjectively determined by the attending physi
`cian.
`In addition, the compounds of this invention may be
`employed as a solution, cream, or lotion by formulation
`with pharmaceutically acceptable vehicles containing
`0.1-5 percent, preferably 2%, of active compound
`which may be administered to a fungally affected area.
`The following examples illustrate the preparation of 55
`representative compounds of this invention.
`
`20
`
`25
`
`30
`
`40
`
`45
`
`50
`
`EXAMPLE 2
`Rapamycin 42-ester with phenylcarbamic acid
`A solution of 100 mg of rapamycin in 0.8 ml of pyri
`dine was treated at 0° under anhydrous conditions with
`60 mg of phenyl isocyanate in 0.5 ml of pyridine. After
`stirring at 0° anhydrous for 5 hours, the reaction mix
`ture was diluted with 20 ml of 2N HCl at 0° and ex
`tracted with ethyl acetate. The ethyl acetate extract was
`washed with brine, dried with magnesium sulfate and
`evaporated. The residue was chromatographed on silica
`gel. Elution with 25% ethyl acetate in benzene afforded
`65 mg of the title compound as a white solid, mp
`106°-109".
`IR=KBr max 3400 (OH), 1725 (CzO), 1645 (amide
`CIO), 1600 (aromatic) 1540, 1440, 1315, 1080, 990 and
`750 (aromatic). 1H NMR (CDCI3, 400 MHz), 57.40 (t,
`2H, aromatic. ortho to NH), 7.32 (t, 2H, aromatic, 3.34
`(s, 3H. CH3O), 3.14 (s, 3H, CH3O). MS (neg. ion FAB)
`1032 (M—), 590, 321, 167.
`EXAMPLE 3
`Rapamycin-31.42-diester with
`[4-(trifluoromethyl)phenyl]carbamic acid
`A solution of 0.45 g 4-(tri?uoromethyl)phenylisocya
`mate in 5 mL dry pyridine was added to a solution of
`1.00 g rapamycin in 10 mL dry pyridine at 0° C., and the
`resulting solution was stirred at 0°—5° C. for 3.5 hours.
`Cold 2N HCl (75 mL) was added and the product was
`extracted into ethyl acetate, which was washed with
`brine, dried over magnesium sulfate and evaporated to
`dryness. The residue was chromatographed on silica
`gel. Elution with 10% ethyl acetate in methylene chlo
`ride afforded 0.63 g of 31,42-diester as a white solid,
`m.p. 147°—195‘ C. 1R(KBr): 3400 (OH). 1730 (CIO),
`1615, 1530, 1320. 1210, 1110, 1060 and 840 cm—1. 1H
`NMR (CDCl3, 400 MHz): 7.46-7.57 (complex, 8H,
`aromatic), 3.38 (s, 3H, OCH3), 3.36 (s, 3H. OCH3), 3.16
`(s, 3H, OCH3), MS (neg ion FAB): 1287 (M-), 1081,
`894, 590, 321.
`The following representative compounds can be pre
`pared from rapamycin and the appropriate isocyanate
`by employing the method used to prepare the title com
`pound in Example 3.
`Rapamycin-31,42-diester with (phehylmethyDcarbamic
`
`acid
`
`'
`
`with 2-nitroethylcarbamic
`
`EXAMPLE 1
`Rapamycin 42-ester with (4-fluorophenyl)carbamic
`acid
`-
`A solution of 70 mg of rapamycin in 1 mL of pyridine
`was treated at 0° under anhydrous conditions with 50
`mg of para?uorophenyl isocyanate in 1.5 ml of pyridine.
`After stirring at 0° for 5 hours, the reaction mixture was
`diluted with 20 ml of 2N HCl at 0° and extracted with
`ethyl acetate. The ethyl acetate extract was washed
`with brine, dried with magnesium sulfate and evapo
`rated. The residue was chromatographed on silica gel.
