United States Patent [19]
`Colin et al.
`
`[11] Patent Number:
`[45] Date of Patent:
`
`4,814,470
`Mar. 21, 1989
`
`[54] TAXOL DERIVATIVES, THEIR
`PREPARATION AND PHARMACEUTICAL
`COMPOSITIONS CONTAINING THEM
`
`[75] Inventors: Michel Colin, Thoiry; Daniel
`Guenard, Montrouge; Francoise
`Gueritte-Voegelein; Pierre Potier,
`both of Paris, all of France
`
`[73] Assignee:
`
`Rhone-Poulenc Sante, Courbevoie,
`France
`
`[21] Appl. No.: 73,156
`
`[22] Filed:
`Jul. 14, 1987
`[30]
`Foreign Application Priority Data
`Jul. 17, 1986 [FR]
`France .............................. .. 86 10400
`
`[51] Int. Cl.4 .................. .. A01N 43/02; CO7D 305/00
`[52] US. Cl. ................................... .. 514/449; 549/510
`[58] Field of Search ....................... .. 549/510; 514/449
`
`[56]
`
`References Cited
`U.S. PATENT DOCUMENTS
`
`4,206,221 6/1980 Miller et a1. ...................... .. 549/510
`
`OTHER PUBLICATIONS
`V. Senilh et al, C. R. Acad. Sc. Paris, t. 299, Serie II,
`No. 15 (1984), pp. 1039-1043.
`M. E. Jung et a1, Jour. Chem. Soc., Chem. Comm.
`(1978), pp. 315-316.
`
`R. S. Lott et al, Jour. Chem. Soc., Chem. Comm.
`(1979), pp. 495-496.
`E. Herranz et al, Jour. Am. Chem. Soc., vol. 100:11
`(1978), pp. 3596-3598.
`Theodora W. Greene, Protective Groups in Organic
`Synthesis (1981), pp. 223, 225 and 232.
`Primary Examiner—Glennon H. Hollrah
`Assistant Examiner—Mark W. Russell
`Attorney, Agent, or Firm-Stevens, Davis, Miller &
`Mosher
`ABSTRACT
`[57]
`Taxol derivatives of formula
`
`in which R represents hydrogen or acetyl, one of R1 or
`R2 represents hydroxy and the other represents tert
`butoxycarbonylamino and their isomers are useful anti
`tumor agents.
`
`6 Claims, No Drawings
`
`MYLAN - EXHIBIT 1086
`Mylan Laboratories Limited v. Aventis Pharma S.A.
`IPR2016-00712
`
`

`

`1
`
`4,814,470
`
`2
`
`TAXOL DERIVATIVES, THEIR PREPARATION
`AND PHARMACEUTICAL COMPOSITIONS
`CONTAINING THEM
`
`The present invention provides new taxol derivatives
`of formula:
`
`coo
`
`/
`
`c6H5
`
`OCOOCH2CCI3
`O
`II CH3
`
`(III)
`
`O
`=
`ococa;
`E
`ococéns
`
`R0
`
`OH
`0
`ll CH3
`
`(I)
`
`CO-O
`
`2' CPI-R1
`
`C6H5—-CH-Rz
`3'
`
`OH
`
`E
`
`O
`OCOCH3
`OCOC6H5
`
`in which R represents hydrogen or acetyl and one of
`R1 and R2 represents hydroxy and the other represents
`tert-butoxycarbonylamino and their stereoisomeric
`forms, and mixtures thereof.
`Taxol, which is the formula:
`
`in which R’ represents acetyl or 2,2,2-trichloroethox
`ycarbonyl, in an organic solvent such as acetonitrile in
`the presence of silver nitrate and a tert-butanol solution
`of osmium tetroxide at a temperature between 0° and
`40° C., followed by the replacement of the 2,2,2-tri
`chloroethoxycarbonyl group(s) in the product of for
`mula:
`
`OCOOCH2CCI3
`0
`ll CH3
`
`(IV)
`
`15
`
`20
`
`25
`
`CH3CO~O
`
`O
`
`0H
`
`co-o
`
`CI-I-OH
`
`0
`C6H5—-CH—NI-ICOC6H5 OH 5 Fr
`ococrr;
`g
`ococéns
`
`(11)
`
`30
`
`35
`
`shows remarkable properties, in vitro, of promoting the
`polymerization of tubulin and inhibiting the depolymer
`ization of microtubules. For this reason, it is a particu
`larly valuable antileukaemia and antitumour agent.
`Because of the difficulty of extracting taxol from
`trunk barks of different species of Taxus (yew), the
`preparation of derivatives similar to taxol from 10
`deacetylbaccatin III, which can be extracted relatively
`easily from yew leaves, has been proposed. However,
`the derivatives synthesized until now have shown an
`activity which is lower than that of taxol [V. Senilh et
`al., CR. Acad. Sci., 299, series II, No. 15, p. 1039-1043
`(1984)].
`It has now been found, and this forms the subject of
`the present invention, that the products of formula (I)
`have an activity signi?cantly greater than that of taxol
`and, a fortiori, greater than that of compounds of for
`mula (I) in which R represents hydrogen, one of R1 or
`R2 represents hydroxy, and the other represents ethox
`ycarbonylamino.
`According to a feature of the present invention, the
`products of formula (I) may be obtained by reacting the
`sodium salt of tert-butyl N-chlorocarbamate with a
`comound of formula:
`
`55
`
`in which R’, R1 and R2 are as defined as above, by hy
`drogen using zinc in the presence of acetic acid at a
`temperature between 30° and 60° C.
`The reaction of the sodium salt of tert-butyl N
`chlorocarbamate with a compound of formula (III)
`leads to a mixture of isomers of the products of formula
`(IV), the constituents of which may be separated by
`physicochemical methods such as chromatography.
`The sodium salt of tert-butyl N-chlorocarbamate may
`be prepared from tert-butyl carbamate by the method
`described in J. Amer, Chem. Soc., 100, 3596 (1978).
`The starting materials of formula (III) may be ob
`tained by reacting cinnamoyl chloride, optionally pre
`pared in situ, with the compound of formula:
`
`OCOOCI-I2CCl3
`0
`ll CH3
`
`(V)
`
`HO
`
`ococn;
`
`in which R’ is as de?ned above, operating in an anhy
`drous organic solvent such as toluene in the presence of
`silver cyanide at a temperature between 80° and 120° C.
`The compounds of formula (III) may also be obtained
`by reacting cinnamic acid with a compound of formula
`(V) in which R’ is as de?ned above, in an aromatic
`hydrocarbon such as benzene, toluene or a xylene, in
`the presence of a condensation agent, such as a carbodi
`irnide, e.g. dicyclohexylcarbodiimide, or a reactive car
`bonate such as di(2-pyridyl)carbonate, and an activation
`agent, such as dimethylaminopyridine at a temperature
`between 60° and 90° C.
`
`

