WO2008121029A2 - Use of 9-oxoacridine-10-acetic acid and/or salts and/or its esters thereof for the treatment and prophylaxis of malignant tumors of female reproductive system - Google Patents

Abstract

The present invention relates to the treatment and/or prophylaxis of tumor reccurence in organs of a female reproductive system. According to the invention, a method of combination therapy is provided, the method including the steps of: (a) administering an effective amount of 9-oxoacridine-10-acetic acid and/or pharmaceutically acceptable salts and/or esters thereof to a patient in need thereof, and (b) hormonotherapy aimed at lowering the aromatase enzyme activity. The proposed method is useful in treatment of breast cancer, cancer uteri, ovarian cancer.

Description

USE OF 9-OXOACRIDINE-IO-ACETIC ACID AND/OR SALTS AND/OR ITS

ESTERS THEREOF FOR THE TREATMENT AND PROPHYLAXIS OF

MALIGNANT TUMORS OF FEMALE REPRODUCTIVE SYSTEM

FIELD OF THE INVENTION

The present invention relates to medicine, in particular to the treatment of tumors of female reproductive system and prophylactics of their recurrence.

BACKGROUND OF THE INVENTION The treatment of cancer in female reproductive system organs (mammary gland, ovaries, endometrium) is one of the major problems in modern oncology. Average incidence rate of these neoplasms in Russia is 78 per 100 thousands women (30% of general female cancer morbidity). Practically, at present, the female reproductive system cancer is diagnosed in 70% of patients for the first time at stage III or IV, when the treatment proves to be ineffective. As a result, in the Russian Federation, 442 women per 100 thousands die because of these tumors.

Although malignant tumors of breast, uterus and ovaries have estrogen receptors transferring estrogen signals, it has been reported that the treatment of ovarian carcinoma using antiestrogens and progestins has no noticeable clinical effect (Ho. S.- M., Estrogen, Progesteron and Epithelial Ovarian Cancer, 2003, Reproductive Biology and Endocrinology, 2003, vol. 1, p.73).

Furthermore, it has been assumed that, at least as far as the breast cancer concerned, the disappearance or reduction in number of estrogen receptors causes the tumor escape from the therapeutic control and further cancer progression. Recently, it has been shown that estrogens can stimulate growth not only through the activation of estrogen-dependent genes, but also through nongenomic mechanisms, such as the regulation of proteine-kinase activation (Lim, K.T., et al, Nongenomic estrogen signaling in estrogen receptor negative breast cancer cell: a role for the angiotensin II receptor ATI, Breast Cancer Research, 2006, vol.8, p R33 (doi: 10.1186/bcrl509).

Thus, medications inhibiting the action of enzyme aromatase could be useful in treating these tumors. The aromatase enzyme is responsible for the conversion of androstendione and testosterone into estrogens (estrone and estradiol) in some peripherical and tumor tissues, i.e. for the synthesis of extragonadal estrogens. While estrogens have stimulating (mitotic) effect on malignant tumor cells of the reproductive system. Aromatase (more precisely, a specific enzymatic complex of cytochrome P- 450, which is known as aromatase) is the key enzyme in estrogen biosynthesis. Aromatase is present in the ovaries of pre-menopausal women, in the placenta of pregnant women, in the adipose tissue of post-menopausal women, in the tissues of female reproductive system tumors, i.e. in the tumors of the mammary gland, the uterus, the ovaries. Aromatase is connected with endoplasmatic reticulum and consists of two basic proteins.

One of these proteins is cytochrome P-450 (P450_arom)- It converts C₁₉ steroids (androgenes) into C₁₈ steroids (estrogenes), containing a phenol ring A.

The second protein is NADPH-dependent cytochrome P-450 reductase, which transfers reduced energy equivalents to cytochrome P-450ar_Om-

The decrease of aromatase activity causes the reduction of estrogen synthesis and, as a result, lowering the estrogen (stimulating) effect on tumor cells, that provides a significant therapeutic efficacy of aromatase inhibitors (see Table No. 1).

Table No. 1. Aromatase inhibitors used in clinical practice.

According to the mechanism of action, aromatase inhibitors (AI) are divided into concurrent, causing temporary block of the enzyme, and «suicidal», that irreversibly binds aromatase active sites.

Irreversible inhibitors have entirely steroidal nature (they are chemical structural analogues of androstendione) and form an irreversible bond with the enzyme, inhibiting its biocatalyst activity completely. They are also called as "aromatase inactivators". After the effect of such inhibitor is finished, some period of time is required for the enzyme to be newly synthesized in the tissues.

Competitive inhibitors can be of steroidal and non-steroidal structure. The non-steroidal inhibitors are piperidines (aminoglutethimide, rogletimide), imidazoles (liarozole, fadrozole), triazoles (anastrozole, letrozole, vorozole), and other non-steroidal compounds. They reversibly interact with cytochrome part of P-450 enzymatic system.

Non-steroidal aromatase inhibitors can have selective (acting on the aromatase enzyme only) and nonselective action (they inhibit the synthesis of other hormones, in particular, gluco- and mineral corticoids).

Among steroidal aromatase inhibitors, plomestane, formestane and exemestane are known (see Fig. 1). Among non-steroidal aromatase inhibitors, aminoglutethimide, fadrozole, anastrozole, letrozole, vorozole are known (see Fig. 2).

STEROIDALAROMATASE INHIBITORS

Formestane Examestane

Figure 1. Steroidal aromatase inhibitors

NON-STEROIDAL AROMATASE INHIBITORS

Vorozole Anastrozole

Fig 2. Non-steroidal aromatase inhibitors

The above mentioned aromatase inhibitors bind the enzyme's cathalytic site. For example, anastrozole (2,2'[5-(lH-l,2,4-triazole-l-ylmethyl)- l,3phenylene]bis(2-methylpropionitrile) is the competitive aromatase inhibitor and causes only temporary block the enzyme activity.

Another non-steroidal aromatase inhibitor, letrozole ((4,4'-[(lH-l,2,4-triazole-l- yl)-methylene]bis-benzonitrile)), is a 200 times more potent aromatase inhibitor in in vitro system than aminoglutethimide, and is 19 times more potent than arimidex (anastrozol) .

Vorozole, more precisely (+)-vorozole, or (+)-6-((4-chlorophenyl)- IH- 1,2,4- triazole-l-ylmethyl)-l -methyl- lH-benzotriazole, is the (+)-enantiomer of R76713 aromatase inhibitor. For example, exemestane (6-methylenandrosta-l,4-diene-3,17-dione) is a structural analogue of androstendione and by its mechanism of action relates to aromatase inactivators of «suicidal» type, because it irreversibly inactivates the enzyme by binding to its active site. The use of aromatase inhibitors (as opposed to progestins and antiestrogens) is expedient only in women with absent ovarian function, because the decrease of estrogenes originating from the peripheral tissues will automatically cause the increase of gonadal (ovarian) estrogene synthesis.

At the same time, all these classes of the preparations (antiestrogens and progestins) are widely used as adjuvant therapy and also for the treatment of advanced breast cancer and advanced uterine cancer, which are non-operable. Neither antiestrogens, nor progestins have influence on the intratumor estrogen production, while this production can be effectively inhibited using aromatase inhibitors. At present, antiestrogenes and aromatase inhibitors are prescribed both as adjuvant therapy of operable breast cancer and as therapy of nonresectable advanced cancer in patients with estrogen-receptor-positive tumor status, because it is considered that the tumor sensitivity to hormone therapy is directly associated with the presence of estrogen receptors in the tumor tissue.

The administration of aromatase inhibitor letrozole in patients with endometrial carcinoma was reported to have certain positive effect. However, the clinical studies

(tumor volume reduction) did not show any correlation with the initial steroid receptor level and its dynamics during the treatment period (Berstein, L., et al., Neoadjuvant therapy of endometrial cancer with aromatase inhibitor letrozole: endocrine and clinical effect. European Journal of Obstetric & Ginecology and Reproductive Biology, 2002, vol. 105, pp. 161-165.).

Further, some ineffective attempts to treat cases of recurrent or advanced endometrial cancer using aromatase inhibitor (letrozole) were reported, showing the response rate as low as 9.4%, although in 86% of patients, the tumors were estrogen- receptor-positive, and 86% were progestin-receptor-positive (see Ma, B. B., The activity of letrozole in patient endometrial cancer and correlation with biological markers - a study of the National Cancer Institute of Canada Clinical Trial Group, International Journal of Gynecology Cancer, 2004, vol. 14, pp. 650-658). At that, progestins are commonly known to be effective in such patients. On the contrary, the attempts to treat endometrial stromal sarcoma with aromatase inhibitors, such as amidoglutethimide (Spano, J.P., Long-term survival of patients given hormonal therapy for metastatic endometrial stromal sarcoma, Medical Oncology, 2003, vol.20, pp.87-93), and letrozole (Leunen, M. et al., Low grade endometrial stromal sarcoma treated with the aromatase inhibitor letrosole, Gynecologic Oncololgy, 2004, vol. 95, pp. 769-771) were significantly more successful.

It was also reported for aromatase inhibitors that the development of resistance of estrogen-dependent tumor to one aromatase inhibitor does not automatically result in undoubted resistance to another inhibitor (Carlson R. W., Sequencing of endocrine therapies in breast cancer- integration of recent data, Breast Cancer Research and

Treatment, 2002, vol. 75, Supplement 1, pp. S27-32).

In the in vitro tissue culture of ovarian cancer, endometrial cancer and breast cancer, it was shown that aromatase inhibitors could decrease cell proliferative activity, regardless of receptor status and/or aromatase activity in the tumor tissue (Sasano, H. et.al., Effect of aromatase inhibitors on the pathobiology of human breast, endometrial and ovarian carcinoma, Endocrine-related cancer, 1999, vol.6, p.197-204.).

