United States Patent [191
`Kao
`
`lllllllllllllllllllllllllllIlllllllllllllllllllIllllIllllllllllllllllllllll
`[11] Patent Number:
`5,194,447
`[45] Date of Patent:
`Mar. 16, 1993
`
`US005194447A
`
`-
`
`[54] SULFONYLCARBAMATES OF RAPAMYCIN
`[75] Inventor: Wenling Kao, Chester, Pa.
`[73] Assignee: American Home Products
`Corporation, New York, N.Y.
`[21] Appl. No.: 837,048
`[22] Filed:
`Feb. 18, 1992
`[51] Int. 61.5 ............... .. 017D 491/16; A61K 31/395;
`A61K 31/685; 007E 7/04
`[52] US. Cl. ................................... .. 514/542; 514/ 546
`[58] Field of Search .............................. .. 540/452, 456
`[56]
`References Cited
`U.S. PATENT DOCUMENTS
`
`3,992,992 12/1975 Sehgal et a1. ..................... .. 424/192
`3,993,749 11/1976 Sehgal et a1. .
`4,316,885 2/1982 Rakhit ..... ..
`4,401,653 8/1983 Eng ......... ..
`4,650,803 3/ 1987 Stella et a1.
`4,885,171 12/1989 Sehgal et a1. .
`5,078,999 l/ 1992 Warner et a1. . . . .
`
`. . . . .. 424/122
`
`5,080,899 1/ 1992 Sturm et a1. . . . . . .
`5,091,389 2/1992 Ondeyka et a1. ..
`
`. . . . .. 424/122
`.... .. 514/291
`
`5,100,883 3/1992 Schiehser . . . . . . . . . .
`
`. . . . .. 514/183
`
`5,1“),889 3/1992 Calne ................................. .. 514/291
`
`OTHER PUBLICATIONS
`U.S. Ser. No. 07/686,727 ?led Apr. 17, 1991-Kao, Vo
`gel, Musser.
`Vezina, C. J. Antibiot. 28, 721-726 (1975).
`Sehgal, S. N., J. Antibiot. 28, 727-732 (1975).
`Baker, 11., J. Antibiot. 31, 539-545 (1978).
`Martel, Q, Can. J. Physiol. Pharmacol. 55, 48 (1977).
`Staruch et a1., FASEB 3, 3411 (1989).
`Dumont, F, FASEB 3,5256(1989).
`Calne et al., Lancet, 1183-1185 (1978).
`Morris, R. Med. Sci. Res. 17:877 (1989).
`Primary Examiner-Robert T. Bond
`Attorney, Agent, or Firm-Arnold S. Milowsky
`
`ABS'I'RACI‘
`[57]
`A compound of the. structure
`
`wherein
`R1 and R2 are each, independently, hydrogen or
`—CONHS01-Ar; and
`Ar is phenyl, naphthyl, pyridyl, quinolyl, isoquinolyl,
`quinoxalyl, thienyl, thionaphthyl, furyl, benzofuryl,
`benzodioxyl, benzoxazolyl, benzoisoxazolyl, or ben
`zodioxolyl; wherein the Ar group may be optionally
`mono-, di, or tri-substituted; with the provision that
`R1 and R2 are not hydrogen; or
`or a pharmaceutically acceptable salt thereof which by
`virtue of its 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 antifungal activity is useful
`in treating fungal infections.
`
`7 Claims, No Drawing
`
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`1
`
`2
`
`SULFONYLCARBAMATES OF RAPAMYCIN
`
`25
`
`5,194,447
`wherein
`R1 and R2 are each, independently, hydrogen or
`--CONHSO2-Ar; and
`Ar is phenyl, naphthyl, pyridyl, quinolyl, isoquinolyl,
`quinoxalyl, thienyl, thionaphthyl, furyl, benzofu
`ryl, benzodioxyl, benzoxazolyl, benzoisoxazolyl, or
`benzodioxolyl; ‘wherein the Ar group may be op
`tionally 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 l-6 carbon
`atoms per alkyl group, alkylthio of l-6 carbon
`atoms, -—SO3H, —PO3H, and —-CO;H;
`with the proviso that R1 and R2 are not both hydrogen;
`or a pharmaceutically acceptable salt thereof when the
`Ar group contains a basic nitrogen or when the Ar
`group is substituted by dialkylamino of l-6 carbon
`atoms per alkyl group, —-SO3H, -—PO3H, or COZH.
