WO1996024602A1

WO1996024602A1 - Epipodophyllotoxin triaryloxyacetyl derivatives, method for preparing same, and use of said derivatives as cancer drugs

Info

Publication number: WO1996024602A1
Application number: PCT/FR1996/000201
Authority: WO
Inventors: Thierry Imbert; Yves Guminski; Francis Colpaert; Jean-Pierre Tarayre; Bridget Hill
Original assignee: Pierre Fabre Medicament
Priority date: 1995-02-08
Filing date: 1996-02-07
Publication date: 1996-08-15
Also published as: AU4722996A; EP0808319A1; FR2730233B1; CA2212956A1; FR2730233A1; JPH10513469A

Abstract

Derivatives of general formula (I), wherein R' is methyl or 2-thienyl, R is an acyl group of formula R1OCH2CO, wherein R1 is a phenyl ring ortho-, meta- or para-substituted by one or more substituents X, where X is a formyl, acetyl, cyano or trifluoroacetyl group, and pharmaceutically acceptable salts thereof, are disclosed. A method for preparing said derivatives, their use as a drug, and pharmaceutical compositions containing them, are also disclosed.

Description

TRIARYLOXYACETYL DERIVATIVES OF EPIPODOPHYLLOTOXINS,

THEIR PREPARATION PROCESS AND THEIR USE

AS ANTI-CANCER DRUGS

The present invention relates to new acylated derivatives of etoposide, their preparation process and their use as medicaments.

In the field of chemical compounds originating from natural lignans, there are hemisynthetic derivatives constituting the class of epipodophylloids, of which ethylidene glucoside from demethylepipodophyllotoxin (etoposide, R = H, R '= CH ₃ ) and thenethylidene glucoside of demethyl-epipodophyllotoxin (teniposide R = H, R '= 2-thienyl).

These two compounds are commonly used clinically as anticancer agents. Etoposide has anti-tumor properties to treat different forms of cancer, especially small cell lung cancer and testicular cancer. Both etoposide and teniposide are considered inhibitors of the DNA-linked topoisomerase II enzyme.

Topoisomerase II is a nuclear enzyme that has the ability to cut and reweld the two strands of DNA. Its inhibition by etoposide involves the formation and stabilization of a complex called a cleavable complex where DNA and the enzyme are linked by covalent bond. This stabilized complex (DNA-enzyme-drug) thus prevents the DNA from connecting and leads subsequently to cell death (DNA topoisomerase in cancer, Milan POTMESIL, Kurt M. OHN, Oxford University Press 1991, p. 230) .

There appear, during the treatment of patients with etoposide, a certain number of drawbacks as with most anticancer drugs in terms of undesirable side effects, poor tolerance, or even toxicity, related to the dose used, such as for example, hematological disorders (neutropenia, thrombocytopenia), cardiovascular disorders such as hypotension, or also nausea, vomiting or alopecia, and muscites in high doses. The anticancer efficacy of etoposide is therefore limited by these side effects.

Furthermore, it is shown that the therapeutic efficacy of etoposide is also limited in terms of survival by increasing the doses administered to mice in the P 388 leukemia model (see table). It is therefore necessary to look for new products which cause better survival, whatever the dose used, compared to etoposide. Particularly unexpectedly, this better survival is observed with the compounds of the present invention in the in vivo test in the model of leukemia P 388, and thus shows their better therapeutic anticancer efficacy.

The present invention therefore relates to the triacylated derivatives of ethylidene or of thenylidene glucoside of demethyl-epipodophyllotoxin of formula I

in which R represents an acyl group of phenoxy acetyl type of formula R10CH? CO, in which RI is a phenyl ring substituted by one or more substituents X, X representing a formyl, acetyl, cyano or trifluoroacetyl group, in ortho, meta or para, and R 'represents a methyl or 2-thienyl group, and their pharmaceutically acceptable salts.