As for the treatment of advanced ovarian carcinomas with aromatase inhibitor, the clinical trials showed no correlation between the response to aromatase inhibitor

(letrozole) treatment and tumor receptor features (Papadimitriou,C.A. et. al., Hormonal therapy with letrozole for relapsed epithelial ovarian cancer. Long-term results of a phase II study. Oncology, 2004; vol. 66, pp. 112-117).

There was also shown that estrogen-positive cells pre-incubated with aromatase inhibitor (formestane) demonstrate the increased sensitivity to antibody-dependent cell- mediated response, that leads to accelerated cytolysis (Braun D. R et al, Aromatase inhibitors increase the sensitivity of human tumor cells to monocyte-mediated, antibody-dependent cellular cytotoxicity, The American Journal of Surgery, 2005, vol. 190, pp. 570 - 571). Therefore, molecular mechanisms responsible for carcinoma sensitivity to antiestrogenes and progestines, on one hand, and to aromatase inhibitors, on the other hand, are completely different.

The nuclear factor kappa B (NF- KB) family is composed of specific cytoplasmatic proteins activated in eukaryote cells under unfavorable conditions, in particular under chemotherapy and radiation. Non-active form of the protein is present in a complex with its own inhibitor. Once activated (with specific phosphorylkinases), the complex decomposes and protein NF KB is translocated into the nucleus and switches on the target genes.

Activation of these genes is associated with proliferation, angiogenesis, and apoptosis suppression which is a key link in development of resistance of tumor cells to chemotherapy.

The agents inhibiting NF KB activity may act on any step of activation of the factor; e.g. they can bind NF KB, inhibit translocation NF KB to nucleus, inhibit regulation of YY transcription factor with NF KB, inhibit apoptosis promotion, inhibit formation of complex of NF KB with other factors regulating NF KB activity. The search for new NF KB inhibitors is carried out extensively, including screening among known compounds. For example, dehydroxymethylepoxyquinomycin (DHMEQ) (a structural analog of antibiotic epoxyquinomycin C and its derivatives) (see WO 2006/060819), curcumin, its derivatives (see WO 03/090681) and other compounds were found to possess the ability to inhibit NF KB activity.

9-oxoacridine-lO-acetic acid is a substance with following structure:

According to another nomenclature, it also can be named lθ-(carboxymethyl)-

9(10H)acridone (CMA), in CAS base it has a number 38609-97-1, its international unproprietary name is cridanimod.

It shall be appreciated that, as used in the present specification, when the 9- oxoacridine-10-acetic acid is mentioned, its pharmaceutically acceptable salts and esters are also meant, if not specified otherwise.

Similarly, as used throughout the present specification, the abbreviation CMA designates 9-oxoacridine-lO-acetic acid itself as well as its pharmaceutically acceptable salts and esters, if otherwise is not specifically indicated or other sense is not readily apparent from the context.

9-oxoacridine-10-acetic acid derivatives were proposed as powerful antiviral agents by Hoffman La Roche Inc. employees (see USA patent No. 3,681,360).

Nowadays, medicaments on the basis of 9-oxoacπdine-10-acetic acid and its pharmaceutically acceptable salts, such as Neovir^®, are used for the treatment and prevention of a wide range of diseases. In particular, its immunomodulatory, interferonogenous, antibacterial, anti-promoter and radioprotective properties are well known.

However, while multiple research experiments have been conducted to enhance the efficacy of cancer therapy, the problem of increasing the efficacy of treatment of female reproductive system tumors is still unresolved.

The object of the present invention is to provide a new effective method for treatment of female reproductive system tumors, i.e. tumors derived from estrogen- dependent tissues.

DETAILED DESCRIPTION OF THE INVENTION

The invention is based on the unexpected finding that CMA, its salts and esters, exhibit a dual activity, not only markedly increasing the ability of aromatase inhibitors to inhibit aromatase enzyme activity, but also acting to depress NF KB activity, the activation of which, as is known, not only protects the tumor cell from harmful effects, but also leads to the same aromatase enzyme activation. Further, it was surprisingly established that the combined action of CMA, its salts and/or esters with aromatase inhibitor on the female reproductive system malignant tumors leads to suppression of tumor cells proliferation, even when the intratumor estrogene steroid synthesis with aromatase contribution is insignificant, absent or has no influence on the tumor growth. No similar effect was observed when these classes of compounds were administered separately (i.e. when either solely CMA, or solely aromatase inhibitors were administered).

Furthermore, the inventors of the present invention have now found that the newly discovered effect of CMA persists for a long time even after administration of CMA is discontinued, in other words, there is an effect of tumor «sensitization» to the subsequent or parallel action of aromatase inhibitors.

Due to these newly discovered features, CMA increases markedly the ability of aromatase inhibitors to diminish the proliferative activity of human tumors derived from the estrogen-dependent tissues, in particular, from the organs of female reproductive system. Thus, the inventors of the present invention have now found a new feature of 9- oxoacridine-10-acetic acid that expands the medical use of 9-oxoacridine-lO-acetic acid, its pharmaceutically acceptable salts and esters. That is, in treatment of tumors derived from the estrogen-dependent tissues, with aromatase inhibitors, 9-oxoacridine- 10-acetic acid, its pharmaceutically acceptable salts and esters allow to highly increase the antitumoral effect of aromatase inhibitors.

Also, the inventors of the present invention have now surprisingly found that if 9-oxoacridine-lO-acetic acid or its pharmaceutically acceptable salts or esters are administered before the hormone therapy commencement, this lead to sensitization of the tumor to further action of aromatase inhibitors, and this effect persists after the administration of 9-oxoacridine-l 0-acetic acid or said derivative is discontinued.

Further, according to the present invention, though 9-oxoacridine-l 0-acetic acid itself was reported to have no considerable cytostatic activity, the experiments on animals conducted according to the present invention have demonstrated that the tumor growth inhibition with conventional hormone therapy was more effective in the presence of 9-oxoacridine-l 0-acetic acid and its salts.lt is known that 9-oxoacridine-10- acetic acid possesses interferon-inducing properties. However, in cell culture studies, the inventors of the present invention observed no measurable increase of interferon level, and thus, it shall be concluded that the newly-discovered property of CMA does not results from interferon activity and is not mediated by interferon. Further, in vivo studies conducted by the inventors of the present inventors have demonstrated that in respect of growth of malignant tumors of female reproductive system, 9-oxoacridine-l 0-acetic acid continues to reveal dose-dependent effect on the tumor growth in doses which exceed maximal interferon-inducing dose, i.e. the threshold dose of 9-oxoacridine-lO-acetic acid, further increase of which does not lead to increased tissue and serum interferon levels.

Moreover, in case of depletion of interferon system (it is typically observed upon repeated administration of any interferon inducer including 9-oxoacridine-10-acetic acid) 9-oxoacridine-10-acetic acid and/or its pharmaceutically accepted salts and its esters continue to reveal dose-dependent influence on the tumor growth inhibition when combined with aromatase inhibitors, anti-tumor and enzyme-inhibitory activity of which, as a result, markedly increases. Without being bound by theory, it is believed that this inhibition can be caused by either decrease in the cell proliferative activity, or accelerated cell apoptosis, or both.

Thus, a new effect of 9-oxoacridine-10-acetic acid has been found by the inventors of the present invention though at present the mechanism of this effect is not entirely clear. However, realization of this mechanism leads to a decrease of active NF KB level and opens new frontiers in the use of 9-oxoacridine-10-acetic acid and/or its pharmaceutically accepted salts and esters.

The term "pharmaceutically acceptable salt" as used herein, means those salts, which maintain the above mentioned properties of 9-oxoacridine-10-acetic acid and which are not unacceptable biologically or unacceptable in some other way. The pharmaceutically acceptable salts derived from the salt forming bases could be obtained with inorganic or organic bases.

The salts with inorganic bases include, but are not limited to, sodium, potassium, lithium, ammonium, calcium and magnesium salts.

The salts with organic bases include, but are not limited to, salts of primary, secondary, tertiary and quaternary amines, such as alkylamines, dialkylamines, trialkylamines, substituted alkylamines, di(substituted alkyl)amines, tri(substituted alkyl)amines, alkenylamines, dialkenylamines, trialkenylamines, substituted alkenylamines, di(substituted alkenyl)amines, tri(substituted alkenyl)amines, cycloalkylamines, di(cycloalkyl)amines tri(cycloalkyl)amines, substituted cycloalkylamines, di(substituted cycloalkyl)amines, tri(substituted cycloalkyl)amines, cycloalkenylamines, di(substituted cycloalkenyl)amines, di(substituted cycloalkenyl)amines, arylamines, diarylamines, triarylamines, heteroarylamines, diheteroarylamines, triheteroarylamines, heterocyclylamines, diheterocyclylamines, triheterocyclylamines, mixed di- and tri-amines, where at least one of the substitutes on amine differs and is selected from the group, including alkyl, substituted alkyl, alkenyl, substituted alkenyl, aryl, heteroaryl, heterocyclyl, etc. Amines, in which two or three substitutes together with the nitrogen atom to which they are connected, form a heterocyclyl or a heteroaryl, also are included here.

Specific examples of appropriate amines include, in particular, isoprpylamine, trimethylamine, diethylamine, tri(isopropyl)amin, tri(«-propyl)amine, ethanolamine, 2- dimethylaminoethanol, tromethamine, lysine, arginine, histidine, caffeine, procaine, choline, betaine, ethylendiamine, glucosamine, N-alkylglucamine, theobromine, purines, piperazine, piperidine, morpholine, N-ethylpiperidine etc.

