WO1995025728A1

WO1995025728A1 - Taxane derivatives with antitumor activity

Info

Publication number: WO1995025728A1
Application number: PCT/EP1995/000918
Authority: WO
Inventors: Nicola Mongelli; Giovanni Biasoli; Maria Menichincheri; Domenico Fusar Bassini; Marina Ciomei
Original assignee: Pharmacia S.P.A.
Priority date: 1994-03-18
Filing date: 1995-03-13
Publication date: 1995-09-28
Also published as: AU1950495A; GB9405400D0; IL112998A0

Abstract

Taxane derivatives of formula (I) are disclosed, characterized by the presence at position 2' and/or 7 of the taxane structure of an alkylating residue, optionally with a spacer consisting of an amino acid or a peptide residue. These compounds are useful as antitumor agents. A process for their preparation is also described.

Description

Taxane derivatives with antitumor activity

The present invention is .directed to taxane derivatives endowed with antitumor activity, to a method for their preparation and to pharmaceutical compositions containing them.

The taxane family of diterpenes includes Paclitaxel (also named taxol in several publications) , isolated and characterized from an extract of bark of Taxus brevifolia L. , and Cephalomannine (see J. Chem. Soc. Chem. Commun. 102, 1979) ; other taxane analogues are also known and were prepared by semisynthesis starting from 10-deacetyl baccatin III, extracted from the needles of Taxus baccata L. , see ani et al. in JACS, 93, 2325 (1971) and ovelle et al., Proc. Am. Assoc. Cancer Res. 31, p. 417 (1990) . These compounds have been shown to possess antitumor activity, but there is a continuous need for more potent derivatives with better pharmacodistribution. The present invention provides a taxane derivative of the formula I:

wherein

A and B, which are the same or different, are each, independently, a single bond, an amino acid residue or a peptide spacer; R is a phenyl, t-butoxy or 1-methyl-l-propenyl group; R₃ is hydrogen or an acetyl group; and either

(i) R_! is hydrogen and R₂ is an alkylating residue having a formula selected from: a) -CO-(CH₂)_m-R₄ where m is 0, 1, 2, 3 or 4 and R₄ is halogen, oxiranyl, methyloxiranyl, aziridinyl.

cyclopropyl, β-lactone, ) or 7-lactone

b) -CO-CX=CH, where X is Cl, Br, N0₂ or CN; c) -CO-Ph-p-N(CH₂CH₂Cl)₂; d) -CO-CH(NH₂) CH₂-Ph-p-N(CH₂CH₂C1)₂; e) -CO-(CH₂)₃-Ph-p-N(CH₂CH₂Cl)₂; f) -CO(CH₂)_nN(CH₂CH₂Cl)₂ wherein n=0-5; or

(ii) R₂ is hydrogen and R, is an alkylating residue as defined above; or (iii) R, and R₂, which may be the same or different, are both an alkylating residue as defined above with the exclusion of 2'-chloroacetyltaxol. The peptide spacer may be from 1 to 10, for example 2 to 4, amino acid residues long. The spacer should be susceptible to intracellular lysosomal hydrolysis. The spacer may be resistant to extracellular hydrolysis. Examples of preferred peptide spacers include βAla, Gly, Phe-Gly, Phe-Phe-, Leu- Gly, Val-Ala, Phe-Ala, LeurPhe, Leu-Ala, Phe-Leu-Gly, Phe- Phe-Leu, Leu-Leu-Gly, Phe-Tyr-Ala, Phe-Gly-Phe, Phe-Phe-Gly, Phe-Leu-Gly-Phe, Gly-Phe-Leu-Gly-Phe, Gly-BAla, Phe-Gly-BAla, Phe-Phe-βAla, Leu-Gly-βAla, Val-Ala-βAla, Phe-Ala-βAla, Leu- Phe-βAla, Leu-Gly-βAla, Phe-Leu-Gly-βAla, Phe-Phe-Leu-βAla, Leu-Leu-Gly-βAla, Phe-Tyr-Ala-βAla, Phe-Gly-Phe-βAla, Phe- Phe-Gly-βAla, Phe-Leu-Gly-Phe-βAla and Gly-Phe-Leu-Gly-Phe- βAla.

The wavy line in formula (I) denotes that the oxygen linked at position 7 of the taxane structure may be in either configuration, i.e. β (natural) or α. Preferably R represents a phenyl group, R₃ is an acetyl group, A and B represent independently the amino acid residue β-Ala or the peptide residue Leu-Gly, Leu-Gly-βAla, Phe-Leu- Gly or Phe-Leu-Gly-βAla. The preferred alkylating residues which R, and R₂ may represent include:

b') -CO-CBr=CH₂, C0-CC1=CH₂

C) -CO-Ph-p-N(CH,CH₂Cl)₂

More particularly preferred compounds according to the present invention are the following ones:

1) 2'-α-bromoacryloyltaxol

2) 2'-p-(N,N-bis-(2-chloroethy1)amino)benzoyltaxol 3) 7-chloroacetyltaxol

4) 2'-/?-(α-bromoacryloyl)alanyltaxol

5) 7-α-bromoacryloyltaxol

6) 7-p-(N,N-bis-(2-chloroethyl) amino)benzoyltaxol 7) 7-/3-(α-bromoacrylogyl) alanyltaxol

8) 7-3-(chloroacetyl)alanyltaxol

9) 7-/3-(glicydyl)alanyltaxol

10) 7-(N,N-bis-(2-chloroethyl)glicyl)taxol

11) 7-/3-(N,N-bis-(2-chloroethyl)glicyl)alanyltaxol 12) 7-bromoacetyl-taxol

13) 7-cyclopropylcarbonyl-taxol

The present invention also provides a process for preparing a compound of the formula I as above defined, which process comprising reacting an optionally protected taxane derivative of the formula T

wherein R, R₃, A and B are as defined above and R', and R'₂ are each, independently, hydrogen or a protecting group, with a compound of the formula III

X-R_Λ (III) wherein X is a leaving group and R_A is an alkylating residue as defined above for R, and R , and then removing, if necessary, the protecting group.

X may be any conventional leaving group, for instance a halogen such as Br or Cl, preferably Cl.

Examples of process embodiments are as follows: 1) The reaction of a compound of the formula II

where R and R₃ are as defined above, with a compound of formula III, as defined above, gives a compound of formula

I wherein A and B are single bonds and R_t is either an alkylating residue as defined above and R₂ is hydrogen, or both R, and R₂ are alkylating residues.

