EP0873347A2

EP0873347A2 - Novel amine derivatives of epipodophyllotoxin 2", 3"-dideoxyglycosides, preparation method therefor and use thereof as a drug and for treating cancer

Info

Publication number: EP0873347A2
Application number: EP96934895A
Authority: EP
Inventors: Claude Monneret; Laurent Daley; Yves Guminski; Thierry Imbert; Bridget Hill
Original assignee: Pierre Fabre Medicament SA
Current assignee: Pierre Fabre Medicament SA
Priority date: 1995-10-12
Filing date: 1996-10-11
Publication date: 1998-10-28
Also published as: CA2234484A1; FR2739857B1; JP2000505053A; BR9610851A; FR2739857A1; WO1997013776A3; CN1202173A; MX9802933A; NZ320323A; KR19990064166A; WO1997013776A2; AU7304096A

Abstract

A compound of general formula (I), wherein X, Y, R1 and R2 are as defined in the description, a preparation method therefor and the use thereof as a drug and for treating cancer, are disclosed.

Description

New amino derivatives of 2 ", 3" dideoxygiycosides of epipodophyllotoxin, their preparation process, their use as a medicament and their use intended for anticancer treatments.

The present invention relates to new amino derivatives 2 ", 3" dideoxygiycosides of epipodophyllotoxin, their preparation process, their use as a medicament and their use intended for anticancer treatments.

Among the derivatives derived from natural lignans, there is the class of epipodophylloids, having the basic skeleton of podophyllotoxin. It includes hemi-synthetic derivatives such as Etoposide or Teniposide which are commonly used in the preparation of drugs for the treatment of cancer. They are considered major products in this area.

Etoposide has anti-tumor properties and is used to treat in particular small cell lung cancer and testicular cancer.

The disadvantage of these products is their lack of water solubility and therefore encounter difficulties in terms of formulation and administration.

The object of the present invention is to show that derivatives of 4'-demethyl epipodophyllotoxin, having in position 4, a substitution of structure 2 "-deoxyglycoside, allows, by the incorporation of one or more nitrogen, to form compounds whose addition salts have an aqueous solubility which makes it possible to respond to the problem and show the desired anticancer activity.

Patent EP-0 196 618 relates to water-soluble derivatives of 4'-demethyl epipodophyllotoxin of formula:

where R ₁ = Me, X ₁ = NH ₂ , NMe ₂ , X ₂ = OH Patent EP-0 415 453 mentions β-D-altroside derivatives of 4'-demethyl epipodophyllotoxin of formula

where R ₁ = Me R ₂ , R ₃ = OH and NH ₂ or F and NH ₂ as well as JP 0161, 423.

Other publications mention analogous derivatives (Carbohydr. Res. 1990, 206, 219; Chem. Pharm. Bull. 1986, 34, 3733; Chem. Pharm. Bull 1986, 34, 3741; Chem. Lett. 1987, 799) .

The fact of using a glycoside having a 2 "-deoxy position is quite particular. It makes it possible to obtain more lipophilic compounds than the hydroxyl analogs and, consequently, to have a broad spectrum of anti-tumor activity. This allows them to have better membrane penetration and to be able to more easily reach the biological target such as, for example, solid tumors with little irrigation. In addition, the advantage of having an amino function, for example in the 3 "position, gives the possibility of salification therefore sufficient water solubility for better formulation and better administration. The present invention therefore relates to a compound of general formula I

in which the grouping at 3 "N (R ₁ R ₂ ) is in position β (2-deoxy D Arabino series) or α (2-deoxy D ribo series) relative to the ring, R ₁ and R ₂ identical or different represent a hydrogen atom, a C ₁ to C ₆ alkyl group, which can form a ring, this ring possibly comprising a heteroatom such as oxygen or nitrogen, a C ₁ to C ₆ aminoalkyl group or cyanomethyl.

X and Y can be the same or different and represent an OH, CH ₃ , CH ₂ - NH ₂ , X and Y can also be linked and form a ring, such as, for example, 2-methyl 1,3 dioxane, thus forming a 4,2-ethylidene 3-amino-2,3-dideoxy-β-D arabino or ribo-hexo pyranoside type bicyclic osidic skeleton.

It also relates to their addition salts with mineral or organic acids salifying the nitrogen atom (s), in particular the hydrochlorides.

Advantageously, the NR ₁ R ₂ group is an NH ₂ or N (CH ₃ ) _{2 group} .

The NR ₁ R _{2 group} can also be an amino group substituted once or twice with methyl, CH ₂ CN, CH ₂ -CH ₂ -NH ₂ , forming a ring such as morpholine.

Advantageously, the compounds of general formula I will be chosen with a glycoside for which X and Y form a ring with a chain OCH (CH ₃ ) OCH ₂ , such as 4,6 ethylidene-3-amino-2,3 -dideoxy-β-D arabino-hexo pyranoside or else 4.6 ethylidene-3-amino-2,3-deιoxy-β-D-ribo-hexopyranoside. In particular, the compounds according to the invention are selected from the following compounds:

• 4'-Demethyl-4-0 (3-amino-4,6-ethylidene-2,3-dideoxy-β-D-arabinohexopyranosyI) epipodophyllotoxin,

• 4'-Demethyl-4-0 (3-amino-4,6-ethylidene-2,3-dideoxy-β-D-ribohexopyranosyl) epipodophyllotoxin,

• 4'-Demethyl-4-0 (3-dimethylamino-4,6-ethylidene-2,3-dideoxy-β-D-arabinohexo- pyranosyl) epipodophyllotoxin,

• 4'-Demethyl-4-0 (3-dimethylamino-4,6-ethylidene-2,3-dideoxy-β-D-ribohexo-pyranosyl) epipodophyllotoxin,

• 4'-Demethyl-4-0 (3-cyanomethylamino-4,6-ethylidene-2,3-dideoxy-β-D-ribohexo-pyranosyl) epipodophyllotoxin,

• 4'-Demethyl-4-0 (3- (N-morpholino) -4,6-ethylidene-2,3-dideoxy-β-D-ribohexo-pyranosyl) epipodophyllotoxin,

• 4'-Demethyl-4-0 [3 (2-aminoethylamino) -4,6-ethylidene-2,3-dideoxy-β-D-ribohexo- pyranosyl)] epipodophyllotoxin,

• 4'-Demethyl-4-0 (3-amino-2,3,6-tridesoxy-β-D-ribohexopyranosyl)

epipodophyllotoxin,

• 4'-Demethy 1-4-0 (3, 6-diamino-2,3,6-tridesoxy-β-D-ribohexopyranosyl)

epipodophyllotoxin.