`

`4,814,470
`4.
`3
`For‘example, while operating in the presence of dicy
`Speci?c rotation: [orb23 = — 38.4° (c= 1, chloro
`clohexylcarbodiimide and dimethylaminopyridine, it is
`form).
`Ultraviolet spectrum: Amax=231 nm (15150),
`particularly advantageous to use a molar excess of cin
`namic acid relative to the product of formula (V), the
`hmax=275 nm (1200), hmax=283 nm (1035).
`Infrared spectrum: main characteristic absorption
`dicyclohexylcarbodiimide being used in a stoichiomet
`ric quantity relative to the cinnamic acid and the dime
`bands at 3580, 3440, 2960, 1770 and 1730 cm-1.
`Proton nuclear magnetic resonance spectrum
`thylaminopyridine being used in a stoichiometric quan~
`(CDC13, 400 MHz, shifts in ppm): 1.21 (s, 3H); 1.27 (s,
`tity relative to the starting material of formula (V). In
`3H); 1.36 (s, 9H); 1.86 (s, 3H); 1.96 (s, 3H); 2.39 (s, 3H);
`general, at least 4 moles of cinnamic acid are used per
`mole of the compound of formula (V).
`2.62 (m, 1H); 3.90 (d, J=7, 1H); 4.17 and 4.32 (2d, J=9,
`2H); 4.63 (d, J=3, 1H); 4.59 and 4.90 (2d, J: 12, 2H);
`The compounds of formula (V) in which R’ is as
`de?ned above may be obtained by reacting 2,2,2-tri
`4.77 (s, 2H); 4.96 (d, J=9, 1H); 5.27 (dd, J=9 and J=3,
`1H); 5.42 (d, J=9, 1H); 5.55 (m, 1H); 5.69 (d, J=7, 1H);
`chloroethyl chloroformate with baccatin III or with
`lO-deacetylbaccatin III, in a basic organic solvent such
`6.21 (t, J=9, 1H); 6.23 (s, 1H); 7.39 (5H); 7.51, 7.62 and
`8.09 (5H).
`as pyridine, at a temperature between 0° and 50° C.
`A product (2’S, 3'R) of general formula (IV) (250 mg)
`Baccatin III and IO-deacetylbaccatin III are natural
`in which R’ represents a 2,2,2—trichloroethoxycarbonyl
`products which can be extracted from yew (Taxus bac
`radical, R1 represents a hydroxy radical and R2 repre
`cata L) leaves or bark.
`The products of general formula (I), and especially
`sents a tert-butoxycarbonylamino radical, the charac
`those in which R represents hydrogen atom, R1 repre
`teristics of which are as follows:
`Speci?c rotation: [a] p23: -43.5° (c= 1, chloro
`sents hydroxy and R2 represents tert-butoxycar
`bonylamino, have valuable biological activities.
`form).
`Their biological activity may be determined in vitro,
`Ultraviolet spectrum: hmax=231 nm (15300),
`using tubulin extracted from pig brain, by the method of
`hmax=275 nm (1035), Amax=283 nm (905).
`Infrared spectrum: characteristic absorption bands at
`M. L. Shelanski et al., Proc. Natl. Acad. Sci. U.S.A., 70,
`765-768 (1973). The depolymerization of the microtu
`3400, 3000, 1770 and 1730 cm—1.
`bules in tubulin may be studied by the method of G.
`Proton nuclear magnetic resonance spectrum
`(CDC13, 400 MHz, shifts in ppm): 1.18 (s, 3H); 1.23 (s,
`Chauviere et al., CR. Acad. Sci., 293, series II, 501-503
`3H); 1.40 (s, 9H); 1.86 (s, 3H); 2.08 (s, 3H); 2.24 (s, 3H);
`(1981). In this study, the products of formula (I) were
`2.64 (m, 1H); 3.98 (d, J=7, 1H); 4.17 and 4.32 (d, J=9,
`found to be approximately twice as active as taxol.
`In vivo, the products of formula (I) were found to be
`2H); 4.48 (d, J =3, 1H); 4.60 and 4.92 (2d, .1: 12, 2H);
`4.78 (s, 2H); 4.97 (d, J=9, 1H); 5.22 (dd, J=9 and 1:3,
`active in mice grafted with leukaemia L 1210 or with
`1H); 5.32 (d, J=9, 1H); 5.58 (m, 1H); 5.70 (d, J=7, 1H);
`leukaemia P 388, at doses of between 1 and 10 mg/kg,
`administered intraperitoneally. At equitoxic doses, the