`
`65
`
`Rapamycin-31,42-diester with cyclohexylcarbamic acid
`' Rapamycin-31,42-diester with methylcarbamic acid
`Rapamycin-31,42-diester with 2-propylcarbamic acid
`60
`Rapamycin-31,42-diester with tri?uoromethylcarbamic
`acid
`Rapamycin-31,42-diester
`acid
`Rapamycin-31,42-diester with [4-(N,N-dimethyl)butyl]
`carbamic acid
`Rapamycin-31,42-diester
`thyl]carbamic acid
`
`with [2-(carbomethoxy)e
`
`NOVARTIS EXHIBIT 2130
`Par v Novartis, IPR 2016-00084
`Page 4 of 7
`
`

`
`5,118,678
`7
`Rapamycin-3 1 ,42-diester with [4-(methylthio)butyl]car
`bamic acid
`with
`Rapamycin-3l,42-diester
`phenyl)butylcarbamic acid
`
`[3-ethyl-4-(4-nitro
`
`EXAMPLE 4
`Rapamycin-42-ester with
`[4-(trifluoromethyl)phenyl]carbamic acid
`Continued elution of the residue described in Exam
`ple 3 with 30% ethyl acetate in methylene chloride
`afforded 0.20 g 42-monoester as a white solid, mp
`144°-l72° IR (KBr): 3450 (OH), 1730 (C20), 1650,
`1615, 1535, 1445, 1315, 1210, 1110, 1050, 980 and 835
`cm~1. 1H NMR (CDC13, 400 MHz): 67.55 (d, 2H, aro
`matic), 7.50 (d, 2H, aromatic), 3.39 (s, 3H, OCI-I3), 3.34
`(s, 3H, OCH3), 3.14 (s, 3H, OCH3). MS (neg ion FAB):
`1100 (M—), 590,321, 160.
`The following representative compounds can be pre
`pared from rapamycin and the appropriate isocyanate
`by employing the method used to prepare the title com
`pound in Example 4.
`Rapamycin-42-ester with (phenylmethyl)carbamic acid
`Rapamycin-42-ester with cyclohexylcarbamic acid
`Rapamycin~42-ester with methylcarbamic acid
`Rapamycin-42-ester with 2-propylcarbamic acid
`Rapamycin-42-ester with tri?uoromethylcarbamic acid
`Rapamycin-42-ester with 2-nitroethylcarbamic acid
`Rapamycin-42-ester with [4-(N,N-dimethyl)butyl]car
`bamic acid
`-
`Rapamycin-42-ester with [2-(carbomethoxy)ethyl]car
`bamic acid
`Rapamycin-42-ester with
`bamic acid
`Rapamycin-42-ester with [3-ethylz4-(4-nitrophenyl)
`butylcarbamic acid
`
`20
`
`30
`
`[4-(methylthio)butyl]car
`
`EXAMPLE 5
`Rapamycin-3l,42-diester with (4-nitrophenyl)carbamic
`acid
`40
`A solution of 0.38 g 4-nitrophenylisocyanate in 4 mL
`pyridine was added to a solution of 1.16 g rapamycin in
`11 mL pyridine at 0° and stirred at 0° for 15 minutes,
`then warmed to 20° over 15 minutes and stirred at 20°
`for 1.5 hours. Cold 2N HCl (75 mL) was added. the
`products were extracted into ethyl acetate, washed with
`brine, dried over MgSO4 and evaporated to dryness.
`The residue was chromatographed on silica gel; elution
`with 10% ethyl acetate in methylene chloride afforded
`160 mg of the diester as a yellow solid, mp 164°-l71°.
`IR (KBr): 3380 (OH), 2910, 1725 (C20), 1635, 1590,
`1500, 1320, 1200 and 850 cm-1. 1H NMR (CDC13, 400
`MHz): 68.20 (d, 4H, aromatic), 7.57 (d, 2H, aromatic),
`7.53 (d, 2H, aromatic), 3.38 (s, 3H, OCH3), 3.37 (s, 3H,
`OCH3), 3.16 (s, 3H, OCH3). MS (neg ion FAB): 1241
`(M—), 1058, 590.
`
`45
`
`55
`
`EXAMPLE 6
`Rapamycin-42-ester with (4-nitropheny1)carbamic acid
`Further elution of the residue described in Example 5
`with 30% ethyl acetate in methylene chloride afforded
`380 mg of the 42-monoester as a yellow solid, mp
`137°-144°. IR (KBr): 3420 (OH), 2910, 1725 (C=O),
`1500, 1320, 1205 and 840 cm-1. NMR (CDCI3, 400
`MHZ): 68.20 (d, 2H, aromatic), 7.56 (cl, 2H, aromatic),
`3.39 (s, 3H, OCH3), 3.34 (s, 3H, OCH3), 3.14 (s, 3H,
`OCH3). MS (neg ion FAB): 1077 (M—), 590, 321.