The present invention also relates to pharmaceutical compositions comprising at least one compound of general formula I according to the invention and an appropriate excipient.

The pharmaceutical compositions can be presented in a suitable manner by administration by the injectable route or by the oral route in the form of capsules, capsules, tablets at the dosage of 1 to 200 mg / m ² by injection and from 5 to 500 mg. / m ² orally per 24 h period.

These derivatives can thus be administered in human clinic to treat different forms of cancer such as small cell lung cancer, testicular cancer, embryonic tumors, neuroblastomas, kidney cancer, Hodgkin's and non-Hodgkin's lyphomas, acute leukemias, colorectal cancers, melanomas, placental choriocarcinomas and breast adenocarcinomas.

The present invention also relates to processes for the preparation of the compounds of formula I as well as their addition salts with pharmaceutically acceptable mineral or organic acids.

The present invention therefore relates to the processes for preparing the compounds of general formula I according to the invention, in which a compound of formula III or IV or V is reacted

with 4'-Demethyl 4'-benzyloxycarbonyl epipodophyllotoxin with BF ₃ etherate, or trimethyl silyl trifluoromethanesulforate in an inert solvent at low temperature;

in formulas III and IV, the substituent in position 3 can be α or β, NR ₁ R ₂ can be an amino protected by a group Z,

in formula V, P represents an alcohol protecting group and the products resulting from this condensation are deprotected and hydrogenated to provide the compounds of formula I,

the primary amines in position 3 of the glycosyl are methylated by formalin and sodium cyanoborohydride.

The intermediate of formula IV is prepared by reacting a mixture of diacetoxy azido glycoside VI

with tert-butyl dimethylsilyl chloride in the presence of imidazole, in that the products resulting from this reaction are separated, in that each of these products are deacetylated, cyclized to 4,6-ethylidene with the acetal of the acetaldehyde in a catalytic acid medium.

Other characteristics of the process according to the invention will appear in the light of the description which follows, in particular of the synthesis process shown in diagram I.

It is done according to a described methodology (JC Florent and C. Monneret, J. Chem. Soc. Chem. Comm. 1987, 1171 and B. Abbaci, JC Florent and C. Monneret, Bull. Soc. Chim. Fr. 1989 , 667) from glucal 1. The azide ion is condensed to provide a glycosidic intermediate 2 whose anomeric OH is protected by a silyl group exclusively in the β position according to the technique described (C. Kolar and G. Kneissl Angew. Chem Int. Ed 29, 809 (1990)) to provide the mixture of the 2 epimeric azides: 3 and 4 separable chromatographically at this stage. By basic deacetylation in the presence of sodium methylate, the diols in position 4, 6 are obtained: compounds 5 and 12. The azide diol compound β 5 is cyclized to ethylidene conventionally using the acetaldehyde acetal in catalysis acid to lead to compound 6 whose azide is reduced to amino 7 to be protected by a benzyloxycarbonyl group (Z) in compound 8, necessary for coupling with demethyl epipodophyllotoxin, itself protected on its phenol in 4 'by a benzyloxycarbonyl group, this intermediate will be called DMEPT4'-OZ. This coupling is carried out initially by cleavage of the protective silyl by the F "ions followed by the treatment with BF3 etherate at low temperature in the same medium. The deprotection of the Z groups by hydrogenolysis provides the compound 10 of general formula I (NR ₁ R ₂ = βNH ₂ ; XY = OCH (CH ₃ ) OCH ₂ ). Compound 1 1 of general formula I (NR ₁ R ₂ = βNMe ₂ ; XY = OCH (CH ₃ ) OCH ₂ ) s' obtained by methylation of the preceding primary amine by the action of formalin and sodium cyanoborohydride.

The glycosidic intermediate azide α 12 follows the same reaction sequence as its epimer and provides in an identical manner the coupling derivative with DMEPT4'OZ from the azide 3 "α 13, in this case the coupling is carried out more easily according to two techniques.

The first technique consists in treating the ribohexopyranoside derivative 13 with trimethyl silyl trifluoromethane sulfonate (TMSOTf) at - 40 ° C in CH ₂ Cl ₂ . The second technique consists in using the etherate of BF ₃ in CH ₂ Cl ₂ at - 15 ° C. The intermediate obtained 14 is then catalytically reduced to lead to 15 corresponding to the general formula I where NR ₁ R ₂ = αNH ₂ and XY = OCH (CH ₃ ) OCH ₂ . The same methylation as for the compound βNH ₂ 10, with formalin and cyanoborohydride leads to the corresponding dimethylamino derivative: general formula I (NR ₁ R ₂ = αNMe ₂ ; XY = OCH (CH ₃ ) OCH ₂ ). From primary amine 15, it is possible to alkylate nitrogen with a halogen derivative such as iodoacetonitrile under weakly basic conditions with triethylamine in DMF and lead to derivative 16 of general formula I where NR ₁ R ₂ = αNHCH ₂ CN; XY = OCH (CH ₃ ) OCH ₂ . Likewise, derivative 17 is obtained by forming the morpholine ring by cyclization of diiodinated ether on the same primary amine intermediate 15. The same alkylation with iodoethylamine protected by a group Z on compound 15 provides derivative 18 which is catalytically reduced to 19 diamino derivative corresponding to the general formula I with NR ₁ R ₂ = α-NHCH ₂ CH ₂ NH ₂ ; XY = OCH (CH ₃ ) OCH ₂ .