`6.07 (t, J =9, 1H); 6.27 (s, 1H); 7.33-7.45 (5H); 7.48, 7.61
`products of formula (I) showed an antitumour ef?cacy
`and 8.04 (5H).
`A product (2’R, 3'S) of general formula (IV) (250 mg)
`greater than that of taxol (i.e. an increased survival time,
`with the animals surviving long-term).
`in which R’ represents a 2,2,2-trichloroethoxycarbonyl
`The following Examples illustrate the invention.
`radical, R1 represents a tert-butoxycarbonylamino radi
`cal and R2 represents a hydroxy radical, the characteris
`tics of which are as follows:
`Speci?c rotation: [a]1)23= ——37.8° (c= 1, chloro—
`form).
`Ultraviolet spectrum: hmax=231 nm (14500),
`hmax=274 nm (1730), Amax=282 nm (1520).
`Infrared spectrum: characteristic absorption bands at
`3590, 3440, 3000, 1770 and 1730 cm"1.
`Proton nuclear magnetic resonance spectrum
`(CDC13, 400 MHz, shifts in ppm): 1.20 (s, 3H); 1.27 (s,
`3H); 1.37 (s, 9H); 1.87 (s, 3H); 2.02 (s, 3H); 2.42 (s, 3H);
`2.64 (m, 1H); 3.96 (d, J=7, 1H); 4.19 and 4.32 (2d, J=9,
`2H); 4.59 (wide d, J: 12, 2H); 4.78 (s, 2H); 4.91 (d,
`J: 12, 11-1); 5.00 (cl, J=9, 1H); 5.40 (s, 1H); 5.51 (d, J=9,
`1H); 5.58 (m, 1H); 5.69 (d, J=7, 1H); 6.25 (s, 1H); 6.31
`(t, J=9, 1H); 7.36, 7.40 and 7.46 (5H); 7.48, 7.68 and 8.06
`(5H).
`And a product (2’S, 3'R) of general formula (IV) (180
`mg) in which R’ represents a 2,2,2-trichloroethoxycar
`bonyl radical, R1 represents a tert-butoxycar
`bonylamino radical and R2 represents a hydroxy radi
`cal, the characteristics of which are as follows:
`Speci?c rotation: [a]D23= —32° (c= 1, chloroform).
`Ultraviolet spectrum: hmax=231 nm (14900),
`hmax=275 mm (1180), 7tmax=282 nm (1050).
`Infrared spectrum: characteristic absorption bands at
`3600, 3440, 3000, 1770 and 1730 cm-1.
`Proton nuclear magnetic resonance spectrum
`(CDC13, 400 MHz, shifts in ppm): 1.18 (s, 3H); 1.27 (s,
`3H); 1.38 (s, 9H); 1.89 (s, 3H); 2.02 (s, 3H); 2.32 (s, 3H);
`
`EXAMPLE 1
`A solution of tert-butyl N-chlorocarbamate sodium
`salt (0.5 g) and silver nitrate (l g) in acetonitrile (20 cc)
`is stirred vigorously for 5 minutes. A solution (0.2 cc) of
`osmium tetroxide in tert-butyl alcohol (0.1 mole per
`liter solution), the product of formula (III) in which R’
`represents a 2,2,2-trichloroethoxycarbonyl radical (2 g)
`and water (0.16 cc) are then added. After stirring for 20
`hours at a temperature in the vicinity of 20° C. and in
`the absence of light, tert-butyl N-chlorocarbamate so
`dium salt (0.5 g), the osmium tetroxide solution (0.1 cc)
`and water (0.06 cc) are added. After stirring vigorously
`for 48 hours, the reaction mixture is ?ltered through
`Celite. The ?lter is rinsed with acetonitrile and the
`?ltrate is concentrated to dryness. The product ob
`tained is puri?ed by chromatography on silica (Merck
`7736 silica), eluting with an etherzhexane (50:50 by vol
`ume) mixture and operating under slight pressure. The
`unreacted product of formula (III) (900 mg) and the
`oxyaminated products are isolated in this way and the
`latter are puri?ed and separated by thick layer chroma
`tography, eluting with a methylene chloridemethanol
`(98:2 by volume) mixture.
`The following are thereby obtained:
`A product (2’R, 3'S) of general formula (IV) (295 mg)
`in which R’ represents a 2,2,2-trichloroethoxycarbonyl
`radical, R1 represents a hydroxy radical and R1 repre
`sents a tert-butoxycarbonylamino radical, the charac
`teristics of which are as follows:
`
`60
`
`65
`
`10
`
`45
`
`50
`
`55
`
`

