Drugs for estrogen receptor beta (ERβ)-positive breast tumors

Abstract

The present invention relates to the field of medicine and to the steroid compound 4-hydroxyandrost-4-ene-3,17-dione (4-OHA) or its esters as a prodrug for use in the treatment of patients with estrogen receptor beta (ERβ)-positive breast tumors. The present invention further relates to pharmaceutical compositions comprising said steroid compound for topical application and to the use of said steroid compound in the preoperative topical treatment of ERβ-positive breast cancer tissue.

Description

[Technical field]

[0001] The present invention relates to the field of medicine and to the steroid compound 4-hydroxyandrost-4-ene-3,17-dione (4-OHA) as a prodrug for use in the treatment of patients suffering from estrogen receptor beta (ERβ) positive breast tumors. The present invention further relates to pharmaceutical compositions comprising said steroid compound for topical application and to the use of said steroid compound in the pre-operative topical treatment of ERβ positive breast cancer tissue. [Background technology]

[0002] After colorectal cancer, breast cancer is the most frequent cancer disease in Western countries, despite the fact that it affects almost exclusively women. In Germany, breast cancer accounts for approximately 20% of all cancers diagnosed in women.

[0003] To date, treatment in the majority of cases consists of surgical removal of the tumor (lumpectomy) or, if the size of the tumor precludes breast-conserving therapy, complete removal of the affected breast. In both cases, surgery (surgical resection) is followed by drug therapy. A particular complication of breast cancer is the high probability of metastatic spread to other organs, especially the liver, brain, bone and skin. Surgery is therefore followed by drug therapy (adjuvant therapy). This therapy also aims to prevent local recurrence of the disease. If the resected tumor presents estrogen receptors, anti-estrogen drugs (tamoxifen or aromatase inhibitors) are used. If estrogen receptors are absent, postoperative drug therapy with cytostatic drugs is performed.

[0004] Tamoxifen binds to the estrogen receptor (ER) and blocks the growth-promoting effects of estrogen. Aromatase inhibitors block the final step in the production of estrogen from androgen precursors (c-19-steroids). They either bind irreversibly to the active site of the enzyme (aromatase inactivators, i.e. steroidal aromatase inhibitors: 4-hydroxyandrostenedione (formestane = Lentaron®) or exemestane (Aromasin®)) or competitively inhibit the cytochrome p450 portion of the enzyme (non-steroidal aromatase inhibitors: Anastrozol (Arimidex®) and Letrozol (Femara®).

[0005] If the breast cancer shows overexpression of the Her2neu gene, antibodies against this cell surface protein, such as trastuzumab (Herceptin®), are administered intravenously.

[0006] Basic and clinical data indicate that the aromatized metabolites of androgens, i.e., estrogens, are hormones involved in the pathogenic cellular changes associated with the development of several hormone-dependent cancers, such as breast, endometrial, and ovarian cancers. Aromatase has recently been recognized as an enzyme capable of directly synthesizing estradiol in tumors.

[0007] Endogenous estrogens are finally formed with androstenedione or testosterone as direct precursors. Local production of estrogens in breast tissue, especially in the tumor cells themselves, was thought to be of great importance in the treatment of this disease. In this context, the aromatization of steroid ring A, which is carried out by the enzyme aromatase, and its inhibition, were thought to play an important role in anti-breast cancer approaches.

[0008] Therefore, conventional therapeutic approaches against female breast cancer have focused on aromatase inhibition per se, as well as other concepts based on estrogen receptor antagonists or therapeutic antibodies. Thus, steroid substances that have been reported to have aromatase inhibitory properties, such as Δ 1 -testrolactone [U.S. Pat. No. 2,744,120], 4-hydroxy-androst-4-ene-3,17-dione and its esters [see, for example, U.S. Pat. No. 4,235,893], 10-(1,2-propadienyl)-estr-4-ene-3,17-dione (U.S. Pat. No. 4,289,762), 10-(2-propynyl)-estr-4-ene-3,17-dione [J. Amer. Chem. Soc., 103 ,3221 (1981) and U.S. Pat. No. 4,322,416], 19-thioandrostene derivatives (European Patent Application No. 100566), androsta-4,6-diene-3,17-dione, androsta-1,4,6-triene-3,17-dione [British Patent Application No. 2,100,601A], androsta-1,4-diene-3,17-dione [Cancer Res. (Suppl.) 42 , 3327 (1982)] is described.

[0009] However, clinical studies carried out using estrogen receptor (ER) antagonists and aromatase inhibitors have shown that these compounds are unable to exert a significant and sufficient inhibitory effect on tumor proliferation and / or growth. For example, Tamoxifen®-based therapy suffers from drawbacks such as tachyphylaxis (treatment failure) and its effect in some cells is estrogen-like (e.g. risk of thrombosis and endometrial cancer), which is why Tamoxifen® cannot be administered for longer than 3-5 years. Newer drugs, such as aromatase inhibitors, can in principle only be used in postmenopausal women and are only effective against ER-positive tumors.

[0010] It has been shown that stably transfected cells expressing the androgen receptor (AR) acquire the ability to respond to androgens by inducing cell proliferation arrest (J. Szelei et al., “Androgen-Induced Inhibition of Proliferation in Human Breast Cancer MCF7 Cells Transfected with Androgen Receptor”, Endocrinol., 138, no. 4, 1406-1412, 1997). A more recent report (J. Ortmann et al., “Testosterone and 5α-dihydrotestosterone inhibit in vitro growth of human breast cancer cell lines”, Gynecol. Endocrinol. 2002; 16: 113-120) shows that testosterone and 5α-dihydrotestosterone (DHT) can inhibit cell proliferation in certain cell lines. Testosterone itself has a similar effect. However, its clinical usefulness is hindered by the fact that it is rapidly metabolized to either estradiol or dihydrotestosterone (DHT). Local formation of estradiol from testosterone within breast cancer cells by local aromatase activity promotes cancer growth.