An example of a salt of 9-oxoacridine-lO-acetic acid with an alkali metal is the sodium salt:

An example of a salt with amino compound is the salt with 1-deoxy-l- (methylamino)-D-glucitol (i.e. with meglumine, or, the same, withN-methylglucamine):

H₂-CH₂-(CH₂OH)₄-CH₂OH

Other examples of salts with various complex quaternary ammonium bases include salts with amine-substituted carbohydrates, for example, with 2-deoxy-2- amino(or 2-alkylamino)-D-glucose, where R is H or a lower alkyl:

with 1 -deoxy- 1 -methylamino-D-glucose:

as well as salts with various esters of carbohydrates and aliphatic amino alcohols, for example

R₂

where R₁, R₂ are independently alkyl, aryl, heteryl

The examples of appropriate cations are, in particular, cations of 3-O-(N,N-dimethylamino-n-propyl)-l,2:5,6-di-O-isopropyliden-α,D- glucofuranose,

a τaκ>κe κaτH0H&i: as well as following cations of:

1-deoxy -l-(ethylamino)-D-glucitol (i.e. eglumine), 1 - deoxy - 1 -(propylamino)-D- glucitol,

1- deoxy -l-(butylamino)-D- glucitol, 1- deoxy -l-(methylamino)-L- glucitol, 1- deoxy -l-(ethylamino)-L- glucitol, 1- deoxy -l-(propylamino)-L- glucitol, and 1 - deoxy - 1 -(butylaniino)-L- glucitol.

According to the present specification, esters of 9-oxoacridine-lO-acetic acid include compounds obtained by hydrogen atom substitution in acid OH-group with an organic group R. Examples of suitable esters include but are not limited to, esters of 9-oxoacridine-

10-acetic acid with lower alkyls (namely with (CrC₁₂)alkyls, in particular ethyl, propyl, isopropyl, butyl and amyl esters), as well as with choline and other lypophilic alcohols.

After rapid penetration through biological membranes in vivo, these compounds are easily hydrolyzed to free 9-oxoacridine-10-acetic acid.

According to the present invention, pharmaceutically acceptable salts of the 9- oxoacridine-10-acetic acid can be used in a single dose from 0.5 to 100 mg/kg body weight (calculated based on 9-oxoacridine-10-acetic acid), preferably from 4 to 20 mg/kg body weight. The daily dose can vary from 2 to 1000 mg/kg, preferably from 2 to 200 mg/kg. In each particular case, the preparation dosage can be calculated by a specialist on the basis of the specification and the examples.

It is appreciated that the used amounts of 9-oxoacridine-10-acetic acid or of its pharmaceutically acceptable salt or of its ester and aromatase inhibitors are synergistically effective.

The term "estrogene-dependent tissues" in this specification means the tissues, the growth of which (growth due to proliferation and/or cells hypertrophy included) is mainly stimulated by estrogens. To this type of tissues belong the tissues of the organs of female reproductive system, such as tissues of the uterus, the ovary and mammary glands. Therefore, within this specification, breast cancer, endometrial cancer (both epithelial and stromal) and ovarian cancer refer to the malignant tumors, derived from estrogen-dependent tissues.

On the basis of these new findings about CMA properties, a novel method for the treatment of female reproductive system tumors is provided, which comprises the steps of simultaneous or consecutive administration of (a) CMA and (b) aromatase inhibitors to a patient in need of such a treatment, or a combination of these modes. The proposed method according to the invention provides the enhancement of antitumoral treatment efficacy in clinical practice.

None of the above described newly-discovered properties of CMA, its salts and its esters, as far as to the knowledge of the present inventors, have been disclosed in the prior art. Based on the these findings, the present inventors in a further aspect of the invention, provide the use of CMA, its salts and its esters in combination with aromatase inhibitors in the treatment of tumors of female reproductive system organs.

Also, the inventors of the present invention are unaware any mentioning of evidence of attempts to administer aromatase inhibitors and CMA preparations in combination or subsequently in treatment of female reproductive system neoplasm in clinics, including adjuvant and non-adjuvant regimens.

The inventors of the present invention have now surprisingly found that 9- oxoacridine-10-acetic acid and/or its pharmaceutically acceptable salts and/or esters can have a dual action, not only enhancing the aromatase inhibitor effect, but also increasing the ant-tumor effect of various cytostatic agents from different classes, when used in treatment of female reproductive system malignant tumors.

If the chemotherapy was added at various stages of such combined hormone therapy or was used after such combined treatment (aromatase inhibitor plus CMA), the use of CMA provides much more effective inhibiting influence on the tumor growth. The same effect was observed when hormone-refractive (hormone-resistant) variants of the tumors were treated with the inventive method. The above described sensitizing effect of a combined administration of aromatase inhibitors and 9-oxacridine-10-acetic acid and its derivatives together with cytostatic chemical preparations was not observed when tumor cells underwent exposure to aromatase inhibitors or to CMA separately. It was surprisingly found that such "sensitizing" effect persisted upon withdrawal of the combined treatment with 9- oxoacridine-10-acetic acid (or its pharmaceutically acceptable salts and esters) and hormone preparations.

According to the present invention, a method for treatment of tumors of organs of female reproductive system, is provided, wherein the tumors are derived from estrogen-depended tissue, the method comprising the steps of: (a) administering an effective amount of a compound selected from the group including 9-oxoacridine-10- acetic acid and/or its pharmaceutically acceptable salts and/or its esters to a patient in need thereof, and (b) hormonotherapy aimed to diminish aromatase enzyme activity including administration of aromatase inhibitors. In still further aspect, a method of prophylaxis of reccurence of tumors of organs of female reproductive system is provided, wherein the tumor is derived from estrogen- depended tissue, the method comprising the steps of: (a) administering an effective amount of a compound selected from the group including 9-oxoacridine-lO-acetic acid and/or its pharmaceutically acceptable salts and/or its esters to a patient in need thereof, and (b) hormonotherapy aimed to diminish aromatase enzyme activity including administration of aromatase inhibitor.

Thus, according to the present invention, prophylaxis of tumors recurrence (i.e. prevention of tumor relapse) is provided in relation to the tumor, regression or elimination of which was previously achieved with surgical manipulation or with chemotherapy, hormonotherapy, x-ray therapy, biotherapy, or with combination mentioned methods.

In some preferred embodiment of the invention, a method of treatment and/or prophylaxis of tumors of organs of female reproductive system and/or their relapse with use of 9-oxoacridine — 10 — acetic acid and/or its pharmaceutically acceptable salts and/or its esters is provided comprising administering to a patient in need thereof 9- oxoacridine-10-acetic acid and/or its pharmaceutically acceptable salts and/or its esters in combination with an aromatase inhibitor.

In still further aspect, a method of increasing the sensitivity of malignant tumor derived from estrogen-dependent tissue to aromatase inhibitor (inhibitors), is provided comprising administration of an effective amount of a compound selected from the group including 9-oxoacridine 10-acetic acid and/or its pharmaceutically acceptable salts and/or its esters.

As used herein, the terms "to treat," "treating," and "treatment" refer to administering or introduction (prescription) a therapy, an agent, a compound, or composition to a subject having a disease or pathologic condition (for example to a human being, for example to a patient or to a person with risk of the pathology condition development, for example, malignant tumor). In general, a treatment is provided to a subject suffering from a disorder (for example, malignant tumor, in particular, maliganant tumor as described in this specification), a symptom or symptoms of a disorder, increased risk of a disorder, or predisposition to a disorder, to cure, to recover, to improve the life quality; to alleviate symptoms; to diminish the extent of a disorder, symptoms of a disorder, or a degree of predisposition to a disorder, to induce stabilization (i.e., to prevent worsening) of the state of disorder, to delay or slow down the disorder progression, to induce amelioration or palliation of the disorder state, and remission (whether partial or total), whether detectable or undetectable.

The term "treatment" can also mean a prolonged survival as compared to expected survival in the absence of treatment. The term "treatment" can also mean administration, introduction, prescription or applying otherwise a dose of a therapeutic agent, a composition, a compound alone or in its combination with one or more other agents, compounds or compositions.

As used herein, the terms "in combination", "combination" refer to a simultaneous or consecutive administration, introduction, prescription or application by any other route of a dose of various therapeutic agents, compositions, or compounds (in particular, CMA and an aromatase inhibitor). If the agents or compounds are used consequently, preferably, at the time of the commencement of use of a second agent, the first agent already presents in effective concentration in the target area (tissue(s)) or its pharmacodynamic action (pharmacological effect) persists at least till the time of the commencement of the use of the second agent.

Further, the term "effective amount of a compound/drug" relates to an amount of this compound/drug effective to induce a specified action. The effective amount of one and the same compound/drug can vary depending on particular effect and particular combination, e.g. the amounts of CMA effective to induce the antitumor activity of cisplatine can differ from the amounts of CMA effective to induce the antitumor activity of paclitaxel. Similarly, the amounts of CMA effective to induce the antitumor activity of cisplatine can differ from the amounts of CMA effective to reduce the level of active NF KB factor. According to the present invention, preferred salts of 9-oxoacridine- 10- acetic acid are selected from the group including sodium, meglumine, eglumine salts and the salt with 3-O-(N,N-dimethylamino-«-propyl)-l,2:5,6-di-O-isopropyliden- a,D - glucofuranose.

According to present invention, preferable esters of 9-oxoacridine-lO-acetic acid are selected from the group including ethyl, propyl, butyl, isopropyl, amyl esters.

In some preferred embodiments of the invention, 9-oxoacridine - 10 - acetic acid and/or its pharmaceutically acceptable salts and/or its esters are administered every day or every second day during 30 to 180 day at a daily dose from 4 to 300 mg/kg body weight (calculated based on 9-oxoacridine- 10-acetic acid).

In some embodiments of the invention as steroidal as well as non-steroidal aromatase inhibitor can be used. According to present invention, preferably aromatase inhibitor is selected from the group including formestane, exemestane, aminoglutethimide, fadrozole, anastrozole, letrozole, vorozole.

In still another embodiment of the invention, a method for treating a tumor of an organ of a female reproductive system is provided, the method comprising the steps of: (a) administering an effective amount of a compound selected from the group including

9-oxoacridine- 10-acetic acid, its pharmaceutically acceptable salts and esters to a patient in need thereof, and (b) hormonotherapy aimed to diminish aromatase enzyme activity including administration of aromatase inhibitors, wherein the method further comprises administering one or more chemotherapeutic agents.

As the present inventors have now found, the efficacy of this combined action (aromatase inhibitors, CMA, chemotherapeutic agent (or agents)) does not depend on tumor receptor status and/or aromatase activity in the tumor tissue.