`

`10
`
`15
`
`25
`
`30
`
`35
`
`20
`
`4,814,470
`6
`5
`Ultraviolet spectrum: 7tmax=217 nm (26800),
`2.62 (m, 1H); 3.87 (d, J=7, 1H); 4.15 and 4.32 (2d, J=9,
`2H); 4.60 (wide (1, J =12, 2H); 4.77 (s, 2H); 4.91 (d,
`)tmax=222 nm (26900), Xmax=232 nm (16100),
`J=l2, 1H); 4.96 (d, J=9, 1H); 5.16 (d, J=3, 1H); 5.34
`kmax=276 nm (23600), Xmax=283 nm (24400).
`Infrared spectrum: main characteristic absorption
`(d, J=9, 1H); 5.57 (m, 1H); 5.67 (d, J=7, 1H); 6.16 (t,
`bands at 3420, 1760, 1725, 1710 and 1635 cm—1.
`J=9, 1H); 6.23 (s, 1H); 7.39 (5H); 7.53, 7.66 and 8.07
`Proton nuclear magnetic resonance spectrum
`(5H).
`(CDC13, shifts in ppm): 5.73 (d, J =7, CZH); 3.99 (d,
`Zinc powder (150 mg) is added to a solution of the
`J=7, C3H); 5.02 (d, J=9, C5H); 1.88 and 2.68 (m,
`product (2'R, 3’S) of general formula (IV) (150 mg) in
`2><C6H); 5.62 (m, C7H); 6.30 (s, C10H); 6.21 (t, J=8,
`which R’ represents a 2,2,2-trichloroethoxycarbonyl
`C13H); 2-48 (In, C14H2); 1-29 (S, C16H3); 1-23 (S, C17H3);
`radical, R1 represents a hydroxy radical and R2 repre
`2.16 (s, C13H3); 1.88 (s, C19H3); 4.20 and 4.34 (d, J=9,
`sents a tert-butoxycarbonylamino radical, in acetic acid
`2><C20H); 2.31 (acetate); 7.45, 7.60 and 8.07 (benzoate);
`(5 cc). The reaction mixture is stirred for 2 hours at 50°
`6.53 (d, J=16, C2, H); 7.89 (d, J: 16, C3, H); 7.45 (4H);
`C. and it is then ?ltered and concentrated to dryness.
`7.60 (1H); 4.62 to 4.93 (d, J: 12); 4.79 (s, 2H).
`The residue is taken up with water and extraction is
`Mass spectrum (chemical ionization) m/z 1023
`carried out with ethyl acetate. The combined organic
`(MH+), 1005, 831, 813, 683, 665, 491, 431, 369, 309, 291,
`phases are concentrated to dryness and the residue is
`149, 131 and 123.
`puri?ed by thick layer chromatography, eluting with a
`(2) Cinnamic acid (35.52 g; 240 mmols), anhydrous
`methylene chloridezmethanol (97:3 by volume) mixture.
`toluene (1 liter), dicyclohexylcarbodiimide (49.44 g; 240
`A product (2'R, 3’S) of general formula (I) (94 mg) in
`mmols), product of general formula (V) in which R’
`which R represents a hydrogen atom, R1 represents a
`represents a 2,2,2-trichloroethoxycarbonyl radical (53.5
`hydroxy radical and R2 represents a tert-butoxycar
`g; 60 mmols) and dimethylaminopyridine (7.32 g; 60
`bonylamino radical, the characteristics of which are as
`mmols) are introduced into a 2 liter three-necked round
`follows, is thereby obtained:
`bottomed ?ask, equipped with a stirrer and a thermome
`Speci?c rotation: [a]D23= ~36“ (c=0.74; ethanol).
`ter, under an argon atmosphere. The mixture is heated
`Ultraviolet spectrum: kmax=230 nm (14800),
`for 18 hours at 70° C. under an argon atmosphere. After
`Amax=275 nm (1730), kmax=283 nm (1670).
`cooling at 0° C. for 4 hours, the precipitate formed is
`Infrared spectrum: main characteristic absorption
`separated by ?ltration and then washed with cold tolu
`bands at 3590, 3440, 1740-1700 cmrl.
`ene (100 cc).
`Proton nuclear magnetic resonance spectrum
`The ?ltrate is concentrated to dryness and it is then
`(CDC13, 400 MHz, shifts in ppm): 1.12 (s, 3H); 1.24 (s,
`taken up with methylene chloride (1 liter). The solution
`3H); 1.35 (s, 9H); 1.77 (s, 3H); 1.87 (s, 3H); 2.28 (m, 2H);
`in methylene chloride is washed with an aqueous 3%
`2.37 (s, 3H); 2.58 (m, 1H); 3.91 (d, J=7, 1H); 4.19 and
`(w/v) hydrochloric acid solution (3x150 cc). After
`4.32 (2d, J =9, 2H); 4.26 (m, 1H); 4.62 (d, .1 =2, 1H); 4.94
`concentrating the organic phase, the residue (92 g) is
