FI78109C - FOER FARING FOER FRAMSTAELLNING AV ANTRACYKLINGLYKOSIDER. - Google Patents

Description

7 ö 1 u y

METHOD FOR THE PREPARATION OF ANTHRACYCLINE GLYCOSIDES - FORM OF ANTACACYCLINGLYCOSIDES

The invention relates to an analogous process for the preparation of a therapeutically active anthracycline glycoside of general formula (A).

O OH

COCH2X

l | f %% * OH

O

* 2 KH2 in the formula X is a hydrogen atom or a hydroxy group, R1 is a hydrogen atom or a methoxy group, and R2 is a bromine, chlorine or fluorine atom. The invention also encompasses pharmaceutically acceptable acid addition salts of these anthracycline glycosides.

The anthracycline glycosides of formula (A) are prepared according to the invention by carrying out a substitution reaction of 4'-epi-4'-O-trifluoromethanesulfonyl-1-N-protected daunorubicin I 1 of general formula (II).

H3C-J

cf3so20-NH1 »a 78109 having a trifluoroacetyl or trichloroethoxycarbonyl group as defined above and dissolved in anhydrous methylene chloride (R5-COCF2) or acetonitrile (R8S-CO-O-CH2Cl2g), and tetra (n-butyl), ammonium tetra (n-butyl) ammonium chloride or cesium fluoride to give the N-protected glycoside of general formula III

0 OH

| C0CH3 L O OH! ! (III)

NHIU

"a wherein R is a hydrogen atom or a methoxy group, Rg is a bromine or chlorine atom and R1 is a trifluoroacetyl group, or R8 is a fluorine atom and R3 is a trichloroethoxycarbonyl group, and is subjected to mild alkaline hydrolysis (R3 = -COCF3) with 0.1 N aqueous sodium hydroxide. or (R 3 = -CO-O-CH 2 Cl 3) reductive deprotection with zinc powder and acetic acid in acetone at 0 ° C to give the free glycoside base of formula (A) (XH) which is isolated as the hydrochloride by treatment with methanolic hydrogen chloride and, if desired. this is reacted with bromine dissolved in chloroform to give the corresponding 14-bromo derivative, which is hydrolyzed with aqueous sodium formate to give the free glycoside base of formula (A) (X <0H), which is isolated as the hydrochloride.

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It should be noted that said preparation of the 14-bromo derivative and its treatment with an aqueous solution of sodium formate correspond to the procedure described in U.S. Patent No. 3,803,124.

When there is a bromine or chlorine atom, a substitution reaction is carried out between the compound of formula (II) and tetra- (n-butyl) ammonium bromide or chloride, and the amine protecting group R is preferably a trifluoroacetyl group. When R 2 is a fluorine atom, a substitution reaction is carried out between the compound of formula (II) and cesium fluoride and the amine protecting group R is preferably

O

trichloroethoxycarbonyl group.

The starting materials (II) are either previously known compounds, such as 4'-epi-4'-O-trifluoromethanesulfonyl-N-trifluoroacetyldaunorubicin (II: R 1 = OCH 9, R 2 = COCFg, see European Patent Application 8117618.1), or they can be prepared from previously known 4'-epi-daunorubicin derivatives by reaction with trifluoromethanesulfonic anhydride.

The compounds of the invention and their pharmaceutically acceptable acid addition salts can be used as antitumor drugs. These novel compounds of formula (A) are used in the treatment of various experimental tumors in mice, such as L1210, P388, P388 / 04 and the like, in the same way as their parent compounds (i.e. daunorubicin and doxorubicin). Screening experiments found that in many cases these new compounds are more active than the parent compounds. Accordingly, the invention also relates to pharmaceutical compositions comprising an anthracycline glycoside of the invention or a pharmaceutically acceptable salt thereof in admixture with a pharmaceutically acceptable diluent or carrier.