2) The reaction of a compound of the formula IV

where R and R₃ are as defined above and D is an aminoacid residue or a peptide spacer as defined above, with a compound of formula III, as defined above, gives a compound of formula I wherein A is an aminoacid residue or a peptide spacer as defined above, R, is an alkylating residue as defined above, B is a single bond and R₂ is hydrogen. 3) The reation of a compound of the formula V

where R and R₃ are as defined above, G is a hydroxy protecting group or a group G' which is a protected aminoacid residue or a peptide spacer as defined above, with a compound of formula III as defined above, and removal of the hydroxy protecting group or the protected amino acid residue or peptide, gives a compound of formula I wherein A and B are single bonds, R, is hydrogen and R₂ is an alkylating residue as defined above. 4) The reaction of a compound of formula VI

where R and R₃ are as defined above, E is an amino acid residue or a peptide spacer as defined above, with a compound of formula III, as defined above, gives a compound of formula I wherein A is a single bond, Rj is hydrogen, B is an amino acid residue or a peptide spacer as defined above and R₂ is an alkylating residue as defined above. 5) The reaction of a compound of formula V, where R and R₃ are as defined above, G is G' as defined above, with a compound of formula III, as defined, and subsequent removal of the amino acid or peptide protecting group, gives a compound of formula I wherein A is an amino acid residue or a peptide spacer as defined above, Rj is hydrogen, B is a single bond and R₂ is an alkylating residue as defined above. 6) The reaction of a compound of the formula VII

where R, G', E and R₃ are as defined above, with a compound of formula III, as defined above, and subsequent removal of the amino acid or peptide protecting group, gives a compound of formula I wherein A and B are amino acid residues or peptide spacers as defined above, R, is hydrogen and R₂ is an alkylating residue as defined above. 7) The reaction of a compound of formula VIII

where D, R and R₃ are as defined above, E is an amino acid residue or a peptide spacer such as defined above, with a compound of formula III, as defined above, gives a compound of formula I wherein A and B are amino acid residues or peptide spacers as defined above, and Rj and R₂ are alkylating residues as defined above. Compounds prepared according to process embodiment 1) are obtained by reacting a compound of formula II (e.g. taxol) with the activated acyl derivative of the corresponding alkylating agent (e.g. acyl chloride, anhydride, mixed anhydride, imidazolide, acyl isourea, etc.) in a suitable organic solvent (e.g. methylene chloride, acetonitrile, dioxane, etc.) in the presence of a base (e.g. triethylamine, pyridine, dimethylaminopyridine, sodium or potassium bicarbonate, etc.). A large excess of the alkylating agent and of the base is necessary, in order to obtain the taxane derivative bearing two alkylating residues.

The reaction is typically carried out at a temperature varying from 0°C to 70°C and the reaction time may vary from about 1 to about 24 hours. Compounds prepared by process embodiment 2) are obtained by reacting a compound of formula IV with the activated acyl derivative of the corresponding alkylating agent (e.g. acyl chloride, anhydride, mixed anhydride, imidazolide, acyl isourea, etc.) in a suitable organic solvent (e.g. methylene chloride, acetonitrile, dioxane, etc.) in the presence of a base (e.g. triethylamine, pyridine, dimethylaminopyridine, sodium or potassium bicarbonate, etc.). The reaction is typically carried out at a temperature varying from 0°C to 40°C and the reaction time may vary from about 1 to 7 hours. The compounds of formula IV may be prepared by reacting taxol or a taxol analog with protected amino acid or peptide in the presence of a condensating reagent, and with or without the additional presence of a base (e.g. triethylamine, pyridine, dimethylaminopyridine etc) , preferably at room temperature, followed by the removal of the protecting group with known methods. Any known amino protecting groups can be utilized and commercially available protected amino acids or peptides can be utilized as the starting materials. For example, amino acids or peptides protected with t-Boc, trityl, FMOC or carbobenzyloxy (CBZ) can be utilized. Amino acids or peptides protected with t-Boc, trityl or FMOC groups are preferred. The protecting group may be removed by conventional techniques, for instance hydrolysis or hydrogenolysis. Compounds prepared by process embodiment 3) are obtained by reacting a compound of formula V with the activated acyl derivative of the corresponding alkylating agent (e.g. acyl chloride, anhydride, mixed anhydride, imidazolide, acyl isourea, etc.) in a suitable organic solvent (e.g. methylene chloride, acetonitrile, dioxane, dimethylformamide, etc.) in the presence of a base (e.g. triethylamine, pyridine, dimethylaminopyridine, sodium or potassium bicarbonate, etc. ) .

Compounds of the formula V wherein G is a hydroxy protecting group are already known in the literature (e.g. triethyl silyl group, acetyl, allyloxycarbonyl) , those wherein G is a protected aminoacid or peptide may be prepared as described above for the compounds of the formula IV (of case 2) , without the final deprotect.ion.

Finally the removal either of the hydroxy protecting group or of the protected amino acid or peptide residue affords the expected compounds of case 3) .

For example the 2'-triethylsilyl group may be removed with diluted hydrochloric acid in the tetrahydrofuran or with pyridine-hydrofluoric acid in the tetrahydrofuran or with piridine-hydrofluoric acid in acetonitrile etc.

Removal of the amino acid or peptide in 2' may be carried out with NaHC0₃ in H₂0/MeOH for 2-5 hours

Compounds prepared by process embodiment 4) are obtained by reacting a compound of formula VI with a mildly activated acyl derivate of the corresponding alkylating agent (e.g. anhydride, mixed anhydride, acyl isourea) in a suitable organic solvent (e.g. methylene chloride, acetonitrile, dioxane, etc.) in the presence of a base (e.g. triethylamine, pyridine, etc.). The reaction is typically carried out at a temperature varying from 0°C to 30°C. The reaction time may vary from about 1 to 7 hours.

Compounds of formula VI are prepared by acylation of 2'- protected taxol or a taxol analog with a protected amino acid or peptide in the presence of a condensing agent, then by removal of the protecting groups both of the amino acid or peptide residue and of the 2'-position residue as described above. Compounds prepared according to process embodiment 5) are obtained as described for compounds of process embodiment 3) without final removal of the aminoacid or peptide residue, but just removing the aminoacid or peptide protecting group as described above.

Compounds prepared according to process embodiment 6) are obtained by reacting a compound of formula VII with an activated acyl derivative of the corresponding alkylating agent (e.g. acyl chloride, anhydride, mixed anhydride, imidazolide, acyl isourea, etc.) in a suitable organic solvent (e.g. methylene chloride, acetonitrile, dioxane, etc.) in the presence of a base (e.g. triethylamine, pyridine, dimethylaminopyridine, sodium or potassium bicarbonate, etc.). The reaction is typically carried out at a temperature ranging from 0°C to 40°C and the reaction time may vary from about 1 to 4 hours.

Finally the removal of the amino acid protecting group at the 2'-position affords the expected compounds of process embodiment 6) . Compounds prepared according to process embodiment 7) are obtained by reacting a compound of formula VIII with an activated acyl derivative of the corresponding alkylating agent (e.g. acyl chloride, anhydride, mixed anhydride, imidazolide, acyl isourea, etc.) in a suitable organic solvent (e.g. methylene chloride, acetonitrile, dioxane, etc.) in the presence of a base (e.g. triethylamine, pyridine, dimethylaminopyridine, sodium or potassium bicarbonate, etc.). The reaction is typically carried out at a temperature ranging from 0°C to 40°C and the reaction time may vary from about 1 to 4 hours.

The starting compounds of the formulae II, III, VII and VIII are known compounds or may_, be prepared by known or analog procedure.

Biological activity

The cytotoxic activity of the compounds has been evaluated on B16-F10 murine melanoma cell line, which was responsive to taxol (1) . The mode of action of the compound was tested on tubulin assembly-disassembly assay in comparison with taxol (2) . Because the compound here described can differ from taxol by forming irreversible binding to the tubulin, a new assay that can discriminate between reversible and irreversible binding, has been set up (3) .