`(d, J=9, 1H); 5.22 (s, 1H); 5.26 (dd, J=9 and J=2, 1H);
`taken up with ethyl ether (500 cc). The solution is al
`5.46 (d, J=9, 1H); 5.68 (d, J=7, 1H); 6.22 (t, J=9, 1H);
`lowed to stand at a temperature in the vicinity of 0° C.
`7.38 (5H); 7.50, 7.60 and 8.12 (5H).
`for 48 hours. The precipitate formed is separated by
`Mass spectrum (FAB) rn/z: 808 (MH+), 790, 752,
`?ltration and washed with ethyl ether at 0° C. The
`734, 708, 690, 527, 509, 449, 405, 387, 345, 327, 282, 226
`?ltrate is concentrated to dryness. A product (89 g) is
`and 185.
`thereby obtained, which is chromatographed on Merck
`The product of general formula (III) in which R’
`7734 silica (2.7 kg), eluting with a toluene:methanol
`represents a 2,2,2-trichloroethoxycarbonyl radical may
`(95:5 by volume) mixture. The product of general for
`be prepared according to one of the following methods:
`mula (III) (58 g) in which R’ represents a 2,2,2-tri
`(1) Oxalyl chloride (11.92 cc) is added to a solution of
`chloroethoxycarbonyl radical is thereby obtained
`cinnarnic acid (9.84 g; 66.5 mmols) in anhydrous toluene
`(yield=94.6%).
`(150 cc). The reaction mixture is stirred for 1 hour at 60°
`The product of general formula (V) in which R’ rep
`C. and the excess oxalyl chloride is then removed by
`resents a 2,2,2-trichloroethoxycarbonyl radical may be
`distillation. The cinnamoyl chloride obtained is taken
`prepared as follows:
`up with anhydrous toluene (300 cc) and the product of
`A solution of 10-deacetylbaccatin III (30 g; 55
`general formula (V) (12 g) in which R’ represents a
`mmols) in anhydrous pyridine (480 cc) is cooled under
`2,2,2-trich1oroethoxycarbonyl radical and silver cya
`an argon atmosphere to 3° C. 2,2,2-Trich1oroethyl
`nide (7.9 g) are then added. The reaction mixture is
`chloroformate (25.5 cc; 184 mmols) is added in the
`heated for 10 hours at 110° C., with vigorous stirring.
`course of 3 minutes. The reaction mixture is stirred for
`After cooling, the reaction mixture is ?ltered and the
`3 minutes at 20° C. and then for 6 minutes at 28° C. The
`precipitate is rinsed with ethyl acetate. The combined
`solution is then cooled using an ice bath and it is then
`?ltrates are poured into ice-cold water. Extraction is
`quickly poured into ice-cold water (1 liter). The aque
`carried out with ethyl acetate. The combined organic
`ous phase is extracted 3 times with methylene chloride
`phases are concentrated to dryness and then taken up
`(1 liter in total). After concentrating, the pyridine is
`with ether (200 cc). A stream of ammonia is passed
`removed by exhaustive extraction with 1,2-dichloroe
`through this solution until the ammonium cinnamate
`thane. The crude product obtained (61.9 g) is puri?ed
`formed precipitates. After ?ltering, the ethereal solu
`by chromatography on silica (Merck 7736 silica; 1.2 kg),
`tion is concentrated and the residue is chromatographed
`eluting with a methylene chloridezmethanol (99:1 by
`on silica.(Merck 7736 silica), eluting with methylene
`volume) mixture.
`chloride under pressure. The product of general for
`The product of general formula (V) (45.6 g) in which
`mula (III) (7.6 g) in which R’ represents a 2,2,2-tri
`R’ represents a 2,2,Z-trichloroethoxycarbonyl radical,
`chloroethoxycarbonyl radical, the characteristics of
`the characteristics of which are as follows, is thereby
`obtained (yield=93%):
`which are as follows, is thereby obtained (yield: 55%):
`Melting point: 233°—234° C.
`[a]D=-56° (c=0.567; chloroform).
`
`45
`
`50
`
`65
`
`