Preferably, R 'represents a methyl group. Advantageously, RI represents a group chosen from 2-formyl phenyl, 4-formyl phenyl, 3-formyl phenyl, 4-acetyl phenyl, 4-cyano phenyl, 4-trifluoro acetyl phenyl and 2,4-diformyl phenyl.

Trisubstituted derivatives of formula I where R '= CH ₃ and R = aryloxyacetyl, are described in patent application FR-A-2 699 535. These compounds, which have shown improved anticancer activity compared to etoposide, do not not show as great a survival as the new derivatives according to the present invention. The particularly interesting activity observed for the derivatives according to the invention appears to come from the substitution of the aromatic nucleus of the phenoxyacetyl group R as defined above.

The present invention also relates to the processes for preparing a compound of formula I, for which the derivative of formula I above is reacted for which R represents a hydrogen atom and R 'is defined above, with a precursor of the acyl R defined previously, such as for example the activated acid RjOCF COOH, in particular an acid halide

R ι OCH ₂ COX, in which X represents a halogen such as chlorine or bromine, preferably chlorine.

They are preferably chlorides of acyls whose RI groups are defined above.

The acid chlorides used are prepared by the action of oxalyl chloride in CH ₂ CI ₂ at 0 ° C in the presence of traces of DMF. Coupling with the derivatives of formula I for which R = H is carried out directly in the same solvent with a molar ratio (acid chloride / derivative) of 5 to 8, in the presence of pyridine.

The derivatives of general formula I are thus obtained. The examples below describe the preparation of various compounds of general formula I according to the invention.

EXAMPLE 1:

4'-Demethyl 4- (2-formyl phenoxy acetyl) -4-0- (2,3-bis- (2-formyl phenoxy acetyl) -4,6-ethylidene-β-D-glucosyl) epipodophyllotoxin. In a 100 ml three-necked flask, 2.3 g of 2-formyl phenoxy acetic acid (12.7 mmol) are placed in solution in 50 ml of CH ₂ Cl? under argon away from humidity. An ice bath maintains the temperature of the reaction medium at 0 ° C., then 1 ml of DMF and 1.78 g of oxalyl chloride (14 mmol) are introduced dropwise into 5 ml of CH ₂ CI ₂ . The medium is stirred for 2 h at this temperature. This solution is then introduced dropwise into the reaction medium of a second 250 ml three-necked flask in which are placed 1.5 g of etoposide (2.55 mmol), 2.1 ml of pyridine in 50 ml of CH _2. CI ₂ with stirring at 0 ° C. The reaction medium is left stirring for 24 h with return to ambient temperature. The reaction medium is poured onto an HCl N solution (100 ml) then the organic phase is decanted, washed with saturated NaCl solution, then dried over Na ₂ S0 ₄ and evaporated. The residue is chromatographed on Siθ ₂ and eluted with a petroleum ether / AcOEt mixture: 60/40 then 50/50 to obtain 710 mg (Rd 25%) of triacylated derivative which is crystallized from isopropyl ether. Anal. : C ₅₆ H ₂₀ O ₂₂ PM = 1074.994 CH

Hold. 62.57 4.69

Tr. 62.32 4.82

F ~ 135 ° C

IRv (KBr) cm-i = 1774, 1687, 1601, 1485, 1460. Mass Spectrum (DCI / NH ₃ ) m / e = 1092 (M + NH ₃ ) iHRMN200MHzCDCl ₃ δ: 1.32 (3H, d, J = 4.9 Hz, H ₈ "); 2.8 (IH, m, H ₃ ); 3.15 (IH, dd, J = 14.1 Hz and 5.1 Hz, H ₂ ); 3, 64 (6H, s, OCH ₃ x 2); 5.33 (1H, t, J = 8.6 Hz, H ₃ ");

5.65 (1H, s, OCH _A O); 5.86 (1H, s, OCH _B 0); 6.24 (2H, s, H ₂ 'H ₆ '); 6.49 (1H, s, H ₈ ');

6.46 (1H, d, J = 8.2 Hz, OAr: H ortho); 6.76 (1H, s, H ₅ '); 6.75 (1H, d, J = 8.2 Hz, OAr: H ortho); 6.96-7.01 (4H, m, OAr: H ortho, 3H para); 7.33-7.55 (3H, m, OAr: 3H meta); 7.79-7.86 (3H, m, OAr: 3H meta); 10.42 (1H, s, CHO); 10.49 (1H, s, CHO); 10.54 (1H, s, CHO).

