KR20010079665A - New use of taxoid derivatives - Google Patents

Abstract

The present invention relates to novel uses of taxoid derivatives. More specifically, it relates to a method of treating abnormal cell proliferation of different resistant cell lines expressing the complex-tolerant P-glycoprotein. This type of cell is typical of colon cancer cells. The products of the present invention can also be used to treat colon cancer in mammals, including humans. Preferred compounds are 4α-acetoxy-2α-benzoyloxy-5β, 20-epoxy-1β-hydroxy-7β-dimethoxy-9-oxo-11-taxen-13α-yl (2R, 3S) -3-tert- Butoxycarbonylamino-2-hydroxy-3-phenyl-propionate.

Description

New Uses of Taxoid Derivatives {NEW USE OF TAXOID DERIVATIVES}

The present invention relates to novel uses of taxoid derivatives. More specifically, it relates to a method of treating abnormal cell proliferation of different resistant cell lines expressing the complex-tolerant P-glycoprotein. This type of cell is typical of colon cancer cells. The present invention relates to the treatment of abnormal cell proliferation of different resistant cell lines expressing complex-tolerant P-glycoprotein and also to the treatment of abnormal cell proliferation of different resistant cell lines expressing complex-tolerant P-glycoprotein. The present invention relates to the treatment of colon tumors in mammals, including humans, by administration of the compound of 1 or a pharmaceutically acceptable salt or solvent compound thereof.

In the above formula,

Z represents a hydrogen atom or a radical of formula (2),

R ₁ is

Benzoyl radicals optionally substituted with one or more identical or different atoms or radicals selected from halogen atoms and alkyl radicals containing 1 to 4 carbon atoms, alkoxy radicals containing 1 to 4 carbon atoms or trifluoromethyl radicals,

A tennoyl or furoyl radical or a radical R ₂ -O-CO-,

R ₂ is

Alkyl radicals containing 1 to 8 carbon atoms,

Alkenyl radicals containing 2 to 8 carbon atoms,

Alkynyl radicals containing 3 to 8 carbon atoms,

Cycloalkyl radicals containing 3 to 6 carbon atoms,

Cycloalkenyl radicals containing 4 to 6 carbon atoms or

Bicycloalkyl radicals containing 7 to 10 carbon atoms (these radicals being halogen and hydroxyl radicals, alkoxy radicals containing 1 to 4 carbon atoms, each alkyl moiety containing 1 to 4 carbon atoms) Dialkylamino radicals, piperidino or morpholino radicals, 1-piperazinyl radicals (alkyl radicals containing 1 to 4 carbon atoms or phenylalkyl radicals in which the alkyl moiety contains 1 to 4 carbon atoms). Optionally substituted at position), cycloalkyl radicals containing 3 to 6 carbon atoms, cycloalkenyl radicals containing 4 to 6 carbon atoms, phenyl radicals (halogen atoms and alkyl radicals containing 1 to 4 carbon atoms) Or optionally substituted with one or more atoms or radicals selected from alkoxy radicals containing 1 to 4 carbon atoms ), A cyano or carboxyl radical or alkyl moiety is optionally substituted with one or more substituents selected from alkoxycarbonyl radicals containing 1 to 4 carbon atoms),

Phenyl or α- or β-naphthyl radicals, optionally substituted with one or more atoms or radicals selected from halogen atoms and alkyl radicals containing 1 to 4 carbon atoms or alkoxy radicals containing 1 to 4 carbon atoms, or

5-membered aromatic heterocyclic radicals, preferably selected from furyl and thienyl radicals,

Or a saturated heterocyclic radical optionally substituted with one or more alkyl radicals containing 4 to 6 carbon atoms and containing 1 to 4 carbon atoms,

R ₃ is

Unbranched or branched alkyl radicals containing 1 to 8 carbon atoms,

Unbranched or branched alkenyl radicals containing 2 to 8 carbon atoms,

Unbranched or branched alkynyl radicals containing 2 to 8 carbon atoms,

Cycloalkyl radicals containing 3 to 6 carbon atoms,

Halogen atoms and alkyl, alkenyl, alkynyl, aryl, aralkyl, alkoxy, alkylthio, aryloxy, arylthio, hydroxyl, hydroxyalkyl, mercapto, formyl, acyl, acylamino, aroylamino, alkoxycarbo Optionally with one or more atoms or radicals selected from the group consisting of Nylamino, amino, alkylamino, dialkylamino, carboxyl, alkoxycarbonyl, carbamoyl, alkylcarbamoyl, dialkylcarbamoyl, cyano, nitro and trifluoromethyl radicals Substituted phenyl or α- or β-naphthyl radicals,

Or a halogen atom and alkyl, aryl, amino, alkylamino, dialkylamino, alkoxycarbonylamino, acyl, arylcarbonyl, cyano, containing at least one same or different hetero atom selected from nitrogen, oxygen and sulfur atoms A 5-membered aromatic heterocycle optionally substituted with one or more identical or different substituents selected from carboxyl, carbamoyl, alkylcarbamoyl, dialkylcarbamoyl or alkoxycarbonyl radicals,