The chemotherapeutic agent according to the invention can be selected from the group including but no limited: alkylating agents, for example, cyclophosphamide, chlorambucil, cisplatin, carboplatin, busulphan, melphalan, streptosotozine, triethylenmelamine, mitomycin C, dacarbasine, procarbasine, and others; antimetabolites, for example methotrexate, etoposide, 6-mercaptourine, 6-thioguanin, citarabine, 5-fluorouracil, capacetabine, fludarabine and others; antitumoral antibiotics, for example actinomycine D, doxorubicine, daunorubicin, bleomycine, mitramycine and others; vinca alcaloids, such as vincristine and vinblastine; taxans (taxoids), for example, paclitaxel and docetaxel and their derivatives; glucocorticoids, for example prednisone; nitrosourea and its derivatives, such as lomustine, carmustine, and others; antitumoral enzymes, for example, asparaginase and its pegylated forms or ots forms, conjugated with non-polypeptide polymers; topoisomerase inhibitors, such as topotecane and irinotecane, and others; microtubule tubulin polymerization inhibitors, for example, vinorelbine.

According to the present invention, the inventive method of the combined or subsequent administration of CMA and aromatase inhibitors can be also supplemented with administration of any of the following, either solely, or in combination: estrogen receptors antagonists (aritiestrogenes, for example, tamoxifen, toremifen) progestin receptor agonists (progestines, for example, progesterone; medroxyprogesterone acetate, megestrole acetate, hydroxyprogesterone capronat)

- LHRH-antagonists (antagonists of luteinizing hormone releasing hormone), which also can be used in treatment of malignant tumors of female reproductive system (for example, goserelin, buserelin)

- therapeutic antibodies against growth factor receptors, for example, trastuzumab, that represents monoclonal antibodies against HER2/neu receptors of human epidermal growth factor.

In some preferred embodiments of the present invention, 9-oxoacridine- 10- acetic acid, its salt and/or ester can be administered in the following manner:

- as a treatment course before commencement of treatment with aromatase inhibitor.

- as a treatment course simultaneously with treatment with aromatase inhibitor.

- the treatment with 9-oxoacridine-lO-acetic acid and/or its salt and/or ester commences before the beginning of treatment with an aromatase inhibitor and then

CMA is administered simultaneously along with the treatment with the aromatase inhibitor.

In some embodiments of the invention, a method of treatment comprises a course of administration of 9-oxoacridine-10-acetic acid, its pharmaceutically acceptable salts or esters thereof prior to the treatment with aromatase inhibitor.

In another embodiment of the invention, 9-oxoacridine-10-acetic acid, its pharmaceutically acceptable salt or ester is administered concurrently along with the administration of aromatase inhibitor or it can be initiated prior to administration of an aromatase inhibitor and continued along with an aromatase inhibitor administration.

Further, according to the present invention, the use of 9-oxoacridine — 10 — acetic acid and/or its acceptable salts and/or its esters is provided for the preparation of a medicament for the treatment of malignant tumors of female reproductive system in combination with an aromatase inhibitor. According to the invention, along with 9-oxoacridine - 10 - acetic acid and/or its acceptable salts and/or its esters, the medicament can further comprises other components, such as: various excipients and additives, including: solubilizers, for example, aminosugars (or amino alcohols) and theirs esters, cyclodextrins, for example, hydroxypropyl-β-cyclodextrin; emulsifiers, for example, tweens; stiffeners; photo (light) filters, for example, methylene blue; complexing agents; stabilizers, for example, trilon B; prolongators, for example, methylcellulose and polyvinylpyrrolidone; corrigents, for example, sorbitol; dyes; preservatives; as well as organic- and non- organic buffer systems aimed to maintain a constant pH. This medicament can be produced in various pharmaceutical forms such as: a solution for injection, tablets, an enteric coated tablet, powder or a granulate in capsules or in sachets, suppositories, aerosol or solution for inhalation, prolonged form for oral use or in the form suitable for abdominal implantation, on the basis of solid or semi-solid or a polymer matrix.

According to the present invention, the preferred salts of 9-oxoacridine-lO-acetic acid for preparation of a medicament for the treatment of malignant tumors of female reproductive system in combination with an aromatase inhibitor, are selected from the group including sodium, meglumine, eglumine salts and the salt with 3-0-(N₅N- dimethylamino-o-propyl)-l,2:5,6-di-0-isopropyliden- a,O -glucofuranose.

Further, according to the present invention, a kit for the treatment of malignant tumors of female reproductive system is provided, comprising 9-oxoacridine - 10 - acetic acid and/or its acceptable salts and/or its esters in amounts effective in potentiating the action of the said aromatase inhibitor. According to the present invention, preferred salts of 9-oxoacridine- 10-acetic acid for use in the kit for the treatment of malignant tumors of female reproductive system, are selected from the group including sodium, meglumine, eglumine salts and the salt with 3-O-(N,N-dimethylamino-«-propyl)-l,2:5,6-di-O-isopropyliden- a,O glucofuranose. According to the present invention, an aromatase inhibitor for use in a kit for the treatment of malignant tumors of female reproductive system according to the presnt invention is selected from the group including formestane, exemestane, aminoglutethimide, fadrozole, anastrozole, letrozole, vorozole.

The unit dosage forms of the kit according to the invention can be presented as a solution for injection in ampoules or vials, or as tablets, or as a enteric coated tablet, or as powder or a granulate in capsules, in flasks or in sachets, or as suppositories, or as aerosol or solution for inhalation, or as prolonged form for oral use or can be formed on the basis of a solid or semi-solid polymer matrix for abdominal implantation.

It shall be also appreciated that each of the inventive compositions and medicaments may alternatively include, compise, or be substantially composed of any suitable components disclosed in the present specification, and such compositions and medicaments, including pharmaceutical compositions, and a kit according to the invention, may additionally or alternatively be prepared in such a way that a component, a material, an ingredient or an object could be excluded therefrom, which was used in a corresponding medicament or composition known in the prior art, or which is not necessary to achieve the technical effect of the present invention. The same refers to a method of treatment according to the invention, which alternatively may include, comprise, or be substantially composed of any matching stages disclosed in the present specification, and such inventive methods may additionally or alternatively exclude some steps or objects, which is used in a method, known in the prior art, or which is not necessary to achieve the technical effect of the present invention.

Further, the invention is illustrated by particular examples of implementation, not limiting the scope of the present invention.

Commercially available preparations of CMA salts, for example sodium CMA salt (preparation Neovir, Pharmsynthez, Russia), meglumine CMA salt (preparation Cycloferon, NTFF Polysan, Russia), salt of CMA with 3-O-(N,N-dimethylamino-«- propyl)-l,2:5,6-di-O-isopropyliden-α,D-glucofuranose (hereinafter named "CMA N- propylglucamine salt") (preparation Anandin, Mediter, Russia) as well as commercially available CMA (Sigma, USA, cat. # 17927, catalogue of year 2005), among others, were used in the experiments and clinical studies carried out by the present inventors.

Esters and some other CMA salts were synthesized by known, relatively simple methods (see for example: Inglot A.D. et al., Archivum Immunologiae et Therapiae Experimentalis, 1985, vol. 33, pp. 275-285; RU 2135474; RU 2036198; RU 2033413). The amounts of CMA esters and salts in the Examples and tables were calculated based on the mass of one mole of 9-oxoacridine- 10-acetic acid.

In some cases, for example, for clinical per rectum administration, suppositories were prepared according to the invention, containing CMA or its pharmaceutically acceptable salt or ester. Such suppositories were prepared on the base of widely used for such purposes suppositorial masses, such as in particular Witepsol (Witepsol W 35, E 75), as it is illustrated by the examples. The following commercially available chemically pure aromatase inhibitors were used in the in vitro experiments: formestane (4-hydroxyanrost-4-ene-3 , 17-dion); aminoglutethimide (3 -(4-aminophenyl)-3 -ethyl-2,6- piperidinedione); fadrozole (4-(5,6,7,8-tetrahydro-imidazole[l,5- α]piridine-5- yl)benzonitrile); anastrozole (α,α, α', α'--tetramethyyl-5-(lH-l,2_s4-triazole-l-yl)methyl]- ;w-benzendiacetonnitrile), exemestane (10,13-dimethyl-6-methylidene-

7,8 ,9, 10, 11 , 12, 13 , 14, 15 , 1 ό-decahydrocyclopentaneM-phenantrien-S , 17,-dione), (Sigma-Aldrich company, USA; Shaanxi Dafeng Scientific, Industrial and Trading Co., China), and others.

The following commercially available aromatase inhibitors of various chemical structure and of various generations of this class of preparations, were uses in the example experiments relating to the treatment of patients with female reproductive system cancer according to the present invention: aminoglutethimide (Mamomit®, Pliva company, Croatia), anastrozole (Arimidex®, Novartis, Switzerland), exemestane (Aromasin®, Pharmacia Italia, Italy), and other.

The method of estimation of tumor response on the preparation in histoculture was used to determine the effect of various aromatase inhibitors in combination with CMA, its salts and ester on human tumors.

In the experiments, explants obtained from various human tumors, derived from estrogene-dependent tissues, including breast cancer, epitelial endometrium cancer, ovarian cancer, stromal endometrium cancer, were investigated. Explants obtained during surgery were prepared and incubated in a media with various agents (CMA salts, aromatase inhibitors and combinations). In examples listed below, which illustrate the invention, aromatase inhibitors were added in the incubation media in concentration equal or fold to the IC₅₀ of corresponding aromatase inhibitor. ICs₀ index is widely used in pharmacology to characterize the potency of aromatase inhibitors to inhibit activity of aromatase enzyme (for example, see Goss, P., Gween, K.M.E.H., Current perspectives on aromatase inhibitors in breast cancer, Journal of Clinical Oncology, 1994, vol. 12, pp. 2460-2470). IC₅₀ represents the concentration of aromatase inhibitor, which inhibits the human aromatase activity by 50% in determined model system that contains a source and a substrate for aromatase (an androgenic steroid). In one series of experiments according to the invention, the tumor tissue was exposed simultaneously to the CMA and aromatase inhibitors. In other experiments, cells were at first exposed to the CMA, then the CMA exposure was stopped, and cells were transferred to the culture medium with aromatase inhibitors. In some experiments, the tumor tissue was firstly exposed to the CMA, and then aromatase inhibitors were added.