EXAMPLE 2: 4'-Demethyl 4 '- (4-formyl phenoxy acetyl) -4-0- (2,3-bis- (4-form> 1 phenoxy acetyl) -4,6-ethylidene-β-D-glucosyl ) epipodophyllotoxin.

By the same method as for Example 1, but using 4-formyl phenoxy acetic acid in place of 2-formyl phenoxy acetic acid, the desired triacylated compound is obtained by chromatography with elution in an ether mixture. petroleum / AcOEt 50/50 then 40/60 and crystallization from ethyl ether with a yield of 60%. Anal. : C ₅₆ H ₅₀ Q ₂₂ 0.55 H ₂ 0 PM = 1084.884

CHH ₂ 0 Wedge. 62.57 4.69

Tr. 61.54 4.69 0.92% adjusted 62.00 4.75 0.92%

F ~ 140 ° C

IR v (KBr) cm- '= 1774, 1691, 1508 Mass spectrum (DCI / NH ₃ ) m / e = 1092 (M + NH ₃ ) • H NMR 200 MHz CDC1 ₃ δ: 1.31 (3H, d, J = 4.8 Hz, H ₈ "); 2.8 (IH, m, H ₃ ); 3.12 (IH, dd, J = 14.1 Hz and 5.08 Hz, H ₂ ); 3.62 (6H, s, 2 x OCH ₃ ); 5.30 (IH, t, J = 8.9 Hz, H ₃ '); 5 , 68 (1H, s, OCH _A O); 5.87 (1H, s, OCH _B 0); 6.22 (2H, s, H ₂ 'H ₆ '); 6.49 (1H, s, H ₈ '); 6.75 (1H, s, H ₅ '); 6.77 (2H, d, J = 7.49 Hz, OAr: H ortho); 6.91 (2H, d, J = 8, 6 Hz, OAr: H ortho); 7.04 (2H, d, J = 8.6 Hz, OAr: H ortho); 7.7-7.8 (6H, m, OAr: H meta); 9.77 ( 1H, s, CHO); 9.80 (1H, s, CHO); 9.85 (1H, s, CHO).

EXAMPLE 3:

4'-Demethyl 4 '- (3-formyl phenoxy acetyl) -4-0- (2,3-bis- (3-formyl phenoxy acetyl) -4,6-ethylidene-β-D-glucosyl) epipodophyllotoxin.

By the same method as for Example 1, but using 3-formyl phenoxy acetic acid in place of 2-formyl phenoxy acetic, the triacylated derivative is obtained with a yield of 33% by chromatography on Siθ ₂ and elution with a mixture of CH ₂ CI ₂ / Acetone 99/1 then 95/5 and crystallization from ethyl acetate. F ~ 125 ° C.

Anal. : C ₅₆ H5o θ22 PM = 1074.994

C H

Hold. 62.57 4.69 Tr. 62.09 4.70

EXAMPLE 4:

4'-Demethyl 4 '- (4-cyano phenoxy acetyl) -4-0- (2,3-bis- (4-cyano phenoxy acetyl) -4,6-ethylidene-β-D-glucosyl) epipodophyllotoxin. By the same method as for Example 1, but using 4-cyano-phenoxy acetic acid instead of 2-formyl phenoxy acetic acid, the following is obtained after chromatography on Si0 ₂ and elution with a petroleum ether / AcOEt 50/50 and crystallization from isopropyl ether of the triacylated derivative with a yield of 75%. F ~ 153 ° C