1) In vitro drug sensitivity assay.

Exponentially growing murine melanoma cells (2 x 10⁴/ml B16- F10 and 4 x 10⁴/ml B16-F10/TAX, which are resistant to taxol and maintained in the presence of 50nM taxol) were seeded in RPMI 1640 medium supplemented with 10% heat-inactivated fetal calf serum and 2mM glutamine in 24 well-plates (Costar) . UV2237 murine fibrosarcoma cells were seeded (2 x IO⁴ /ml) in E-MEM medium with non-essential aminoacids supplemented with 1% vitamins, 10% heat-inactivated fetal calf Serum and 2mM glutamine in 24 well-plates. Scalar concentrations of tested compounds were added immediately after seeding. The inhibition of cell growth was evaluated by counting cells with a coulter counter after 72 hours incubation. For each tested compound concentration triplicate cultures were used. The antiproliferative activity of the tested compounds was calculated . from dose-response curves and expressed as IC₅₀ (dose causing 50% inhibition cell growth in the treated cultures relative to untreated controls) . The results are shown in Table 1. 2) Microtubule assembly and disassembly assay Calf brain tubulin was prepared by two cycles of assembly- disassembly (Shelanki M.L., Gaskin F. and Cantor CR. , Proc. Natl. Acad. Sci. U.S.A. 70, 765-768, 1973) and stored in liquid nitrogen in MAB (0.1 M MES, 2.5 mM EGTA, 0.5 mM MgSO₄,Mg80₄ 0.1 mM EDTA, 0.1 mM DDT pH 6.4).

All the experiments were carried out on protein stored for less than 4 weeks. Before each experiment, the tubulin was kept 30 min at 4°C. Assembly was monitored by method of Gaskin et al (Gaskin F. , Cantor CR. and Shelanski M.L. J. Molec. Biol. 89, 737-758, 1974) . The cuvette (1 cm path) containing tubulin (lmg/ml) and 1 mM GTP was shifted to 37°C and continuous turbidity measurements were made at 340 nm on Perkin-Elmer 557 Double Wavelength Double Beam Stectrophometer equipped with an automatic recorder and a ter ostatically regulated sample chamber. After 30 minutes 4 mM CaCl₂ was added and depolymerisation was measured for 10 minutes as decreased turbidity. At regular intervals of 15 minutes scalar doses of tested compounds were added and variations in the turbidity were monitored. Data are expressed as Dose Effecting tubulin ripolymerisation by 50% (ED₅₀) induced by the tested compounds. The results are shown in Table 2. As already reported in literature, compounds with moieties bound to position 2' resulted inactive in this assay and are not presented in the Table.

3) Irreversible tubulin binding assay.

Calf brain tubulin was left to assembly in MAB plus 1 mM GTP for 20 min at 37°C as described above. At the end, disassembly of microtubules was obtained by adding 4 mM CaCl₂.

The tested compound was then added at the concentration that permits 40-60% ripolymerization in the assembly-disassembly assay and the microtubules formation was monitored for 20 min.

The sample was kept 40 min at 0°C and then centrifuged at 4°C for 60 min at 5'000 x g in Ultrafree-MC filter Units, PLGC low-protein binding regenerated cellulose membrane NMWL: 10,000. The ability of the unfiltered material to riassembly in microtubules was monitored at 37°C in MAB plus ImM GTP. After 20 min, the sensitivity microtubules to CaCl₂ addition was controlled.

CaCl₂ was able to disassembly the microtubules only if the compound has been removed by centrifugation. If no effect was observed after CaCl₂ addition, it was in agreement with an irreversible binding of the tested compound to the tubulin. Results are reported in table 2. Table 1: Citotoxicity on B16-F10, B16F10/TAX and UV2237 cells

IDso(nM)²

Compound" R,-A R₂-B B16F10 B16F10/ UV2237 TAX

Taxol H H 35±3 629±80 452±78

FCE 28567 bromoacryloyl H 24±1 255 141+15

FCE 28616 chloroacetyl H 9+1 67±13 51±2

FCE 28642 chloroacetyl chloroacetyl 9±2 18+0 40±7

FCE 28514 H chloroacetyl 27±4 137+18 56±10

FCE 28573 H α-bromoacryloyl- 17+4 38±13 57±14 βalanyl

FCE 28975 H bromoacetyl 46±5 361±23 295+9

FCE 29050 H cyclopropyl- 49±5 — — carbonyl

Table 2: Activity in the tubulin assembly test

Compound¹ R.-A R₂-B ED₅₀(μM)²⁾ TYPE OF BINDING

Taxol H H 0.8 REV.

FCE 28514 H chloroacetyl 1.8 IRREV.

FCE 28975 H bromoacetyl 0.8 IRREV.

FCE 29050 H cyclopropyl- 0.6 IRREV. carbonyl

1) R₂=CH₃CO, R=Ph

2) ID₅₀=Dose Effecting Cell Growth after 72 h treatment ED₅₀=Dose Effecting tubulin riassembly by 50% at 37°C REV=Reversible IRREV=Reversible

A human or animal suffering from a tumour may thus be treated by a method which comprises the administration thereto of a taxane derivative of formula (I) as defined above or 2'- chloroacetyltaxol. The condition of the human or animal may thereby be improved.

The use of 2'-chloroacetyltaxol as therapeutic agent is another object of the invention. The invention also provides a pharmaceutical composition containing a compound of formula (I) as defined above or 2'-chloroacetyltaxol as an active substance in association with one or more pharmaceutically acceptable excipients. The composition of the invention is usually prepared following conventional methods and is administered in a pharmaceutically suitable form. Administration can be made by any of the accepted ways for administration of antitumor agents such as, e.g. intravenous, intramuscular or subcutaneous injection, or topical application. For systemic injection the active compound may be, e.g. dissolved in a vehicle consisting of polyoxyethylated castor oil (Cremophor EL) 50% and ethanol 50% and then diluted with glucose 5% solution at the desidered concentration, or other pharmaceutically suitable carriers. The amount of the active compound administered depends on the treated subject, for example age, weight and sex, and the severity of affliction. The method of administration depends on the judgement of the prescribing physician. A suitable dosage for an average 70 kg human may range from about 0.01 g to about 1000 mg per day.

The following examples illustrate the invention but they are not intended to limit it thereto. Example 1 2'-(carbobenzyloxy-βalanyl) ,7-chloroacetyl-taxol (FCE 28464).

To a solution of 106 mg of 2 '- (carbobenzyloxy-βalanyl) taxol (0.1 mmols) in 10 ml of dichloromethane, were added 24 mg of dimethylaminopyridine (0.2 mmols) , 0.042 ml of triethylamine (0.3 mmols) and 0.024 ml of chloroacetylchloride (0.3 mmols) . The reaction mixture was stirred for 4 hours at room temperature, washed with diluted hydrochloride acid (0.1 N) , water and dried on anhydrous sodium sulphate.

The dichloromethane was evaporated under vacuum and the residue was chromatographed on silica gel with ethyl acetate- hexane 1:1 as eluant, affording 60 mg of the title compound.