`The pharmaceutically acceptable salts are those de
`rived from such inorganic cations such as sodium, po
`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
`bon atoms per alkyl group, and the like; and organic and
`inorganic acids as: acetic, lactic, citric, tartaric, suc
`cinic, maleic, malonic, gluconic hydrochloric, hydro
`bromic, phosphoric, nitric, sulfuric, methanesulfonic,
`and similarly known acceptable acids and.
`Of these compounds, preferred members are those in
`which Ar is optionally mono-, di-, or tri- substituted
`phenyl. When Ar is substituted with arylalkyl of 7-10
`carbon atoms, it is preferred that aryl portion of the
`arylalkyl moiety be a phenyl group.
`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
`
`BACKGROUND OF THE INVENTION
`This invention relates to carbamates of rapamycin
`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 hygroscopieus, 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); US. Pat. No. 3,929,992; and US. Pat. No.
`3,993,749].
`Rapamycin alone (US. 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 a1.
`[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, 3411 (1989). Cyclosporin A
`and Eli-506, other macrocyclic molecules, also have
`30
`been shown to be effective as immunosuppressive
`agents, therefore useful in preventing transplant rejec
`tion [FASEB 3, 3411 (1989); FASEB 3, 5256 (1989); R.
`Y. Calne et al., Lancet 1183 (1978); and European Pa
`tent Application 0401747 A2].
`Mono- and diacylated derivatives of rapamycin (es
`teri?ed at the 28 and 43 positions) have been shown to
`be useful as antifungal agents (115. 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.
`
`35
`
`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
`
`gen]
`
`\OMe
`
`'
`
`55
`
`65
`
`under neutral conditions or in the presence of a base,
`such as pyridine.
`The 3l-carbamylated compounds of this invention
`can be prepared by protecting the 42-a1cohol of rapa
`mycin with a protecting group, such as with a tert-butyl
`dimethylsilyl group, followed by carbamylation of the
`3l-position with an isocyanate with the general struc
`ture shown above. Removal of the protecting group
`provides the 3l-carbamylated compounds. In the case
`of the tert-butyl dimethylsilyl protecting group, depro
`tection can be accomplished under mildly acidic condi
`tions. .
`Having the 31-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.
`The isocyanates used to prepare the compounds of
`the invention are commercially available or can be pre
`pared by methods that are disclosed in the literature.
`March [Advanced Organic Chemistry, 3d ed., pp. 391,
`452, and 479 (1985)] describes a general method for
`preparing arylsulfonyl isocyanates, that can be used
`
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`
`5,194,447
`3
`where the arylsulfonylisocyanate is not commercially
`available. The following scheme is illustrative of one
`method starting from an aryl moiety. Other methods of
`preparing arylsulfonyl isocyanates are known in the
`literature.
`
`4
`
`3
`H-PLN cells control C3H mouse —
`N" I," v1’I-Il'<'§r u‘
`H-PLN cells control C31! mouse —
`3H-PLN cells test compound-treated CBH mouse
`
`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.
`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
`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
`daysi-SD.) of the drug treatment group is compared
`with the control group.
`The following table summarizes the results of repre
`sentative compounds of this invention in these three
`standard test procedures.
`TABLE 1
`LAF
`PLN
`(ratio)
`(ratio)
`0.91
`0.84 (p.o.)
`1.03 (i.p.)
`0.47
`+
`1.00
`
`Compound
`Example 1
`
`Example 3
`Example 4
`Rapamycin
`
`0.74
`0.10
`LCD
`
`also; or
`Ar-H CISOZOH
`
`ArSOgOH
`
`V
`
`ArSO1Cl Q5299 ArSOgNCO
`
`Immunosuppressive activity was evaluated in an in 15
`vitro standard pharmacological test procedure to mea
`sure lymphocyte proliferation (LAF) and in two in vivo
`standard pharmacological test procedures. The ?rst in
`vivo procedure was a popliteal lymph node (PLN) test
`20
`procedure which measured the effect of compounds of
`this invention on a mixed lymphocyte reaction and the
`second in vivo procedure evaluated the survival time of
`a pinch skin graft. The ability to prevent or treat trans
`plantation rejection was also demonstrated in an in vivo
`heart allograft standard test procedure.
`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 are cultured for 72 hours with FHA and
`IL-1 and pulsed with tritiated thymidine during the last
`six hours. Cells are cultured with and without various
`concentration 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 according to the following ratio.