`

`5
`
`25
`
`30
`
`35
`
`4, 814,470
`8
`7
`Speci?c rotation: [0.]D23=—58° (c=0.465; chloro
`The total yield of lO-deacetylbaccatin III is 300 mg
`per kg of leaves.
`form).
`Ultraviolet spectrum: 7tmax=232 nm (19000),
`kmax=276 nm (990), 7tmax=283 nm (810).
`Infrared spectrum: characteristic absorption bands at
`3420, 1765, 1730 and 1720 cm~1.
`Proton nuclear magnetic resonance spectrum
`(CDC13, shifts in ppm): 1.12 (s, 3H); 1.16 (s, 3H); 1.85 (s,
`3H); 2.16 (s, 3H); 2.30 (s, 3H); 2.30 (m, 2H); 2.05 and
`2.65 (2m, 2H); 4.00 (cl, J =7, 1H); 4.18 and 4.35 (2d,
`J =9, 2H); 4.63 and 4.92 (2d, J= 12, 21-1); 4.76 and 4.80
`(2d, J=12, 2H); 4.92 (t, J=9, 1H); 5.00 (d, J=9, 1H);
`5.61 (m, 1H); 5.66 (d, J=7, 1H); 6.30 (s, 1H); 7.50, 7.64
`and 8.13 (2: and 1d, J=7, 5H)
`Mass spectrum (chemical ionization) m/z 893
`(MH+), 875, 701, 683, 579, 387, 327, 309 and 123.
`The IO-deacetylbaccatin III may be obtained as fol
`lows:
`Ground non-dried Taxus baccata L leaves (100 kg)
`are subjected to accelerated percolation, in a rotary
`20
`device, with 95° alcohol (the true alcohol content of
`which changes to 80°—85° because of the water con
`tained in the leaves). The ?rst maceration is carried out
`with alcohol (300 liters) and the subsequent macerations
`(4>< 200 liters) are carried out with alcohol recovered
`by distillation and the alcohol level of which is main
`tained at 85°. Each percolation lasts for 10 hours and is
`carried out at a temperature in the vicinity of 20" C.
`Mixing is ensured by circulating the solvent using a
`pump.
`Each ethanolic phase is concentrated under receded
`pressure (50-60 mm Hg; 5.4 kPa). The concentrates
`from each operation (approximately 70 liters), with high
`water content, are combined and concentrated again to
`a volume of 20 liters in order to remove the residual
`alcohol.
`The extract, which is not evaporated to dryness,
`remains in an aqueous medium (20 liters) in the form of
`a solid suspension. It is taken up with methylene chlo
`ride (9 extractions with a total of 100 liters of methylene
`40
`chloride).
`The solution in methylene chloride thus obtained (87
`liters), containing the dry extract (2 kg), is concentrated
`to a volume of 5 liters.
`Chromatography is carried out.in a 24 cm diameter
`column containing silica (10.3 kg) (Zeosil: 8 kg; Celite:
`2.3 kg).
`Successive elutions are carried out, at a ?ow rate of 8
`to 9 liters/hour, with:
`methylene chloride (150 liters) (fraction 1);
`a methylene chloridezmethanol (99.5:0.5 by volume)
`mixture (150 liters) (fraction 2);
`a methylene chloride:methanol (99:1 by volume) mix
`ture (170 liters) (fraction 3) and
`a methylene chloride=methanol (98:2 by volume) mix
`ture (130 liters) (fraction 4).
`The ?rst two fractions are combined to give 1.74 kg
`of dry extract. The third fraction gives 390 g of dry
`extract. The fourth fraction gives 20 g of dry extract.
`The third fraction (390 g), which contains essentially
`the IO-deacetylbaccatin III, is chromatographed again
`on silica, eluting with a methylene chloride:methanol
`(99:1 by volume) mixture, at a ?ow rate of 4 liters/hour.
`4 fractions are thereby obtained, the most useful of
`which (154 g) gives, after concentrating and digesting in
`65
`methylene chloride, pure lO-deacetylbaccatin III (22 g).
`The mother liquors (132 g), puri?ed by chromatogra
`phy on silica, give IO-deacetylbaccatin III (8 g).
`
`EXAMPLE 2
`Operating as in Example 1, but starting with the prod
`uct (2'S, 3’R) of general formula (IV) in which R’ repre
`sents a 2,2,2-trichloroethoxycarbony1 radical, R1 repre
`sents a hydroxy radical and R2 represents a tertbutox
`ycarbonylamino radical, the product (2'S, 3’R) of gen
`eral formula (I) in which R represents a hydrogen atom,
`R1 represents a hydroxy radical and R2 represents a
`tertbutoxycarbonylamino radical, the characteristics of
`which are as follows, is obtained:
`speci?c rotation: [a]D23= ~29" (c=0.69; ethanol).
`Ultraviolet spectrum: kmax=229 nm (14700),
`kmax=275 nm (2350), hmax=282 nm (2280).
`Infrared spectrum: characteristic absorption bands at
`3580, 3440, 1740 and 1700 cm-1.
`Proton nuclear magnetic resonance spectrum
`(CDCl3-CD3OD, 400 MHz, shifts in ppm): 1.14 (s, 3H);
`1.20 (s, 3H); 1.40 (s, 9H); 1.75 (s, 3H); 1.97 (s, 3H); 2.27
`(s, 3H); 2.53 (m, 1H); 3.90 (d, J=7, 1H); 4.22 and 4.31
`(2d, J=9, 2H); 4.24 (m, 1H); 4.50 (d, J=2, 1H); 5.01 (d,
`J=9, 1H); 5.19 (d, J=2, 1H); 5.32 (s, 1H); 5.67 (d, J=7,
`1H); 6.17 (t, J=9, 1H); 7.26-7.45 (5H); 7.48, 7.62 and
`8.07 (5H).
`Mass spectrum (FAB) m/z: 808 (MH+), 752, 734,
`690, 527, 509, 449, 405, 387, 345, 327, 299, 266 and 185.
`EXAMPLE 3
`Operating as in Example 1, but starting with the prod
`uct (2’R, 3’S) of general formula (IV) in which R’ repre
`sents a 2,2,2-trichloroethoxycarbonyl radical, R1 repre
`sents a tert-butoxycarbonylamino radical and R2 repre
`sents a hydroxy radical, the product (2’R, 3’S) of gen
`eral formula (I) in which R represents a hydrogen atom,
`R1 represents a tert-butoxycarbonylamino radical and
`R2 represents a hydroxy radical, the characteristics of
`which are as follows, is obtained:
`Speci?c rotation: [a]D23= ~29“ (c=0.47; ethanol).
`Ultraviolet spectrum: Arnax=229 nm (16300),
`hmax=274 nm (2570), Amax=282 mm (2380).
`Infrared spectrum: main characteristic absorption
`bands at 3590, 3440, 2990, 1740-1700 cm“1.
`Proton nuclear magnetic resonance spectrum
`(CDC13, 400 MHz, shifts in ppm): 1.12 (s, 3H); 1.22 (s,
`3H); 1.35 (s, 9H); 1.77 (s, 3H); 1.91 (s, 3H); 2.27 (m, 2H);
`2.38 (s, 3H); 2.59 (m, 1H); 3.96 (d, J=7, 1H); 4.19 and
`4.31 (2d, J=9, 2H); 4.25 (m, 1H); 4.58 (dd, 1:9 and
`J=2, 1H); 4.97 (d, J=9, 1H); 5.22 (s, 1H); 5.35 (d, J=2,
`1H); 5.48 (d, J=9, 1H); 5.67 (d, J=7, 1H); 6.26 (t, J=9,
`1H); 7.35, 7.40 and 7.46 (5H); 7.49, 7.62 and 8.07 (5H).
`Mass spectrum (FAB) m/z: 808 (MH+), 790, 752,
`734, 708, 527, 509, 449, 405, 387, 345, 327, 282, 226 and
`185.
`
`45
`
`55
`
`60
`
`EXAMPLE 4
`Operating as in Example 1, but starting with the prod- I
`uct (2’S, 3’R) of general formula (IV) in which R’ repre
`sents a 2,2,2-trichloroethoxycarbony1 radical, R1 repre
`sents a tert-butoxycarbonylamino radical and R2 repre
`sents a hydroxy radical, the product (2’S, 3’R) of gen
`eral formula (I) in which R represents a hydrogen atom,
`R1 represents a tert-butoxycarbonylamino radical and
`R2 represents a hydroxy radical, the characteristics of
`which are as follows, is obtained:
`Speci?c rotation: [a]D23= —33° (c=0.81; ethanol).
`
`