In substituents of phenyl, α- or β-naphthyl and aromatic heterocyclic radicals, the alkyl moieties of the alkyl radicals and other radicals contain 1 to 4 carbon atoms, and the alkenyl and alkynyl radicals contain 2 to 8 carbons Containing atoms, aryl radicals are phenyl or α- or β-naphthyl radicals,

R ₄ is

Alkoxy radicals containing 1 to 6 carbon atoms in the unbranched or side chain,

Alkenyloxy radicals containing 3 to 6 carbon atoms in the unbranched or branched chain,

Alkynyloxy radicals containing 3 to 6 carbon atoms in the unbranched or branched chain,

Cycloalkyloxy radicals containing 3 to 6 carbon atoms or

A cycloalkenyloxy radical containing 4 to 6 carbon atoms,

These radicals include one or more halogen atoms or alkoxy radicals containing 1 to 4 carbon atoms, alkylthio radicals containing 1 to 4 carbon atoms or carboxyl radicals, alkyloxy wherein the alkyl moiety contains 1 to 4 carbon atoms. A carbonyl radical, cyano or carbamoyl radical or each alkyl moiety contains 1 to 4 carbon atoms or optionally contains a second hetero atom selected from oxygen, sulfur or a nitrogen atom together with the nitrogen atom to which it is attached, 1 N-alkylcarbamoyl or N, N-dialkylcarbamoyl radicals or phenyl radicals or alkyl moieties which form saturated 5- or 6-membered heterocyclic radicals optionally substituted with alkyl radicals containing from 4 to 4 carbon atoms Optionally substituted with phenylalkyl radicals containing from 4 to 4 carbon atoms,

R ₅ is

Alkoxy radicals containing 1 to 6 carbon atoms in the unbranched or side chain,

Alkenyloxy radicals containing 3 to 6 carbon atoms,

Alkynyloxy radicals containing 3 to 6 carbon atoms,

Cycloalkyloxy radicals containing 3 to 6 carbon atoms or

A cycloalkenyloxy radical containing 3 to 6 carbon atoms,

These radicals include one or more halogen atoms or alkoxy radicals containing 1 to 4 carbon atoms, alkylthio radicals or carboxyl radicals containing 2 to 4 carbon atoms, alkyloxy wherein the alkyl moiety contains 1 to 4 carbon atoms. A carbonyl radical, cyano or carbamoyl radical or each alkyl moiety contains 1 to 4 carbon atoms or optionally contains a second hetero atom selected from oxygen, sulfur or a nitrogen atom together with the nitrogen atom to which it is attached, 1 N-alkylcarbamoyl or N, N-dialkylcarbamoyl radicals or phenyl radicals or alkyl moieties which form saturated 5- or 6-membered heterocyclic radicals optionally substituted with alkyl radicals containing from 4 to 4 carbon atoms Optionally substituted with phenylalkyl radicals containing from 4 to 4 carbon atoms.

Preferably, aryl radicals which may be represented by R ₃ are halogen atoms (fluorine, chlorine, bromine, iodine) and alkyl, alkenyl, alkynyl, aryl, arylalkyl, alkoxy, alkylthio, aryloxy, arylthio, Hydroxyl, hydroxyalkyl, mercapto, formyl, acyl, acylamino, aroylamino, alkoxycarbonylamino, amino, alkylamino, dialkylamino, carboxyl, alkoxycarbonyl, carbamoyl, dialkylcarbamoyl, sia Is a phenyl or α- or β-naphthyl radical optionally substituted with one or more atoms or radicals selected from no, nitro and trifluoromethyl radicals, and the alkyl portion of the alkyl radical and other radicals contains 1 to 4 carbon atoms Alkenyl and alkynyl radicals contain 2 to 8 carbon atoms and the aryl radicals are phenyl or α- or β-naphthyl radicals.

Preferably, the heterocyclic radical, which may be represented by R ₃ , contains one or more identical or different atoms selected from nitrogen, oxygen and sulfur atoms, halogen atoms (fluorine, chlorine, bromine, iodine) and 1 to 4 Alkyl radicals containing 6 carbon atoms, aryl radicals containing 6 to 10 carbon atoms, alkoxy radicals containing 1 to 4 carbon atoms, aryloxy radicals containing 6 to 10 carbon atoms, amino radicals, 1 Alkylamino radicals containing from 4 to 4 carbon atoms, dialkylamino radicals containing from 1 to 4 carbon atoms in each alkyl moiety, acylamino radicals containing from 1 to 4 carbon atoms in the acyl moiety, from 1 to 4 carbon atoms Alkoxycarbonylamino radicals containing carbon atoms, Acyl radicals containing 1 to 4 carbon atoms, 6 to 10 aryl moieties Arylcarbonyl radicals containing carbon atoms, cyano, carboxyl or carbamoyl radicals, alkylcarbamoyl radicals in which the alkyl moieties contain 1 to 4 carbon atoms, and di-containing each alkyl moiety containing 1 to 4 carbon atoms. Alkylcarbamoyl radicals or alkoxy moieties are 5-membered aromatic heterocyclic radicals optionally substituted with one or more identical or different substituents selected from alkoxycarbonyl radicals containing 1 to 4 carbon atoms.