The compounds of formula I where XY does not form a ring are obtained in the following manner, represented in diagram 2:

the silylated α azide 12 is selectively tosylated with tosyl chloride in pyridine at 21, the primary alcohol tosylate is exchanged for iodo derivative 22. At this stage, the secondary alcohol in position 4 is protected by a chloroacetate to provide the functionalized 2-desoxysugre 23 ready to be condensed with DMEPT 4'-OZ under the usual conditions with the etherate of BF ₃ in methylene chloride at low temperature. Intermediate 24 obtained, allows by passage over the amberlite resin IRA410 treated in basic medium to deprotect the chloroacetate group to compound 25 and the final catalytic hydrogenation allows in a state to reduce the azide function to amino 3 "α, to deprotect the Z function in position 4 'and reduce the carbon 6 "to methyl to provide the derivative 26 corresponding to the general formula I where NR ₁ R ₂ = 3" αNH ₂ ; X = OH ; Y = CH ₃ . Intermediate 24 obtained previously can react with the azide ion in DMF at room temperature to provide 3 "αN ₃ , 6" -N ₃ comprising an azidoacetate at position 4 "27, which can be cut into alcohol 4" by a treatment similar to the previous one: passing over the Amberlite IRA 410 exchange resin to supply the diazidoalcohol 28. The final catalytic reduction step makes it possible to obtain the diamine derivative 29 corresponding to the general formula I (where NRjR ₂ = 3 "αNH ₂ ; X = OH; Y = CH ₂ NH ₂ ) The salts formed from the nitrogen compounds are, for example, hydrochlorides and are formed conventionally by treating a methanolic solution of the nitrogen compound with a stoichiometric solution with respect to sites to be salified with hydrochloric methanol previously dosed The crystallized hydrochloride can optionally be obtained by precipitation in the reaction medium by addition of ethyl ether.

By way of indication and to illustrate the various stages of the synthesis but, without limitation, the following examples are given.

EXAMPLE 1

General formula I: NR ₁ R ₂ = βNH ₂ ; XY = OCH (CH ₃ ) OCH ₂

4'-Demethyl-4-0 (3-amino-2,3-dideoxy-4,6-ethylidene-β-D-arabinohexopyranosyl) epipodophyllotoxin (compound 10)

Stage 1

A solution of tri-O-acetyl-D-glucal 1 (50 g; 183 mmol) in water (400 ml) is heated for 3 h at 80 ° C. The reaction medium is then cooled to 20 ° C. before addition of sodium azide (17.9 g; 275 mmol) and acetic acid (38 ml; 600 mmol). After stirring at room temperature for 24 h, the medium is neutralized with NaHCO ₃ (salt). The phase is extracted with ethyl acetate (3 × 500 ml). The organic phases are combined, dried over MgSO ₄ and then concentrated under reduced pressure. This provides 51 g of crude product 2 immediately treated as follows. Stage 2

Tertbutyldimethylsilyl-3-azido-2,3-dideoxy-4,6-di-O-acetyl-β-D-arabinohexopyranoside 3 and terbutyldimethylsilyl-3-azido-2,3-dideoxy-4,6-di-O-acetyl -β-D-ribohexopyranoside 4

To a solution of the crude mixture 2 (16.1 g; 58 mmol) obtained in stage 1 in anhydrous dichloromethane (200 ml) previously cooled to 0 ° C., 6 g of imidazole (87, 8 mmol) and 13.24 g of tert-butyldimethylsilyl chloride (87.8 mmol). After stirring for 15 min at 0 ° C and 19 h at 20 ° C, the reaction medium is poured into 500 ml H ₂ O. The aqueous phase is extracted with CH ₂ Cl ₂ (200 ml) then after drying over MgSO ₄ , the organic phase is concentrated under reduced pressure and the residue (18.7 g) is chromatographed on silica gel (cyclohexane / AcOEt: 9/1). 10.7 g of 3 (syrup; 48%) and 4.7 g of 4 (syrup; 21%) are thus isolated; however, intermediate fractions contain a mixture of 3 and 4 (3.3 g; 15%).

Stage 3

Tertbutyldimethylsilyl-3-azido-2,3-dideoxy-β-D-arabino-hexopyranoside 5

To a solution of derivative 3 (3 g; 7.7 mmol) obtained in stage 2 in anhydrous methanol (40 ml) is added, under argon, a solution of 1M sodium methanolate (1.9 ml). After reaction for 1 h 30 at 20 ° C, the reaction medium is brought to pH = 7 by addition of H ⁺ resin (amberlite IRC 50 S). The reaction mixture is filtered and the filtrate concentrated under reduced pressure resulting in 2.27 g of 5 (97%).

Stage 4

Tertbutyldimethylsilyl-3-azido-2,3-dideoxy-4,6-O-ethylidene-β-D-arabinohexopyranoside (6)

To a solution of 5 (0.20 g; 0.6 mmol) obtained in stage 3 in 5 ml of acetonitrile, 0.94 ml (6.6 mmol) of the acetaldehyde diethylacetal is added, followed by 15 mg of paratoluenesulfonic acid (0.08 mmol). The reaction medium is stirred at room temperature for 1 h and then diluted with ethyl acetate (20 ml) before washing with a sodium hydrogen carbonate solution (pH = 9) (20 ml) then with water (20 ml). The organic phase is dried over MgSO ₄ and then concentrated under reduced pressure to provide 0.25 g of crude product. Purification on silica gel (cyclohexane / AcOEt: 95/5) allows 0.19 g of pure 6 (86%) to be isolated.