`
`60
`
`8
`
`EXAMPLE 7
`Rapamycin-3l.42-diester with
`(4-methylphenyl)carbamic acid
`A solution of 1.04 g 4-methylphenylisocyanate in 3
`mL pyridine was added to a solution of 2.20 g rapamy
`cin in 8 mL pyridine at O‘ and stirred at 0° for 3.5 hours.
`Cold 2N HC] (180 mL) was added, the products were
`extracted into'ethyl acetate, washed with brine, dried
`over M gSO4 and evaporated to dryness. The reside was
`chromatographed on silica gel; elution with 10% ethyl
`acetate in methylene chloride afforded 300 mg of 31,42~
`diester as a yellow solid, mp 129°-145°.
`IR(KBr): 3400 (OH), 2920, 1725 (C:O), 1645, 1520,
`1450, 1215, 1200 and 812 cm-1. NMR (CDC13, 400
`MHz): 57.27 (complex, 4H, aromatic), 7.10 (complex,
`4H, aromatic), 3.38 (s, 31-1, OCH3), 3.35 (s, 3H, OCH3),
`3.16 (s, 3H. OCH3), 2.30 (s, 3H, ArCH3), 2.28 (s, 3H,
`AI‘CH3). MS (neg ion FAB): 1179 (M—), 1027.
`
`EXAMPLE 8
`Rapamycin-42-ester with (4-methylphenyl)carbamic
`acid
`Further elution of the residue described in Example 7
`with 30% ethyl acetate in methylene chloride afforded
`200 mg of the 42-monoester as a yellow solid, mp
`123°—135°. IR (KBr): 3420 (OH). 2930. 1720 (C_—_O),
`1640, 1520, 1440, 1220, 1203, and 1055 cm— 1. NMR
`(CDCI3, 400 MHz): 57.27 (d. 2H, aromatic). 7.10 (d, 2H,
`aromatic), 3.40 (s, 3H, OCI~I3), 3.34 (s. 3H. OCH3), 3.14
`(s, 3H. OCH3), 2.30 (s, 3H, ArCHg). MS (neg ion FAB):
`1046 (M—), 590, 321.
`
`EXAMPLE 9
`Rapamycin~42~ester with (2,4-difluorophenyl)carbamic
`acid
`A solution of 0.24 g rapamycin and 0.10 g 2,4
`difluorophenylisocyanate in 6 mL pyridine was stirred
`for 45 minutes at 0°. Cold 2N HCl (50 mL) was added,
`the product was extracted into ethyl acetate, washed
`with brine, dried over MgSO; and evaporated to dry
`ness. Chromatography on silica gel using 25% ethyl
`acetate in methylene chloride afforded 69 mg of 42
`ester as a white solid, mp 109°—114°. IR (KBr): 3420
`(OH), 2910, 1720 (C_—_O), 1635, 1520, 1215, 1085 and
`975 cm-l. NMR (CDC13, 400 MHz): 66.84 (complex,
`2H, aromatic), 6.69 (complex, 1H, aromatic), 3.39 (s,
`3H, OCH3), 3.33 (s, 3H, OCH3), 3.14 (5, 31-1, OCH3). MS
`(neg ion FAB): 1068 (M—), 590, 321.
`EXAMPLE 10
`Rapamycin-3l,42-diester with
`(2,4-difluorophenyl)carbamic acid
`A solution of 0.20 g rapamycin and 0.14 g 2,4~
`di?uorophenylisocyanate in 10 mL pyridine was stirred
`5 hours at 20°. Cold 2N HCl (50 mL) was added, the
`product was extracted into ethyl acetate, washed with
`brine and dried over MgSO4. The solvent was evapo
`rated off and the residue chromatographed on silica gel
`using 10% ethyl acetate in methylene chloride yielded
`020 g of title compound as a pale yellow solid, mp
`ll5°—123°. IR (KBr): 3420 (01-1), 2910. 1725 (C:O),
`1638, 1523, 1212, 1090 and 835 cm-1. NMR (CDC13,
`400 MHz): 56.85 (complex, 4H, aromatic), 6.70 (com
`plex, 2H, aromatic), 3.39 (s. 3H, OCH3), 3.36 (s, 3H,
`
`NOVARTIS EXHIBIT 2130
`Par v Novartis, IPR 2016-00084
`Page 5 of 7
`
`

`
`10
`OC1-13). 3.36 (s. 31-1. OCH3), 3.15 (s. 3H, OCH3). 0.88
`(pl-1, t-butyl), 0.075 (31-1. SiCH3). 0.061 (311, SiCH3). MS
`(neg ion FAB): 1182 (M‘), 1027, 1008. 590, 435.