Preferably, the R ₄ and R ₅ radicals, which may be the same or different, contain 1 to 6 carbon atoms and are methoxy, ethoxy, ethylthio, carboxyl, methoxycarbonyl, ethoxycarbonyl, cya Furnace, carbamoyl, N-methylcarbamoyl, N-ethylcarbamoyl, N, N-dimethylcarbamoyl, N, N-diethylcarbamoyl, N-pyrrolidinocarbonyl or N-piperidinocarbonyl radicals To an unsubstituted or branched alkoxy radical optionally substituted.

In more detail, the invention relates to the product of formula 1 wherein Z represents a hydrogen atom or a radical of formula 2, wherein R ₁ represents a benzoyl radical or the radical R ₂ -O-CO-, and R ₂ is tert -Butyl radical, R ₃ is an alkyl radical containing 1 to 6 carbon atoms, an alkenyl radical containing 2 to 6 carbon atoms, a cycloalkyl radical containing 3 to 6 carbon atoms, a halogen atom ( Fluorine, chlorine) and alkyl (methyl), alkoxy (methoxy), dialkylamino (dimethylamino), acylamino (acetylamino), alkoxycarbonylamino (tert-butoxycarbonylamino) or trifluoromethyl radicals Phenyl radicals or 2- or 3-furyl, 2- or 3-thienyl or 2-, 4- or 5-thiazolyl radicals optionally substituted with one or more identical or different atoms or radicals selected from R ₄ and R _5, which may be the same or different, represent unbranched or branched alkoxy radicals each containing 1 to 6 carbon atoms.

In more detail, the invention relates to the product of formula 1 wherein Z represents a hydrogen atom or a radical of formula 2, wherein R ₁ represents a benzoyl radical or the radical R ₂ -O-CO-, and R ₂ is tert -Butyl radical, R ₃ isobutyl, isobutenyl, butenyl, cyclohexyl, phenyl, 2-furyl, 3-furyl, 2-thienyl, 3-thienyl, 2-thiazolyl, 4-thiazolyl Or 5-thiazolyl radical, R ₄ and R _5, which may be the same or different, each represent a methoxy, ethoxy or propoxy radical.

Even more particular concern is that Formula _{1 wherein} R ₃ represents a phenyl radical and R ₁ represents a tert-butoxycarbonyl radical, and R ₄ and R ₅ , which may be the same or different, represent a methoxy, ethoxy or propoxy radical. Is the product of.

In a higher concern, the present invention provides 4α-acetoxy-2α-benzoyloxy-5β, 20-epoxy-1β-hydroxy-7β, 10β-dimethoxy-9-oxo-11-taxen-13α-yl of Formula 1a. (2R, 3S) -3-tert-butoxycarbonylamino-2-hydroxy-3-phenylpropionate.

Process for the preparation of the derivative according to the invention in two ways from patent WO 96/30355

This is known. Of formula 3 According to the first multi-step method, starting with 10-deacetylbaccatin, it is optionally protected at positions 7 and 13, for example in the form of silyl diether, followed by RX of Formula 4 wherein R is The same radical and X represents a reactive ester moiety or a halogen atom, such as sulfuric acid or sulfonic acid ester moiety) to give the product having -OR units at position 10 and silyl groups at positions 7 and 13. Subsequently, the silyl protecting group is replaced with a hydrogen atom to obtain a compound having an -OR group at the 10 position and a 0H group at the 7 and 13 positions. The latter derivative is optionally etherified at position 7 by reaction with a derivative of formula 4 to yield a derivative of formula 1 wherein Z corresponds to hydrogen.

The final step is in the presence of or in the presence of β-lactam according to the method described in the patent EP 617,018, for example at position 13 of the formula 1a derivative in which Z represents hydrogen, according to a method known per se. Esterification in the presence of oxazolidine as described in one patent WO 96/30355. After deprotection of the protecting group at positions 7 and 10, an ester of formula 1a in which Z is other than hydrogen and R represents hydrogen is thus obtained. The next step is carried out at positions 7 and 10 by the action of a reactant (Pummerer type reaction) formed in situ from R-SO-R of formula 5, where R has the same meaning as described above, sulfoxide and acetic anhydride. Are reacted simultaneously to form alkylthioalkyloxy intermediates at the 7 and 10 positions.

The final step of obtaining the desired compound of formula 1a is carried out on the intermediate compound obtained above by the action of activated Raney nickel.

In general, the action of the reactants formed in situ from the sulfoxides of the formula (5), preferably dimethyl sulfoxide and acetic anhydride, is carried out at temperatures of 0 to 50 ° C. in the presence of acetic acid derivatives such as acetic acid or haloacetic acid.

In general, the action of activated Raney nickel in the presence of aliphatic alcohols or ethers is carried out at temperatures of -10 to 60 ° C.