[0011] On the other hand, AR itself was considered as a regulator of processes involved in differentiation, homeostasis and development of male secondary sexual characteristics. Therefore, treatment schemes directed at AR focused on male androgen-related diseases such as hypogonadism, male infertility, prostate cancer, delayed puberty, hirsutism, androgen deficiency disorders, androgen replacement in aging men and male pattern baldness. In this context, the disclosure of US 2,762,818 A does not go beyond the use of 4-hydroxytestosterone and its esters to treat androgen deficiency conditions per se, based on its androgenic and anabolic properties. On the other hand, there was no objective or finding that suggested activity against breast cancer cells or an effect that could infer efficacy against pathologies. Moreover, US 2003 / 0229063 A is concerned with the low androgen to estrogen ratio in men (leading to endocrine disorders) and attempts to use 4-hydroxytestosterone for this purpose only, based solely on its alleged aromatase inhibitory effect.

[0012] WO2007 / 131737 is inter alia focused on the treatment of breast cancer and in this context refers to targeting the androgen receptor (AR) using steroid compounds.

[0013] As already discussed above, some breast cancer tumors are sensitive to estrogen, which exerts its action through specific nuclear protein receptors, namely the estrogen receptor alpha (ERα) and the estrogen receptor beta (ERβ).

[0014] Therefore, many studies have been carried out to elucidate the role and properties of ERα and ERβ. In this regard, it can be shown that in the majority of breast cancer cases, the expression of ERα dominates that of ERβ. During disease progression, the expression of ERβ decreases in parallel with the increase in malignancy.

[0015] Based on cell culture studies, it has been shown that these two estrogen receptor subtypes mediate the action of estrogen, and that the relative levels of ERβ and ERα in breast cancer may affect cell proliferation, the activity of various signaling pathways, and their response to ER ligands (Endocrinology, Volume 147, Issue 10, 1 October 2006, Pages 4831-4842). Furthermore, it has been shown that ERβ acts as a tissue-specific tumor suppressor with antiproliferative properties (Clin. Pharmacol. Ther. 2011 Jan;89(1):44-55). Moreover, activation of ERβ can further lead to the disruption of signaling pathways mediated by ERα by the generation of heterodimers containing ERα and ERβ (ERα:ERβ heterodimers). Expression of ERα predominates in normal, i.e., unaffected, breast tissues, whereas expression of ERβ predominates in diseased, i.e., breast cancer tissues. It is now believed that ERα drives estrogen-mediated proliferation, whereas ERβ is a counterpart of ERα that limits or completely prevents this estrogen-mediated proliferation (Mol. Endocrinol. 2003, Feb;17(2):Pages 203-8; Lindberg et al.).

[0016] As can be seen from the above, the therapeutic concepts known so far in the field of breast cancer focus either on aromatase inhibition alone or on binding to the androgen receptor (AR).Moreover, in addition to treating breast cancer by aromatase inhibition or AR binding, the therapeutic method also includes in the majority of cases surgical removal of the tumor or, in cases where the size of the tumor prevents breast-conserving therapy, complete removal of the affected breast.

[0017] Such treatments, and in particular the partial or complete removal of the affected breast, are extremely taxing on the patient, both physically and mentally. [Prior art documents] [Patent documents]

[0018] [Patent Document 1] U.S. Patent No. 2,744,120 [Patent Document 2] U.S. Patent No. 4,235,893 [Patent Document 3] U.S. Patent No. 4,289,762 [Patent Document 4] U.S. Patent No. 4,322,416 [Patent Document 5] European Patent Application No. 100566 [Patent Document 6] UK Patent Application No. 2,100,601A [Patent Document 7] US2,762,818A [Patent Document 8] US2003 / 0229063A [Patent Document 9] WO2007 / 131737 [Non-patent literature]

[0019] [Non-Patent Document 1] J.Amer.Chem.Soc.,103,3221(1981) [Non-Patent Document 2] Cancer Res.(Suppl.)42,3327(1982) [Non-Patent Document 3] J. Szelei et al., “Androgen-Induced Inhibition of Proliferation in Human Breast Cancer MCF7 Cells Transfected with Androgen Receptor”, Endocrinol., 138, no. 4, 1406-1412, 1997 [Non-Patent Document 4] J. Ortmann et al., “Testosterone and 5α-dihydrotestosterone inhibit in vitro growth of human breast cancer cell lines”, Gynecol. Endocrinol. 2002;16:113-120 [Non-Patent Document 5] Endocrinology,Volume 147,Issue 10,1 October 2006,Pages 4831-4842 [Non-Patent Document 6] Clin.Pharmacol.Ther.2011 Jan;89(1):44-55 [Non-Patent Document 7] Mol.Endocrinol.2003,Feb;17(2):Pages 203-8;Lindberg et al. Summary of the Invention [Problem to be solved by the invention]

[0020] There is therefore a need, and it is therefore an object of the present invention, to provide an effective therapeutic approach for treating patients suffering from certain types of breast cancer, in particular ERβ-positive breast cancer. In particular, it is an object of the present invention to provide an effective therapeutic approach that allows the reduction of the size of the tumor to a size that results in a breast-conserving therapy, i.e. a therapy that prevents the complete surgical removal of the affected breast. [Means for solving the problem]