`
`30
`
`35
`
`25
`
`3H-control thmus cells — H3-raparnycin-treated thymus cells
`3H-corrtrol thymus cells — H3-test compound-treated cells
`
`50
`
`A mixed lymphocyte reaction (MLR) occurs when 45
`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 of tritiated 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,
`55
`irradiated spleen cells from BALB/c donors are in
`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
`PLN from the injected hind foot. Percent suppression is
`calculated using the non-drug treated animals as allo
`genie 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:
`
`65
`
`Skin Graft
`(days 1- SD)
`H2 1- 0.4
`
`+
`+
`12.0 i 1.7
`
`+ Not evaluated.
`
`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.
`The ability of the compounds of this invention to
`prevent or treat transplantation was demonstrated in a
`heterotropic heart allograft standard pharmacological
`test procedure that emulates transplantation rejection
`that occurs in humans. The following brie?y describes
`the procedure that was used. Male BN rat neonate do~
`nors (less than 5 days of age) were humanely sacri?ced,
`the thymus was dissected away from the heart. All
`connections with the thoracic cavity were severed and
`the heart was removed from the chest cavity and placed
`in cooled RPMI media where all adherent fat and fascia
`were removed. The heart was bisected in half, along the
`midline from the apex to the root of the aorta, to gener
`ate two approximately equal halves each containing
`atrial and ventricular tissue. Recipient male Lewis rats
`were anesthetized with phenobarbitol (50 mg/mL; i.p.),
`the left inner car was swabbed with povidine iodine, and
`1 mL RPMI was injected subcutaneously above the
`cartiledge plate to produce a ?uid ?lled sac. A stab
`incision was made to the sac, into which was inserted a
`single half heart fragment. The pocket was sealed with
`
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`Treatment Group
`Control
`Example 1
`Rapamycin
`
`20
`
`25
`
`35
`
`45
`
`5, 194,447
`6
`5
`and tablets preferably contain up to 99% of the active
`a single drop-of Vet-Seal (3M Animal Care Products).
`ingredient. Suitable solid carriers include, for example,
`Recipients were divided into treatment groups of 10 rats
`calcium phosphate, magnesium stearate, talc, sugars,
`each and the compounds to be evaluated were adminis
`tered at a dosage of 225 lag/day following the transplan
`lactose, dextrin, starch, gelatin, cellulose, methyl cellu
`tation procedure until graft failure occurred. Adminis
`lose, sodium carboxymethyl cellulose, polyvinylpyr
`tration was i.p., either by manual injection or via an
`rolidine, low melting waxes and ion exchange resins.
`Azlet osmotic pump that was implanted into the perito
`Liquid carriers are used in preparing solutions, sus
`neum of the recipient rat. Grafts were inspected for loss
`pensions, emulsions, syrups, elixirs and pressurized
`of cardiac activity on day 7 post-transplant and subse
`compositions. The active ingredient can be dissolved or
`quently on alternate days. Graft survival time is de?ned
`suspended in a pharmaceutically acceptable liquid car
`as the post-transplant day on which the heart graft has
`rier such as water, an organic solvent, a mixture of both
`lost all contractile activity by visual inspection and/or
`or pharmaceutically acceptable oils or fats. The liquid
`cardiac monitor. Individual rejection times were aver
`carrier can contain other suitable pharmaceutical addi~
`aged to produce a mean survival time for each treated
`tives such as solubilizers, emulsi?ers, buffers, preserva
`group. The following table shows the results obtained
`tives, sweeteners, ?avoring agents, suspending agents,
`for a representative compound of this invention. The
`thickening agents, colors, viscosity regulators, stabiliz
`control group represents rats that received the allograft
`ers or osmo-regulators. Suitable examples of liquid car
`and no test compound. Rapamycin was evaluated for
`riers for oral and parenteral administration include
`the purpose of comparison.
`water (partially containing additives as above, e.g. cel
`TABLE 2
`lulose derivatives, preferably sodium carboxymethyl
`cellulose solution), alcohols (including monohydric
`Mean Survival Time (days)
`alcohols and polyhydric alcohols, e.g. glycols) and their
`1 1.5
`derivatives, and oils (e.g. fractionated coconut oil and
`34.4
`38
`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.
`The compounds of this invention may be adminis
`tered rectally in the form of a conventional suppository.