The effect on the human tumor tissues was determined using the following methods: by estimating the changes in tumor tissue proliferation (it was estimated by rate of radioactive label uptale in the newly synthesized DNA) and by estimating the changes of NF KB activity. Human tumor cell survival rate in hystological culture under combined exposure to CMA, aromatase inhibitors and chemotherapeutical agents of various generations was estimated by the quantity of formazan (produced under influence hydrogenases of survived cells when the cell were incubated with 3-(4,5- dimethyl-2-yl)-2,5-diphenyltetrazolium bromide (MTT-assay)) (for example, see

Hoffman, R.M. The clinical benefit of the histoculture drug response assay. Japanese

Journal of Cancer Chemotherapy, 2000, vol. 27, Supplement 11, pp. 321-322).

Various aromatase inhibitors used in clinics, were administered (under the patient informed consent) with simultaneous or consequent administration of salts and

CMA esters, to estimate the clinical efficacy of the inventive method of treatment.

Clinical responses were estimated according to criteria of World Health Organization, as well as according to changes in serum level of tumor markers during the therapy.

Further, the invention is illustrated in more detail by way of the following non- limiting examples:

EXAMPLE 1.

IC₅₀ determination for aromatase inhibitors of various chemical classes.

Before the beginning of experiments, the IC₅₀ was determined on each aromatase inhibitor by the traditional method (Thompson, E.A and Siiteri, P. K., The involvement of human placental microsomal cytochrome P-450 in aromatization, Journal Biological Chemistry, 1974, vol. 249, pp. 5373-5378). Human placentary microsomes were used as a source of human aromatase (Ryan, K.J., Biological aromatization of steroids., Journal Biological Chemistry, 1959, vol. 234, pp. 268-272). As aromatase substrate, [lβ³H]Δ4-androstendione (1 niKi 100 nm) (Institute of molecular genetics, RAS) was used. Each experiment was repeated three times. The results are presented in the Table No. 1.

Table NoJ. IC₅₀ of various aromatase inhibitors

Comments: SI — steroidal inhibitor, NSI — non-steroidal inhibitor.

EXAMPLE 2. Histoculture system of malignant tumor tissue for determination the effect of aromatase inhibitors, as well in combination with CMA, its pharmaceutically acceptable salts and esters on cell proliferation and NF-κB activity in female reproductive system malignant tumors.

Samples of female reproductive system malignant tumor tissues, obtained during surgery, were cut into 1-1.5 mm pieces. Pieces were put on the surface of porous gelatine sponge and were incubated in Eagle's basal medium supplemented with 5% fetal calf serum. The tissue culture was cultivated during 10 days at 37°C in atmosphere with 5% CO₂. At that, one part of pieces of the same tumor of the same patient was incubated in presence of 10^'6 M testosterone (aromatase substrate), and the other part of the pieces was cultivated without testosterone.

In the defined cases, simultaneously with testosterone, to the tissue culture were added: - various aromatase inhibitors.

- various salts of CMA and its esters.

- combination of aromatase inhibitor on the one hand and salts of CMA and its esters on the other hand.

The pieces of tumor tissue were incubated during 10 days. The pieces were transferred on the new gelatinous sponge and were incubated hi the fresh medium in presence of [³H]thymidine (5 mkKi/ml) during 72 hours to estimate the proliferation rate. Upon the time expiiy, the tissue pieces were weighted, processed with enzymes to separate the cells, and DNA was isolated. For this purpose, the tissue was place into the 0.1 mg/ml collagenase solution and incubated for one night at 37°C, then 0,1 mg/ml of proteinase K was added, and the tissue was incubated for 3 hours at 37°C . DNA isolation was performed according to the routine method in lysis buffer containing 5 M guanidine isothiocyanate, with subsequent extraction by phenol/chloroform mixture and precipitation by 96% ethanol in presence of 3 M sodium acetate. Isolated DNA was diluted with 50 mid of TE-buffer (0.01 M Tris-HCl 0.001 M EDTA pH 7.8). The amount of DNA was determined spectrophotometrically (as optical density (OD) at 260 nm). Radioactivity of the label ([³H]-timidine) incorporated into DNA was estimated with a scintillation counter.

The decrease of tissue DNA specific radioactivity was the measure of antitumoral action of used agents (or their combination). At that all investigated tumor explants were divided in two categories:

Category A: Explants, in which DNA specific radioactivity increased in presence of testosterone. It testified that aromatase is active enough in this tumor and convert testosterone into estradiol which stimulates the tumor growth. In this case, aromatase inhibitors, which inhibit this conversion, had to depress the tumor cell proliferation. In thids case the level in presence of testosterone only was considered as the initial DNA specific radioactivity (proliferation) level. The ratio «Specific radioactivity after exposure)) /«Initial specific radioactivity» was determined. The ratio less than 1 certified the decrease of proliferation. The ratio equal to 1 meant that there are no changes of estrogene-induced proliferation under the influence of investigated agents and their combinations.

Category B: Explants, in which DNA specific radioctivity did not change in presence of testosterone. This meant that aromatase way of intratumor estrogen synthesis has no significance for the growth of the tumor in this patient. In this case, the possible antiproliferative (antitumoral) effect of aromatase inhibitors, as well in combination with CMA, has other mechanisms. In this case, the initial DNA specific radioactivity level (=proliferation) was the level of DNA specific radioactivity without any hormone/antihormone influences. The ratio (R) = «Specific radioactivity after exposure/Initial specific radioactivity)) was determined. The ratio less than 1 showed the decrease of proliferation. The ratio equal to 1 meant that there were no changes of estrogene-induced proliferation under the influence of investigated agents and their combinations.

NF KB activity before and after incubation was determined with the kit

StressXpress NF-κB, p50 ELISA Kit (StressGen company, catalogue N° EKS-445) with the luminescent spectrophotometer. Changes in NF KB activity were expressed as percent ratio to the control, that is to the basal (without any influences) activity of NFKB factor in tumor cells.

EXAMPLE 3. The ability of CMA, its salts and its esters to increase the antiproliferative effect of aromatase inhibitors on the breast cancer.

The ability of CMA, its salts and esters to increase the antiproliferative effect of aromatase inhibitors on the breast cancer was estimated on the tumor explants obtained from 5 female patients with breast cancer:

Study of influence on tumor proliferation was performed according to the method, as described in Example 2.

The results are presented in the Table No. 2 (for category A explants) and in the Table No. 3 (for category B explants). Table No. 2.

Changes of breast cancer tissue proliferative activity, previously stimulated by addition of testosterone to the incubation medium

(category A) under action of CMA in presence of aromatase inhibitors.

Human breast cancer tissue explant (Category A) code

(J)

C BCl A BC2 A BC3 A DO Aromatase inhibitor (AT) EXM AMG VRL VCL FMS FAD ANL VCL EXM FAD LET VCL (J)

AI concentration (IC₅0X 1), riM 5 1800 1,2 30 6 15 - 5 6 10 -

R

C

H VCL 0,74 0,65 0,44 1,0 0,84 0,44 0,52 1,0 0,21 0,64 0,32 1,0 m CMA, its salt or ester (10 -^"9^y - M) (J)

I CMA 0,35 0,24 0,18 1,0 0,30 0,10 0,30 1,0 0,12 0,41 0,18 1,0 m m NaCMA 0,39 0,26 0,21 1,0 0,27 0,12 0,33 1,0 0,12 0,38 0,20 1,0

MegCMA 0,32 0,23 0,22 1,0 0,29 0,15 0,32 1,0 0,09 0,40 0,21 1,0

Ti EgCMA 0,29 0,19 0,20 1,0 0,26 0,14 0,36 1,0 0,11 0,37 0,21 1,0 ι— PropylCMA 0,36 0,25 0,17 1,0 0,27 0,15 0,34 1,0 0,15 0,36 0,22 1,0 m EthylEt CMA 0,38 0,28 0,19 1,0 0,28 0,13 0,31 1,0 0,14 0,38 0,23 1,0

IO PropylEt CMA 0,40 0,24 0,22 1,0 0,32 0,11 0,29 1,0 0,12 0,41 0,19 1,0

Comments: examestane (EXM); aminoglutethimide (AMG); vorozole (VRL); formestane (FMS); fadrozole (FAD); letrozole (LET), anastrozole (ANL); VCL — vechicle (carrier); 9-oxoacridine-lO-acetic acid (CMA); sodium salt CMA (NaCMA), meglumine (that is N- methylglucamine) CMA salt (MegCMA); eglumine (that is N-ethylglucamine) CMA salt (EgCMA); N-propylglucamine CMA salt (PropylCMA); CMA ethyl ester (EthylEt CMA); propyl ester CMA (PropylEt CMA).

Z8

Data, presented in the Table No.3, show, that CMA, CMA salts and CMA esters strongly increase the ability of aromatase inhibitors to inhibit the aromatase activity, preventing the conversion of androgen into estrogen, and so depressing the hormone stimulated proliferation in human breast cancer tissue. These CMA features are manifested independently from chemical structure of an aromatase inhibitor as well as from its mechanism of action. At that, CMA salts and CMA esters themselves have no inhibitory activity regarding aromatase: during the action of the CMA (or its salt and ester) only there is no blocking (or depressing) of cell proliferation stimulated by estrogen converted from the androgen in the tumor tissue under influence of aromatase.

Table 3.

Changes of breast tumor tissue proliferative activity, previously stimulated by addition of testosterone in the incubation medium (category B) under action of CMA in presence of aromatase inhibitors.