Biological activity tests

The compounds of Examples 1, 2 and 3 were compared in vitro to 4,710,109 daunorubicin (ONR) and doxorubicin (DX) on HeLa cells and DX-sensitive P388 cells as well as DX-resistant P388 cells (P388 / DX). The results are shown in Table I. All new derivatives appear to be more cytotoxic to HeLa cells and DX-sensitive P388 cells than their parent compound. This increased cytotoxicity is even more striking when considering the activity of these compounds on P388 / DX cells. In this case, a 100- to 250-fold increase in the cytotoxicity of the activity of these derivatives can be observed compared to the parent compound. These compounds were tested in vivo for three different experimental leukemias. The activity against ascites P388 leukemia is shown in Table II. The activity shown by the compounds of Example 3 is the same as that shown by the DNA, whereas the second DNA derivative, which is the compound of Example 1, has a significantly better antitumor effect than the DNA.

The activity of the highest tolerated dose of the compound of Example 2 (4.15 mg / kg) is approximately the same as that of DX. All of the new compounds are active against P388 / DX leukemia, as shown in Table III, whereas DNR and DX show no activity. These three new compounds were also tested for advanced Gross leukemia, the results of which are shown in Table IV. In this system, the compound of Example 3 is as active as the DNA, while the compound of Example 1 appears to be more active than its parent compound. The compound of Example 2 has approximately the same effect as DX. Two of these compounds were also tested when administered orally, showing interesting activity, while DNR and DX are inactive when administered orally.

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Table I

Cytotoxic activity of the compounds of Examples 1, 3 and 2 _________------------ _______ Compound _________ HeLa * _____ P388 ** ___ P388 / DX ^ ___ DNR 12.3 2.8 980

Example 1 6.8 0.5 4

Example 3 5.8 2.4 8.8 DX 12.5 4.25 1250

Example 2 3 0.2 7 - »1» M il. I · - .Ι.Ι · ΜΜΜ ··. Ι 1 · ^^ · .. - »|. · Ι - + Colony inhibition test with drug» 24 hours exposed ++ Cytotoxicity, assessed after 48 hours of drug initialization.

The results have been obtained from several experiments.

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Table II

a)

Effect on ascites P388 leukemia

Dosage ^ T / CC ^ LTS ^ Toxic

Compound (mg / kg)% of deaths-> cases

DNR \ ~ 2 ~ 9 Ϊ65 ~ * Ϊ55 Ö? 2Ö Ö? 2Q

I 4.4 179 135 0/18 0/18 j 6.6 130 115 0/20 12/20

Example 1 2.9 145 0/10 0/10 4.4 160 0/10 0/10 6.6 195 0/10 0/10 10 280 3/10 0/10

Example 3 2.9 130 0/10 0710 4.4 155 0/10 0/10 6.6 145 0/10 0/10 10 85 7/8 6/8 DX 4.4 215 0/10 0/10 6.6 235 0/10 0/10

Example 2 2.4 200 0/10 0/10 2.88 230 0/10 0/10 3.46 210 0/10 0/10 _______________ 4_j_15 ______ 325 _________ 4/10 _______ 1/10 ______ a) Experiments were performed on CDF ^ mice by inoculating 10 ^ leukemia cells i.p.

b) The treatment was performed i.p. day no. 1 from tumor inoculation.

c) Mean survival time of treated mice / mean survival time of controls x 100 d) Long-term survival (> 60 days) e) Estimated from necropsy findings.

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Table III

Effect on ascites P388 / DX leukemia3 ^

Dose ^^ T / Cc ^ Toxic

Compound (mg / kg)% of death.

cases0 DNR 474 91 Ö7IÖ 6.6 87 0/10

Example 1 4.4 143 0/10 10 143 0/10

Example 3 6.6 148 0/10 10 122 3/10 DX 6.6 108 1/20 10 117 1/20

Example 2 2.88 122 0/20 3.46 125 0/20 4.15 137 1/20 a) Experiments were performed on BDF. Mice inoculated with 6 cells i.p.

b) The treatment was performed i.p. day no. 1 from tumor inoculation.

c) Mean survival time of treated mice / mean survival time of controls x 100 d) Estimated from necropsy observations.