Further methods are described in the FR 97-14442 application. This invention allows, in one step, direct, selective simultaneous alkylation of the 2-hydroxyl functional groups at positions 7 and 10 of 10-deacetylbaccatin or a derivative thereof esterified at position 13 of the formula (6).

In the above formula,

A represents hydrogen or the side chain of the formula (2a) or the oxazolidine unit of the formula (2b).

(Wherein G represents a protecting group for a hydroxyl functional group, R ₁ and R ₃ have the same meaning as in Formula 2, and R _a and R _b which may be the same or different may be hydrogen or alkyl, aryl, halo, Alkoxy, arylalkyl, alkoxyaryl, haloalkyl, haloaryl, substituents may optionally form a 4-7 membered ring)

Preference is given to using 10-deacetylbaccatin, ie the product of formula (3), which allows significant savings in terms of process as starting material and also avoids the necessary intermediate protection and deprotection steps in the old process.

Among the protecting groups G of the hydroxyl functional group of the formula (2a), generally described in literatures such as Greene and Wuts, Protective Groups in Organic Synthesis, 1991, John Wiley & Sons, and MacOmie, Protective Groups in Organic Chemistry, 1975, Plenum Press For example,

Ethers, preferably benzyl ethers optionally substituted with one or more groups such as methoxymethyl ether, 1-ethoxyethyl ether, benzyloxymethyl ether, p-methoxybenzyloxymethyl ether, methoxy, chloro, nitro, Ethers such as silyl ethers such as 1-methyl-1-methoxyethyl ether, 2- (trimethylsilyl) ethoxymethyl ether, tetrahydropyranyl ether and trialkylsilyl ether,

It is desirable to select all protecting groups which are deprotected under conditions in which the remaining molecules, such as carbonates such as trichloroethyl carbonate, are slightly decomposed or not decomposed at all.

More specifically, the R _a and R _b radicals of formula (2b) are selected from those and derivatives described in patent WO 94/07878, particularly preferred are those in which R _a is hydrogen and R _b is a p-methoxyphenyl radical Do.

Alkylating agents are:

Alkyl halides, preferably alkyl iodine (RI)

Alkyl sulfates such as methyl sulfate

Oxonium, such as trialkyloxonium boron salt, in particular trimethyloxonium tetrafluoroborate (Me ₃ OBF ₄ ).

Methyl iodine is preferably used.

Alkylating agents are used in the presence of an anionic agents such as one or more strong bases in anhydrous medium.

Among the bases that can be used in anhydrous media:

Alkali metal hydrides such as sodium or potassium hydride

Alkali metal alkoxides such as potassium tert-butoxide

Silver oxide Ag ₂ O

1,8-bis (dimethylamino) naphthalene

See, eg, P. Caubere Chem. Rev. 1993, 93, 2317-2334 or M. Schlosser Mod. Synth. Mention may be made of monometallic or bimetallic base mixtures such as those described in the publications such as Methods (1992), 6, 227-271; Particular preference is given to alkyllithium / alkali metal t-butoxide or alkali metal amide / alkali metal t-butoxide blends. One of the two bases may be produced “in situ”.

Of all possible combinations of alkylating and anionic agents, preference is given to using methyl iodine in the presence of potassium hydride.

The reaction is preferably carried out in an organic medium which is inert under the reaction conditions. Among the solvents, preference is given to using the following materials:

Ethers such as tetrahydrofuran or dimethoxyethane

If silver oxide is used, it is preferred to use polar aprotic solvents such as dimethylformamide or aromatic solvents such as toluene.

When 1,8-bis (dimethylamino) naphthalene is used, it is preferable to use an alkyl ester such as ethyl acetate.

For a better implementation of the invention, it is preferred to use at least two, preferably 2 to 20 molar ratios of anionizing agent and substrate.

It is also preferred to use at least two, preferably 2 to 40, molar ratios of alkylating agent and substrate.

Preference is given to using reaction temperatures of -30 ° C to 80 ° C.

The reaction time is advantageously in the range of several hours to 48 hours depending on the reactants selected.

After the alkylation step, if the latter is carried out on 10-deacetylbaccatin, the process then proceeds to the esterification step in a known manner according to the method described, for example, in the aforementioned patent EP 617,018 or WO 96/30355. .

Therefore, according to the first three-step method, the process first begins with the dealkylation of 10-deacetylbaccatin, using an alkylating agent in the presence of a strong base, and in the second step, a diethered 10- at the 7 and 10 positions. Deacetylbaccatin is coupled at 13 position with β-lactam, which is suitably protected in the presence of an active agent selected from tertiary amines and metal bases which ensure the formation of the alkoxide at the 13 position. Deprotection of the side chains is then achieved by the action of inorganic or organic acids.

Therefore, according to the second three-step method, the process first begins with the dealkylation of 10-deacetylbaccatin, using an alkylating agent in the presence of a strong base, and in the second step is a 10-dehydrated diether at positions 7 and 10 Acetylbaccatin is coupled at the 13 position with oxazolidine in the presence of an activator such as dialkylaminopyridine and a coupling agent such as diimide. Ring opening of oxazolidine is achieved by the action of an inorganic or organic acid.