`For administration by intranasal or intrabronchial inha
`lation or insufflation, the compounds of this invention
`may be formulated into an aqueous or partially aqueous
`solution, which can then be utilized in the form of an
`aerosol. The compounds of this invention may also be
`administered transdermally through the use of a trans
`dermal patch containing the active compound and a
`carrier that is inert to the active compound, is non toxic
`to the skin, and allows delivery of the agent for systemic
`absorption into the blood stream via the skin. The car
`rier may take any number of forms such as creams and
`ointments, pastes, gels, and occlusive devices. The
`creams and ointments may be viscous liquid or semi
`solid emulsions of either the oil-in-water or water-in-oil
`type. Pastes comprised of absorptive powders dispersed
`in petroleum or hydrophilic petroleum containing the
`active ingredient may also be suitable. A variety of
`occlusive devices may be used to release the active
`ingredient into the blood stream such as a semipermia
`ble membrane covering a reservoir containing the ac
`tive ingredient with or without a carrier, or a matrix
`containing the active ingredient. Other occlusive de
`vices are known in the literature.
`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
`
`As shown in Table 2, treatment with a representative
`compound of this invention signi?cantly (p<0.05) pro
`longed survival time of the transplanted allograft. The
`untreated allograft was rejected by 11.5 days, whereas
`treatment with a representative compound of this in
`vention prevented rejection until 34.4 days. As such, the
`results of this standard pharmacological test procedure
`demonstrate that the compounds of this invention are
`useful in preventing or treating transplantation rejec
`tion.
`Because the compounds of this invention are structur
`ally similar to rapamycin and have a similar activity
`profile 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.
`As the compound of Example 3 was prepared via its
`42-silylated intermediate (Example 2), the compound of
`Example 2 is therefore a useful as an intermediate of
`compounds of this invention.
`The compounds of this invention can be formulated
`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 flavoring agents, lubricants, solu
`bilizers, suspending agents, fillers, 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
`
`55
`
`60
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`10
`
`20
`
`25
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`5,194,447
`8
`7
`was diluted with 130 mL ethyl acetate and washed with
`ges or sachets containing liquids. The unit dosage form
`brine (2X20 ml). The ethyl acetate solution was dried
`can be, for example, a capsule or tablet itself, or it can be
`with magnesium sulfate and evaporated to dryness to
`the appropriate number of any such compositions in
`package form. The dosage to be used in the treatment
`give 208 mg of the title compound as a white foam.
`must be subjectively determined by the attending physi
`IR (KBr): 3400 (OH), 2930, 1730 (C=O), 1640 (aro
`matic), 1440, 1340, 1160, 1085 cm-1. NMR (CDC13, 400
`clan.
`In addition, the compounds of this invention may be
`MHZ): 6 8.0 (m, 2H, aromatic), 7.59 (m, 3H, aromatic),
`employed as a solution, cream, or lotion by formulation
`3.41 (s, 3H, OCH3), 3.26 (s, 3H, OCH3). MS (neg. ion
`with pharmaceutically acceptable vehicles containing
`FAB): 1095 (M-), 589, 321.
`0.1-5 percent, preferably 2%, of active compound
`EXAMPLE 4
`which may be administered to a fungally affected area.
`The following examples illustrate the preparation of
`v Rapamycin 42-ester with
`representative compounds of this invention.
`(4-chlorophenylsulfony1)carbamic acid
`EXAMPLE 1
`A solution of 400 mg of rapamycin in 10 mL ether/
`toluene (1:4) was treated at —- 10' under nitrogen with
`Rapamycin 42-ester with phenylsulfonylcarbamic acid
`100 mg of 4-chlorobenzenesulfonylisocyanate in 1 mL
`A solution of 192 mg of rapamycin in 1 mL toluene
`toluene. The mixture was stirred at 0' under nitrogen
`was treated at 0' under nitrogen with 37 mg of benzene
`for 3.5 hours, diluted with 120 mL of ethyl acetate,
`sulfonyl isocyanate in 1 mL toluene. After stirring at 0'
`washed with saturated sodium bicarbonate solution,
`under nitrogen for one hour, the reaction mixture was
`followed by a water wash, and dried with magnesium
`diluted with 100 ml of ethyl acetate and washed with
`sulfate. The ethyl acetate solution was evaporated and
`brine (3X20 ml). The ethyl acetate solution was dried
`the residue chromatographed on silica gel. Elution with
`with magnesium sulfate and evaporated. The residue
`ethyl acetate afforded 100 mg of the title compound as
`was chromatographed on silica gel. Elution with ethyl
`a white solid, mp 142'-146’. The title compound was
`acetate/hexane (1:1) afforded 36 mg of the title com
`isolated as a trihydrate.
