CS270411B2 - Method of new 4 halogen anthracycline glycosides production - Google Patents

Abstract

<IMAGE> Compounds of the above formula (X=H or OH, R1=H, OH or OCH3, R2=Br, Cl or F) and their acid addition salts are useful as antitumour agents. Those in which X=H may be prepared by an exchange reaction between a 4'-epi-4'-O-trifluoromethanesulphonyl-N-protected-daunorubicin (or a corresponding 4-demethyl or 4-demethoxy derivative) and (C4H9)4NCl, (C4H9)4NBr or CsF. Those in which X=OH are prepared from those in which X=H by bromination and reaction with aqueous sodium formate.

Description

The present invention relates to a process for the preparation of novel 4'-haloanthracycline glycosides which are useful as antitumor agents.

The present invention relates to a process for the preparation of anthracycline glycosides of the general formula A wherein:

X is hydrogen ^R 2 is bromine,

or a hydroxyl group, chlorine or fluorine, as well as reacting them with a pharmaceutically usable compound of both addition salts of formula II

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with acids,

wherein ^R 3 is an amino protecting group,

With a compound introducing a bromine, chlorine or fluorine atom as a substituent ₽ ₂ , selected from the group consisting of cesium fluoride, tetra- (n-butyl) ammonium bromide and tetra- (n-butyl) ammonium chloride, optionally in the presence of an inert organic solvent, from the compound of formula III obtained

in which

R? and Rj are as defined above, deprotecting to form compounds of formula A wherein X is hydrogen, then optionally brominating the obtained compounds and then treating the obtained 14-bromo derivatives with aqueous sodium formate to form compounds of formula A, wherein X is hydroxy and the compounds of formula (A) are optionally converted to pharmaceutically acceptable acid addition salts.

When preparing compounds of formula A wherein R ₂ represents a bromine or chlorine atom, then the compounds of formula II include preferably reacted with tetra (n-butyDamoniumbromidem optionally with tetra- (n-butyl) ammonium chloride and the protecting group The protecting group can then be removed by mild alkaline hydrolysis, for example by treatment with a 0.1N aqueous sodium hydroxide solution.

When compounds of formula (A) in which R @ ₂ is fluorine are prepared, the compounds of formula (II) are preferably reacted with cesium fluoride and the amino protecting group R @ 1 is preferably trichloroethoxycarbonyl. In this case, the protecting group is preferably removed by hydrogenation, for example with powdered, zinc and acetic acid.

Said reactions may be carried out in an inert organic solvent such as methylene chloride or acetonitrile.

The starting materials of formula II are either known compounds, for example, 4'-epi-4 -0-trifluoromethanesulfonyl-N-trifluoroacetyldaunorubicin (II, R₁ = _{3 C0CF>} cf. European Patent Application 8117618.1), or readily prepared from known derivatives of 4 ' -epidaunorubicin by reaction with trifluoromethanesulfonic anhydride.

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The compounds obtained by the process of the invention and their pharmaceutically acceptable acid addition salts are useful as antitumor agents. These novel compounds of formula (A) are used in the same manner as the parent compounds (i.e., daunorubicin and doxorubicin) to treat various experimental mouse tumors, such as L1210, Θ3Θ8, Ρ3Θ8 / 04, and the like. in many cases more potent than the parent compounds. Accordingly, the present invention also relates to pharmaceutical compositions comprising an anthracycline glycoside produced according to the invention or a pharmaceutically acceptable acid addition salt thereof in admixture with a diluent or carrier that is pharmaceutically acceptable.