[0021] Various aspects, advantageous features and preferred embodiments of the present invention summarized in the following items contribute individually or in combination to solving the object of the present invention. 1. A compound of formula I for use as a prodrug in a method for treating a patient suffering from an estrogen receptor beta (ERβ)-positive and optionally an androgen receptor (AR)-positive breast tumor. [ka] 2. The steroid compound 4-hydroxyandrost-4-ene-3,17-dione (4-OHA) or an ester thereof, wherein the prodrug is applied topically to a patient. In a preferred embodiment, the patient has an ERβ-positive and AR-positive breast tumor. In a further preferred embodiment, the patient suffers from an ERβ positive and AR negative breast tumor. An example of an ERβ positive and AR negative breast cancer is, for example, triple negative breast cancer ("TNBC"). TNBC is defined as estrogen receptor alpha (ERα) negative, progesterone receptor (PR) negative, and HER2 receptor negative. ERβ has been found to be expressed in TNBC. Within the meaning of the present invention, the term "prodrug" defines a drug or compound that, after administration, is metabolized (ie, converted within the body) into a pharmacologically active substance. With respect to the term "ester", reference is made to the following definition. 2. The steroid compound 4-OHA or its ester for use according to item 1, wherein the ERβ-positive breast cancer is triple-negative breast cancer (TNBC). 3. The steroid compound 4-OHA or an ester thereof for use according to items 1 or 2, wherein the treatment is carried out before a planned surgical resection of breast tumor tissue and / or the treatment is for reducing breast tumor tissue. 4. The steroid compound 4-OHA or an ester thereof for use according to any one of items 1 to 3, wherein said prodrug 4-OHA is metabolized to a pharmacologically active substance having hydroxyl groups at least at the C4 and C17 positions. 5. The steroid compound 4-OHA or an ester thereof for use according to any of items 1 to 4, wherein the prodrug 4-OHA is metabolized to a pharmacologically active substance having a higher binding affinity for estrogen receptor beta (ERβ) compared to its binding affinity for estrogen receptor alpha (ERα), either both of these estrogen receptors being expressed by breast tumor tissue or only ERβ being expressed by breast tumor tissue, and wherein said treatment results in agonistic binding of ERβ, and wherein preferably said pharmacologically active substance exhibits a binding affinity for ERβ that is at least 3-fold, preferably at least 4, 5, or 6-fold, more preferably at least 9-fold, most preferably at least 10-fold higher than its binding affinity for ERα. If the breast tumor tissue expresses an androgen receptor, it is further desirable that the pharmacologically active agent have binding affinity for the androgen receptor (in addition to ERβ) expressed by the breast tumor tissue. Also, breast tumor tissue may not express the androgen receptor, so long as it expresses ERβ, for example, if the breast tumor tissue is TNBC, the androgen receptor is not expressed. 6. The steroid compound 4-OHA or an ester thereof for use according to any one of items 1 to 5, wherein the pharmacologically active substance comprises 4-hydroxy-5-alpha-androstane-3-beta,17beta-diol (4-OHBA) or 4-hydroxytestosterone (4-OHT), preferably 4-hydroxy-5-alpha-androstane-3β,17β-diol (4-OHBA) or 4-hydroxytestosterone (4-OHT), preferably the pharmacologically active substance is 4-OHBA. In an equally preferred embodiment, the pharmacologically active substance comprises 4-hydroxytestosterone (4-OHT) or a hydroxyester thereof, preferably 4-hydroxytestosterone (4-OHT) or a hydroxyester thereof. Alternatively or in combination, the pharmacologically active substance comprises 4-hydroxy-5α-androstane-3β,17β-diol (4-OHBA) or a hydroxyester thereof, preferably 4-hydroxy-5α-androstane-3β,17β-diol (4-OHBA) or a hydroxyester thereof. 7. 4-Hydroxytestosterone (4-OHT) or a hydroxyester thereof for use in a method of treating a patient suffering from an estrogen receptor beta (ERβ) positive and optionally an androgen receptor (AR) positive breast tumor, which is applied topically to the patient. 8. 4-Hydroxy-5α-androstane-3β,17β-diol (4-OHBA) or a hydroxyester thereof for use in a method of treating a patient suffering from an estrogen receptor beta (ERβ) positive and optionally an androgen receptor (AR) positive breast tumor, which is applied topically to the patient. 9. 4-OHT or 4-OHBA or the respective hydroxyesters for use in the method according to item 7 or 8, wherein the patient is suffering from TNBC. 10. 4-OHT for use in the method according to any one of items 7 to 9, wherein the treatment is carried out prior to planned surgical removal of breast tumor tissue and / or the treatment is for reducing breast tumor tissue. 11. Compounds of formula I as prodrugs for use in treating patients with estrogen receptor beta (ERβ) positive breast tumors and, optionally, androgen receptor (AR) positive breast tumors. [ka] 1. A pharmaceutical composition suitable for topical application comprising the steroid compound 4-hydroxyandrost-4-ene-3,17-dione (4-OHA) or an ester thereof, and a pharma- ceutical acceptable carrier(s) and / or excipient(s), wherein the treatment is carried out prior to planned surgical resection of breast tumor tissue. In a preferred embodiment, the patient has an ERβ-positive and AR-positive breast tumor. In a further preferred embodiment, the patient has an ERβ-positive and AR-negative breast tumor, eg, TNBC. 12. The pharmaceutical composition for use according to item 11, wherein the ERβ-positive breast cancer is triple-negative breast cancer (TNBC). 13. A pharmaceutical composition for use according to any one of the preceding items, wherein the pre-operative treatment comprises topical application of a pharmaceutical composition in an amount effective to reduce breast tumor tissue. 14. A pharmaceutical composition for use according to any one of the preceding items, wherein the preoperative treatment is carried out for a period sufficient to achieve reduction of the breast tumor tissue to a size that allows for breast-conserving surgical resection of the breast tumor tissue. In preferred embodiments, pre-operative treatment is administered for a period of up to 6 months, or for a period of up to 5, 4, 3, or 2 months. It is also possible that reduction of breast tumor tissue to a size that allows for breast-conserving surgical resection can be achieved after 11 weeks, or 10, 9, 8, 7, 6, 5, 4, 3, or 2 weeks of preoperative treatment. 15. A pharmaceutical composition for use according to any one of the preceding items, wherein the tumor tissue as determined by ultrasound, X-ray or mammography is significantly reduced at the completion of treatment compared to the size of the breast tumor tissue at the start of treatment, preferably the tumor tissue is reduced by at least 13% by volume, or at least 14% by volume, or 15% by volume, or at least 20% by volume, preferably at least 30% by volume, more preferably at least 13% by volume or 14% by volume, or at least 40% by volume, even more preferably at least 50% by volume, at least 60% by volume or at least 70% by volume. In a preferred embodiment, the size of the breast tumor tissue is determined by mammography. If the breast cancer is characterized by expression of ERβ but no (or less significant) expression of ERα and additionally no expression of AR, such as TNBC, the tumor tissue as determined by ultrasound, x-ray, or mammography is reduced by at least 15% by volume compared to the size of the breast tumor tissue at the start of treatment. 