According to a third method, the process is carried out by esterifying with β-lactam or oxazolidine in the presence of a coupling agent and / or an active agent as described in the two methods described above at the 13 position of baccatin which is suitably protected at positions 7 and 10. Start with After deprotection at positions 7 and 10, the dietherization at positions 7 and 10 is carried out with an alkylating agent in the presence of a strong base. Deprotection of the side chains is then achieved by the action of inorganic or organic acids.

The product of formula 1 has notable biological properties.

In vivo, the product of formula 1 has been proved to be active in mice transplanted with colon adenocarcinoma C51 or C38 and other liquid or solid tumors at a dosage of 1 to 30 mg / kg.

The compound of formula 1 has anti-tumor properties, more specifically activity against tumors resistant to Taxol ^R and Taxotere ^R. Such tumors include, for example, colon tumors in which overexpression of the complex-tolerant P-glycoprotein has occurred. Multi-tolerability is a common term for the resistance of tumors to various compounds with different structures and mechanisms of action. Taxoids are generally known to be well recognized by experimental tumors such as P388 / DOX, a P388 rat leukemia cell line selected for doxorubicin (DOX) resistance, which expresses P-glycoprotein. Compounds according to the invention are less recognized by P388 / DOX. In more detail, the compound is less recognized than Taxotere ^R.

Compounds of formula (1) are mainly used for the preparation of therapeutic agents for abnormal cell proliferation of cell lines expressing complex-tolerant P-glycoproteins. The compound of formula 1 is mainly used to prepare a therapeutic agent for colon cancer.

Compounds of formula (1) and major compounds, wherein R ₄ and R ₅ are methoxy, respectively, have activity compared to other known taxoids, such as Taxol ^R or Taxotere ^R , which treat cancer cell lines expressing complex-tolerant P-glycoproteins. Have It is also active against tumor cells that are resistant to doxorubicin or resistant to Vincristine.

The product of Formula 1 may be used in combination with at least another therapy. More preferably in combination with other therapies including anti-tumor drugs, monoclonal antibodies, immunotherapy, radiotherapy or biological response modifiers.

The product of formula 1 is preferably administered by parenteral administration, such as intravenous, intraperitoneal, intramuscular or subcutaneous administration.

Introduction

The product of Formula 1a is a potent anticancer agent in pre-clinical models.

Antiproliferative of in vitro resistant cell lines

Docetaxel has been found to be cross-resistant to cell lines expressing the complex-tolerant P-glycoprotein (RIGEL I. and HORWITZ SB, Studies with RP 56976, (Taxotere ^R : Semisynthetic Analogue of Taxol)). , J. Natl. Cancer Inst., 83, 288-291, 1991).

Cytotoxicity of product 1a is tested in comparison to docetaxel for different resistant cell lines expressing the complex-tolerant P-glycoprotein 4 days after continuous exposure using standard methods.

Tolerance index is calculated by dividing the IC ₅₀ value calculated for IC ₅₀ values calculated from the resistant cells in susceptible cells.

The product of Formula 1a was found to be more active than docetaxel in all resistant tumor cell lines evaluated. Interestingly, the product of Formula 1a is usually cross-resistant to docetaxel (resistance index: 4.8 to 23.5) and in P388 / TXT, P388 / VCR, HL60 / TAX and Calc18 / TXT cell lines, which may be even more typical of clinical conditions. It was found to be the minimum cross-resistant (resistance index: 1.8 to 4.3).

In addition, significant reductions in the cross-tolerance of the product of Formula 1a (resistance index: 7.6 and 10) were also found in the two cell lines KB V1 and P388 / DOX (resistance index: 59 and 50) that are most resistant to docetaxel.

The product of Formula 1a is also evaluated in a murine P388 / CPT5 cell line which acquired resistance to camptothecins that do not express P-glycoprotein. Significant decreases in cross-tolerance have been found in the case of the product of Formula 1a and docetaxel (resistance indices: 4.8 and 10.5) compared to camptothecin (resistance index: 286). This result illustrates the possible application of the formula 1a product to the treatment of tumors that acquired resistance to camptothecin derivatives.

Comparison of Resistance of Docetaxel and Products of Formula 1a to Resistant Cell Lines Expressing P-glycoprotein Resistant cell line Resistance Index for mdr1 mRNA level ^* Docetaxel Product of Formula 1a P388 / DOX 50.7 10.0 +++ P388 / TXT 4.8 1.8 ++ P388 / VCR 5.8 1.8 ++ HL60 / TAX 8.1 2.5 ++ Calc18 / TXT 23.5 4.3 ++ KB V1 59.0 7.6 ++++ * Comparison of expressions obtained from Northern blot experiments using human mdr1 gene as probe

CaCo2

The antiproliferative product of Formula 1a and docetaxel are tested in human colon adenocarcinoma CaCo-2 cell line (ATCC HTB37), which appears to cause expression of the base level of P-glycoprotein encoded by the mdr1 gene. Methodology is used: 96 well plates, contact time with the agent (96 hours) and neutral red staining of viable cells. The results, expressed in IC ₅₀ , are summarized in the table below.