Anal. : C ₅₆ H ₄₇ N ₃ O _i9 PM = 1066.01

C H N

Hold. 63.09 4.44 3.94

Tr. 62.77 4.63 3.82 IR v (KBr) cm-i = 2226, 1774, 1606, 1506, 1485 Mass spectrum (FAB) m / e = 1088 (M + Na) + iH 200 MHz N NMR: 1.36 (3H, d, J = 4.9 Hz, H ₈ "); 2.88 (IH, m , H ₃ ); 3.15 (1H, dd, J

= 5.1 Hz and 14 Hz, H ₂ ); 5.31 (1H, t, J = 8.8 Hz, H ₃ "); 5.76 (1H, s, OCH _A 0); 5.94 (1H, s, OCH _B O); 6.27 ( 2H, s, H ₂ 'H ₆ '); 6.55 (IH, s, H ₈ '); 6.79 (IH, s, H ₅ '); 6.75 (2H, d, j = 9 Hz , OAr: H ortho); 6.90 (2H, d, J = 9 Hz, OAr: H ortho); 7.04 (2H, d, J = 9 Hz, OAr: H ortho); 7.5 1- 7.62 (6H, m, OAr: H meta).

EXAMPLE 5:

4'-Demethyl 4 '- (4-acetyl phenoxy acetyl) -4-0- (2,3-bis- (4-acetyl phenoxy acetyl) -4,6-ethylidene-β-D-glucosyl) epipodophyllotoxin.

By the same method as for Example 1, but using 4-acetyl phenoxy acetic acid instead of 2-formyl phenoxy acetic acid, one obtains with a yield of 77% by chromatography on Si0 ₂ and elution by a heptane / AcOEt 60/40 mixture and crystallization from ethanol, the triacylated derivative of etoposide. F ~ 125 ° C Anal. : C ₅₉ H ₅₆ O ₂₂ H ₂₀ PM = 1135,105

C H

Hold. 63.44 5.05 Tr. 62.43 5.15

IR v (KBr) cm-ι = 3624, 2920, 1774, 1880, 1601 iH NMR 200 MHz DMSO δ: 1.24 (3H, d, J = 4.8 Hz, H ₈ "); 2.47 (3H , s, COCH ₃ ); 2.51

(3H, s, COCH ₃ ); 2.53 (3H, s, COCH ₃ ); 2.9-3, 1 (2H, m, H ₂ H ₃ ); 5.20 (2H, s, OCH ₂ CO);

5.3-5.4 (2H, m, H ₂ "H ₃ "); 5.79 (1H, s, OCH _A 0); 5.98 (1H, s, OCH _B 0); 6.29 (2H, s, H ₂ "H ₆ '); 6.52 (1 H, s, H ₈ '); 7.1 (1H, s, H ₅ '); 7.81 (2H, d, J = 8.7 Hz, OAr: H ortho);

6.97 (2H, d, J = 8.7 Hz, OAr: H ortho); 7.06 (2H, d, J = 8.7 Hz, OAr: H ortho); 7.83-7.97 (6H, d, J = 8.7 Hz, OAr: H meta).

By the same method the following compounds are prepared:

• 4'-Demethyl 4 '- (4-trifluoro acetyl phenoxy acetyl) -4-0- (2,3-bis- (4-tri- fluoro acetyl phenoxy acetyl) -4,6-ethylidene-β-D-glucosyl ) epipodophyllotoxin;

• 4'-Demethyl, 4 '- (2,4-diformyl phenoxy acetyl) -4-0- (2,3-bis- (2,4-diformyl phenoxy acetyl) -4,6-ethylidene-β-D- glucosyl) epipodophyllotoxin;

• 4'-Demethyl-4 '- (4-formyl phenoxy acetyl) -4-0- (2,3-bis- (4-formyl phenoxy acetyl) -4,6-thenylidene-β-D-glucosyl) epipodophyllotoxin. ORGANIC PART

The products of the present invention are tested in the model of 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 the test not very discriminating.