Stage 5

Tertbutyldimethylsilyl-3-amino-2,3-dideoxy-4,6-O-ethylidene-β-D-arabino-hexopyranoside (7)

To a solution of 6 (2 g; 6 mmol) obtained in stage 4 in 30 ml of ethyl acetate are added 50 μl of triethylamine then 0.5 g of 10% palladium on carbon. The reaction medium is placed under a hydrogen atmosphere (atmospheric pressure). After stirring for 6 h at room temperature, the catalyst is removed by filtration and the organic phase concentrated under reduced pressure to provide 1.82 g of pure 7 (98%).

Stage 6

Tertbutyldimethylsilyl-3-aminobenzyloxycarbonyl-2,3-dideoxy-4,6-O-ethylidene-β-D- arabino-hexopyranoside (8) Benzyloxycarbonyl chloride (1.12 ml; 7.88 mmol) is added, under argon, to a solution previously cooled to 0 ° C of acetal 7 (1.82 g; 6 mmol) obtained in step 5 in a mixture of anhydrous dichloromehane (30 ml) and triethylamine (1.27 ml; 9.1 mmol). After stirring for 8 h, the reaction medium is poured into 100 ml H ₂ O and the aqueous phase is extracted with CH ₂ C1 ₂ (100 ml). The organic phase is dried over MgSO ₄ , concentrated under reduced pressure and the residue is purified by chromatography on silica gel (cyclohexane / AcOEt: 6/1 and 4/1) in order to isolate 1.9 g of 8 (72% ).

Stage 7

3-aminobenzyloxycarbonyl-2,3-dideoxy-4,6-O-ethylidene-β-D-arabinohexopyranoside of 4'-benzyloxycarbonyl-epipodophyllotoxin (9)

To the sugar 8 (2.0 g; 4.57 mmol) obtained in stage 6 in solution in anhydrous dichloromethane (100 ml) are added 5.06 ml of tetrabutylammonium fluoride (1.1 M solution in THF; 5, 5 mmol). When the total disappearance of 8 is observed by TLC (2 h of stirring), the reaction medium is cooled to -20 ° C. DMEPT 4'-OZ (2.57 g; 4.8 mmol) is then added successively, then 8.44 ml of BF _3. And ₂ O (68.6 mmol). After reaction for 1 h at -20 ° C., the reaction medium is poured into 200 ml of a saturated NaHCO ₃ solution (addition of NaHCO ₃ salts) (pH = 9). The organic phase is dried over MgSO ₄ , concentrated under reduced pressure, then the crude residue (6.2 g) is chromatographed on silica gel (CH ₂ Cl ₂ / Acetone: 98/2 then 97/3) to provide 9 ( 2.1 g; 54%).

Stage 8

To a solution of 9 (0.28 g; 0.33 mmol) in 20 ml of ethyl acetate are added 30 μl of triethylamine then 150 mg of 10% palladium on carbon. The reaction medium is placed under a hydrogen atmosphere (atmospheric pressure). After stirring for 1 h 30 at room temperature, the catalyst is removed by filtration and the organic phase concentrated under reduced pressure and then chromatographed on silica gel (CH ₂ Cl ₂ / MeOH: 97/3 then 95/5) to provide 172 mg pure compound (90%). (Recrystallization from CH ₂ Cl ₂ / pentane).

Preparation of the hydrochloride

To amine 10 (61 mg; 0.10 mmol) dissolved in anhydrous dichloromethane (6 ml) is added a 0.098 M hydrochloric methanol solution (1.09 ml;

0.106 mmol). The reaction medium is stirred for 10 minutes. The expected product is precipitated after addition of ether (20 ml). 56 mg (86%) of the hydrochloride are collected.

10.

EXAMPLE 2

General formula I (NR ₁ R ₂ = βNMe ₂ ; XY = OCH (CH ₃ ) OCH ₂ )

4'-Demethyl-4-0 (3-dimethylamino-2,3-didesoxy-4,6-ethylidene-β-D- arabinohexopyranosyl) epipodophyllotoxin (Compound 11)

To a solution of 10 (0.19 g; 0.33 mmol) in anhydrous dichloromethane (15 ml) are added successively formaldehyde (13.5 μl) and sodium cyanoborohydride (85 mg). After stirring for 45 min at room temperature, these same reagents are added and the reaction is continued for 45 min. The reaction medium is diluted with CH ₂ Cl ₂ (30 ml) and washed with water (40 ml). The organic phase is dried over MgSO ₄ and concentrated under reduced pressure. The residue is chromatographed on silica gel (CH ₂ Cl ₂ / MeOH: 97/3). This provides 101 mg of 1 1 (51%).

;

Preparation of the hydrochloride

To the amine 11 (101 mg; 0.17 mmol) dissolved in anhydrous dichloromethane (7 ml) is added a 0.098 M hydrochloric methanol solution (1.72 ml; 0.17 mmol). The reaction medium is stirred for 10 minutes. The expected product is precipitated after addition of ether (20 ml). 86 mg (81%) of the 1 L hydrochloride are collected.

EXAMPLE 3

General formula I: NR ₁ R ₂ = α-NH ₂ ; XY = OCH (CH ₃ ) OCH ₂

4'-Demethyl-4-O- (3-amino-2,3-dideoxy-4,6-ethylideneβ-D-ribohexopyranosyl) epipodophyllotoxin (compound 19)

Stage 1

Tertbutyldimethylsilyl-3-azido-2,3-dideoxy-β-D ribo hexopyranoside (compound 12)

Similarly, in stage 3 of Example 1, but using compound 4, compound 12 is obtained which is used directly in stage 2.