16. A pharmaceutical composition for use according to any one of the preceding items, wherein the preoperative treatment is initiated up to 6 months, or up to 5, 4, 3, or 2 months prior to the planned surgical procedure, i.e., breast-conserving surgical removal of the breast tumor tissue. Preoperative treatment can also begin up to 11 weeks, or up to 10, 9, 8, 7, 6, 5, 4, 3, or 2 weeks prior to the scheduled surgery. Preoperative treatment may also begin up to 8 weeks prior to the scheduled surgery, preferably 6 or 5 weeks prior to surgery. 17. A pharmaceutical composition for use according to any one of the preceding items, wherein the prodrug 4-OHA or ester thereof is present in the pharmaceutical composition in an amount of about 1.5% to about 4% by weight, preferably about 2.0% to about 3.0% by weight, more preferably about 2.0% or about 2.5% by weight, based on the amount of the pharmaceutical composition. In a preferred embodiment, the pharmaceutical composition is in the form of a topically applicable composition, such as a cream, gel, lotion, or ointment, more preferably the pharmaceutical composition is a cream. 18. A pharmaceutical composition for use according to any one of the preceding paragraphs, to be applied topically once or twice daily. 19. A pharmaceutical composition suitable for topical application comprising 4-hydroxytestosterone (4-OHT) and / or 4-hydroxy-5α-androstane-3β,17β-diol (4-OHBA) or a hydroxyester of 4-OHT or 4-OHBA, and a pharma- ceutically acceptable carrier(s) and / or excipient(s), for use in the treatment of patients suffering from estrogen receptor beta (ERβ)-positive breast tumors and optionally androgen receptor (AR)-positive breast tumors, wherein the treatment is preferably carried out prior to planned surgical resection of breast tumor tissue. In a further preferred embodiment, the patient has an ERβ-positive and AR-positive breast tumor. In a further preferred embodiment, the patient has an ERβ-positive and AR-negative breast tumor, eg, TNBC. 20. The pharmaceutical composition for use according to item 19, wherein the ERβ-positive breast cancer is triple-negative breast cancer (TNBC). 21. A pharmaceutical composition for use according to items 19 and 20, wherein the preoperative treatment comprises topical application of a pharmaceutical composition in an amount effective to reduce breast tumor tissue. In preferred embodiments, pre-operative treatment is administered for a period of up to 6 months, or for a period of up to 5, 4, 3, or 2 months. It is also possible that reduction of breast tumor tissue to a size that allows for breast-conserving surgical resection can be achieved after 11 weeks, or 10, 9, 8, 7, 6, 5, 4, 3, or 2 weeks of preoperative treatment. 22. A pharmaceutical composition for use according to any one of the preceding items, wherein the tumor tissue as determined by ultrasound, X-ray or mammography is significantly reduced at the completion of treatment compared to the size of the breast tumor tissue at the start of treatment, preferably the tumor tissue is reduced by at least 13% or 14% by volume, or at least 15% by volume, or at least 20% by volume, preferably at least 30% by volume, more preferably at least 40% by volume, even more preferably at least 50% by volume, at least 60% by volume or at least 70% by volume. In a preferred embodiment, the reduction is measured as a reduction in volumetric size; the volumetric size of breast tumor tissue can be determined by mammography. Where the breast cancer is characterized by expression of ERβ but absent (or insignificant) expression of ERα, such as TNBC, the tumor tissue as determined by ultrasound, x-ray, or mammography can be reduced by at least 15% by volume compared to the size of the breast tumor tissue at the start of treatment. 23. A pharmaceutical composition for use according to any one of the preceding items, wherein the preoperative treatment is initiated up to 6 months, or up to 5, 4, 3, or 2 months prior to the planned surgery, i.e., breast-conserving surgical resection of the breast tumor tissue. Preoperative treatment can also begin up to 11 weeks, or up to 10, 9, 8, 7, 6, 5, 4, 3, or 2 weeks prior to the scheduled surgery. Preoperative treatment may also begin up to 8 weeks prior to the scheduled surgery, preferably 6 or 5 weeks prior to surgery. 24. A pharmaceutical composition for use according to any one of the preceding items, wherein 4-OHT and / or 4-OHBA are present in the pharmaceutical composition in an amount of about 1.5% by weight to about 4% by weight, preferably about 2.0% by weight to about 3.0% by weight, more preferably about 2.0% by weight or about 2.5% by weight, based on the amount of the pharmaceutical composition. In a preferred embodiment, the pharmaceutical composition is in the form of a topically applicable composition, such as a cream, gel, lotion, or ointment, more preferably the pharmaceutical composition is a cream. 25. A pharmaceutical composition for use according to any one of the preceding paragraphs, to be applied topically once or twice daily. 26. Use of 4-OHA or its esters as a prodrug in the preoperative topical treatment of ERβ-positive breast cancer tissue. In a preferred embodiment, the patient has an ERβ-positive and androgen receptor (AR)-positive breast tumor. In a further preferred embodiment, the patient has an ERβ-positive and androgen receptor (AR)-negative breast tumor, eg, TNBC. 27. Use of 4-hydroxytestosterone (4-OHT) in the preoperative topical treatment of ERβ-positive and optionally androgen receptor (AR)-positive breast cancer tissue. In a preferred embodiment, the patient has an ERβ-positive and AR-positive breast tumor. In a further preferred embodiment, the patient has an ERβ-positive and AR-negative breast tumor, eg, TNBC. 28. A pharmaceutical composition for use according to any of the preceding items, wherein the ERβ positive breast cancer is triple negative breast cancer (TNBC) and the pharmaceutical composition further comprises, in addition to the prodrug 4-OHA or an ester thereof, and / or in addition to a compound selected from 4-OHT, 4-OHBA and esters of 4-OHT and 4-OHBA, a further active ingredient different from the prodrug 4-OHA and different from a compound selected from 4-OHT, 4-OHBA and their esters, which binds to ERβ and thereby activates ERβ. 29. A pharmaceutical composition for use according to item 28, wherein the further active ingredient is selected from antiestrogens and anticancer indole compounds. In a preferred embodiment, the anti-estrogen drug is, for example, fulvestrant or tamoxifen.The anti-cancer indole compound is preferably an indole derivative that alters estrogen metabolism or is effective in stopping the growth of cancer cells.An example of an anti-cancer indole compound is indole-3-carbinol. [Brief description of the drawings]