`

`4,814,470
`10
`The following Example illustrates a composition
`according to the invention.
`
`COMPOSITION EXAMPLE
`The product of formula I obtained in Example 1 (40
`mg) is dissolved in Emulphor EL 620 (1 cc) and ethanol
`(1 cc) and the solution is then diluted by adding physio
`logical saline (18 cc).
`This composition may be administered by introduc
`tion into an intravenous perfusion of physiological sa
`line given over a period of 1 hour.
`We claim:
`1. A taxol derivative of formula:
`
`RO
`
`5
`
`15
`
`20
`
`9
`Ultraviolet spectrum: kmax=230 nm (14250),
`kmax=275 nm (1380), kmax=282 nm (1270).
`Infrared spectrum: characteristic absorption bands at
`3580, 3440, 2900, 1740-1700 cm-l.
`Proton nuclear magnetic resonance spectrum
`(CDCl3, 400 MHZ, shifts in ppm): 1.12 (s, 3H); 1.22 (s,
`3H); 1.36 (s, 9H); 1.72 (s, 3H); 1.94 (s, 3H); 2.32 (s, 3H);
`2.51 (m, 1H); 3.85 (d, J=7, 1H); 4.20 and 4.29 (2d, J=9,
`2H); 4.22 (m, 1H); 4.58 (dd, 1:2 and J=9, 1H); 4.97 (d,
`J=9, 1H); 5.14 (d, J=2, 1H); 5.22 (s, 1H); 5.65 (d, J=7,
`1H); 5.81 (d, J=9, 1H); 6.17 (t, J=9, 1H); 7.37 (5H);
`7.50, 7.63 and 8.07 (5H)
`Mass spectrum (FAB) m/z: 808 (MH'l'), 752, 740,
`708, 690, 549, 527, 509, 449, 405, 387, 345, 327, 299, 226
`and 185.
`The present invention also provides pharmaceutical
`compositions containing a compound of formula (I) in
`combination with one or more pharmaceutically ac
`ceptable, inert or physiologically active, diluents or
`adjuvants.
`These compositions may be presented in any form
`appropriate for the administration route envisaged. The
`parenteral route, and especially the intravenous route, is
`25
`the preferential route for administration.
`The compositions according to the invention for
`parenteral administration may be aqueous or nonaque
`ous sterile solutions, suspensions or emulsions. Propy
`lene glycol, vegetable oils, especially olive oil, and in
`jectable organic esters, e. g. ethyl oleate, may be used as
`the solvent or the vehicle. These compositions may also
`contain adjuvants, especially wetting agents, emulsi?ers
`or dispersants. The sterilization may be carried out in
`several ways, e.g. using a bacteriological ?lter, by in
`corporating sterilizing agents into the composition, by
`irradiation or by heating. They may also be in the form
`of sterile solid compositions which may be dissolved or
`dispersed in sterile water or any other injectable sterile
`medium.
`The products of general formula (I) are more particu
`larly used in the treatment of actue leukaemias and solid
`tumours, at daily doses which are generally between 1
`and 2 mg/kg by the intravenous (perfusion) route for an
`adult.
`
`in which R represents hydrogen or acetyl and one of
`R1 and R2 represents hydroxy and the other represents
`tert-butoxycarbonylamino, and the stereoisomeric
`forms thereof and their mixtures.
`2. A taxol derivative according to claim 1, in which R
`is hydrogen, R1 is hydroxy and R2 is tert-butoxycar
`bonylamino, and having the 2’R, 3’S configuration.
`3. A taxol derivative according to claim 1, in which R
`is hydrogen, R1 is hydroxy, and R2 is tert-butoxycar
`bonylamino, and having the 2'S, 3’R con?guration.
`4. A taxol derivative according to claim 1, in which R
`is hydrogen, R1 is tert-butoxycarbonylarnino, and R2 is
`hydroxy, and having the 2’R, 3'S configuration.
`5. A taxol derivative according to claim 1, in which R
`is hydrogen, R1 is tert-butoxycarbonylamino, and R2 is
`hydroxy and having the 2'S, 3’R con?guration.
`6. A pharmaceutical composition which contains a
`taxol derivative as claimed in claim 1 combined with
`one or more pharmaceutically acceptable, inert or phys
`iologically active diluents or adjuvants.
`1.!
`i * 1‘
`*
`
`30
`
`35
`
`40
`
`45
`
`50
`
`55
`
`60
`
`65
`
`