Comments: examestane (EXM); amino glutethimide (AMG); vorozole (VRL); formestane (FMS); fadrozole (FAD); letrozole (LET), anastrozole (ANL); VCL - vechicle (carrier); 9-oxoacridine-10-acetic acid (CMA); sodium salt CMA (NaCMA), meglumine (that is N-methylglucamine) CMA salt (MegCMA); eglumine (that is N- ethylglucamine) CMA salt (EgCMA); N-propylglucamine CMA salt (PropylCMA); CMA ethyl ester (EthylEt CMA); propyl ester CMA (PropylEt CMA). Data presented in the Table No. 3, show that CMA, its salts and its esters strongly increase the ability of aromatase inhibitors to inhibit the proliferation activity in human breast cancer tissue. This effect occurs even if aromatase activity is low (or cells poorly respond or do not respond to the estrogen produced as a result of testosterone aromatisation).

Listed CMA features are manifested independently from chemical structure of an aromarase inhibitor and its mechanism of action. At that CMA salts and esters themselves have no inhibitory activity regarding the tissue proliferation. At the action of CMA alone (or its salt and ester) there is no blocking (or depressing) of tumor proliferation stimulation effect, caused by estrogene, produced in tissue under aromatase influence. It is necessary to mention the fact that the most lipophilic CMA compounds (i.e. CMA esters) and non-steroidal inhibitors of the triazole group have more potent synergistic inhibiting effect on the tumor tissue proliferative activity.

EXAMPLE 4. The ability of CMA, its salts and its esters to increase the antiproliferative effect of aromatase inhibitors on the uterine malignant tumors.

The ability of CMA, its salts and its esters to increase the antiproliferative effect of aromatase inhibitors on the uterine malignant tumors was studied in tumoral explants obtained from 3 patients with uterine cancer:

Studies of influence on tumor proliferation were performed according to the method, described in Example 2.

The results are presented in the Table No.4.

Table No. 4.

Change of proliferative activity (PA) of human breast cancer tissue, that initially was not changed in presence of testosterone in incubation medium (Category B) under the action of CMA in combination with aromatase inhibitors.

C DO

m

(/>

X m m

O

73

C ι— m ro σ>

Comments: examestane (EXM); aminoglutethimide (AMG); vorozole (VRL); formestane (FMS); fadrozole (FAD); letrozole (LET)₃ anastrozole (ANL); VCL - vechicle (carrier); 9-oxoacridine-lO-acetic acid (CMA); sodium salt CMA (NaCMA), meglumine (that is N- methylglucamine) CMA salt (MegCMA); eglumine (that is N-ethylglucamine) CMA salt (EgCMA); N-propylglucamine CMA salt (PropylCMA); CMA ethyl ester (EthylEt CMA); propyl ester CMA (PropylEt CMA).

10

Data, presented in the Table No.4, show, that CMA salts and esters strongly increase the ability of aromatase inhibitors to inhibit the human uterine cancer growth. This effect of CMA occurs even if aromatase activity is low (or cells poorly respond or do not respond to the estrogene produced as a result of testosterone aromatisation).

Listed CMA features are manifested independently from chemical structure of an aromatase inhibitor and its mechanism of action. At that, CMA salts and esters themselves have no inhibitory activity regarding the uterine cancer tissue proliferation. At the action of CMA alone (or its salt and ester) there is no blocking (or depressing) of tumor proliferation stimulation, caused by estrogens, produced in tissue under aromatase influence.

EXAMPLE 5. The ability of CMA, its salts and its esters to decrease the NFKB activity in presence of aromatase inhibitors in female reproductive system malignant tumor tissues.

Experiments were performed as is described in Example 2. Changes of NFKB activity are presented as percent ratio to the control, that is to the basal (without any influences) activity of NF-κB in tumor cells. The results of experiments are presented in the Table No. 5.

Table No. 5.

The ability of CMA, its salts and esters to decrease the NF-κB factor activity in presence of aromatase inhibitors in female reproductive system malignant tumor tissues.

(J)

C OU

m

(/)

I m

m

Comments: examestane (EXM); aminoglutethimide (AMG); vorozole (VRL); formestane (FMS); fadrozole (FAD); letrozole (LET),

Ti anastrozole (ANL); VCL - vechicle (carrier); 9-oxoacridine-lO-acetic acid (CMA); sodium salt CMA (NaCMA), meglumine (that is N- c ι— methylgmcamine) CMA salt (MegCMA); CMA ethyl ester (EthylEt CMA). m ro

B σ> 10

Data, presented in the Table No.4, show, th at CMA salts and esters strongly increase (potentiate) the ability of aromatase inhibitors to decrease the NF KB activity in cells of maligmamt tumors of female reproductive system. CMA and its derivatives themselves have no influence on NF KB activity, but combined action (aromatase inhibitor + CMA) resulted in significant decrease of level of active NF KB.

EXAMPLE 6. The ability of CMA, its salts and its esters to «sensitize» female reproductive system malignant tumor tissues to the subsequent antitumoral action of aromatase inhibitor.

Breast cancer explants (BCa), ovarian cancer (CaOV), and uterine stromal cancer (SCU) were used. Experiments were carried out as is described in Example 2. During first 5 days of incubation, CMA, its salt and its ester (or vechicle = control) were added to the incubation medium. Then CMA was replaced at the cultivating medium with one or another aromatase inhibitor and the tissue was cultivated for next 5 days.

The initial DNA specific radioactivity level (^proliferation level) was the level of DNA specific radioactivity without any hormone/antihormone influences. The ratio «Specific radioactivity after exposure/Initial specific radioactivity)) was determined. The ratio less than 1 showed the decrease of proliferation. The ratio equal to 1 meant that there are no changes in estrogene-induced proliferation under the action of investigated agents and their combinations. The results of experiments are presented in the Table No. 6.

Table No. 6.

The ability of CMA, its salts and esters to «sensitize» female reproductive system malignant tumor tissues to the subsequent antitumoral action of aromatase inhibitor.

(J)

C DO

m (/) m m

Ti c ι— m

ro σ>

Comments: examestane (EXM); aminoglutethimide (AMG); vorozole (VRL); formestane (FMS); fadrozole (FAD); letrozole (LET), anastrozole (ANL); VCL - vechicle (carrier); 9-oxoacridine-lO-acetic acid (CMA); sodium salt CMA (NaCMA), meglumine (that is N- methylglucamine) CMA salt (MegCMA); eglumine (that is N-ethylglucamine) CMA salt (EgCMA); N-propylglucamine CMA salt (PropylCMA); CMA ethyl ester (EthylEt CMA); propyl ester CMA (PropylEt CMA).

10

Data presented in Table No. 6, show that CMA₅ its salts and its esters strongly increase the sensitivity of female reproductive system cancer cells to the subsequent influence of an aromatase inhibitor. It is also evident, that the «sensitization» degree varies depending on chemical nature and mechanism of action of the aromatase inhibitor.

EXAMPLE 7. The ability of CMA, its salts and its esters in combination with aromatase inhibitors to potentiate the effect of chemotherapeutic agents of various classes on female reproductive system malignant tumors.

The histological culture method, like that described in the Example 2, was used to estimate the ability of CMA, its salts and its esters in combination with AI to potentiate the effect of chemotherapeutic agents of various classes on female reproductive system malignant tumors. The explants (obtained during surgery) of the malignant stromal uterine tumor (patient identification SAK), ovarian cancer (patient identification ARR), breast cancer (patient identification ATT) were investigated.

In the present example, instead of proliferative activity, the survival rate was estimated. The hormones were not added to the cultivation medium. The RPMI 1640 medium with addition of 20% embrional veal serum was used to cultivate tumor explants. The tissue samples were incubated during 7 days at 37°C in 5% CO₂ atmosphere. The each tissue piece was incubated on the surface of gelatin sponge submerged in the medium in a well of a 24-wells plastic tissue culture plate.

The effect of each chemotherapeutic agent (CTA) was estimated upon his IC ₅₀ regarding each tumor of each patient. IC₅₀ is the CTA concentration that leads to the decrease of tumor cells survival rate by 50%. To determine the IC₅₀ for each CTA 5 concentrations with «2 x» increment were used. That is, each subsequent CTA concentration was twice as much than previous one. For each CTA concentration no less than three wells (=one experimental point) were used. In this experiment, to the culture medium of several samples AI in concentration 1/10 IC₅₀ of the corresponding AI (see Table 1) and/or CMA in concentration 10^"12 M, were added. Thus, the IC₅₀ of each CTA could be compared (regarding each tumor) in presence or absence of CMA and/or any given AI. Control samples were incubated without AI, CMA and/or CTA.

After a 7 day incubation, the amount of surviving cells was estimated using the

MTT-assay. For this purpose, the cells were isolated and into each well with the cells 3-

(4,5-dimethyltiazole-2-yl)-2,5-diphenyltetrazolium bromide (MTT) (Sigma) was added, and the plated were incubated for 6 hours at 37⁰C in 5% CO₂ atmosphere. After that, samples were transferred to new well. 100% dimethylsulphoxide (DMSO) was added in each well to dilute the newly formed formazan crystalls. Optical density (OD) of formed formazan for each cup was determined at 540 nm wavelength. Wet weight of each incubated tissue piece was estimated to determine the specific absorption that is OD for 1 g of tissue. The inhibition rate (IR, %) for each CTA concentration was determined using following formula:

1 — OD for 1 g of incubated with CTA tumor tissue IR (%) = x

100 % OD for 1 g tumor tissue incubated without any influence

The average IR concentration for each CTA concentration was determined (three wells). The «dose-response» curve was polotted (average IR versus CTA concentration). According to the plotted curve, the CTA, causing the 50% tumor inhibition (specified in the present example as ChemICso), was determined.

The results are presented in the Table No. 7 (for uterine cancer), in the Table No. 8 (for ovarian cancer), in the Table No. 9 (for breast cancer).