[0022] [Figure 1] FIG. 1 shows the specific binding [%] according to the present invention. [Diagram 2] FIG. 1 shows MDA MB-231 cells incubated with increasing concentrations of 4-OHT(K+) at two different concentrations. [Diagram 3]FIG. 1 shows the mean tumor volumes before and after 2.0% SAVE cream treatment. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0023] Definition of Terms Within the meaning of the present invention, the term "ester" in any instance of its reference denotes a suitable ester group, such as a substituted or unsubstituted hydrocarbon group, the carbon atoms of which may also be substituted by 1-3 heteroatoms (O, S, N). In particular, the ester group may be or contain a linear, branched, cyclic and / or aromatic hydrocarbon group, or an acyl group or an aroyl group, such as, but not limited to, formyl, acetyl, propionyl, butyryl, pentanoyl, hexanoyl, heptanoyl and benzoyl. The hydrocarbon group may have 1-12 carbon atoms and may be substituted or unsubstituted with substituents, such as halogens. The carbon atoms of the hydrocarbon group may also be substituted by 1-3 heteroatoms (O, S, N), forming, for example, a heteroaromatic group.

[0024] In the case of the compound of formula I, the ester can be formed with the 4-OH (hydroxyl) group. Whenever 4-OHA is mentioned in the present invention, the ester is also included in this term. In the case of 4-OHT, the ester can be formed with the 4-OH group and / or the 17-OH group. In the case of 4-OHBA, the ester can be formed with the hydroxyl group selected from the 3-OH group, the 4-OH group and the 17-OH group of the 4-OHBA structure. An ester according to any one of the above definitions can be used as a prodrug; upon administration, the ester group can be metabolized leaving the respective OH group free.

[0025] Detailed Description of the Invention The present invention will now be described in more detail by preferred embodiments and examples which are presented for illustrative purposes only and are not to be understood as in any way limiting the scope of the invention.

[0026] In the context of ERβ-positive breast cancer, the inventors have surprisingly found that a significant reduction in tumor size can be achieved by topically administering steroid compounds as prodrugs which, when metabolized, show high selectivity for ERβ and at the same time no or very low selectivity for ERα and, in addition, bind to the androgen receptor (AR) which may be expressed by breast cancer tissue.

[0027] The pharmacologically active substance results from the metabolism of the prodrug, 4-hydroxyandrost-4-ene-3,17-dione (4-OHA) of formula I or an ester thereof, when the prodrug is topically applied to diseased ERβ positive cancerous tissue (breast cancer tissue).

[0028] Since the pharmacologically active substance resulting from the metabolism of the prodrug 4-OHA exerts its function mainly through the activation of ERβ expressed by breast cancer tumor tissue, breast cancer tissue may be AR positive in addition to being ERβ positive, but this is not necessarily the case.In the present invention, it has been found that the prodrug 4-OHA (or the pharmacologically active substance resulting from the metabolism of the prodrug, such as 4-hydroxytestosterone (4-OHT)) can also be used to treat ERβ positive breast cancer, which is therefore AR negative.A highly relevant example of such ERβ positive, AR negative breast cancer is triple negative breast cancer (TNBC).