FAB-MS: m/z 1133, M- tT^", 489. Η-NMR (400 MHz, CDC1-,) : δ 1.15 (s, 3H, 16) ; 1.21 (s, 3H,

17); 1.75 (Ξ, 1H, OH-1) ; 1.82 (s, 3H, 19) ; 1.98

(d, J=1.2 Hz, 3H, 18); 1.8-2.0 (m, 1H, 6β) ; 2.16

(s, 3H, CH₃CO-IO) ; 2.17 (dd, J=9.1 Hz, J=15.4 Hz,

1H, 14) ; 2.37 (dd, J=9.1 Hz, J=15.4 Hz, 1H, 14); 2.47 (s, 3H, CH₃CO-4) ; 2.4-2.6 ( , 3H, 6α +

NHCHCHoCO) ; 3.4-3.7 (m, 2H, NHCH₂CH₂CO) ; 3.95 (d,

J=7.0 Hz, 1H, 3); 3.98-4.21 (two doublets, J=15.1

Hz, 2H, C0CH₂C1) ; 4.19 (d, J=8.5 Hz, 1H, 20β) ; 4.33

(d, J=8.5 Hz, 1H, 20 ) ; 4.88 (s, 2H, COOCH₂Ph) ; 4.97 (d, J=8.5 Hz, 1H, 5) ; 5.17 (t, J=6.3 Hz, 1 H,

NHCH₂CH₂CO) ; 5.51 (d, J=3.3 Hz, 1 H, 2'); 5.67 (m,

2K, 2+7) ; 6.01 (dd, J=3.3 Hz, J=9.1 Hz, 1H, 3'); 6.18 (s, IH, 10) ; 6.22 (ddd, J=9.1 Hz, J=9.1 Hz, J=1.2 Hz, IH, 13) ; 7.2-8.2 (m, 21H, 4-| (~ \+ NH- 4')

By analogous procedure the following compounds can be obtained:

2'-(carbobenzyloxy-βalanyl) ,7-(α bromoacryloyl) -taxol (FCE 28483) .

FAB-MS: m/z 1189, M-H^"1", 657, 489. Η-NMR (400 MHz, CDC1₃) : <S 1.16 (s, 3H, 16) ; 1.20 (s, 3H, 17) ; 1.87 (s, 3H, 19) ; 1.8-2.0 (m, IH, 6β) ; 2.00 (s, 3H, 18) ; 2.13 (s, 3H, CH₃CO-10) ; 2.17 (dd, J=15.5, J=9.1 Hz, IH, 14) ; 2.38 (dd, J=15.5, J=9.1 Hz, IH, 14) ; 2.47 (s, 3H, CH₃CO- 4) ; 2.5-2.8 ( , 3H, 6a + OCOCH₂CH₂NH) ; 3.4-3.7 (m, 2H, OCOCH_?CH_:,NH. ;

3.98 (d, J=7.0 Hz, IH, 3) ; 4.21 (d, J=8.5 Hz, IH, 20β) ; 4.34 (d, J=8.5 Hz, IH, 20α) ; 4.89 (ε, 2H, COOCH₂ Ph) ; 4.97 (d, J=9.1 Hz, IH, 5) ; 5.17 (t, J=6.2 Hz, IH, OCOCH₂CH₂NH) ; 5.51 (d, J=3.2 Hz, IH, 2') ; 5.62 (dd, J=7.3 Hz, J=10.5 Hz, IH, 7) ; 5.71

(d, J=7.0, IH, 2) ; 6.02 (dd, J=3.2 Hz, J=8.8 Hz, IH, 3') ; 6.22 (t, J=9.1 Hz, IH, 13) ; 6.29 (s, IH, 10) ; 6.25-6.83 (two doublets, J=1.8 Hz, 2H,=CH₂) ; 7.2-8.2 (m, 21H, 4-Ph + NH-4 ' ) . Example 2 2'-(β alanyl) ,7-chloroacetyl-taxol (FCE 28482)

To a solution of 50 mg of 2' (carbobenzyloxy-βalanyl) 7- chloroacetyl taxol in 6 ml of methanol and 4.5 ml of formic acid, were added 80 mg of Pd/C 5%.

The reaction mixture was stirred for 4 hours at room temperature. The catalyst was filtered, washed with methanol and the solvents were evaporated to dryness under vacuum. The residue was dissolved in methanol, diluted with toluene and evaporated, affording 39 mg of the title compound. FAB-MS: m/z 1001, M+H^{"1 +}; 585; 357.

Η-NMR (400 MHz, CDC1₃) : δ 1.13 (s, 3H, 16) ; 1.18 (s, 3H,

17) ; 1.80 (s, 3H, 19); 1.92 (d, J=1.5 Hz, 3H, 18); 1.8-2.0 (m, IH, 6β) ; 2.14 (s, 3H, CH₃CO-10) ; 2.44

(s, 3H, CH₃CO-4); 2.2-2.8 (m, 5H, CH₂-14 + 6a +

NH₂CH₂CH,CO) ; 3.0-3.3 (m, 2H, NH₂CH₂CH₂CO) ; 3.88 (d,

J=6.7 Hz, IH, 3); 3.97-4.19 (two doublets, J=14.9

Hz, 2H, C0CH₂C1) ; 4.17 (d, J=8.5 Hz, IH, 20β) ; 4.31 (d, J=8.5 Hz, IH, 20α) ; 4.95 (d, J=9.4 Hz, IH, 5);

5.52 (d, J=5.0 Hz, IH, 2') ; 5.5-5.7 (m, 2H, 7+2) ;

5.93 (dd, J=5.0 Hz, J=8.5 Hz, IH, 3') ; 6.0-6.2 (m,

2H, 13 + 10) ; 7.2-8.2 (m, 15H, 3-Ph) ; 8.14 (d,

J=8.5 Hz, IH, NH-4'); 8.25 (bs, IH, HCOOH) . Example 3 7-chloroacetyl-taxol (FCE 28514)

A solution of 23 mg of 2'-(βalanyl) , 7-chloroacetyl-taxol in 10 ml of methanol was stirred for 3 hours at room temperature. The methanol was evaporated under vacuum and the residue was chromatographed on silica gel with ethyl acetate- hexane 1:1 as eluant, affording 14 mg of the title compound.

FAB-MS: m/z 930, M+H^{"1 +} ; 357. Η-NMR (400 MHz, CDC1₃) : δ 1.15 (s, 3H, 16) ; 1.21 (s, 3H,

17) ; 1.81 (d, J=1.2 Hz, 3H, 18) ; 1.82 (s, 3H, 19) ;

1.88 (m, IH, 6β) ; 2.17 (s, 3H, CH₃CO-10) ; 2.2-2.4

(m, 2H, CH₂-14) ; 2.38 (s, 3H, CH₃CC~4) ; 2.64 (m,

IH, 6a ) ; 2.6 (bs, IH, OH-2' ) ; 3.91 (d, J=6.7 Hz, IH, 3) ; 3.99,4.21 (two doublets, J=14.9 Hz, 2H,

C0CH₂C1) ; 4.18 (d, J=8.2 Hz, IH, 20β) ; 4.32 (d,

J=8.2 Hz, IH, 20α) ; 4.80 (d, J=2.6 Hz, IH, 2') ;

4.95 (d, J=8.2 Hz, IH, 5) ; 5.61 (dd, J=7.3 Hz,

J=10.5 Hz, IH, 7); 5.67 (d, J=6.7 Hz, IH, 2); 5.80 (dd, J=2.6 Hz, J=8.8 Hz, IH, 3'); 6.14 (s, IH.,

10); 6.18 (m, IH, 13); 7.06 (d, J=8.8 Hz, IH, NH-

4') ; 7.3-8.2 (m, 15 H, 3-Ph) . Example 4 2'-(α bromo-acryloyl) -taxol (FCE 28567)

To a solution of 85 mg of taxol (0.1 mmols) in 6 ml of acetonitrile, were added 30 mg of α bromo-acrylic acid (0.2 mmols), 41 mg of N,N'-dicyclohexylcarbodiimide (0.2 mmols) and 5 mg of dimethylaminopyridine.