`The following representative compounds can be pre
`pared from rapamycin and the appropriate isocyanate
`by employing the method used to prepare the title com
`pound in Example 14.
`Rapamycin-42-tert-butyldimethylsilyl
`with (phenylmethyl)carbamic acid
`Rapamycin-42-tert-butyldimethylsilyl
`with hexylcarbamic acid
`Rapamycin~42-tert-butyldimethylsilyl
`with di?uoromethylcarbamic acid
`ether-3 l -ester
`Rapam ycin -42-tert-butyldimethylsilyl
`with [4-(N,N-dimethyl)butyl]carbamic acid
`Rapamycin-42-tert-butyldimethylsilyl
`ether-31-ester
`with [4-(carbomethoxy)buty1]carbamic acid
`Rapamycin-42-tert-butyldimethylsilyl
`ether-3 l -ester
`with [2-(phenylmethyl)butyl]carbamic acid
`
`ether-3l-ester
`
`ether-3 l -ester
`
`ether-3 l -ester
`
`20
`
`5,118,678
`9
`0CH:).- 3.15 (s. 3H, OCHg). MS: (neg ion FAB): 1223
`(M—), 1049, 590. 321.
`EXAMPLE ll
`Rapamycin-31,42-diester with phenylcarbamic acid
`A solution of 0.50 g rapamycin and 0.19 g
`phenylisocyanate in 10 mL pyridine was stirred at 20°
`for 16 hours. Cold 2N 11C] (100 mL) was added and the
`product was extracted into ethyl acetate, washed with
`brine and dried over M gSO4. The solvent was removed
`and the residue chromatographed on silica gel using
`10% ethyl acetate in methylene chloride, yielded 0.32 g
`title compound as a white solid, mp 139°-148°.
`1R(KBr): 3330 (OH), 2920, 1720 (C20), 1670, 1590,
`1520, 1430, 1205, 745 and 685 cm—1. NMR (CDCI3, 400
`MHz): 67.27-740 (complex, 81-1. aromatic), 7.06 (com
`plex, 2H. aromatic), 3.39 (s. 3H, OC1-13). 3.37 (s, 3H,
`OCI-13), 3.16 (s, 3H, OC1-I3). MS (neg ion FAB): 1151
`(M—), 1013, 590, 321.
`EXAMPLE 12
`Rapamycin-3l.42-diester with
`(4-?uorophenyl)carbamic acid
`A solution of 1.54 g rapamycin and 0.68 g 4-fluoro
`phenylisocyanate in 10 mL pyridine was stirred at 0° for
`4.5 hours. Cold 2N HCl (100 mL) was added, the prod
`uct was extracted into ethyl acetate, washed with brine,
`dried over MgSO4, evaporated to dryness, and chro
`matographed on silica gel using 10% ethyl acetate in
`methylene to afford 0.65 g product as a white solid, mp
`l29°—l40°. IR (KBr): 3430 (OH). 2920. 1725 (C20),
`1640, 1505. 1200 and 825 cmrl. NMR (CDCI3, 400
`MHz): 67.33 (complex, 41-1, aromatic). 6.99 (t. 41-1. aro
`matic). 3.38 (s, 3H, OCH3), 3.35 (s. 3H. OCHg). 3.16 (s,
`3H. OC1-13). MS (neg ion FAB): 1187 (M_), 1031, 590,
`321.
`
`EXAMPLE l5
`Rapamycin-3l-ester with (2,4-difluorophenyl)carbamic
`acid
`Rapamycin-42-tert-butyldimethylsilyl ether-3l-ester
`with (2,4-di?uorphenyl)carbamic acid (0.60 g) was
`stirred for 16 hours at 20n in a solution ot'4 mL tetrahy
`drofuran, 4 mL water and 12 mL acetic acid. After
`dilution with 100 mL water, the product was extracted
`into methylene chloride, washed with brine, dried over
`MgSO4 and evaporated to an oil, which was chromato
`graphed on silica gel using 30% ethyl acetate in methy
`lene chloride to afford 220 mg of title compound as a
`white solid, mp 103°-1l7°. IR (KBr): 3440, 2920, 1735
`(C20), 1640, 1525, 1090 and 840 cm*‘. NMR (CDCI3,
`400 MHz): 86.85 (complex, 3H, aromatic), 3.40 (s, 3H,
`OCH3), 3.36 (s, 3H, OCH3), 3.15 (s, 3H, OC1-13). MS
`(neg ion FAB): 1068 (M-), 894, 590, 128.