[0029] The present inventors have found that topical application of 4-OHA (or its esters) results in the metabolism of 4-OHA in breast adipose tissue and tumor tissue. The metabolism is mediated, for example, by aldo-keto-reductase, which reduces the keto group present in the prodrug. As found by the present inventors, the enzyme that reduces 4-OHA to 4-OHT is AKR1C3 (aldo-keto reductase family 1 member C3). Due to the presence of (only) one OH group (compared to at least two OH groups of the metabolites), the percutaneous permeability of 4-OHA or its esters is improved compared to the percutaneous permeability of the metabolite itself. Examples of metabolites are 4-OHEPI (4-β-hydroxy-epiandrosterone), 4-OHBA, or 4-OHT.

[0030] By applying the prodrug (steroid compound) or its ester topically to ERβ-positive and optionally AR-positive breast cancer tissue, and / or by applying 4-OHT or 4-OHBA or an ester of 4-OHT or 4-OHBA (particularly 4-OHT or its ester), a reduction in tumor size can be achieved. As a result of this size reduction, surgical removal of breast tumor tissue can be performed while simultaneously maintaining the affected breast larger than without treatment. In other words, by applying the prodrug or 4-OHT or 4-OHBA or an ester of 4-OHT or 4-OHBA (particularly 4-OHT or its ester), complete removal of the affected breast can often be avoided. Instead, breast-conserving surgery can be performed. Therefore, in the context of the present invention, the prodrug is applied before the planned surgical removal of breast tumor tissue.

[0031] Furthermore, because the number of OH groups of the prodrug is smaller compared to the number of OH groups of the pharmacologically active substance, topical application of the prodrug 4-OHA (or its ester) to the affected area (breast) is improved compared to topical application of the respective pharmacologically active substance, since the skin permeability of the prodrug (or its ester) is enhanced.

[0032] In particular, it has been discovered by the present inventors that the hydroxyl group at C4 position and the hydroxyl group at C17 position in the prodrug of formula I or its ester metabolite(s), respectively, play essential roles in achieving the objectives of the present invention.

[0033] The hydroxyl group at C4 in the metabolized steroid compound allows the resulting pharmacologically active substance to more accurately distinguish between ERα and ERβ expressed by breast tumor tissues, as compared to, for example, β-adiol, which does not present a hydroxyl group at C4. As found, the metabolite of the present invention presenting a hydroxyl group at C4 shows a significantly higher binding affinity for ERβ compared to its binding affinity for ERα. For example, the affinity of metabolite 4-OHBA for ERβ is about 10 times higher than its affinity for ERα. In addition, it has been found that binding to ERβ enhances the expression of ERβ in breast cancer tumor tissues. This enhanced ERβ expression results in increased tumor suppression.

[0034] Besides the beneficial effects of the OH group at C4 position with respect to improved affinity for ERβ (but not for ERα) disclosed above, there are further beneficial properties based on this particular position of the OH group in the steroid compound of formula I. The steroid compound 4-OHA is not a substrate for the enzyme 5α-reductase, which catalyzes the conversion of testosterone to dihydrotestosterone (DHT), thus strongly accelerating its metabolism, enhancing its affinity for the androgen receptor (AR) and inhibiting its binding to SHBG (sex hormone binding globulin).

[0035] It has been found that binding to ERβ as well as to androgen receptor (AR) leads to increased expression of ERβ. Furthermore, binding to AR can lead to a decrease or even inhibition of cell proliferation. This is particularly beneficial when AR is expressed by tumor cells, such as breast cancer tumor cells.

[0036] The binding of the steroid compound of the present invention to AR is mediated by the OH group at C17. In other words, the hydroxyl group at C17 has the effect of giving the steroid compound the additional ability to bind to androgen receptor (AR). This results in the activation of AR-mediated signal transduction pathway, which ultimately induces the arrest (or at least the reduction) of cell proliferation.

[0037] However, even more surprisingly, in the present invention, it has been found that 4-OHA also exerts beneficial effects when used to treat patients with ERβ-positive breast cancer that does not express AR. A very important example of such breast cancer is TNBC. Surprisingly, in the present invention, it has been found that treatment with the prodrug 4-OHA or 4-OHT can reduce the tumor volume of TNBC. This is surprising and noteworthy, especially since TNBC is known as a type of breast cancer that can be very aggressive. Since it does not express ERα, PR, and HER2 / neu, the effects of the treatments known so far are not satisfactory. Therefore, there is a need for improved treatment of ERα-negative breast cancer, especially TNBC.

[0038] 4-OHBA acts correspondingly on the same principle. Esters of these compounds can also be used since such esters are metabolized to the respective compounds 4-OHA, 4-OHT and 4-OHBA, respectively.

[0039] Thus, the present invention provides a compound of formula I for use as a prodrug in a method of treating a patient suffering from an estrogen receptor β (ERβ)-positive and / or androgen receptor (AR)-positive breast tumor, or suffering from an ERβ-positive and AR-negative breast cancer. [ka] The present invention relates to the steroid compound 4-hydroxyandrost-4-ene-3,17-dione (4-OHA) or an ester thereof, wherein said prodrug is applied topically to a patient. Preferably, the treatment is performed before the planned surgical removal of breast tumor tissue (pre-operative treatment).

[0040] As further found in the present invention, 4-OHT can be used in place of the prodrug 4-OHA in the methods of treatment disclosed herein. Similarly, 4-OHBA and esters of 4-OHA, 4-OHT and 4-OHBA can also be used.

[0041] The prodrug is a pharma- ceutical active compound(s) having hydroxyl groups at at least the C4 and C17 positions, e.g., 4-hydroxytestosterone: [ka] (4-OHT), and [ka] It is thought to be metabolized to (4-OHBA).