We therefore modified the test protocol to make it more selective. The administration of tumor cells is done intravenously and not intraperitoneally. The tumor cells then spread throughout the body very quickly. The administration of the test compound is then done intraperitoneally. Two parameters are thus defined to highlight the activity of the compounds:

- Determination of the effective dose 50, which represents the minimum dose of the compound to be administered to obtain significant survival of the animals compared to the control animals; - determination of the maximum survival time of the animals, whatever the dose administered. The fact of being able to administer large doses of the compound, and of observing a significant survival, makes it possible to obtain a measure of the maximum therapeutic efficacy of the product which can be achieved. It has been unexpectedly found that the compounds of the present invention cause exceptionally high survival compared to other commonly used topoisomerase II inhibitors. These results were sufficiently surprising to justify the comparative study with numerous reference compounds, whatever their chemical structure such as etoposide, etopofos, teniposide, amonafide, amsacrine, intoplicin, bisantrene, daunorubicin, and doxorubicin. 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 CDF1 hybrid female mice (Balb / c females x DBA / 2 males) of 20 ± 2 g (Cancer Chemother. Rep. 3: 9,

1972).

The tumor cells are implanted intravenously (106 cells / mouse) on day 0. The animals are randomized and divided into groups of two 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 survival of the treated animals. 86% of the control mice die on the 7th day after the tumor transplant. A substance will be considered active if it induces survival longer than eight days.

The biological results are summarized in the following table

S urvie

P 388 Maximum T / C *

^DE so mg / kg (J)%

Controlled animals with iv administration of (6-8) tumor cells Median 7

Eloposide 10 19 271

Etopofos 28 17 242

Teniposide 20 15 214

Intoplicin 28 10 142

Iiiaclif amonafid (10-160)

Amsacrine iiiaclif (10-160)

Bisantrene inactive (10-160) toxic at 640

Daunorubicin inactive (2.5-10) toxic to 40

Doxorubicin 7 13 185

Composed of 1 "example 18 - - 285 (FR-A-2 699 535)

Compound of example 19 - - 164 (FR-A-2 699 535)

Compound of example 1 - - 500

Compound of ^Example 2 20 48 685

Compound of Example 5 - - 357

* The T / C value represents the ratio between the average of the group of animals treated and the average survival of the group of animals checked.

It is thus demonstrated that, on the one hand, among the reference products acting on topoisomerase II, the compound causing the best survival is etoposide, and on the other hand, the compound of Example 2 according to l The invention causes a maximum survival 2.5 times greater than etoposide. The compounds according to the invention also exhibit a maximum survival up to 4 times greater than that of the compounds described in FR-A-2 699 535.

These results therefore show the advantage of the products of the invention and their usefulness as anticancer therapeutic agents causing very high survival, over a very wide dose range. It may be appreciated in the light of these results the usefulness of these compounds as a medicament for treating the various forms of cancer in which topoisomerase II inhibitors are used, such as in particular small cell lung cancer, cancer of the testes, embryonic tumors, neuroblastomas, kidney cancer, Hodgkin and non-Hodgkin lymphomas, acute and relapsed leukemias, placental choriocarcinomas, breast adenocarcinomas, as well as to increase the therapeutic efficacy of topoisomerase inhibitor compounds the treatment of tumors refractory to usual therapies: colorectal cancers and melanomas.

The present invention therefore also relates to the derivatives of general formula I defined above for their use as a medicament and their use for the preparation of a medicament intended for anticancer treatment, in particular of the various forms of cancer in which the topoisomerase inhibitors They are used.

The present invention finally relates to pharmaceutical compositions comprising at least one compound of general formula I according to the invention, and a suitable excipient. The pharmaceutical compositions can be adapted for administration by the injectable or oral route in the form of capsules, capsules, tablets, at a dosage of 2 to 200 mg / iM per 24 h by injection and from 5 to 400 mg. / m2 per 24 hours for the oral route.