Stage 2

Tertbutyldimethylsilyl-3-azido-2,3-dideoxy-4,6-O-ethylidene-β-D-ribo hexopyranoside (13) To a solution of 12 (0.25 g; 0.8 mmol) obtained in stage 1 in 10 ml of acetonitrile, 1.1 ml (8 mmol) of the acetaldehyde diethylacetal and then 52 mg of camphorsulfonic acid are added (0.24 mmol). The reaction medium is stirred at ambient temperature for 9 h then diluted with ethyl acetate (30 ml) before washing with a sodium hydrogen carbonate solution (pH = 9) then with water (30 ml) . The organic phase is dried over MgSO ₄ and then concentrated under reduced pressure to provide 0.3 g of crude product. Purification on silica gel (cyclohexane / AcOEt: 15/1) makes it possible to isolate 0.15 g of pure 13 (55%).

Stage 3

3-azido-2,3-dideoxy-4,6-O-ethylidene-β-D-ribo-hexopyranoside of 4'-benzyloxycarbonyl-epipodophyllotixne (compound 14)

1st synthetic route:

To a mixture of DMEPT 4'-OZ (438 mg; 0.82 mmol), of 13 (270 mg; 0.82 mmol) obtained in stage 2 and of 4 A molecular sieve (1.5 g) in anhydrous dichloromethane (30 ml) cooled to -40 ° C., trimethylsilyltrifluoromethanesulfonate (TMSOTf) (446 μl; 2.46 mmol) is added. After reacting for 1 h 15 at -40 ° C., the reaction medium is neutralized with triethylamine (342 μl), filtered and then washed with a saturated NaCl solution (20 ml). The organic phase is dried over MgSO ₄ , concentrated under reduced pressure and then the crude residue is chromatographed on silica gel (cyclohexane / AcOEt: 65/35) to provide 14 (260 mg; 45%).

2nd synthetic route:

To a mixture of DMEPT 4'OZ (1.85 g; 3.46 mmol), of 13 (1.20 g; 3.64 mmol) obtained in stage 2 in anhydrous dichloromethane (100 ml) cooled to - 15 ° C, is added boron trifluoride etherate (BF3-Et ₂ O) (425 μl; 3.46 mmol). After reaction for 2 h at -15 ° C, the reaction medium is diluted with 100 ml of CH ₂ C1 ₂ and then poured into 200 ml of a saturated NaHCO ₃ solution. The organic phase is dried over MgSO ₄ , concentrated under reduced pressure and then the crude residue (2.9 g) is chromatography on silica gel (cyclohexane / AcOEt: 65/35) to provide 14 (1.19 g; 47%) (recrystallization Et ₂ O / hexane).

Stage 4

3-amino-2,3-dideoxy-4,6-O-ethylidene-β-D-ribo-hexopyranoside

epipodophyllotixin 15

To a solution of 14 (1 10 mg; 0.15 mmol) obtained in stage 3 in a mixture of 10 ml of ethanol and 5 ml of ethyl acetate are added 20 μl of triethylamine then 20 mg of palladium on carbon at 10%. The reaction medium is placed under a hydrogen atmosphere (atmospheric pressure). After stirring for 2 h at room temperature, the catalyst is removed by filtration and the organic phase concentrated under reduced pressure and then chromatographed on silica gel (CH ₂ Cl ₂ / MeOH: 97/3 then 95/5) to provide 63 mg of 15 pure (72%).

HT and H _{6 '} ); 6.51 (1H, s, H ₈ ); 6.86 (1H, s, H ₅ ).

Preparation of the hydrochloride

To amine 15 (50 mg; 0.087 mmol) obtained in stage 4 are added 890 μl of a 0.098 M hydrochloric methanol solution (0.087 mmol). After stirring for 10 minutes and addition of ether (10 ml), the crystals or hydrochloride (52 mg, 98%) are obtained by filtration.

EXAMPLE 4

General formula I: NR ₁ R ₂ = α-NMe ₂ ; XY = OCH (CH ₃ ) OCH ₂

4'-Demethyl-4-O (3-N, N-dimethylamino-2,3-dideoxy-4,6-O-ethylidene-β-D-ribohexopyranosyl) epipodophyllotoxin 20

To a solution of 15 (29 mg: 0.05 mmol) obtained in stage 4 of Example 3 in

1 ml of acetonitrile are added successively formaldehyde (10.3 μl) and sodium cyanoborohydride (12 mg). After stirring for 2 h at room temperature, the reaction medium is diluted with CH ₂ Cl ₂ (20 ml) and washed with water (20 ml). The organic phase is dried over MgSO ₄ and concentrated under reduced pressure. The residue is reacted under the same conditions and after identical treatment chromatography on silica gel (CH ₂ Cl ₂ / MeOH: 97/3). This provides 29 mg of 20 (95%).

Preparation of the hydrochloride

To amine 20 (59 mg; 0.1 mmol) dissolved in anhydrous methanol (2 ml) is added a solution of 0.098 M hydrochloric methanol (1 ml; 0.1 mmol). The reaction medium is stirred for 10 minutes. The expected product is precipitated after addition of ether (20 ml). 33 mg (53%) of crystals are collected.

EXAMPLE 5

General formula I: NR ₁ R ₂ = α-NHCH ₂ CN; XY = OCH (CH ₃ ) OCH ₂

4'-Demethyl-4-O- (3-cyanomethylamino-2,3-dideoxy-4,6-ethylidene-β-D-ribohexopyranosyl) epipodophyllotoxin (Compound 16)

To a solution of 15 (120 mg; 0.21 mmol) obtained in stage 4 of Example 3 in 4 ml of dimethylformamide are added 200 μl of triethylamine (1.47 mmol) then 100 μl of iodoacetonitrile (1.47 mmol). The reaction medium is stirred for 20 h at room temperature then diluted with ethyl acetate (30 ml) before washing with water (4 × 30 ml). The organic phase is dried over MgSO ₄ , concentrated under reduced pressure and then chromatographed on silica gel (CH ₂ Cl ₂ / acetone: 92/8) to provide 71 mg of pure 16 (55%).