In vitro Growth Inhibitory Effects of the Product of Formula 1a and Docetaxel on CaCo-2 Cell Lines

cell drugs IC ₅₀ (μg / ml) ^* CaCo-2 Product of Docetaxel Formula 1a 108 ± 17 (3) 22 ± 3 (3) * Value in parentheses: number of experiments

The product of Formula 1a is cytotoxic in CaCo-2 cells with IC ₅₀ falling within the range described for other tested human cell lines, HL60, Calc18 and KB (see preceding paragraph). In contrast, docetaxel exhibits a larger IC ₅₀ corresponding to 4.9 times the value observed for the product of Formula 1a in CaCo-2 cells. Therefore, the antiproliferative high activity of the product of Formula 1a, compared to docetaxel in CaCo-2 cells, is consistent with the fact that the product of Formula 1a is less recognized than docetaxel by the P-glycoprotein originally expressed by this cell line.

In vivo antitumor activity

In vivo dosing regimens and anti-tumor efficacy studies are performed using preclinical standard formulations of the product of Formula 1a in ethanol: polysorbate 80 (50:50, v / v). After dilution, the final concentration is 5% ethanol, 5% polysorbate, 90% glucose-5% in water.

Next, a clinical formulation is developed in which the product of Formula 1a is dissolved in Polysorbate 80. After dilution, the final concentration of polysorbate 80 is 5%. In the following summary, in vivo studies are conducted using preclinical standard formulations, unless otherwise noted.

The product of Formula 1a is administered by the intravenous route (i.v.). Experiments and data analysis are performed according to standard protocols.

The active endpoints used to assess subcutaneous transplanted solid tumors are:

Tumor growth inhibition, T / C, where T and C are the median tumor weight in mg of the treatment and control groups, respectively. T / C> 42%,-is inactive, T / C according to NCI standard 42%, + is the minimum level of activity. T / C <10%, ++ is thought to be the antitumor high activity level (DN-2 level) that is the basis for further growth.

In the case of antitumor high activity, the following two endpoints are also used:

Tumor growth delay, T-C, where T and C are the median (days) time required for tumors in treatment and control groups to reach a predetermined size (750-1500 mg).

Logarithmic cell death, the logarithm of the total number of cells killed by the treatment. This value can be converted into activity ratio according to Southern Research Institute (SRI): logarithmic cell death (total) <0.7 =-inactive; 0.7-1.2 = +; 1.3-1.9 = ++; 2.0-2.8 = +++; > 2.8 = ++++ high activity.

For advanced stage tumors, degeneration can be partial (50% or more reduction in tumor weight) or complete. Full regression is included in partial regression.

The endpoint used to assess the activity of the intraperitoneal transplanted leukemia is the percent extension of host life (% ILS) calculated in relation to the median day of death.

The NCI criteria for activity are as follows:

-P388: high activity, ++ 75% ILS; Active, + 27-74% ILS; Inert,-<27% ILS

Toxicity is due to drug deaths ( 10%) or weight loss trough> 20%.

Docetaxel is used for comparative experiments if necessary.

Colon tumor

Colon 51

The product of Formula 1a was converted to i.v. Administration. At HNTD (9.3 mg / kg per injection) it was found to be highly active with 0% T / C and 2.6 logarithmic cell death. Doses below this value result in 2.2 apoptosis. Docetaxel was also found to be highly active with 3.1 log apoptosis.

I.v. of product 1a for colon adenocarcinoma C51 in BALB / c female mice. Dose evaluation

Formulation (Batch) Dosage (mg / kg / dosage) Plan (days) T / C (% on 18th) T-C (day) Algebraic cell death (total) evaluation Product of Formula 1a 24.215.09.35.8 4,6,8 --00 --25.821.9 --2.62.2 Toxicity (5/5 DD) Toxicity (1/5 DD) HNTD Activity Docetaxel 24.215.09.35.8 4,6,8 -004 -31.122.48.9 -3.12.20.9 Toxicity 22.8% bwlHNTD Highly active Median time tumors reach 750 mg in control group = 15.1 days Abbreviations used: HNTD = highest nontoxic dose, bwl = weight loss, DD = death due to drug

Progression Stage Colon 38

The product of Formula 1a was administered to mice with 200 to 400 mg of i.v. Administration. At HNTD (20 mg / kg / dose), it was found to be highly active and result in 5/5 complete regression and all mice survived tumor free at 139 days. Doses below this level (12.4 mg / kg / dose) lead to the same number of complete regression and 80% tumor free live animals. In comparison, docetaxel HNTD causes only 40% partial regression in the same experiment.