`The following representative compounds can be pre
`pared from rapamycin and the appropriate isocyanate
`by employing the method used to prepare the title com
`pound in Example 15.
`Rapamycin-3l-ester with (phenylmethyDcarbamic acid
`Rapamycin-3l-ester with hexylcarbamic acid
`Rapamycin-31-ester with di?uoromethylcarbamic acid
`Rapamycin-31-ester with [4-(N,N-dimethyl)butyl]car
`bamic acid
`Rapamycin-31-ester with [4-(carbomethoxy)butyl]car
`bamic acid
`Rapamycin-3l-ester with [Z-(phenylmethyl)butyl]car
`bamic acid
`What is claimed is:
`1. A compound of the formula
`
`40
`
`45
`
`EXAMPLE 13
`Rapamycin-42-tert-butyldimethylsilyl ether
`Rapamycin (0.914 g) was added to a solution of 0.204
`g imidazole and 0.165 g tert-butyldimethylsilyl chloride
`in 4 mL dimethylformamide at 0° and stirred under
`nitrogen for 16 hours at 20“. Brine (100 mL) was added,
`the product was extracted into ether, washed with
`brine, dried over MgSO4 and evaporated. Chromatog
`raphy on silica gel afforded 0.65 g ofthe title compound
`as a white solid. 1R(KBr): 3430 (OH), 2920, 1715
`(C20), 1645, 1105, 985, 870, 842 and 775 cm-1. NMR
`(CDC13, 400 MHz): 63.41 (s, 3H, OCl-lg), 3.34 (s, 3H,
`50
`OCH3), 3.14 (s, 3H, OC1-13), 0.886 (s, 91-1, t-Bu), 0.074 (s,
`3H, SiCH3), 0.059 (s, 3H, SiCH3). MS (neg ion FAB):
`1027 (M'), 590, 435.
`EXAMPLE 14
`Rapamycin-42-tert-butyldimethylsilyl ether-Sl-ester
`with (2,4-difluorophenyl)carbamic acid
`A solution of 0.38 g of rapamycin-42-tert-butyl-dime
`thylsilyl ether and (2,4-ditluorophenyl)isocyanate in 6
`60
`mL pyridine was stirred at 20 for 16 hours. Cold 2N
`HC] (75 mL) was added, the product was extracted into
`ethyl acetate, washed with brine, dried over MgSO4,
`evaporated and the residue chromatographed on silica
`gel using 2% ethyl acetate in methylene chloride to
`65
`afford 150 mg of title compound as a white solid, mp
`l12°—122°. 1R(KBr): 3440 (OH), 2910, 1745 (C20),
`1640, 1520, 1090, 840 and 765 cm-‘. NMR (CDC13, 400
`MHz): 86.84 (complex, 3H, aromatic), 3.41 (s, 3H,
`
`55
`
`NOVARTIS EXHIBIT 2130
`Par v Novartis, IPR 2016-00084
`Page 6 of 7
`
`

`
`11
`
`5,118,678
`12
`14. A compound of claim 1 which is rapamycin-42
`ester with (4-methylphenyl)carbamic acid.
`15. A compound of claim 1 which is rapamycin-42
`ester with (2,4-difluorophenyl)carbamic acid.
`16. A compound of claim 1 which is rapamycin
`31,42-diester with (2,4-difluorophenyl)carbamic acid.
`17. A compound of claim 1 which is rapamycin
`31,42-diester with phenylcarbamic acid.
`18. A compound of claim 1 which is rapamycin
`31,42-diester with (4-?uorophenyl)carbamic acid.
`19. A compound of claim 1 which is rapamycin-31
`ester with (2,4-di?uorophenyl)carbamic acid.
`20. A compound which is rapamycin-42-tert-butyl
`dimethylsilyl ether-31-ester with