`

`UNITED STATES PATENT AND TRADEMARK OFFICE
`CERTIFICATE OF CORRECTION
`
`PATENTNO. :
`DATED
`;
`
`4,814,470
`March 21, 1989
`
`INVENTOR(S) :
`
`Michel COLIN et al.
`
`Page 1 of 4
`
`ll ls certi?ed that error appears in the above-indenti?ed patent and that said Letters Patent is hereby
`corrected as shown below:
`
`Column 1, Formula (l):
`
`:10
`
`0
`
`OH
`
`(I)
`
`Column 1, Formula (II):
`
`cmco-o
`
`0H
`0
`ll CH) Y
`
`(m
`
`4 OCOCH;
`
`

`

`UNITED STATES PATENT AND TRADEMARK OFFICE
`CERTIFICATE OF CORRECTION
`
`PATENTNO. :
`DATED
`;
`
`4,814,470
`March 21, 1989
`
`INVENTOR(S) :
`
`Michel COLIN et al.
`
`Page 2 of 4
`
`It is certi?ed that error appears in the above-indenti?ed patent and that said Letters Patent is hereby
`corrected as shown below:
`
`Column 2, Formula (Ill):
`
`R'O
`
`O
`
`OCOOCHICCI]
`
`(III)
`
`(111')
`
`3Com,
`
`Coiumn 2, Formula (IV):
`
`

`

`UNITED STATES PATENT AND TRADEMARK OFFICE
`CERTIFICATE OF CORRECTION
`
`PATENTNO. ;
`DATED
`
`4,814,470
`March 21, 1989
`
`lNvEN-roms) I
`
`Michel COLIN et a1.
`
`Page 3 of 4
`
`It is certified that error appears in the above-indentified patent and that said Letters Patent is hereby
`corrected as shown below:
`
`Column 2, Formula (V):
`
`R‘O
`
`O
`
`OCOOCHICCI;
`
`(V)
`
`

`

`UNITED STATES PATENT AND TRADEMARK OFFICE
`CERTIFICATE OF CORRECTION
`
`PATENT NO. :
`DATED
`:
`
`4,314,470
`March 21, 1989
`
`INVENTOFKS) 2
`
`Michel COLIN et a1.
`
`Page 4 of 1;
`
`it is certified that error appears in the above-indentified patent and that said Letters Patent is hereby
`corrected as shown below:
`
`IN THE CLAIMS:
`
`Claim 1, please amend the formula as follows:
`
`E
`UC
`
`ococn,
`
`Signed and Sealed this
`
`Twenty-ninth Day of October 1996
`
`Am“
`
`60%! W
`
`AUESIII’ZS‘ O?icer
`
`(‘vmmimmner (if PHIL’I‘HS and Trademarks
`
`BRL'CE LEHMAN
`
`

`

`UNITED STATES PATENT AND TRADEMARK OFFICE
`
`CERTIFICATE EXTENDING PATENT TERM
`UNDER 35 U.S.C. § 156
`
`PATENT NO.
`
`ISSUED
`
`INVENTOR(S)
`
`1
`
`2
`
`:
`
`4,814,470
`
`March 21, 1989
`
`Michel Colin et al.
`
`PATENT OWNER :
`
`Rhone-Poulenc Rorer S.A.
`
`This is to certify that there has been presented to the
`
`COIVIMISSIONER OF PATENTS AND TRADEMARKS
`
`an application under 35 U.S.C. § 156 for an extension of the patent term. Since it
`appears that the requirements of the law have been met, this certi?cate extends the term of
`the patent for the period of
`
`1,035 days
`
`from July 14, 2007, the original expiration date of the patent, with all rights pertaining
`thereto as provided by 35 U.S.C. § 156(b).
`
`I have caused the seal of the Patent and Trademark
`Office to be af?xed this 2_Qth_day of HOJLQLRILGLIQQZ.
`
`@(q/ % I (067k
`
`Bruce A. Lehman
`Assistant Secretary of Commerce and
`Commissioner of Patents and Trademarks
`
`