Following acronymes are used in the tables:

EXM - examestane; AMG - aminoglutethimide; VRL - vorozole; FMS - formestane; FAD - fadrozole; LET - letrozole; ANL - anastrozole; VCL - vechicle (carrier); CMA - 9-oxoacridine-lO-acetic acid; NaCMA - sodium CMA salt, MegCMA - meglumine (that is N-methylglucamine) CMA salt; EgCMA - eglumine (that is N-ethylglucamine) CMA salt; PropylCMA - N-propylglucamine CMA salt (that is salt wit 3 -0-(N₃N- dimethylamino-n-propyl)- 1 ,2 : 5 ,6-di-O-isopropylidene-α,D-glucofuranose) ; EthylEt CMA - CMA ethyl ester; PropylEt CMA - propyl CMA ester. Table No. 7.

Effect of CMA and AI combination on the sensitivity of human uterine cancer to the chemotherapeutic agents of various classes.

CO

C CU CO

m

CO

I m m

c ι— m σ>

*See the Table No. 1.

Table No. 8.

Effect of CMA and AI combination on the sensitivity of human ovarian cancer to the chemotherapeutic agents of various classes.

(/>

C DO (/>

m

(/>

X m m OO

73

C ι— m ro σ>

* See table 1

Table No. 9.

Effect of CMA and AI combination on the sensitivity of human breast cancer to the chemotherapeutic agents of various classes.

(/> C DD (/>

H C m

U)

I m ¹O m

TH c ι- m ro

σ> *see the Table No. 1

Data presented in Tables Nos. 7,8,9 show that the addition of CMA and AI combination to the culture medium leads to a significant increase of the sensitivity of human female reproductive system cancers to the antitumoral action of CTA of various classes. In cases where the potentiating effect of CMA and AI added separately was not revealed, the combination of CMA and IA, on the contrary, shows an intense potentiating effect on the antitumoral effect of CTA.

This effect was also observed in others experiments with CTA, including other antitumoral agents: for example, other alcylating agents, including chlorambucil, melphalem, mitmycin C, dacarbazine; for example, other antimetabolites, in particular methotrexate, etoposide, cytarabine, capacetabine; for example, other antitumoral antibiotics, in particular, actinomycin D, daunorubicin, bleomycin; for example, vinca alkaloids, vincristine, in particular; for example, glucocorticoids, in particular, prednisone; for example, nitrosourea, in particular, lomustine; for example, antitumoral enzymes, in particular, asparaginase; for example, another topoisomerase inhibitors, in particular, irinotecane; another preparations, that damage the microtubule tubulin assembly, for example, vinorelbine.

Thus, CMA, its salts and esters in combination with aromatase inhibitors could potentiate the action of chemotherapeutic agents of various classes on the female reproductive system malignant tumors, in particular on uterine, ovarian and breast tumors. It shall be noticed that:

- CMA and AI separately do not exerce such effect;

- The observed effect is revealed for AI having various mechanism of action and various chemical structure;

- The organic and non-organic CMA salts and various CMA esters have demonstrated the same effect;

- This effect is observed in relation of CTA having various chemical structure and mechanism of action.

EXAMPLE 8. Preparation of the medication for rectal and intravaginal administration

250 g of finely crystalline CMA were mixed during heating up to 60 degrees Celsius with a suppository mass, consisting from a widely used basis for suppositories Witepsol W 35 and twin 80 emulsifier, the mixture was homogenized in a mixer and suppository mass with total weight of 1.5 kg was obtained; 970 suppositories were molded, weighing 1.5 g each and with following composition: 9-oxoacridine-lO-acetic acid - 0.250 g; twin 80 - 0.45 g; Witepsol W 35 - 1.2 g. The yield was 97%. EXAMPLE 9. Treatment of female reproductive system cancer (in particular, advanced breast cancer) performed according to this invention with simultaneous administration of aromatase inhibitor and CMA.

Female, 56 years old, with advanced breast cancer (Tl Nl Mo) received tamoxifen during 8 months. The resistance to tamoxifen has developed, and the tumor continued to grow. By informed consent, AI (exemestane) was administered in dose 25 mg daily intramuscularly in combination with CMA sodium salt (4 mg/kg body weight once daily as calculated on the basis of CMA residue). Preparations were administered during 180 days. By the 2^nd month from the commenecement of treatment, the tumor process was considered as "stabilization", according to the WHO criteria. Stabilization was observed during the whole period of treatment. The treatment according to the proposed method was well tolerated: main adverse effects were "hot flashes" and nausea, which are specific for exemestane. There were no effects observed caused by the CMA sodium salt. Thus, the inventive method is highly effective and safe in treatment of female reproductive system cancer (in particular of breast cancer), and particularly, in the variants, when CMA and AI are administered simultaneously.

EXAMPLE 10. Treatment of female reproductive system cancer (in particular, diffuse ovarian cancer) performed according to this invention with consequent administration of aromatase inhibitor and CMA.

In female, 65 years old, 3 years after successful surgical treatment, a recurrence of ovarian cancer in small pelvis was observed. Surgical treatment was not performed because of concomitant diseases and significant spread of the tumor process. The treatment with the inventive method was performed by the patient's informed concent. Thereafter, the meglumine CMA salt (Cycloferon, Russia) was administered intravenously during 30 days twice daily (single dose 500 mg (14 mg/kg body weight)calculated on the basis of CMA residue). After that, the CMA treatment was discontinued, and from the day 31 of treatment, the patient received aromatase inhibitor letrozole 2,5 mg/daily per os. The response to treatment was estimated according to the UICC (International Union Against Cancer Conference) criteria, and according to the serum level of tumor marker CAl 25. The response to treatment was estimated every 4 weeks. By the week 8 from the beginning of therapy, a partial response to treatment was observed, the CA 125 level dropped by 50%. This effect persisted during consequent 30 months of treatment.

Thus, the inventive method is highly effective in treatment of female reproductive system cancer (in particular of ovarian cancer) in variant, when 9- oxoacridine-10-acetic acid or its salt is administered before the introduction of aromatase inhibitor.

EXAMPLE 11. Treatment of female reproductive system cancer (in particular, advanced endometrium cancer) when administration of the 9-oxoacridine-lO-acetic acid or its salts is initiated prior to an aromatase inhibitor administration and continued along with an aromatase inhibitor administration.

68 years old female suffered from advanced breast cancer (T2 Nl Ml). Administered progestins (Depo-Provera) did not inghibit the tumor growth. By patient informed concent, eglumine CMA salt was administered (100 mg/kg body weight one time daily in CMA equivalent) in 5% glucose solution with drip intravenous infusion once a day.

This CMA salt was administered for 3 weeks. Since the week 2, anastrozole was administered at dose of 1 mg/daily, and from the week 6; the ethyl CMA ester (as suppositories on the base of Witepsol) was administered per rectum at dose 250 mg twice a day. By the month 3 from the beginning of treatment, a full remission according to the WHO criteria was observed. Remission was observed during the whole period of treatment. The treatment was well tolerated. There were no adverse effects observed caused by the CMA salt. Thus, the inventive method is highly effective and safe in treatment of female reproductive system cancer (in particular, of uterine cancer) in the variant, when treatment with CMA or its salt or its ester is started before the AI and continued simultaneously with the aromatase inhibitor therapy.

EAXMPLE 12. Treatment of female reproductive system cancer when the combination of 9-oxoacridine-10-acetic acid and its salt, aromatase inhibitor, and chemotherapeutic agent is used. In a female, 67 years old, breast cancer (adenocarcinoma) is diagnosed (T2 Nl Ml). Under patient informed consent, aromatase inhibitor letrozole at dose 2,5 mg/daily and eglumine CMA salt (600 mg in CMA equivalrent once a day in tablets per os) were administered. Simultaneously, the patient received three «classical» polychemotherapy CMF courses (cyclophosphamide, methotrexate, 5-fluorouracil) during the 1^st, 2" and 3 ^rd months of treatment. By the month 3 from the beginning of treatment, a partial regression of tumor process was observed, according to the WHO criteria. There were no adverse effects observed, caused by the CMA salt. Thus, the inventive method is highly effective and safe in treatment of female reproductive system cancer in this variant (when addition of chemotherapeutic agents is added to combination with 9- oxoacridine-10-acetic acid, its salt with aromatase inhibitor).

EXAMPLE 13. Prophylaxis of recurrence of malignant tumor of reproductive system with combination 9-oxoacridine-lO-acetic acid salt and an aromatase inhibitor.

63 old female suffered from breast cancer (T₂ N₁ M₀) was treated with mastectomy with following anasrtrozol administration (1 mg/day) in combination with peroral CMA meglumine salt (as gelatine capsules of 250 mg calculated based on CM A residue) once a day. 5-year (120-months) follow-up did not show the relapse of the treated tumor or development new (second) tumor in contralateral breast. Thus, the inventive method is highly effective in prophylaxis of recurrence of female reproductive system cancer, in particular, breast cancer.

EXAMPLE 14. The kit for female treatment of malignant tumor of reproductive system, containing CMA salt and an aromatase inhibitor.

A) Sodium CMA salt tablets (250 mg) are prepared as follows. 125 g of CMA sodium salt are mixed with 83 g of dry polyglucin, 1 g of polyvinylpirrolidone and 272 g of sodium chloride. Tablets are granulated, and the granulate is then sprayed with talc and pellitized in tablet-press. 970 tablets are received (yield 97%). Then the tablets are covered with enteric coating on the base of polymethacrylate, dried and put in 10-tablet polyethylene terephthalate blister and covered with aluminium foil.