The reaction mixture was stirred for 2 hours at room temperature, the precipitate was filtered and the solvent evaporated under vacuum to dryness.

The residue was chromatographed on silica gel with ethyl acetate-hexane 1:1 as eluant, affording 51 mg of the title compound.

FAB-MS: m/z 984, M-H]^"; 525, 284.

Η-NMR (400 MHz, CDCl₃) : <5^" 1.14 (s, 3H, 16); 1.23 (ε, 3H, 17) ; 1.68 (s, 3H, 19) ; 1.94 (d, J=1.2 Hz, 3H, 18) ; 1.8-2.0 (m, IH, 6β) ; 2.23 (s, 3H, CH₃CO-10) ; 2.1- 2.4 ( , 2H, CH₂-14) ; 2.45 (s, 3H, CH₃CO-4); 2.57 (m, IH, 6a) ; 3.81 (d, J=7.0 Hz, IH, 3) ; 4.20 (d,

J=8.5, IH, 20B) ; 4.33 (d, J=8.5 Hz, IH, 20α) ; 4.44 (dd, J=6.5 Hz, J=11.0 Hz, IH, 7) ; 4.98 (dd, J=2.3 Hz, J=9.7 Hz, IH, 5) ; 5.47 (d, J=3.0 Hz, IH, 2') ; 5.69 (d, J=7.0 Hz, IH, 2) ; 5.99 (dd, J=3.0 Hz, J=9.4 Hz, IH, 3') ; 6.27 (m, IH, 13); 6.29 (ε, IH,

10) ; 6.35,7.01 (two doubletε, J=2.1 Hz, 2H_;Z- ) ; 6.96 (d, J=9.4 Hz, IH, NH-4') ; 7.3-8.2 ( , 15H, 3-Ph) . Example 5 7-(N-(α bromo-acryloyl) -βalanyl) -taxol (FCE 28573)

To a solution of 30 mg of a bromo-acrylic acid (0.2 mmolε) in 10 ml of acetonitrile, 40 mg of N,N'-dicyclohexylcarbodiimide were added and the mixture was εtirred for 10 minutes at room temperature.

To the mixture were added 48 mg of 7-(βalanyl) -taxol formate (0.05 mmols) and 0.007 ml of triethylamine (0.05 mmols), and the whole was εtirred for 4 hours at room temperature.

The precipitate was filtered and the solvent evaporated under vacuum to dryness.

The residue was chromatographed on εilica gel with ethyl acetate-hexane 3:2 aε eluant, affording 34 mg of the title compound.

FAB-MS: m/z 1057, M-H]^"; 730, 284.

Η-NMR (400 MHz, CDC1₃) : δ 1.16 (ε, 3H, 16); 1.21 (s, 3H,

17); 1.81 (ε, 3H, 19); 1.83 (d, J=1.4 Hz, 3H, 18);

1.8-2.0 (m, IH, 6β) ; 2.15 (ε, 3H, CH₃CO-10) ; 2.32 ( , 2H, CH₂-14) ; 2.39 (ε, 3H, CH₃CO-4); 2.4-2.7 (m,

3H, OCOCH₂CH₂NH) ; 3.6 (m, 2H, 0C0CH₇CH-,NH) ; 3.75 (d,

J=5.3 Hz, IH, OH-2'); 3.92 (d, J=6.7 Hz, IH, 3);

4.19 (d, J=8.5 Hz, IH, 20β) ; 4.32 (d, J=8.5 Hz,

IH, 20α) ; 4.80 (dd, J=2.6 Hz, J=5.3 Hz, IH, 2'); 4.94 (d, J=7.9 Hz, IH, 5); 5.61 (dd, J=7.0 Hz,

J=10.5 Hz, IH, 7); 5.67 (d, J=6.7 Hz, 2); 5.81

(dd, J=2.6 Hz, J=9.1 Hz, IH, 3'); 6.0,6.95 (two doublets, J=1.5 Hz, 2H, C=CH₂) ; 6.19 (s, IH, 10) ; 6.19 (m, IH, 13); 7.12 (d, J=9.1 Hz, IH, NH-4' ) ; 7.2-8.2 (16 H, 3-Ph + OCOCH₂CH₂NH) .

By analogouε procedure the following compounds can be obtained:

7-(N-(glycidyl) -βalanyl) -taxol (FCE 28599) . FAB-MS: m/z 995, M+H^{"1 +}; 668, 286.

Η-NMR (400 MHz, CDC1₃) : δ 1.16 (s, 3H, 16) ; 1.22 (ε, 3H, 17); 1.6-1.8 (m, 8H, 18+19+6β+OH-l) ; 2.20 (s, 3H,

CH₃CO-10) ; 2.34 (m, 2H, 14); 2.39 (s, 3H, CH₃CO-4) ;

2.2-2.4 (m, 3H, 6α+ NHCH-,CH-,CO) ; 2.7-3.0 ( , 2H, epoxide) ; 3.4-3.7 (m, 4H, NHCH₂CH₂CO+ epoxide + OH-

2'); 3.92 (d, J=6.7 Hz, IH, 3); 4.19 (d, J=8.5 Hz, IH, 20β) ; 4.32 (d, J=8.5 Hz, IH, 20α) ; 4.80 (m,

IH, 2') ; 4.94 (d, J=8.8 Hz, IH, 5) ; 5.61 ( , IH,

7); 5.67 (d, J=6.7 Hz, IH, 2); 5.81 (dd, J=8.8 Hz,

J=1.3 Hz, IH, 3') ; 6.20 (m, 2H, 13 + 10) ; 6.87

(bε, IH, NHCH₂CH₂CO) ; 7.04 (d, J=8.8 Hz, IH, NH- 4') ; 7.3-8.2 (m, 15H, 3-Ph ) .

Example 6

2'-chloroacetyl-taxol (FCE 28616)

To a εolution of 170 mg of taxol (0.2 mmolε) in 20 ml of dichloromethane, were added 24 mg of dimethylaminopyridine (0.2 mmols), 0.056 ml of triethylamine (0.4 mmols) and 0.024 ml of chloroacetylchloride (0.3 mmolε) . The reaction mixture waε εtirred for 1 hour at room temperature, waεhed with diluted hydrochloride acid (0.1N), water and dried on anhydrouε εodium εulphate. The dichloromethane was evaporated under vacuum and the residue was chromatographed on silica gel with ethyl acetate- hexane 1:1 as eluant, affording 160 mg of the title compound. FAB-MS: m/z 930, M+H~]-; 509; 362; 344.