B6D2F _One I.v. of product 1a for advanced colon adenocarcinoma C38 in female mice. Dose evaluation

Formulation (Batch) Dosage (mg / kg / dosage) Plan (days) T-C (day) Algebraic cell death (total) Complete degeneration evaluation Product of Formula 1a 32.220.012.47.7 14,17,20 --- 25.0 --- 2.7 -5/55/52/5 Toxicity (1/5 DD) HNTD High Activity (5 / 5TFS) High Activity (4 / 5TFS) Activity Docetaxel 32.220.012.47.7 14,17,20 -29.16.12.5 -3.10.70.3 -0/50/50/5 Toxicity 20.6% bwlHNTD High Activity Limit Activity Inactivity Median time for tumor to reach 1000 mg in control group = 18.5 days Abbreviation used: HNTD = nontoxic highest dose, bwl = weight loss, DD = drug-induced death, TFS = no tumor survivor observed at 139 days Number

Claims (9)

Of a compound of Formula 1 for the preparation of a therapeutic agent for abnormal cell proliferation of a cell line expressing a complex-tolerant P-glycoprotein comprising administering to a mammal an effective amount of a compound of Formula 1 or a pharmaceutically acceptable salt or solvent compound thereof Usage. Formula 1 Formula 2 In the above formula, Z represents a hydrogen atom or a radical of formula (2), R ₁ is Benzoyl radicals optionally substituted with one or more identical or different atoms or radicals selected from halogen atoms and alkyl radicals containing 1 to 4 carbon atoms, alkoxy radicals containing 1 to 4 carbon atoms or trifluoromethyl radicals, A tennoyl or furoyl radical or a radical R ₂ -O-CO-, R ₂ is Alkyl radicals containing 1 to 8 carbon atoms, Alkenyl radicals containing 2 to 8 carbon atoms, Alkynyl radicals containing 3 to 8 carbon atoms, Cycloalkyl radicals containing 3 to 6 carbon atoms, Cycloalkenyl radicals containing 4 to 6 carbon atoms or Bicycloalkyl radicals containing 7 to 10 carbon atoms (these radicals are halogen atoms and hydroxyl radicals, alkoxy radicals containing 1 to 4 carbon atoms, each alkyl moiety containing 1 to 4 carbon atoms Alkylamino radicals, piperidino or morpholino radicals, 1-piperazinyl radicals (alkyl radicals containing 1 to 4 carbon atoms or phenylalkyl radicals in which the alkyl moiety contains 1 to 4 carbon atoms); Optionally substituted), cycloalkyl radicals containing 3 to 6 carbon atoms, cycloalkenyl radicals containing 4 to 6 carbon atoms, phenyl radicals (halogen atoms and alkyl radicals containing 1 to 4 carbon atoms, or Optionally substituted with one or more atoms or radicals selected from alkoxy radicals containing 1 to 4 carbon atoms) , A cyano or carboxyl radical or alkyl moiety is optionally substituted with one or more substituents selected from alkoxycarbonyl radicals containing 1 to 4 carbon atoms), Phenyl or α- or β-naphthyl radicals, optionally substituted with one or more atoms or radicals selected from halogen atoms and alkyl radicals containing 1 to 4 carbon atoms or alkoxy radicals containing 1 to 4 carbon atoms, or 5-membered aromatic heterocyclic radicals, preferably selected from furyl and thienyl radicals, Or a saturated heterocyclic radical containing 4 to 6 carbon atoms and optionally substituted with one or more alkyl radicals containing 1 to 4 carbon atoms, R ₃ is Unbranched or branched alkyl radicals containing 1 to 8 carbon atoms, Unbranched or branched alkenyl radicals containing 2 to 8 carbon atoms, Unbranched or branched alkynyl radicals containing 2 to 8 carbon atoms, Cycloalkyl radicals containing 3 to 6 carbon atoms, Halogen atoms and alkyl, alkenyl, alkynyl, aryl, aralkyl, alkoxy, alkylthio, aryloxy, arylthio, hydroxyl, hydroxyalkyl, mercapto, formyl, acyl, acylamino, aroylamino, alkoxycarbo Optionally with one or more atoms or radicals selected from the group consisting of Nylamino, amino, alkylamino, dialkylamino, carboxyl, alkoxycarbonyl, carbamoyl, alkylcarbamoyl, dialkylcarbamoyl, cyano, nitro and trifluoromethyl radicals Substituted phenyl or α- or β-naphthyl radicals, Or a halogen atom and alkyl, aryl, amino, alkylamino, dialkylamino, alkoxycarbonylamino, acyl, arylcarbonyl, cyano, containing at least one same or different hetero atom selected from nitrogen, oxygen and sulfur atoms A 5-membered aromatic heterocycle optionally substituted with one or more identical or different substituents selected from carboxyl, carbamoyl, alkylcarbamoyl, dialkylcarbamoyl or alkoxycarbonyl radicals, In substituents of phenyl, α- or β-naphthyl and aromatic heterocyclic radicals the alkyl moieties of the alkyl radicals and other radicals contain 1 to 4 carbon atoms, and the alkenyl and alkynyl radicals have 2 to 8 carbon atoms Wherein the aryl radical is a