EXAMPLE 6

General formula I: NR ₁ R ₂ = α-morpholino; XY = OCH (CH ₃ ) OCH ₂

4'-Demethyl-4-O- (3-N-morpholino-2,3-didesoxy-4,6-ethylidene-β-D-ribohexopyranosyl) epipodophyllotoxin (Compound 17)

To a solution of 15 (60 mg; 0.10 mmol) obtained in stage 4 of Example 3 in 2 ml of dimethylformamide are added 58 μl of triethylamine (0.42 mmol) then 512 mg of di-iodoethyl ether (1, 57 mmol). The reaction medium is stirred for 96 h at room temperature and in the dark, then diluted with ethyl acetate (30 ml) before washing with water (4 x 30 ml). The organic phase is dried over MgSO ₄ , concentrated under reduced pressure and then chromatographed on silica gel (CH ₂ Cl ₂ / acetone: 92/8) to provide 46 mg of pure 17 (68%).

EXAMPLE 7

General formula I: NR ₁ R ₂ = α-NH ₂ (CH ₂ ) ₂ NH ₂ ; XY = OCH (CH ₃ ) OCH ₂

4'-Demethyl-4-O [3- (2-Aminoethylamino) -2,3-dideoxy-4,6-ethylidene-β-D-ribohexopyranosyl] epipodophyllotoxin 18

To a solution of 15 (173 mg; 0.30 mmol) obtained in stage 4 of Example 3 in 10 ml of dimethylformamide are added triethylamine (127 μl; 0.91 mmol) and N-benzyloxycarbonyl-2- iodoethylamine (0.28 g; 0.91 mmol). The reaction medium is stirred for 5 days at room temperature then, diluted with water (30 ml). After extraction of ethyl acetate (30 ml) washing with water (5 × 20 ml), the organic phase is dried over MgSO ₄ , concentrated under reduced pressure, then chromatographed on silica gel (CH ₂ Cl ₂ / MeOH: 97/3) to provide 155 mg of pure 18 (68%).

which is directly involved in the following debenzylation stage:

To a solution of 18 (0.15 g; 0.20 mmol) in a mixture of ethyl acetate and ethanol (10 ml, 1/1) are added triethylamine (30 μl) then palladium on carbon at 10% (0.1 g). The reaction medium is placed under a hydrogen atmosphere (atmospheric pressure).

After stirring for 1 h 30 at room temperature in the presence of hydrogen at atmospheric pressure, the catalyst is removed by filtration and the organic phase concentrated under reduced pressure and chromatographed on silica gel (CH ₂ Cl ₂ / MeOH (NH ₃ ): 97/3) to provide 107 mg (84%) of 19.

Preparation of the hydrochloride

To the di-amine 19 (64 mg; 0.10 mmol) dissolved in anhydrous methanol (3 ml) is added a 0.098 M hydrochloric methanol solution (2.13 ml; 0.21 mmol). The reaction medium is stirred for 10 minutes. The expected product is precipitated after addition of ether (20 ml). 50 mg (73%) of the hydrochloride are collected.

EXAMPLE 8

General formula I: NR ₁ R ₂ = α-NH ₂ ; X = OH; Y = CH ₃

4'-Demethyl-4-0 (3-amino-2,3,6-tridesoxyβ-D-ribo-hexopyranosyl) epipodopoyllotoxin (compound 26)

Stage 1

Tertbutyldimethylsilyl-3-azido-2,3-dideoxy-6-O-tosyl-β-D-ribo-hexopyranoside 21

A solution of tosyl chloride (1.55 g; 8.15 mmol) in pyridine (10 ml) is added dropwise to a solution of diol 12 (2.06 g; 6.79 mmol) obtained stage 1 of Example 3 previously cooled to 0 ° C. After stirring at the same temperature for 1 h, then for 18 h at 20 ° C, the reaction medium is diluted with dichloromethane (100 ml). The organic phase is washed with water (2 × 100 ml), dried over MgSO ₄ and concentrated under reduced pressure. The residue is purified on silica gel (cyclohexane / EtOAc: 8/2). This provides 2.02 g of 21 (65%).

Stage 2

Tertbutyldimethylsilyl-3-azido-6-iodo-2,3,6-trideoxy-β-D-ribo-hexopyranoside 22

Compound 21 (2.02 g; 4.42 mmol) obtained in stage 1 in solution in 120 ml of acetone is heated at reflux for 72 h in the presence of sodium iodide (2.65 g;

17.68 mmol). After cooling, the reaction medium is concentrated under reduced pressure (30 ml) then diluted with dichloromethane (100 ml). After washing with an aqueous solution of sodium thiosulfate at 10%, then drying over MgSO ₄ and evaporation under reduced pressure, a residue is obtained which is purified on silica (cyclohexane / EtOAc: 8/2) providing 1.5 g (82 %) of 22.

Stage 3

Tertbutyldimethylsilyl-3-azido-6-iodo-4-O-chloroacetyl-2,3,6-tridesoxy-β-D-ribohexopyranoside (compound 23).

Chloroacetyl chloride (396 ml; 5 mmol) is added to a solution of 22

(1.03 g; 2.5 mmol) obtained in the previous stage in a mixture of dichloromethane (20 ml) and pyridine (404 μl; 5 mmol). After 1 h of stirring at -10 ° C., the reaction medium is diluted with CH ₂ Cl ₂ (30 ml) and washed with water (3 × 20 ml). A usual treatment, followed by chromatography on silica (cyclohexane / EtOAc: 10/1) delivers 1.1 g (90%) of compound 23.