Η-NMR (400 MHz, CDC1₃) : δ 1.14 (s, 3H, 16) ; 1.23 (s, 3H,

17) ; 1.68 (ε, 3H, 19) ; 1.76 (s, IH, OH-1) ; 1.89 (m, IH, 6β) ; 1.93 (d, J=1.2 Hz, 3H, 18) ; 2.20 (dd,

J=8.8 Hz, J=15.2 Hz, IH, 14); 2.23 (ε, 3H, CH₃CO-

10) ; 2.39 (dd, J=9.4 Hz, J=15.2 Hz, IH, 14) ; 2.46

(ε, 3H, CH₃CO-4) ; 2.50 (d, J=4.1 Hz, IH, OH-7) ;

2.57 (m, IH, 6α) ; 3.82 (d, J=7.0 Hz, IH, 3) ; 4.14,4.19 (two doubletε, J=15.3 Hz, 2H, C0CH₂C1) ;

4.21 (d, J=8.5 Hz, IH, 20β) ; 4.32 (d, J=8.5 Hz,

IH, 20α) ; 4.44 (m, IH, 7) ; 4.98 (dd, J=2.0 Hz,

J=8.5 Hz, IH, 5) ; 5.55 (d, J=2.9 Hz, IH, 2'); 5.69

(d, J=7.0 Hz, IH, 2) ; 6.02 (dd, J=2.9 Hz, J=9.4 Hz, IH, 3') ; 6.27 (m, IH, 13) ; 6.29 (ε, IH, 10);

6.85 (d, J=9.4 Hz, IH, NH-4' ) ; 7.3-8.2 (m, 15H,

3-Ph) . Example 7

2' ,7-bis-chloroacetyl-taxol (FCE 28 642)

To a solution of 170 mg of taxol (0.2 mmols) in 20 ml of dichloromethane, were added 61 mg of dimethylaminopyridine (0.5 mmols), 0.28 ml of triethylamine (2 mmols) and 0.16 ml of chloroacetylchloride (2 mmols) .

The reaction mixture was stirred for 4 hours at room temperature, washed with diluted hydrochloride acid (0.1 N) , water and dried on anhydrous sodium sulphate.

The dichloromethane was evaporated under vacuum and the residue was chromatographed on silica gel with ethyl acetate- hexane 1:1 as eluant, affording 143 mg of the title compound. FAB-MS: m/z 1004, M-H^~|^', 601, 542, 360. Η-NMR (400 MHz, CDC1₃) : δ 1.15 (s, IH, 16) ; 1.21 (s, IH, 17); 1.73 (s, IH, OH-1) ; 1.82 (s, IH, 19); 1.89 (m, IH, 6β) ; 1.97 (d, J=1.2 Hz, 3H, 18); 2.16 (s, 3H, CH₃CO-10) ; 2.23 (dd, J=9.1 Hz, J=15.5 Hz, IH, 14β) ; 2.38 (dd, J=9.1 Hz, J=15.5, IH, 14α) ; 2.45 (s, 3H, CH₃CO-4); 2.66 (m, IH, 6α) ; 3.95 (d, J=6.7

Hz, IH, 3); 3.99,4.21 (two doublets, J=14.9 Hz, 2H, C0CH₂C1) ; 4.15,4.20 (two doublets, J=14.9 Hz, 2H, C0CH₂C1) ; 4.19,4.35 (two doublets, J=8.2 Hz, 2H, CH₂-20) ; 4.98 (d, J=7.9, IH, 5); 5.58 (d, J=3.2 Hz, IH, 2'); 6.66 (m, 2H, 2+7); 6.01 (dd, J=3.2

Hz, J=9.4 Hz, IH, 3') ; 6.18 (s, IH, 10); 6.24 (ddd, 1=9.1 Hz, J=9.1 Hz, J=1.2 Hz, IH, 13); 6.87 ( d , J=9 . 4 Hz , IH , NH-4 ' ) ; 7 . 3 -8 . 2 (m , 15 H , 3 -Ph) .

Example 8

2 '-{p-[N,N-bis-(2-chloroethyl) amino]benzoyl}taxol

To a solution of taxol (85.3 mg, 0.1 mmols) in methylene chloride (2 ml) were added triethylamine (15.2 mg, 0.15 mmols) and p-[N,N-bis- (2-chloroethyl) amino]benzoylchloride (42 mg, 0.15 mmols) and the mixture was stirred under nitrogen at room temperature. After 8 hours the same amounts of triethylamine and p-[N,N-bis- (2-chloroethyl)amino] benzoylchloride were added and the reaction mixture was stirred under reflux for 6 hours. Then the εolvent waε evaporated under vacuum and the crude material purified by flash chromatography over silica gel (eluant: n-hexane:ethyl acetate = 1:1) to yield 28 mg (26%) of the title compound as a white solid.

Rf ~ 0.31 (eluant : n-hexane : ethyl acetate = 1:1) .

FAB-MS: m/z 1097.4 M^~]\ Η-NMR (400 MHZ, CDC1₃) : δ 1.13 (s, 3H, 16) ; 1.23 (ε, 3H,

17) ; 1.67 (s, 3H, 19) ; 1.74 (s, IH, OH-1) ; 1.88

(m, IH, 6β) ; 1.95 (d, J=1.2 Hz, 3H, 18); 2.12 (dd,

J=8.8 Hz, J=15.3, IH, 14); 2.31 (dd, J=9.4 Hz,

J=15.3, IH, 14) ; 2.23 (s, 3H, CH₃CO-10) ; 2.42 (s, 3H, CH₃CO-4); 2.4-2.6 (m, 2H, OH-7 + 6a) ; 3.6-3.9

(m, 9H, 3+N(CH₂CH₂Cl)₂) ; 4.19 (d, J=8.5 Hz, IH,

20β) ; 4.31 (d, J=8.5 Hz, IH, 20α) ; 4.45 (m, IH, 7) ; 4.97 (dd, J=2.3 Hz, J=9.7 Hz, IH, 5) ; 5.66 ( , 2H, 2' + 2) ; 5.97 (dd, J=4.1 Hz, J=8.8 Hz, IH, 3') ; 6.25 (ddd, J=8.8 Hz, J=9.4 Hz, J=1.2 Hz, IH, 13) ; 6.29 (ε, IH, 10) ; 6.68 (m, 2H, CO Ph-N) ; 7.07 (d, J=8.8 Hz, IH, NH-4') ; 7.2-8.2 (m, 17H, 3-Ph+CO-

Ph-N) .

Example 9

7-{p-[N,N-bis-(2-chloroethyl)amino]benzoyl}taxol

A εolution of 2'-triethylsilyltaxol (obtained according to literature procedure, see J. Org. Chem. 5J5, 3798-9, 1993) (95 mg, 0.098 mmols) and triethylamine (0.3 ml, 2.168 mmols) in N,N-dimethylformamide (1 ml) was treated at 80°C with a few crystals of 4-dimethylaminopyridine and portionwise with p- [N,N-bis-(2-chloroethyl)amino] benzoylchloride (180 mg, 0.640 mmols) . After 2 hourε εtirring under nitrogen, the reaction mixture was cooled at room temperature, treated with water and extracted with ethyl acetate. The organic phase was treated with sodium chloride εaturated εolution, dried with anhydrouε εodium εulphate, filtered and evaporated under vacuum.