[0042] By treating patients with breast cancer that is ERβ positive and preferably (but not necessarily) AR positive with said prodrug or with any one selected from 4-OHT, 4-OHBA and 4-OHA, 4-OHT and 4-OHBA esters (especially 4-OHT or its esters), tumor tissue is significantly reduced at the end of treatment.As a result, compared with the size of tumor tissue at the start of treatment, tumor tissue is reduced to the extent that subsequent surgery can be performed by breast-conserving method.In other words, complete removal of affected breast can be avoided.

[0043] Therefore, in one embodiment, pre-operative treatment involves pre-operative reduction of breast tumor tissue.

[0044] If not known from other information or data, the receptor status of the target cells or tissues with respect to AR and / or ERβ (and also ERα) can be determined and, if desired, quantified by standard methods known to those skilled in the art, including immunoassays involving antibodies specific for AR and / or ER, DNA and / or RNA hybridization assays involving nucleic acid probes specific for AR and / or ER, or PCR amplification tests.

[0045] In a preferred embodiment, the tumor tissue of the affected breast is reduced by at least 13% by volume, at least 14% by volume, at least 15% by volume, or at least 20% by volume, preferably at least 30% by volume, more preferably at least 40% by volume, even more preferably at least 50% by volume, at least 60% by volume, or at least 70% by volume, compared to the size of the breast tumor tissue at the beginning of treatment. The size of the tumor tissue can be determined by any suitable means known to those skilled in the art, such as ultrasound, X-ray, or mammography. In one embodiment, the reduction in tumor volume shown above can be reached after 40 days of treatment.

[0046] In a further aspect, the metabolized steroid compound 4-OHA according to the invention exhibits increased binding affinity to estrogen receptor beta (ERβ) expressed by breast tumor tissue when comparing its binding affinity to estrogen receptor alpha (ERα) expressed by breast tumor tissue, respectively. In a preferred embodiment, the pharmacologically active agent exhibits a binding affinity to ERβ that is at least 3-fold, preferably at least 4, 5, or 6-fold, more preferably at least 9-fold, and most preferably at least 10-fold higher than its binding affinity to ERα.

[0047] As such, the metabolized steroid compounds of formula I of the present invention exhibit binding affinity to ERβ mediated by the C4-OH group, and preferably exhibit binding affinity to AR mediated by the C17-OH group.

[0048] In one embodiment, the AR-specific binding affinity is determined by IC 50 ≦500nM, e.g. IC 50 ≦100nM or IC 50 ≤ 50 nM, where IC 50 IC is defined as the concentration of a compound required to reduce the specific binding of the reference compound, 5α-dihydrotestosterone (DHT), by 50%. 50 Values ​​were determined by known methods using radiolabeled DHT as a reference compound, e.g., a reference concentration of 1 nM radiolabeled [ 3 H]-DHT by standard dextran-coated charcoal adsorption as described by Raynaud et al., J. Steroid Biochem. 6, 615-622 (1975) or similar IC methods described in the literature. 50 The differences in binding affinity to the order contemplated in the present invention are important because the concentration of AR in target cells is very low, typically in the nanomolar range.

[0049] The steroid compounds of the present invention, when metabolized to a pharmacoactive substance, may additionally exhibit androgen receptor (AR) binding affinity, which in AR-positive breast cancer cells leads to increased expression of ERβ, with concomitant inhibition or at least reduction, preferably significant reduction, of cell proliferation.

[0050] Moreover, preferably, the compounds of the present invention are not substrates for 5α-reductase, and since these substances cannot be easily metabolized like, for example, testosterone, their inhibitory effect on tumor cells is greatly enhanced, and in addition, they exert an apoptotic effect on the tumor cells themselves.

[0051] The compounds of the present invention are AR-specific compounds but are not metabolized by aromatase, and therefore can fully exert the action and effect associated with the specifically defined therapeutic use via AR.However, as found in the present invention, the expression (existence) of AR on tumor tissue is not a necessary prerequisite for the prodrug and its metabolites, such as 4-OHBA or 4-OHT, to exert the positive effects disclosed herein.Rather, as can be shown in the present invention, the prodrug and its metabolites achieve their beneficial effects mainly through the activation of ERβ expressed on tumor tissue.Therefore, it is not essential that tumor tissue expresses AR in addition to ERβ.

[0052] In a preferred embodiment, the pharmacologically active agent is selected from the group consisting of 4-hydroxy-5α-androstane-3β,17β-diol (4-OHBA) and 4-hydroxytestosterone (4-OHT).

[0053] In the experiments carried out with the compounds of the present invention, it has been possible to show that the steroid compound (4-OHA) according to formula I or its esters have excellent skin penetration capabilities, so that sufficient tumor inhibition or size reduction, respectively, can be achieved by simple topical administration, at the site of the patient in need of treatment, of a therapeutically or prophylactically, preferably therapeutically effective amount of this steroid compound according to the present invention, for example an ointment, lotion or cream. After topical administration, the compound(s) penetrate through the skin and become concentrated in the adipose tissue.

[0054] Therefore, in a further aspect, the present invention provides a compound of formula I as a prodrug for use in the treatment of patients suffering from estrogen receptor β (ERβ) positive and optionally androgen receptor (AR) positive breast tumors. [ka] and a steroid compound 4-hydroxyandrost-4-ene-3,17-dione (4-OHA) or an ester thereof, and a pharma- ceutically acceptable carrier(s) and / or excipient(s), or comprising a compound selected from the group consisting of 4-OHT, 4-OHBA and esters of 4-OHT and 4-OHBA, and a pharma- ceutically acceptable carrier(s) and / or excipient(s), wherein the treatment is performed prior to the planned surgical resection of breast tumor tissue.