B) Tablets of aromatase inhibitor are prepared as follows. The tablets of following composition are prepared

Anastrozole (USP24), mg 1.00

Talc, mg 8.80

Polyvinylpirolydon, mg 5.00

Fine-grained silicon oxide, mg 6.80

Modified starch, mg 42.70

Microcrystallic cellulose Avicel PH 102, mg 76.70

Microcrystallic cellulose Avicel PH 101, mg 40.00

100 mg of aromatase inhibitor anastrozole (USP 24) and 0.38 g of fine-grained silicon oxide are riddled and premixed during 10 minutes. 7.67 g of microcrystallic cellulose Avicel PHl 02, 0.50 g of polyvinylpirolidone, 4.27 g of modified starch, 2.00 g of microcrystallic cellulose Avicel PHlOl and 0.38 g of talc are riddled and added to the received premix. All components are mixed during 20 minutes. Then, the mixture is compressed in the consolidator at 50 IdNT. Consolidated mixture is evacuated with the vibrating sieve from the consolidator. Then 2.00 g of microcrystallic cellulose Avicel PHlOl and 0.50 g of talc are added, and the mixture is mixed during 20 minutes. Then the mixture is pellitized in the tablet-press at 4 IdST. 980 tablets are recieved (yield 98%), each containing from 0.98 to 1.02 mg of anastrozole. Tablets are put in the 10-tablet polyethyleneterephthalate blister and covered with aluminium foil.

C) The kit for female treatment of malignant tumor of reproductive system, containing CMA salt and an aromatase inhibitor anastrozole.

Two prepared blisters with CMA sodium salt tablets and two prepared blisters with the aromatase inhibitor tablets are put in the individual consumer package (carton box) with a patient information leaflet.

EXAMPLE 15. The kit for female treatment of malignant tumor of reproductive system, containing CMA salt and an aromatase inhibitor. A) Preparation of tablets, containing CMA salt as an active ingredient. 278 g of CMA eglumine salt (150 in CMA equivalent) and 32.6 g of eglumine are mixed and granulated in the granulator, adding the methylcelMose solution. After sieving, the granulate is sprayed with magnesium stearate, and the mixture is pellitized, 970 tablet cores are received. Received tablet cores are covered with emulsium, that contains 13% (mass.) of methacrylate copolymer and ethcarylate and 7% (mass.) of 1,2- propylenglicole. The cover mass of each core is 0.015-0.020 g. As a result, the tablets are received, each containing 280 g of eglumine CMA salt, 0.03 g of eglumine and 0.004 g of methylcelMose and magnesium stearate (in total) and 0.015-0.020 g of enteric coating cover. Tablets are put in the 50-tablet polyethylene terephthalate blister and covered with aluminium foil.

B) Preparation of tablets containing aromatase inhibitor exemestane as an active ingredient.

The tablets of following content are prepared:

Exemestane (USP) mg 25.00

Talc, mg 19.80

Polyvinylpirolydon, mg 5.00

Fine-grained silicon oxide, mg 6.80

Modified starch, mg 42.70

Microcrystallic cellulose Avicel PH 102, mg 45.70

Microcrystallic cellulose Avicel PH 101, mg 36.00

2.50 g of aromatase inhibitor exemestane (USP 26) and 0.68 g of fine-grained silicon oxide are riddled and premixed during 10 minutes. 4.57 g of microcrystallic cellulose Avicel PH102, 0.50 g of polyvinylpirolidone, 4.27 g of modified starch, 1.60 g of microcrystallic cellulose Avicel PHlOl and 0.38 g of talc are riddled and added to the received premix. All components are mixed during 20 minutes. Then, the mixture is compressed in the consolidator at 60 IcN. Consolidated mixture is evacuated with the vibrating sieve from the consolidator. Then 2.00 g of microcrystallic cellulose Avicel

PHlOl and 1.60 g of talc are added, and the mixture is mixed during 20 minutes. Then the mixture is pellitized in the tablet-press at 5 IcN. Active compound (exemestane) content in one tablet is from 22.5 to 27.5 mg. Tablets are put in the 50-tablet polyethylene terephthalate blister and covered with aluminium foil.

C) The kit for treatment of the malignant tumors of female reproductive system contains CMA eglumine salt and aromatase inhibitor. Two blisters with CMA eglumine salt and two blisters with aromatase inhibitor exemestane are put in the carton consumer package with a patient information leaflet.

Claims

CLAIMS:

1. A method for the treatment and/or prophylaxis of tumor reccurence in organs of a female reproductive system, the method comprising the steps of: (a) administering an effective amount of 9-oxoacridine-lO-acetic acid and/or pharmaceutically acceptable salts and/or esters thereof to a patient in need thereof, and (b) hormonotherapy aimed to diminish aromatase enzyme activity by administration of aromatase inhibitors.

2. A method of claim 1 wherein said tumors of female reproductive system organs are selected from the group including breast cancer, cancer uteri, ovarian cancer.

3. A method of claim 1 or 2 wherein said salts of 9-oxoacridine-10-acetic acid are selected from the group including sodium, meglumine, eglumine salts and 3-O- (N,N-dimethylamino-72-propyl)- 1 ,2 : 5,6-di-O-isoρropyliden- a, D -glucofuranose salt.

4. A method of claim 1 or 2 wherein said esters of salts of 9-oxoacridine-10-acetic acid are selected from the group including ethyl, propyl, butyl, isopropyl, amyl esters.

5. A method of any one of claims 1 to 4 wherein the said aromatase inhibitor is a steroidal aromatase inhibitor.

6. A method of any one of claims 1 to 4 wherein said aromatase inhibitor is a nonsteroidal aromatase inhibitor.

7. A method of claim 5 wherein said steroidal aromatase inhibitor is selected from the group including formestan and exemestan.

S. A method of claim 6 wherein said non-steroidal aromatase inhibitor is selected from the group including aminoglutetimide, fadrozole, anastrozole, letrozole, vorozole.

9. A method of any one of claims 1 to 8, further comprising administering one or more chemotherapeutic agents.

10. A method of claim 9 wherein said chemotherapeutic agent is selected from the group including alkylating agents; antimetabolites; antitumoral antibiotics; vinca alcaloids; taxans; glucocorticoids; antitumoral enzymes; microtubule tubulin polymerization inhibitors.

11. A method of any one of claims 1 to 10 wherein said 9-oxoacridine-lO-acetic acid and/or its pharmaceutically acceptable salts and/or its esters are administered prior to treatment with an aromatase inhibitor.

12. A method of any one of claims 1 to 10 wherein said 9-oxoacridine-10-acetic acid and/or its pharmaceutically acceptable salts and/or its esters are administered simultaneously with treatment with an aromatase inhibitor.

13. A method of any one of claims 1 to 10 wherein said 9-oxoacridine-10-acetic acid and/or its pharmaceutically acceptable salts and/or its esters are administered before the commencement of treatment with an aromatase inhibitor and then are continued to be administered simultaneously with treatment with an aromatase inhibitor.

14. A method of any one of claims 1 to 13 wherein said 9-oxoacridine-10-acetic acid and/or its pharmaceutically acceptable salts and/or its esters are administered daily or every other day during the period of 1 month to 120 months at a daily dose 4 to 300 mg/kg body weight as calculated based on residue of 9- oxoacridine-10-acetic acid.

15. Amethod of any one of claims 1 to 13 wherein said 9-oxoacridine-10-acetic acid and/or pharmaceutically acceptable salts and/or esters thereof is administered in amounts effective to enhance the inhibitory activity of aromatase inhibitors.

16. Amethod of any one of claims 1 to 13 wherein said 9-oxoacridine-10-acetic acid and/or pharmaceutically acceptable salts and/or esters thereof is administered in amounts effective to effective to reduce active NF kB level.

17. Amethod of any one of claims 1 to 13 wherein said 9-oxoacridine-10-acetic acid and/or pharmaceutically acceptable salts and/or esters thereof is administered in a single dose from 0.5 to 100 mg/kg body weight (calculated based on 9- oxoacridine-10-acetic acid), preferably from 4 to 20 mg/kg body weight.

18. A method of any one of claims 1 to 13 wherein said 9-oxoacridine-lO-acetic acid and/or pharmaceutically acceptable salts and/or esters thereof is administered in a daily dose from 2 to 1000 mg/kg, preferably from 2 to 200 mg/kg.

19. Use of 9-oxoacridine-10-acetic acid and/or its pharmaceutically acceptable salts and/or its esters for the manufacture of a medicament for the treatment of malignant tumors of female reproductive system, wherein 9-oxoacridine-10- acetic acid and/or pharmaceutically acceptable salts and/or esters is used in combination with an aromatase inhibitor.

20. Use of claim 19 wherein said salts of 9-oxoacridine-lO-acetic acid are selected from the group including sodium, meglumine, eglumine salts and 3-0-(N₅N- dimethylamino-n-propyl)-l,2:5,6-di-O-isopropyliden- «,D -glucofuranose salt.

21. Use of 9-oxoacridine-lO-acetic acid and/or salts and/or esters thereof for the preparation of a medicament for treating malignant tumors of female reproductive system in combination with aromatase inhibitors, wherein oxoacridine-10-acetic acid and/or salts and/or esters thereof is used in amounts effective to reduce active NF kB level.

22. A kit for the treatment of malignant tumors of female reproductive system, wherein the kit comprises a unit dosage of 9-oxoacridine-lO-acetic acid and/or acceptable salts and/or esters thereof and a unit dosage of an aromatase inhibitor, wherein 9-oxoacridine-lO-acetic acid and/or pharmaceutically acceptable salts and/or esters thereof is present in amounts effective to enhance the inhibitory activity of the said aromatase inhibitor.

23. A kit of claim 22 wherein the said salts of 9-oxoacridine-lO-acetic acid are selected from the group including sodium, meglumine, eglumine salts and 3-O- (N,N-dimethylamino-/?-propyl)-l,2:5,6-di-O-isopropyliden- α.D-glucofuranose salt.

24. A kit of claim 22 wherein the said aromatase inhibitor is a steroidal aromatase inhibitor selected from formestan or exemestan.

25. A kit of claim 22, wherein the said aromatase inhibitor is a non-steroidal aromatase inhibitor and it is selected from the group including aminoglutethimide, fadrozole, anastrozole, letrozole, vorozole.

26. A kit of any one of claims 22 to 25 wherein said 9-oxoacridine-10-acetic acid and/or pharmaceutically acceptable salts and/or esters thereof is present in unit dosage providing administration of 0.5 to 100 mg/kg body weight as calculated based on P-oxoacridine-lO-acetic acid, preferably from 4 to 20 mg/kg body weight.