The residue (220 mg) in tetrahydrofuran (2 ml) was treated with HCl IN (0.2 ml) at room temperature under nitrogen. After 1 hour stirring, the reaction mixture was treated with water (5 ml) , extracted with ethyl acetate, the organic phase dried over anhydrous sodium sulphate, filtered and evaporated. The crude material was purified by preparative silica gel TLC (eluant : n-hexane: ethyl acetate = 1:1) to yield 12 mg (11%) of the title compound. Rf ~ 0.32 (eluant: n-hexane: ethyl acetate = 1:1) . FAB-MS: m/z 1097.8 M+H^~| ⁺, 1097.6 M-H]^". Η-NMR (400 MHz, CDCl₃) : δ 1.20 (s, 6H, 16 + 17) ; 1.81 (s, IH, OH-1) ; 1.8-2.0 (m, IH, 6β) ; 1.89 (d, J=1.2 Hz, 3H, 18) ; 1.94 (s, 3H, 19) ; 2.01 (s, 3H, CH₃CO-10) ; 2.35 (m, 2H, CH₂-14); 2.39 (ε, 3H, CH₃CO-4); 2.76 (m, IH, 6a ) ; 3.6-3.9 (m, 9H, -N(CH,CH₂C1)₂ + OH-2' ) ; 4.0 (d, J=6.8 Hz, IH, 3) ; 4.24 (d, J=8.5 Hz, IH,

20β) ; 4.34 (d, J=8.5 Hz, IH, 20α) ; 4.81 (dd, J=2.6 Hz, J=4.1 Hz, IH, 2') ; 4.98 (d, J=7.9 Hz, IH, 5) ; 5.68 (dd, J=7.3 Hz, J=10.3 Hz, IH, 7) ; 5.72 (d, J=6.8 Hz, IH, 2) ; 5.82 (dd, J=2.6 Hz, J=9.1 Hz, IH, 3') ; 6.18 (m, IH, 13) ; 6.39 (ε, IH, 10) ; 6.64

(m, 2H, N-Ph-CO) ; 7.07 (d, J=9.1 Hz, IH, NH-4' ) ; 7.3-8.2 ( , 17H, 3-Ph+ N-Ph-CO) .

Example 10

2' ,7-bis-bromoacetyl-taxol

To a solution of 170 mg of taxol (0.2 mmols) in 20 ml of dichloromethane, cooled at 0°, were added 61 mg of dimethylaminopyridine (0.5 mmols) , 0.28 ml of triethylamine (2 mmols) and 0.172 ml of bromoacetylbromide (2 mmols) . The reaction mixture was stirred for two hours at 0° , washed with diluted sulfuric acid, water and dried on anhydrous sodium sulphate. The dichloromethane was evaporated under vacuum, affording 220 mg of the title compound.

By analogous procedure the following compound can be obtained: 2' ,7-bis-(α bromo-acryloyl) -taxol

Example 11

7-bromoacetyl-taxol (FCE 28975)

To a solution of 220 mg of 2' ,7-biε-bromoacetyl-taxol (0.2 mmols) in 15 ml of methanol, 0.05 ml of triethylamine was added and the whole was stirred for 1 hour at room temperature. The solution was treated with 0.05 ml of acetic acid, the methanol was evaporated under vacuum and the residue was cromatographed on silica gel with ethyl acetate- hexane 1:1 as eluant, affording 105 mg of the title compound. FAB-MS: m/z 974, M+H^{"1 +}; 896; 689.

Η-NMR (400 MNz, CDC1₃) : δ 1.15 (s, 3H, 16); 1.21 (s, 3H, 17); 1.81 (d, J=1.5 Hz, 3H, 18); 1.82 (s, 3H, 19); 1.89 (m, IH, 63); 2.16 (ε, 3H, CH₃CO-10) ; 2.34 (m, 2H,

CH₂-14) ; 2.38 (ε, 3H, CH₃CO-4) ; 2.62 (m, IH, 6α) ; 3.60 (bs, IH, OH-2') ; 3.80, 3.96 (two doublets, J=12.6 Hz, 2H, COCH₂Br) ; 3.91 (d, J=7.0 Hz, IH, 3); 4.19, 4.32 (two doublets, J=8.2 Hz, 2H, CH₂-20) ; 4.80 (d, J=2.4 Hz, IH, 2') ; 4.95 (d, J=7.9 Hz, IH,

5) ; 5.60 (dd, J=7.0 Hz, J=10.5Hz, IH, 7) ; 5.66 (d, J=7.0 Hz, IH, 2); 5.80 (dd, J=2.4 Hz), J=8.8 Hz, IH, 3') ; 6.14 (s, IH, 10); 6.18 (m, IH, 13) ;7.05 (d, J=8.8 Hz, IH, NH-4') ;7.3-8.2 (m, 15H, 3-Ph) ; By analogous procedure the following compound can be obtained: 7- (a bromo-acryloyl) -taxol (FCE 28974) FAB-MS: m/z 986,M+IT⁺; 701; 644.

Η-NMR (400 MHz, CDCl₃) : δ 1.17 (s, 3H, 16);1.21 (s, 3H, 17);

1.83 (d, J=1.5 Hz, 3H, 18) ; 1.87 (s, 3H, 19); 1.90

(m, IH, 6_/3); 2.14 (ε, 3H, CH₃CO-10) ; 2.39 (ε, 3H, CH₃CO-4) ; 2.34 ( , 2H, CH₂-14) ; 2.69 (m, IH, 6α) ;

3.59 (d, J=4.9 Hz, IH, OH-2 ' ) ; 3.94 (d, J=6.8 Hz,

IH, 3) ; 4.21- 4.33 (two doubletε, J=8.5 Hz, 2H,

CH₂-20) ; 4.80 (dd, J=4.9 Hz, J=2.6 Hz, IH, 2');

4.95 (d, J=8.2 HZ, IH, 5) ; 5.58 (dd, J=7.3 Hz, J=10.3 Hz, IH, 7) ; 5.69 (d, J=6.8 Hz, IH, 2); 5.80

(dd, J=2.6 Hz, J=9.1 Hz, IH, 3') ; 6.18 ( , IH,

13); 6.26 (m, 2H, 10 + CH=CBrCO) ; 6.83 (d, J=1.8

Hz, IH, CH=CBrCO) ; 7.05 (d, J=9.1 Hz, IH, NH-4' ) ;

7.3-8.2 (m, 15H, 3-Ph)

Example 12

2'-acetyl, 7-cyclopropylcarbonyl-taxol

To a εolution of 89.6 mg of 2 '-acetyl-taxol (0.1 mmols) in 10 ml of acetonitrile, were added 24 mg of dimethylaminopyridine (0.2 mmols), 82 mg of N,N'-dicyclohexylcarbodiimide (0.4 mmolε) and 0.032 ml of cyclopropanecarboxylic acid (0.4 mmolε) .

The reaction mixture was stirred for 6 hours at room temperature, the precipitate was filtered and the solvent evaporated under vacuum to drynesε. The residue was cromatographed on silica gel with ethylacetate-hexane 1:1 as eluant, affording 80 mg of the title compound. FAB-MS: m/z 962,M+K^η+; 593; 326; 284.