Stage 4

4'-Demethyl-4-O (3-azido-6-iodo-2,3,6-tridesoxy-β-D ribohexopyranosyl) epipodophyllotoxin (compound 24)

To a mixture of DMEPT 4'-OZ (1 g; 1.85 mmol), 23 (1 g; 2.04 mmol) of the previous stage in anhydrous dichloromethane (100 ml) cooled to -15 ° C, is added boron trifluoride etherate (BF _{3. and} ₂ O) (455 μl; 3.7 mmol). After reaction for 5 h (- 15 ° C → 0 ° C), the reaction medium is diluted with CH ₂ Cl ₂ (100 ml) then poured into a saturated NaHCO ₃ solution (200 ml). The organic phase is dried over MgSO ₄ , concentrated under reduced pressure. The crude residue is chromatographed on silica gel (cyclohexane / AcOEt: 7/3) to provide 24 (0.8 g; 48%).

Stage 5

4'-Demethyl-4-O (3-Azido-6-iodo-2,3,6-tridesoxy-β-D ribo-hexopyranosyl) 4'-benzyloxy carbonyl epipodophyllotoxin (compound 25)

To a solution of azido-glycoside 24 (257 mg; 0.29 mmol) in a mixture

CH ₂ Cl ₂ / MeOH (15 ml, 2/1) is added with OH- resin (Amberlite IRA 410). After reaction for 3 h at 20 ° C, the reaction medium is filtered and then concentrated under reduced pressure to provide 0.22 g of pure (94%).

Stage 6

4'-Demethyl-4-0 (3-amino-2,3,6-tridesoxy-β-D ribohexopyranosyl) epipodophyllotoxin (compound 26)

To a solution of 25 (0.20 g; 0.25 mmol) of the previous stage in 10 ml of ethyl acetate are added triethylamine (100 μl) then 10% palladium on carbon (0.2 g) . After stirring for 30 h at room temperature in the presence of hydrogen at atmospheric pressure, the catalyst is removed by filtration and the organic phase concentrated under reduced pressure and then chromatographed on silica gel (CH ₂ Cl ₂ / MeOH: 92/8) to provide 50 mg (38%) of 26.

Preparation of the hydrochloride

To the above amine 26 (70 mg; 0.14 mmol) dissolved in anhydrous methanol (6 ml) is added a 0.098 M hydrochloric methanol solution (1.44 ml;

0.14 mmol). The reaction medium is stirred for 10 minutes. The expected product is precipitated after addition of ether (20 ml). 40 mg (53%) of crystallized hydrochloride are collected.

EXAMPLE 9

General formula I: NRιR ₂ = α-NH ₂ ; X = OH; Y = CH ₂ NH ₂

4'-Demethyl-4-0- (3,6-diamino-2,3,6-tridesoxy-β-D ribohexopyranosyl) epipodo- phyllotixin (compound 29)

Stage 1

4'-Demethyl-4'-benzyloxycarbonyl-4-0 (3,6-diazido-4-azidoacetyl-2,3,6-tridesoxy-β-D ribo-hexopyranosyl) epipodophyllotoxin (compound 27)

To a solution of 24 (0.45 g; 0.51 mmol) obtained in stage 4 of Example 8 in

10 ml of dimethylformamide is added sodium azide (0.1 g; 1.5 mmol). The reaction medium is stirred for 64 h at room temperature, diluted with water (30 ml) and ethyl acetate (30 ml). The organic phase is washed with water (4 × 20 ml), dried over MgSO ₄ , concentrated under reduced pressure and chromatographed on silica gel (cyclohexane / AcOEt: 7/3) to provide 0.36 g of 27 ( 90%).

Stage 2

4'-Demethyl-4'-benzyloxycarbonyl-4-0 (3,6-diazido-2,3,6-tridesoxy-β-D-ribohexopyranosyl) epipodophyllotoxin (compound 28)

To the solution of azido-glycoside 27 (70 mg; 0.08 mmol) in 3 ml of a CH2Cl2 / MeOH mixture (2/1, v / v) is added OH- resin (Amberlite IRA 410) .

After reaction for 5 h at 20 ° C, the reaction medium is filtered and then concentrated under reduced pressure to provide 59 mg of pure 28 (94%).

Stage 3

4'-Demethyl-4-0 (3,6-diamino-2,3,6-tridesoxy-β-D-ribo-hexopyranosyl)

epipodophyllotoxin (compound 29)

To a solution of 28 (0.1 1 g; 0.15 mmol) from the previous stage, in a mixture ethyl acetate and ethanol (10 ml, 1/1) are added triethylamine (20 μl) then 10% palladium-on-charcoal (70 mg). After stirring for 16 h at room temperature in the presence of hydrogen at atmospheric pressure, the catalyst is removed by filtration and the organic phase concentrated under reduced pressure to provide 78 mg (95%) of 29.

Preparation of dihydrochloride

To the di-amine 29 (78 mg; 0.14 mmol) above in solution in anhydrous methanol (2 ml) is added a 0.098 M hydrochloric methanol solution (2.92 ml;

0.28 mmol). The reaction medium is stirred for 10 minutes. The expected product is precipitated after addition of ether (20 ml). 70 mg (79%) of crystallized dihydrochloride are collected.

Biological experimentation

The molecules have been tested in biological experiments and have shown their interest as an anticancer agent in tests for P 388 leukemia in vivo in mice. This test is commonly used in the field of anti-cancer substances research (Protocols for screening chemical agents and natural products against animal tumors and other biological Systems, R. Geran, NH Greenberg, MM MacDonald, AM Schumacher and BJ Abbott, Cancer Chemotherapy reports 1972, 3, No. 2).

However, this experimental model is extremely chemosensitive and very many compounds show good activity which makes this test not very discriminating.

We have modified the test protocol to make it more selective. The administration of tumor cells is done intravenously and not by intraperitoneally. They thus spread by circulation throughout the body quickly. The administration of the product to be tested is then done intraperitoneally. Two parameters are defined to highlight the activity of the compounds:

• determination of the effective dose 50 (ED ₅₀ ) which represents the single minimum dose of the compound to be administered in order to obtain significant animal survival compared to the untreated control animals;

• determination of the maximum survival time of the animals whatever the dose administered by single injection. The fact of being able to administer a large dose of the compound and of observing a large survival makes it possible to obtain a measure of the maximum therapeutic efficacy of the product which can be achieved.