phenyl or α- or β-naphthyl radical, R ₄ is Alkoxy radicals containing 1 to 6 carbon atoms in the unbranched or side chain, Alkenyloxy radicals containing 3 to 6 carbon atoms in the unbranched or branched chain, Alkynyloxy radicals containing 3 to 6 carbon atoms in the unbranched or branched chain, Cycloalkyloxy radicals containing 3 to 6 carbon atoms or A cycloalkenyloxy radical containing 4 to 6 carbon atoms, These radicals include one or more halogen atoms or alkoxy radicals containing 1 to 4 carbon atoms, alkylthio radicals containing 1 to 4 carbon atoms or carboxyl radicals, alkyloxy wherein the alkyl moiety contains 1 to 4 carbon atoms. A carbonyl radical, cyano or carbamoyl radical or each alkyl moiety contains 1 to 4 carbon atoms or optionally contains a second hetero atom selected from oxygen, sulfur or a nitrogen atom together with the nitrogen atom to which it is attached, 1 N-alkylcarbamoyl or N, N-dialkylcarbamoyl radicals or phenyl radicals or alkyl moieties which form saturated 5- or 6-membered heterocyclic radicals optionally substituted with alkyl radicals containing from 4 to 4 carbon atoms Optionally substituted with phenylalkyl radicals containing from 4 to 4 carbon atoms, R ₅ is Alkoxy radicals containing 1 to 6 carbon atoms in the unbranched or side chain, Alkenyloxy radicals containing 3 to 6 carbon atoms, Alkynyloxy radicals containing 3 to 6 carbon atoms, Cycloalkyloxy radicals containing 3 to 6 carbon atoms or A cycloalkenyloxy radical containing 3 to 6 carbon atoms, These radicals include one or more halogen atoms or alkoxy radicals containing 1 to 4 carbon atoms, alkylthio radicals or carboxyl radicals containing 2 to 4 carbon atoms, alkyloxy wherein the alkyl moiety contains 1 to 4 carbon atoms. A carbonyl radical, cyano or carbamoyl radical or each alkyl moiety contains 1 to 4 carbon atoms or optionally contains a second hetero atom selected from oxygen, sulfur or a nitrogen atom together with the nitrogen atom to which it is attached, 1 N-alkylcarbamoyl or N, N-dialkylcarbamoyl radicals or phenyl radicals or alkyl moieties which form saturated 5- or 6-membered heterocyclic radicals optionally substituted with alkyl radicals containing from 4 to 4 carbon atoms Optionally substituted with phenylalkyl radicals containing from 4 to 4 carbon atoms. The compound of claim 1, wherein Z represents a hydrogen atom or a radical of formula (2), R ₁ represents a benzoyl radical or radical R ₂ —O—CO—, R ₂ represents a tert-butyl radical, and R ₃ represents 1 to 1 Alkyl radicals containing 6 carbon atoms, alkenyl radicals containing 2 to 6 carbon atoms, cycloalkyl radicals containing 3 to 6 carbon atoms, halogen atoms and alkyl, alkoxy, dialkylamino, acylamino, 2- or 3-furyl, 2- or 3-thienyl or 2-, 4- or 5 or phenyl radical optionally substituted with one or more identical or different atoms or radicals selected from alkoxycarbonylamino or trifluoromethyl radicals R ₄ and R ₅ , which represent thiazolyl radicals, which may be the same or different, each represent an unbranched or branched alkyloxy radical containing 1 to 6 carbon atoms. Usage. The compound of claim 2, wherein Z represents a hydrogen atom or a radical of formula (2), R ₁ represents a benzoyl radical or radical R ₂ —O—CO—, R ₂ represents a tert-butyl radical, and R ₃ is isobutyl , Isobutenyl, butenyl, cyclohexyl, phenyl, 2-furyl, 3-furyl, 2-thienyl, 3-thienyl, 2-thiazolyl, 4-thiazolyl or 5-thiazolyl radical The use of compounds in which R ₄ and R _5, which may be different, each represent a methoxy, ethoxy or propoxy radical. 4α-acetoxy-2α-benzoyloxy-5β, 20-epoxy-1β-hydroxy-7β, 10β-dimethoxy- for the preparation of a therapeutic agent for abnormal cell proliferation of cell lines expressing the complex-tolerant P-glycoprotein. Use of 9-oxo-11-taxen-13α-yl (2R, 3S) -3-tert-butoxycarbonylamino-2-hydroxy-3-phenyl-propionate. Use of a compound according to any one of claims 1 to 4 for the manufacture of a medicament usable in the treatment of colon cancer. Use of a compound according to claim 5 for use in combination with at least another therapy. Use of a compound according to claim 6 wherein the other therapy comprises an anti-tumor drug, monoclonal antibody, immunotherapy, radiotherapy or biological response modulator. Use of a compound according to claim 1, wherein the medicament is administered by parenteral administration. Use of a compound according to any one of claims 1 to 8, wherein the compound of formula 1 is administered by intravenous, intraperitoneal, intramuscular or subcutaneous administration.