Tests illustrating biological efficacy:

The compounds of Examples 1, 2 and 3 were tested in vitro compared to daunorubicin (DNR) and doxorubicin (DX) for potency against HeLa cells and for potency against P388 cells that are sensitive and resistant to doxorubicin (P388 / DX) ). The results are shown in Table I. All new derivatives appear to be more cytotoxic to HeLa cells and P388 cells sensitive to doxorubicin than their parent compound. However, this increased cytotoxicity is significantly more pronounced when the activity of these compounds against P388 / DX is observed. In this case, a 100- to 250-fold increase in the cytotoxicity of these derivatives was found compared to the parent drug. In vivo, these compounds are tested against three different types of experimental leukemia. The anti-tumor activity against P388 ascitic leukemia is shown in Table II. The activity of the compounds of Example 3 is the same as that of daunorubicin (ONR), while another ONR derivative, ie the compound of Example 1, has an anti-tumor activity that is definitely higher than the anti-tumor activity of DNR. *

The compound of Example 2 has about the same potency as DX at the highest tolerated dose (4.15 mg / kg). All novel compounds are active against P388 / DX leukemia as shown in Table III, while daunorubicin (ONR) and doxorubicin (DX) show no effect in this case. These three novel compounds were also tested after i.v. application to augmented Gross Leukemia, the results of which are shown in Table IV. In this system, the compound of Example 3 is as effective as the DNR, while the compound of Example 1 appears to be more potent than the parent compound. The compound of Example 2 shows about the same activity as DX. Two of these compounds have also been tested after oral administration, showing an interesting effect, while DNR and DX are ineffective when administered orally.

Table I ·

Cytotoxic effect of the compounds of Examples 1, 2 3 and ^I0 of 50 <9 ⁿ / ml)

Compound HeLa ⁺ P388 ⁺⁺ P388 / DX ⁺⁺ DNR 12.3 2.8 980 from Example 1 6.8 0.5 4 from Example 3 5.8 2.4 8.8 DX . 12.5 4.25 1250 from Example 2 3 0.2 7

+ colony suppression assay after 24 hours treatment with test compounds ++ cytotoxicity, evaluated after 48 hours treatment with test compounds All data are from several experiments.

CS 270411 02

Table II

, · A) Effect against ascites leukemia Ρ3Θ8 compound dose (mg / kg) T / C ^(c) % LTC ^d) E) death due to toxicity DNR 2.9 165 155 0/20 0/20 4.4 170 135 0/18 0/18 6.6 130 115 0/20 12/20 from the 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 from the example 3 2,9 130 0/10 0/10 .4,4 155 0/10 0/10 6.6 145 0/10 0/10 10 85 0/8 6/8 DX 4.4 215 0/10 0/10 6.6 235 0/10 0/10 10 335 4/10 0/10 from the examples 2 2,4 200 0/10 0/10 2.88 230 0/10 0/10 3.46 210 0/10 0/10 4.15 325 4/10 1/10 Legenda к Table II; (a) experiments are performed on mice (strain COF.) infected i. p. 10? leukemia cells b) i.p. on day 1 after tumor inoculation (c) medium survival time of treated mice / mean survival time control mice x 100 (d) long survival time (more than 60 days) (e) evaluated on the basis of the findings at autopsy.

Table ¹¹¹ Effect against ascitic leukemia P388 / DX ^a) compound dose ^b ) T / C ^(c) death due to (mg / kg) % toxicity DNR 4.4 91 0/10 6.6 87 0/10 from Example 1 4.4 143 0/10 10 143 0/10 from Example 3 6.6 148 0/10 10 122 3/10 DX 6.6 108 1/20 10 117 1/20 from Example 2 2.88 122 0/20 3.46 125 0/20 4.15 137 1/20

a) experiments on ss performed on mice (strain BDF _X ) infected 10 µl cells

b) i.p. on day 1 after tumor inoculation

c) median survival of treated mice / median survival of control mice x 100

(d) evaluated on the basis of autopsy findings

Table IV * a)

Effect against Gross leukemia

compound method of application dose (mg / kg) T / C ^(c) % mortality due to toxicity DNR i. v. 10 142 138 2/18 15 Dec 183 185 1/18 22.5 217 92 9/18 from Example 1 i. v. Ю 217 0/10 15 Dec 150 6/9 22.5 100 ALIGN! 9/9 from Example 3 i. v. 6.6 185 0/10 10 115 7/10 15 Dec 77 10/10 orally 6.6 154 0/10 10 169 0/10 15 Dec 92 7/8 DX i .v. 10 183 . 0/10 13 225 0/10 • 16.9 242 0/10 from Example 2 i * v · 4.05 233 0/10, 5.27 258 0/10 6.85 ‘242 3/10 orally 4.05 114 0/10 5.27 157 0/10

Legend:

a) experiments were performed on mice (strain C ^ H) infected with iv 2 χ 10 ⁶ leukemia cells

b) i.v. on day 1 after tumor inoculation

c) median survival of treated mice / median survival of control mice x 100

(d) evaluated on the basis of autopsy findings.