`und as a white foam.
`1H NMR (CDC13, 400 MHz), 6 8.0 (d, J = 13 cps, 2H,
`IR (KBr) 3440 (OH), 2910, 1735 8: 1710 (C=O), 1635
`aromatic protons meta to sulfonyl group), 7.53 (d, J = 13
`(aromatic), 1440, 1160, 1080 and 980 cm-1. NMR
`cps, 2H, aromatic protons ortho to sulfonyl group), 3.30
`(CDC13, 400 MHz): 6 8.04 (m, 2H, aromatic), 7.65 (m,
`(s, 3H, OCH3), 3.20 (s, 3H, OCH3), 3.13 (s, 3H, OCH3)
`1H, aromatic), 7.56 (m, 2H, aromatic), 3.41 (s, 3H,
`ppm. MS (neg ion FAB): 1129 (MH-). IR KBr max 3430
`OCH3), 3.34 (s, 3H, OCH3), 3.13 (s, 3H, OCH3). MS
`(OH), 2930, 1740, 1730 (both C=O), 1640 (amide
`(neg. ion FAB): 1095 (M-), 590. 321.
`C=O), 1450, 1350 (—SO;—-), 1160 (—SO;—), 1160
`(—SO2—-), 1090, 985 cm-1.
`EXAMPLE 2
`Anal. Calcd. for C5gHg3ClN2016S3 H20: C 58.74; H
`Rapamycin 42-tertbutyldimethylsilyl ether-3l-ester
`7.56; N 2.35 Found: C 58.92; H 7.99; N 2.17
`with phenyl sulfonyl carbamic acid
`Rapamycin (0.914 g) was added to a solution of 0.204
`g imidazole and 0.165 g ten-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 of rapamycin-42-tert
`butyldimethylsilyl ether as a white solid.
`A solution of 308 mg of rapamycin-42-tert-butyldime
`thylsilyl ether in 1 mL toluene was treated at 0° under
`nitrogen with 109 mg of benezenesulfonyl isocyanate in
`1 mL toluene. After stirring at 0‘ under nitrogen for
`three hours, the reaction mixture was diluted with 100
`mL ethyl acetate and washed with brine 2X20 ml). The
`ethyl acetate solution was dried with magnesium sulfate
`and evaporated. The residue was chromatographed on
`silica gel. Elution with ethyl acetate/hexane (1:1) af
`forded 310 mg of the title compound as a white foam.
`IR (KBr): 3400 and 3240 (OH), 2915, 1740 and 1710
`(C=O), 1640, 1440, 1160, 1080 cm-l. NMR (CDCI3,
`400 MHz): 8 8.00 (m, 2H, aromatic), 7.55 (m, 3H, aro
`matic), 3.45 (s, 3H, OCH3), 3.27 (s, 3H, OCH3), 3.13 (s,
`3H, OCH3). MS (neg. ion FAB): 1209 (M-), 590, 156.
`EXAMPLE 3
`Rapamycin-31-ester with phenylsulfonylcarbamic acid
`A solution of 250 mg of Rapamycin-42-tert-butyl
`dimethylsilyl ether-31-ester with phenylsulfonylcar
`bamic acid in 2 mL of tetrahydrofuran was treated with
`2 mL water and 6 mL acetic acid. After stirring at room
`temperature under nitrogen for 20 hours, the mixture
`
`EXAMPLE 5
`Rapamycin 42-ester with (3-methylphenylsulfonyl)
`carbamic acid
`A solution of 400 mg rapamycin in 10 mL ethyl aceta
`te/ether (i) was treated at —l0° under nitrogen with
`127 mg of 3-methylphenylsulfonyl isocyanate in 2 mL
`ether. THe mixture was stirred at 0' under nitrogen for
`4.5 hours, diluted with 120 mL ethyl acetate, washed
`with a saturated sodium bicarbonate solution, and dried
`with magnesium sulfate. The ethyl acetate solution was
`evaporated and the residue chromatographed on silica
`gel. Elution with ethyl acetate/n-hexane (1:1) afforded
`270 mg of the title compound as a white powder, mp
`1 16'-120'.