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Table IV

a)

Effect on Gross leukemia

Administration Dose T / C ° ^ Toxic

Compound mode (mg / kg)% of deaths ^ j DNA DN 1Ö Ϊ42 ~ Ϊ38 2 / Γθ 15 183 185 1/18 22.5 217 92 9/18

Example 1 iv 10 217 0/10 15 150 6/9 22.5 100 9/9

Example 3 iv 6.6 185 0/10 10 115 7/10 15 77 10/10 oral 6.6 154 0/10 via 10 169 0/10 15 92 7/8 DX iv 10 183 0/10 13 225 0 / 10 16.9 242 _ 0/10

Example 2 iv 4.05 233 0/10 5.27 258 0/10 6.85 242 3/10 oral 4.05 114 0/10 5.27 157 0/10 _____________i_______________________________________ a) Experiments were performed on CH mice 2 x 10 6 leukemia cells were transplanted iv

b) Treatment was performed i.v. or orally daily no. 1 from tumor inoculation.

c) Mean survival time of treated mice / mean survival time of controls x 100 d) Estimated from necropsy findings.

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The invention is further illustrated by the following examples.

Example 1: 4'-deoxy-4'-bromidaunorubicin (A: X * H, R 1 = OCH 3, R 2 = Br) 2 g of tetra- (n-butyl) ammonium bromide were added to a solution of 4.0 g of 4'- epi-4'-O-trifluoromethanesulfonyl-N-trifluoroacetyldaunorubicin (II: R1 = OCH2, R3 = COCF3) in 80 ml of anhydrous methylene dichloride. The reaction mixture was kept at room temperature for 1 hour and then washed with water and the organic phase was evaporated to dryness in vacuo. The residue was purified on silica gel using a mixture of methylene chloride and acetone as eluent to give 3.5 g of 4'-deoxy-4'-bromo-N-trifluoroacetyldaunorubicin, m.p. 130 ° C; FD-MS 685 (M +), TLC on silica gel plates (ferck F 254) using methylene chloride-acetone (10: 1 by volume) as solvent system Rf = 0.5.

To a solution of 3 g of 4'-deoxy-N-bromo-N-trifluoroacetyldaunorubicin in 20 ml of acetone was added 160 ml of 0.1 N aqueous sodium hydroxide solution. After 4 hours at 0 ° C, the solution was adjusted to pH 8.6 with 0.1 N hydrochloric acid and extracted with methylene chloride. The solvent was evaporated off and the residue was converted into the hydrochloride by treatment with methanolic hydrogen chloride (2.2 g), m.p. 180 ° C, decomposed, TLC on silica gel plates (Merck F 254) using methylene chloride-methanol-water-acetic acid (80: 20: 7: 3 by volume) as the solvent system gave an Rf value of 0.32.

Example 2: 4'-Deoxy-4'-bromidoxorubicin (A: X = OH, R1 = OCH3, R2 = Br) 2 g of 4'-deoxy-4'-bromodununubicin prepared as described in Example 1 were dissolved in methanol and to a mixture of dioxane. The solution was discussed in U.S. Pat. 3,803,124 to bromine to give the 14-bromo derivative, 10,781,09 followed by aqueous sodium formate to give 4'-deoxy-4'-bromidoxorubicin. This was converted to the hydrochloride by treatment with methanolic hydrogen chloride, m.p. 170 ° C (decomposes), FD-MS 605 (M +), TLC on silica gel plates (Merck F 254) using methylene chloride-methanol-water-acetic acid (80: 20: 7: 3 by volume) as solvent system gave an Rf value of 0 , 20.

Example 3: 41-deoxy-4'-chlorodaunorubicin (A: X = H, R1 = OCH3, R2 = Cl)

When 4'-epi-4'-O-trifluoromethanesulfonyl-N-trifluoroacetyldaunorubicin (II: R 2 = OCH 3, R 2 = COCF 4) was treated with tetra- (n-butyl) ammonium chloride according to Example 1, 4'- deoxy-4'-chloridaunorubicin, m.p. 175 ° C (decomposes), FD-MS 545 (M +), TLC on silica gel plates (Merck F 254) using a methylene chloride-methanol-water-acetic acid mixture (80: 20: 7: 3 by volume) as solvent system gave an Rf value of 0 , 32.