The following examples illustrate the process of the invention in more detail:

EXAMPLE '-deoxy-4 -bromdaunorubicin _(A X = H, R ₂ = Br)

02 g of tetra- (n-butyl) ammonium bromide are added to a solution of 4.0 g of 4-epi-4-O-trifluoromethanesulfonyl-N-trifluoroacetyldaunorubicin (II: R ^ = OCHp = COCF 3) in 80 ml of anhydrous methylene chloride. The reaction mixture is left at room temperature for 1 hour, washed with water and the organic phase is evaporated in vacuo. The evaporation residue is purified on silica gel, eluting with a mixture of methylene chloride and acetone. 3.5 g of 4'-deoxy-4-bromo-N-trifluoroacetyldaunorubicin are obtained, m.p. 130 ° C;

mass spectrometry (field desorption) 685 (M &^lt; ⁺ &^gt;);

thin layer chromatography (silica gel plates Merck F 254) using cozpouštědlového system comprising a mixture of methylene chloride and acetone (10: 1, by volume): _Rf = 0.5.

To a solution of 3 g of 4-deoxy-4-bromo-N-trifluoroacetyldaunorubicin in 20 ml of acetone was added 160 ml of a 0.1N aqueous sodium hydroxide solution. After 4 hours at 0 ° C, the solution is adjusted to pH 8.6 by addition of 0.1N hydrochloric acid solution and extracted with methylene chloride. The solvent was evaporated to give a residue which was converted to the hydrochloride by treatment with methanolic hydrogen chloride (yield 2.2 g); mp 180 ° C (dec.); thin layer chromatography of silica gel plates (Merck F 254) using a solvent system consisting of a mixture of methylene chloride, methanol, water and acetic acid (80: 20: 7: 3, by volume): Rf = 0.32.

Example 2

-deoxy-4'-bromdoxorubicin (A: X with OH, R ₂ = Br) g 4 -deoxy-4-bromounorubicin, prepared as described in Example 1, was dissolved in a mixture of methanol and dioxane. This solution is then first treated with bromine (as described in U.S. Pat. No. 3,883,124) to give the 14-bromo derivative and then with aqueous sodium formate to give 4'-deoxy-4'-bromo-oxorubicin. This material was converted to the hydrochloride by treatment with methanolic hydrogen chloride, m.p. 170 ° C (dec.); Mass Spectrometry (Field Desorption): 605 (M ⁺ ); TLC (silica gel plates Merck F 254) using the solvent system methylene chloride: methanol: water: acetic acid (80: 20: 7: 3 by volume) R _f = 0.20.

Example 3

-deoxy-4 -chlordaunorubicin (А: X - H, _R? = Cl)

Reaction of 4'-epi-4-O-trifluoromethanesulfonyl-N-trifluoroacetyldaunorubicin (II: = nCH 4, R 4 = COCF 4) □ tetra- (n-butyl) ammonium chloride by the procedure described in Example 1 affords 4-deoxy-4 ' ordlordaunorubicin, m.p. 175 ° C (dec.); Mass Spectrometry (Field Desorption): 545 (M ⁺ );

TLC (silica gel plates Merck F 254) using the solvent system methylene chloride: methanol: water: acetic acid (80:20: 7: 3) (by volume): _Rf = 0.32.

Example 4

-deoxy-4-chlorodoxerubicin (A: X = OH, R = Cl)

Following the procedure described in Example 2, 2-deoxy-4-chlorodunorubicin was converted to 4'-deoxy-4-chlorodoxorubicin, which was isolated as the hydrochloride, m.p. 180 DEG C. (decomposition);

Mass spectrometry (field desorption): 561 (M ⁺ ) 1, thin layer chromatography on silica gel plates (Merck F 254) using a solvent system consisting of a mixture of methylene chloride, methanol, water and acetic acid 1 (80: 20: 7: 3 based on volume): _Rf = 0.2.