Claims

1. Compounds of formula I

in which R 'represents a methyl or a 2-thienyl, R represents an acyl group of formula R10CH CO, in which RI is a phenyl ring substituted by one or more substituents X, in ortho, meta or para position, X representing a group formyl, acetyl, cyano or trifluoro acetyl, and their pharmaceutically acceptable salts.

2. Compound of formula I according to claim 1, characterized in that R 'represents a methyl group.

3. Compound of formula I according to claims 1 and 2, characterized in that RI represents a group chosen from 2-formyl phenyl, 4-formyl phenyl, 3-formyl phenyl, 4-acetyl phenyl, 4-cyano phenyl, 4 -trifluoro acetyl phenyl and 2,4-diformyl phenyl.

4. Compound of formula I according to one of claims 1 to 3, characterized in that it is chosen from:

4'-Demethyl-4 '- (2-formyl phenoxy acetyl) -4-0- (2,3-bis- (2-formyl phenoxy acetyl) -4,6-ethylidene-β-D-glucosyl) epipodophyllotoxin, 4 '-Demethyl-4' - (4-formyl phenoxy acetyl) -4-0- (2,3-bis- (4-formyl phenoxy acetyl) -4,6-ethylidene-β-D-glucosyl) epipodophyllotoxin, 4 ' -Demethyl-4 '- (4-formyl phenoxy acetyl) -4-0- (2,3-bis- (4-formyl phenoxy acetyl) -4,6-thenylidene-β-D-glucosyl) epipodophyllotoxin, 4'- Demethyl-4 '- (3-formyl phenoxy acetyl) -4-0- (2,3-bis- (3-formyl phenoxy acetyl) -4,6-ethylidene-β-D-glucosyl) epipodophyllotoxin, 4'-Demethyl-4 '- (4-acetyl phenoxy acetyl) -4-0- (2,3-bis- (4-acetyl phenoxy acetyl) -4,6-eth>' lidene-β-D-glucosyl) epipodophyllotoxin, 4'-Demethyl-4 '- (4-cyano phenoxy acetyl) -4-0- (2,3-bis- (4-cyano phenoxy acetyl) -4,6-ethylidene-β-D-glucosyl) epipodophyllotoxin , 4'-Demethyl-4 '- (4-trifluoro acetyl phenoxy acetyl) -4-0- (2,3-bis- (4-trifluoro phenoxy acetyl) -4,6-ethylidene-β-D-glucosyl) epipodophyllotoxin , 4'-Demethyl-4 '- (2,4-diformyl phenoxy acetyl) -4-0- (2,3-bis- (2,4-diformyl phenoxy acetyl) -4,6-ethylidene-β-D- glucosyl) epipodophyllotoxin.

5. Method for preparing a compound according to one of claims 1 to 4, characterized in that a compound of general formula I is reacted

for which R = H, and R 'is defined in claims 1 or 2, in the presence of pyridine with an acid chloride, derived from the acid of formula R10CH ₂ C0 ₂ H, where RI has the characteristics defined in Claims 1 or 3, prepared by the action of oxalyl chloride in the presence of DMF.

6. As a medicament, the compounds of formula I according to one of claims 1 to 4.

7. Pharmaceutical composition characterized in that it comprises at least one compound of formula I according to one of claims

1 to 4, and a suitable excipient.

8. 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 where topoisomerase II inhibitors are used, such as, for example, small cell lung cancer, testicular cancer, embryonic tumors, neuroblastomas, kidney cancer, Hodgkin and non-Hodgkin lymphomas, acute leukemias, and in relapse, placental choriocarcinomas, mammary adenocarcinomas, as well as to increase the therapeutic efficacy of topoisomerase inhibitor compounds for the treatment of tumors refractory to usual therapies: colorectal cancers and melanomas.