Example 4: 4'-Deoxy-4'-chlorodioxorubicin (A: X = OH, R1 = OCH3, R2 = Cl) 4'-Deoxy-4'-chlorodaunorubicin was converted by the method described in Example 2 to 4'-deoxy-4'-chloro. -doxorubicin isolated as the hydrochloride, m.p. 180 ° C (decomposes), FC mass spectrum 561 (M +), TLC on silica gel plates (Merck F 254) using methylene chloride-methanol-water-acetic acid (80: 20: 7: 3 by volume) as solvent system gave an Rf value of 0.2.

Example 5: 4'-Deoxy-4'-fluoridaunorubicin (A: X = H, R 1 = OCH 3, R 2 = F) in a stirred solution cooled to 0 ° C with 26 g of U.S. Pat. 4,345,068 of 4'-epi-N-trichloroethoxycarbonyldaunorubicin in 650 ml of anhydrous methylene dichloride and 32 ml of anhydrous pyridine (11 ml) were added, a solution of 11 ml of trifluoromethanesulfonic anhydride in 140 ml of anhydrous methylene chloride was added. The reaction mixture was washed with chilled 5% aqueous sodium bicarbonate solution, 0.1 N hydrochloric acid and water, respectively. The organic solution dried over anhydrous sodium sulfate was used in the next step without further purification.

A solution of 2 g of 41-epi-4'-O-trifluoromethanesulfonyl-N-trichloroethoxycarbonyldaunorubicin in 30 ml of acetonitrile was treated with 0.5 g of cesium chloride. The mixture was kept at room temperature for 10 minutes and then poured into water and extracted with methylene chloride. The organic phase dried over anhydrous sodium sulfate was evaporated to dryness in vacuo. The residue was dissolved in 30 ml of ethanol and treated with 4 ml of acetic acid and 0.2 g of zinc dust at 0 ° C. After 1 hour, the mixture filtered through Celite was diluted with water and extracted with methylene chloride. Evaporation of the solvent in vacuo gave a residue which was purified on a silica gel column using methylene dichloride as eluent to give pure 4'-deoxy-4'-fluoridaunorubicin which was isolated as the hydrochloride, FD-MS 529 (M +), TLC on silica gel plates using Merck F ridi-methanol-water-acetic acid mixture (80: 20: 7: 3 by volume) gave an Rf of 0.32.

Example 6: 4'-Deoxy-4'-fluoridoxorubicin (A: X = OH, R 1 = OCH 3, R 2 = F) 4'-Deoxy-4'-fluorodaunorubicin was converted by the method of Example 2 to 41-deoxy-4'-fluorodoxox. rubicin, which was isolated as the hydrochloride.

Claims (5)

12 78109 CLAIM Analogue method for the treatment of a therapeutically active anthracycline glycoside of general formula (A) 0 OH In '' OH i 10 OH! (A) * I 0 NH_ r 2 2 In which X is a hydrogen atom or a hydroxy group, R 1 is a hydrogen atom or a methoxy group and R 1 is a bromine, chlorine or fluorine atom, characterized in that a substitution reaction is carried out according to the general formula (II). 1-epi-41-O-trifluoromethanesulfonyl-N-protected daunorubicin 0 OH 1 ^ OH! ii 0 cf3s ° 2 ° - nhr3 78109 wherein R is as defined above and R is a trifluoroacetyl or X v trichloroethoxycarbonyl group dissolved in anhydrous methylene chloride (R = -COCF) or acetonitrile «3« 3 (RgS-CO-O-CH And tetra (n-butyl) ammonium bromide, tetra (n-butyl) ammonium chloride or cesium fluoride to give the N-protected glycoside of general formula III 0 OH