Example 1 a d 5

-deoxy-4-fluoroaunorubicin (A: X = H, R = F)

To a stirred solution of 26 g of 4'-epi-N-trichloroethoxycarbonyldaunorubicin as described in U.S. Pat. No. 4,345,068 in 650 ml of anhydrous methylene chloride and 32 ml of anhydrous pyridine cooled to 0 DEG C. was added a solution of 11 ml of trifluoromethanesulfonic anhydride. acid in 140 mL of anhydrous methylene chloride. The reaction mixture was washed with cooled 5% aqueous sodium bicarbonate solution, water, 0.1N hydrochloric acid solution and water, respectively. The organic solution dried over anhydrous sodium sulfate was used for the next step without further purification.

To a solution of 2 g of 4'-epi-4'-O-trifluoromethanesulfonyl-N-trichloroethoxycarbonyldaunorubicin in 30 ml of acetonitrile was added 0.5 g of cesium fluoride. After 10 minutes at room temperature, the mixture was poured into water and extracted with methylene chloride. The organic phase dried over anhydrous sodium sulfate was evaporated to dryness in vacuo. To the residue dissolved in 30 ml of ethanol, 4 ml of acetic acid and 0.2 g of zinc powder are added at 0 ° C. After 1 hour, the mixture filtered on Celite was diluted with water and then extracted with methylene chloride. Evaporation of the solvent in vacuo gave the crude product which was chromatographed on a silica gel column using methylene chloride as eluent to give pure 4-deoxy-4-fluoroaunorubicin which was isolated as the hydrochloride.

Irradiated Irrigation (Field Desorption): 529 (M ⁺ ).

Chrum log Ladie on thin layers of silica gel plates (Morek 254 254) using a solvent system consisting of a mixture of methylene chloride, methanol, water and acetic acid (80: 20: 7: 3, based on volume): = 0.32.

Example 6

-deoxy-4 -fluoro-doxorubicin (А: X = OH, R ₂ = F)

Following the procedure described in Example 2, 4-deoxy-4'-fluoroaunorubicin is converted to 4-deoxy-4'-fluordoxorubicin, which is isolated as the hydrochloride.

Claims (8)

A process for the preparation of novel 4-haloanthracycline glycosides of the general formula A in which X represents a hydrogen atom or a hydroxyl group, R ₂ is bromine, chlorine or fluorine, and their pharmaceutically usable acid addition salts, characterized by reacting a compound of formula II known. · Wherein R 1 is an amino protecting group that is a bromine, chlorine or fluorine introducing compound R ₂ selected from cesium fluoride, tetra- (n-butyl) ammonium bromide and tetra- (n-butyl) amo nium chloride, optionally in the presence of an inert organic solvent, from the obtained compound of formula III in which R2 and R1 are as defined above, deprotecting to form compounds of formula (A) wherein X is hydrogen, whereupon the compounds are optionally brominated, and then the obtained 14-bromo derivatives are treated with aqueous sodium formate to form compounds of formula A wherein X is hydroxy and the compounds of formula (A) obtained are optionally converted to pharmaceutically usable acid addition salts. 2. The process of claim 1 wherein the amino protecting group is trichloroethoxycarbonyl. 3. A process according to claim 2, wherein the deprotection is carried out by hydrogenation. . 4. Process according to claim 2 or 3, characterized in that zinc powder and acetic acid are used to remove the protecting group. 5. The process of claim 1 wherein the amino protecting group is trifluoroacetyl. fj 6. A process according to claim 5 wherein the protecting group is removed by mild alkaline hydrolysis. 7 7. A process according to claim 5 or 6, wherein 0.1 N aqueous sodium hydroxide solution is used to remove the protecting group. Process according to one of Claims 1 to 7, characterized in that methylene chloride or acetonitrile is used as the inert organic solvent.