Η-NMR (400 MHz, CDCl₃) : δ 0.7-1.1 (m, 4H, cyclopropyl); 1.17 (s, 3H, 16) ; 1.19 (s, 3H, 17); 1.60 (m, IH, cyclopropyl); 1.74 (ε, IH, OH-1)

1.81 (s, 3H, 19); 1.81 (m, IH, 6_/S);1.99 (d, J=l.4 Hz, 3H, 18); 2.15, 2.16 (two singletε, 6H, CH₃CO- 2'+CH₃CO-10) ; 2.1-2.4 (m, 2H, CH₂-14); 2.42 (s, 3H, CH₃CO-4) ; 2.60 (m, IH, 6α) ; 3.94 (d, J=7.0 Hz, IH,

3); 4.19, 4.32 (two doubletε, J=8.5 Hz, 2H, CH₂- 20); 4.95(m,lH,5) ; 5.55 (m, 2H, 7+2'); 5.69 (d, J=7.0Hz, 1H,2); 5.94 (dd, J=3.5 Hz, J=9.1 Hz, IH, 3'); 6.21 (m, IH, 13); 6.35 (s, IH, 10); 6.91 (d, J=9.1 Hz, IH, NH-4'); 7.3-8.2 (m, 15H, 3-Ph) ;

Example 13

7-cyclopropylcarbonyl-taxol (FCE 29050)

To a solution of 65 mg of 2'-acetyl, 7-cyclopropylcarbonyl- taxol in 10 ml of methanol and 5 ml of ethyl acetate, were added 17 mg of NaHC0₃ and 1.5 ml of H₂0. The reaction mixture was stirred for 4 hours at room temperature, diluted with ethylacetate and washed with brine. The solvents were evaporated under vacuum to drynesε and the residue was cromatographed on silica gel with ethylacetate- hexane 1:1 as eluant, affording 37 mg of the title compound. FAB-MS: m/z 922,M+H^{"1 +}; 637; 577.

Η-NMR (400 MHz, CDC1₃) : δ 0.7-1.1 (m, 4H, cyclopropyl) ; 1.18 (s, 3H, 16) ;1.19 (s, 3H, 17) ;1.54 ( , IH, cyclopropyl) ; 1.81 (s, 3H, 19); 1.84 (d, J=1.5 Hz,

3H, 18) ; 1.80 (m, IH, 6/3) ; 2.17 (s, 3H, CH₃CO-10) ; 2.32 (m, 2H, CH₂-14) ; 2.37 (s, 3H, CH₃CO-4) ; 2.59 (m, IH, 6α) ; 3.63 (d, J=4.7 Hz, IH, OH-2');3.91 (d, J=7.0 Hz, IH, 3) ; 4.19, 4.31 (two doublets, J=8.5, Hz, 2H, CH₂-20) ; 4.79 (dd, J=2.6 Hz, J=4.7

Hz, IH, 2') ;4.92 ( , IH, 5) ; 5.51 (dd, J=7.3 Hz, J=10.5 JHz, IH, 7); 5.68 (d, J=7.0 Hz, IH, 2); 5.81 (dd, J=2.6 Hz, J=9.1 Hz, IH, 3') ; 6.16 (m, IH, 13) ; 6.31 (s, IH, 10) ; 7.07 (d, J=9.1 Hz, IH, NH- 4') ; 7.3-8.2 (m, 15H, Ph)

Claims

1. A taxane derivative of the formula I

wherein A and B, which are the same or different, are each, independently, a single bond, an amino acid reεidue or a peptide spacer;

R is a phenyl, t-butoxy or 1-methyl-l-propenyl group

R₃ is hydrogen or an acetyl group and either

(i) R, is hydrogen and R is an alkylating residue selected from: a) a group -CO-(CH₂)_m-R₄ where m is 0,1,2,3 or 4 and R₄ is halogen, oxiranyl, methyloxiranyl, aziridinyl, cyclopropyl, jS-lactone or γ-lactone or b) -C0-CX=CH₂ where X is Cl, Br, N0₂ or CN or c) -CO-Ph-p-N(CH₂CH₂Cl)₂ or d) -CO-CH(NH₂) CH₂-Ph-p-N(CH₂CH₂C1)₂ or e) -CO-(CH₂)₃-Ph-p-N(CH₂CH₂Cl)₂ or f) - CO(CH₂)_nN(CH₂CH₂Cl)₂ wherein n=0-5, or

(ii) R₂ iε hydrogen and R, iε an alkylating residue as defined above; or (iii) R| and R₂, which may be same or different, are both an alkylating reεidue as defined above, with the excluεion of 2'-chloroacetyltaxol.

2. A compound according to claim 1 wherein A and B are each, independently, an amino acid residue or a peptide spacer selected from _/SAla, Gly, Phe-Gly, Phe-Phe-, Leu-Gly, Val-Ala, Phe-Ala, Leu-Phe, Leu-Ala, Phe-Leu-Gly, Phe-Phe-Leu, Leu-Leu-Gly, Phe-Tyr-Ala,Phe-Gly-Phe, Phe-Phe-Gly,Phe-Leu- Gly-Phe, Gly-Phe-Leu-Gly-Phe, Gly-/3Ala, Phe-Gly-j3Ala, Phe- Phe-/3Ala, Leu-Gly-βAla, Val-Ala-_/3Ala, Phe-Ala-/3Ala, Leu-Phe- jδAla, Leu-Gly-jSAla,Phe-Leu, Gly-jSAla, Phe-Tyr-Ala-_/3Ala, Phe- Gly-Phe-j8Ala,Phe-Phe-Gly-/3Ala, Phe-Phe-Leu-/3Ala, Leu-Leu-Gly- /3Ala, Phe-Leu-Gly-Phe-βAla and Gly-Phe-Leu-Gly-Phe-/3Ala.

3. A compound according to claim 1 or 2 in which R represents a phenyl group, R₃ is an acetyl group, A and B are each, independently, an amino acid residue /3-Ala or a peptide spacer residue Leu-Gly, Leu-Gly- Ala, Phe-Leu-Gly or Phe-Leu- Gly-βAla, and the alkylating residueε which R, and R₂ may represent are selected from: a')-CO-CH₂Cl CO-

b')- CO-CBr=CH₂, CO-CCl=CH₂, and C') -CO-Ph-p-N(CH₂CH₂C1)₂

4. A process for preparing a taxane derivative of formula I, as defined in claim 1, the process comprising reacting an optionally protected taxane derivative of formula T

wherein R', and R'₂ are each, independently, hydrogen or a protecting group and R, R₃, A and B are as defined in claim

1; with a compound of formula III

X-R_A (III) wherein X is a leaving group and R_A is an alkylating residue as defined in claim 1; and removing the protecting groups.

5. A pharmaceutical composition which compriseε, as an active principal, a compound of the formula I as defined in claim 1 or 2 or 2'-chloroacetyltaxol and a pharmaceutically acceptable diluent or carrier.

6. A compound of formula (I) , aε defined in claim 1 or 2, or 2'-chloroacetyltaxol for use in a method of treatment of the human or animal body by therapy.

7. A compound as claimed in claim 6 for uεe as an antitumour agent.