`lH NMR(CDC13, 400 MHz) 8 8.13 (d, J= 13 cps, 1H,
`aromatic proton ortho to sulfonyl group), 7.53 (m, 1H,
`aromatic proton, para to sulfonyl group), 7.39 (m, 1H,
`aromatic protons para to methyl group), 7.33 (d, J = 13
`cps, 1H, aromatic ortho to methyl group), 3.23 (s, 3H,
`OCH3), 3.18 (s, 3H, OCH3), 3.14 (s, 3H, OCH3), 2.67 (s,
`3H, aromatic methyl) ppm. MS (neg. ion FAB): 1109
`(MH- ). IR KBr max 3450 (OH), 2940, 1740, 1720
`(C=O), 1640 (amide C=O), 1450, 1340 (—SO;—),
`1160 (—SO1—), 990 cm-1.
`EXAMPLE 6
`Rapamycin 31,42-diester with phenylsulfonylcarbamic
`acid
`A solution of 200 mg of rapamycin 42-ester with
`phenylsulfonylcarbamic acid in 3 mL ethyl aceta
`
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`te/ether (1:1) was treated at 0' under nitrogen with 200
`
`mg of benzenesulfonyl isocyanate in 0.5 mL of ether.
`
`After stirring at 0' under N; for 18 hours and at 22'
`
`under N; for 3 hours, the reaction mixture was diluted
`
`with 120 mL ethyl acetate, washed with a saturated
`
`sodium bicarbonate solution, and then washed with
`
`brine. The ethyl acetate solution was dried with magne
`
`sium sulfate and evaporate. The residue was chromato
`
`graphed on silica gel. Elution with ethyl acetate af
`
`5
`
`10
`
`15
`
`20
`
`25
`
`forded 75 mg of the title compound as a white foam, mp
`
`104°-108°.
`
`NMR (CDC13, 400 MHz): 6 8.03 (m, 2H, aromatic),
`
`7.55 (m, 3H, aromatic), 3.26 (s, 3H, OCH3), 3.22 (s, 3H,
`
`30
`
`35
`
`OCH3), 3.13 (s, 3H, OCH3) ppm. MS (neg. ion FAB):
`
`1278 (MH-). IR (KBr): 334-0 (OH), 2930, 1730 & 1715
`
`((1:0), 1630 (aromatic), 1445, 1290, 1160, 1095, 990
`
`cm — 1.
`
`What is claimed is:
`
`1. A compound of the formula
`
`wherein
`R1 and R2 are each, independently, hydrogen or
`—CONHSO2-Ar; and Ar is phenyl, naphthyl, pyri
`dyl, quinolyl, isoquinolyl, quinoxalyl, thienyl,
`thionaphthyl, furyl, benzofuryl, benzodioxyl, benz
`oxazolyl, benzoisoxazolyl, or benzodioxolyl;
`wherein the Ar group may be optionally mono-,
`di-, or tri-substituted with a group selected from
`alkyl of 1-6 carbon atoms, phenylalkyl of 7-10
`carbon atoms, alkoxy of l-6 carbon atoms, cyano,
`halo, nitro, carbalkoxy of 2-7 carbon atoms, tri?u
`oromethyl, amino, dialkylamino of 1-6 carbon
`atoms per alkyl group, alkylthio of l-6 carbon
`atoms, —-SO3H, —PO3H, and —COZH;
`with the proviso that R‘ and R2 are not both hydrogen;
`or a pharmaceutically acceptable salt thereof when the
`Ar group contains a basic nitrogen or when the Ar
`group is substituted by dialklyamino of l-6 carbon
`atoms per alkyl group, —S()3H, --PO3H, or —CO2H.
`2. A compound of claim 1 wherein Ar is phenyl; the
`phenyl group being optionally mono-, di-, or tri-sub
`stituted or a pharmaceutically acceptible salt thereof.
`3. A compound of claim 1 which is rapamycin 42
`ester with phenylsulfonylcarbamic acid.
`4. A compound of claim 1 which is rapamycin-31
`ester with phenylsulfonylcarbamic acid.
`5. A compound of claim 1 which is rapamycin 42
`ester with (4-chlorophenylsulfonyl)carbamic acid.
`6. A compound of claim 1 which is rapamycin 42
`ester with (3-methylphenylsulfonyl) carbamic acid.
`7. A compound which is rapamycin 42-tertbutyldime
`thylsilyl ether-31-ester with phenylsulfonylcarbamic
`acid.
`
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`NOVARTIS EXHIBIT 2075
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`Page 6 of 6