Origin of the tumor

P 388 leukemia was chemically induced in 1955 by 3-methylcholanthrene in a DBA / 2 mouse (Am. J. Pathol. 33, 603, 1957).

Pharmacological procedure

The tumors are maintained by weekly passages in the form of ascites in the peritoneum of DBA / 2 mice (original line) and the experiments are carried out on CDF} hybrid female mice (bal b / c female XDBA / 2 males) of 20 ± 2 g (Cancer chemother. Rep. 3, 9, 1972). The tumor cells are implanted intravenously (106 cells per mouse) on day 0. The animals are randomized and divided into groups of 2 for each series.

Anti-tumor substances are administered intraperitoneally (ip) one day after inoculation of the leukemia cells (acute treatment). The solutions are injected at the rate of 10 ml / kg of mouse. The criterion for evaluating anti-tumor activity is the prolongation of the survival of the animals treated. 86% of the mice die on the 7th day after the tumor transplant. A substance will be considered active if it induces survival longer than 8 days.

The following table makes it possible to highlight the aqueous solubility of products of the invention, expressed in mg / ml, the activity of these compounds in terms of ED ₅₀ , survival expressed in days or in T / C%, which represents the ratio between the average survival of the group of animals treated and the average survival of the group of control animals.

It thus appears that the compounds of the invention have retained the level of activity of the reference compounds such as Etoposide and have the additional advantage of having an aqueous solubility which is advantageous for formulation and administration.

Claims

1) A compound of general formula I

in which the grouping at 3 "N (R ₁ R ₂ ) is in position β (2-desoxy D Arabino series) or α (2-desoxy D ribo series) relative to the ring, R ₁ and R ₂ identical or different represent a hydrogen atom, a C ₁ to C ₆ alkyl group, which can form a ring, this ring possibly comprising a heteroatom such as oxygen or nitrogen, a C ₁ to C ₆ aminoalkyl or cyanomethyl group,

X and Y can be the same or different and represent an OH, CH ₃ , CH ₂ -NH ₂ , X and Y can also be linked and constitute a ring, such as, for example, 2-methyl 1,3 dioxane, thus forming a Osidic bicyclic skeleton of 4,6-ethylidene 3 amino 2,3 didesoxy glycoside, and its addition salts with pharmaceutically acceptable mineral or organic acids.

2) A compound of general formula I according to claim 1, characterized in that the group NR ₁ R ₂ is an NH ₂ or N (CH ₃ ) _{2 group} .

3) A compound of general formula I according to claim 1, characterized in that the group NR ₁ R ₂ represents an amino group substituted, once or twice by a methyl, CH ₂ CN or a CH ₂ -CH ₂ -NH ₂ . 4) A compound of general formula I according to one of claims 1 to 3, characterized in that X and Y form a cycle with an OCH (CH ₃ ) OCH ₂ sequence.

5) A compound of general formula I according to one of claims 1 to 4, characterized in that it is chosen from the following compounds:

• 4'-Demethyl-4-0 (3-amino-4,6-ethylidene-2,3-dideoxy-β-D-arabinohexopyranosyl) epipodophyllotoxin,

• 4'-Demethyl-4-0 (3-dimethylamino-4,6-ethylidene-2,3-dideoxy-β-D-arabino-hexopyranosyl) epipodophyllotoxin,

• 4'-Demethyl-4-0 (3-dimethylamino-4,6-ethylidene-2,3-dideoxy-β-D-ribohexopyranosyl) epipodophyllotoxin,

• 4'-Demethyl-4-0 (3-cyanomethylamino-4,6-ethylidene-2,3-dideoxy-β-D-ribohexopyranosyl) epipodophyllotoxin,

• 4'-Demethyl-4-0 (3- (N-morpholino) -4,6-ethylidene-2,3-dideoxy-β-D-ribohexopyranosyl) epipodophyllotoxin,

• 4'-Demethyl-4-0 [3 (2-aminoethylamino) -4,6-ethylidene-2,3-dideoxy-β-D-ribohexopyranosyl)] epipodophyllotoxin,

• 4'-Demethyl-4-0 (3-amino-2,3,6-tridesoxy-β-D-ribohexopyranosyl)

epipodophyllotoxin,

• 4'-Demethyl-4-0 (3,6-diamino-2,3,6-tridesoxy-β-D-ribohexopyranosyl)

epipodophyllotoxin. 6) Compounds according to claim 4, characterized in that they are in the form of hydrochlorides.

7) Process for the preparation of a compound of general formula I according to claims 1 to 4, characterized in that a compound of formula III or IV or V is reacted with 4'-Demethyl 4'-benzyIoxycarbonyl epipodophyllotoxin with BF ₃ etherate, or trimethyl silyl trifluoromethanesulforate in an inert solvent at low temperature;

the primary amines in position 3 of the glycosyl are methylated by formalin and sodium cyanoborohydride. 8) Process for the preparation of a compound of formula IV characterized in that a mixture of diacetoxy azido glycoside VI is reacted

9) As a medicament, the compounds of formula I according to claims 1 to 6. 10) The pharmaceutical compositions characterized in that they contain at least one compound of formula I according to one of claims 1 to 6 and an appropriate excipient.

11) Use of a compound of formula I, according to one of claims 1 to 4, for the preparation of a medicament intended for anticancer treatment and, in particular, for treating forms of cancer, such as, for example, cancer small cell lung cancer, testicular cancer, embryonic tumors, neuroblastomas, kidney cancer, placental choriocarcinomas, breast adenocarcinomas, colorectal cancers, melanomas, Hodgkin and non-Hodgkin lymphomas and acute leukemias.