[0055] In a preferred embodiment, the pre-operative treatment involves topical application of a pharmaceutical composition in an amount effective to reduce breast tumor tissue, which amount can be tailored to the patient individually depending on the size of the tumor at the start of treatment.

[0056] In a further embodiment, the prodrug 4-OHA or an ester thereof, or a compound selected from the group consisting of 4-OHT, 4-OHBA and esters of 4-OHT and 4-OHBA, especially 4-OHT, is present in the pharmaceutical composition in an amount of about 1.5% to about 4% by weight, preferably about 2.0% to about 3.0% by weight, more preferably about 2.0% or about 2.5% by weight, based on the amount of the pharmaceutical composition.

[0057] As disclosed herein, topical administration of prodrug 4-OHA to a patient in need thereof can result in tumor size reduction, so as to achieve breast cancer tissue can be removed while conserving the breast, i.e., avoid complete removal of the affected breast.Therefore, this preoperative treatment is carried out for a sufficient period of time to achieve reduction of breast tumor tissue to a size that allows breast-conserving surgical removal of breast tumor tissue.

[0058] In preferred embodiments, pre-operative treatment is administered for a period of up to 6 months, or for a period of up to 5, 4, 3, or 2 months.

[0059] It is also possible that reduction of breast tumor tissue to a size that allows for breast-conserving surgical resection can be achieved after 11 weeks, or 10, 9, 8, 7, 6, 5, 4, 3, or 2 weeks of preoperative treatment.

[0060] In further embodiments, pre-operative treatment is commenced up to 6 months, or up to 5, 4, 3, or 2 months prior to the planned surgery, ie, breast-conserving surgical resection of breast tumor tissue.

[0061] Preoperative treatment can also begin up to 11 weeks, or up to 10, 9, 8, 7, 6, 5, 4, 3, or 2 weeks prior to the scheduled surgery.

[0062] Preoperative treatment may also begin up to 8 weeks prior to the scheduled surgery, preferably 6 or 5 weeks prior to surgery.

[0063] By topically applying the prodrug (or the pharmaceutical composition containing the prodrug, respectively) or its metabolites, such as 4-OHT and / or 4-OHBA or their esters, to the skin, the prodrug or metabolites are absorbed transdermally and are therefore brought locally to the intended site of action. The pharmacologically active substances are concentrated in the periductal adipose tissue. In long-term treatment, the adipose tissue of the treated breast is significantly reduced. This reduction reduces the amount of estrogen-producing cells with estrogen-producing capacity. The lipophilic and hydrophobic nature of the pharmacologically active substances has the consequence that the pharmacologically active substances are concentrated locally exclusively in adipose tissue and therefore cannot exert any significant systemic effect.

[0064] In the present invention, the pharmaceutical composition or medicinal product, respectively, is applied topically on or near the intended site of action. According to the present invention, the result achieved is an adequate local pharmacologically active substance level in the affected tissue (under treatment) without significant absorption of the pharmacologically active substance occurring in the blood circulation. Therefore, the advantage of this embodiment does not only rely on the topical application itself, but also on the local topical application in such a way that the active compound is immediately concentrated in the tissue at risk and / or in the affected tissue, and not indirectly via the blood circulation.

[0065] For external administration, the pharmaceutical composition or medicament is preferably formulated as an ointment, cream, gel, emulsion or lotion. Formulation as a powder or oil is also conceivable. The base of the formulation is well known to those skilled in the art of cosmetics and pharmaceuticals and does not need to be described in more detail here. For example, vegetable oils and fats, such as almond oil, peanut oil, olive oil, peach kernel oil, castor oil, plant extracts, ethereal oils, etc.; furthermore vegetable waxes and synthetic and animal oils, fats or waxes; lecithin, lanolin alcohol, carotene, fragrances, monohydric or polyhydric alcohols, urea, preservatives and colorants, etc. can be used. Formulation as an oil-in-water or water-in-oil emulsion is preferred.

[0066] The content of the prodrug of the invention in suitable pharmaceutical preparations can be between 0.0001 and 20% by weight, preferably between 0.6% and 10% by weight, more preferably between 1 and 5% by weight of a compound according to the invention. The customary range is 0.6 to 5% by weight.

[0067] When substances are incorporated to enhance skin penetration, the content can be, for example, between 0.01 and 1% by weight, preferably between 0.05 and 0.2% by weight when using hyaluronidase, and between 1 and 25% by weight, preferably between 5 and 10% by weight when using dimethylsorbitol or DMSO.

[0068] Due to the primary target disease of the above treatments, local topical treatments are primarily performed on the skin of female patients, particularly the breasts, although the same therapeutic principles apply to male breast cancer.

[0069] In a further embodiment, the present invention also relates to the use of 4-OHA or an ester thereof as a prodrug in the pre-surgical topical treatment of ERβ positive breast cancer tissue.

[0070] The present invention also relates to the use of a compound selected from the group of 4-OHT, 4-OHBA and esters of 4-OHT and 4-OHBA, especially 4-OHT, in the preoperative external treatment of ERβ-positive and optionally further AR-positive breast cancer tissue.Surprisingly, it has been found in the present invention that the prodrug 4-OHA or its metabolite 4-OHT also shows its beneficial effect when used in the treatment of breast cancer tissue that expresses ERβ but does not express AR.Therefore, in a preferred embodiment, the prodrug 4-OHA or its ester, or its metabolite 4-OHT is used in the preoperative treatment of patients suffering from ERβ-positive and AR-negative breast cancer.4-OHBA is also considered to have the same principle.Esters of these compounds can also be used correspondingly, since such esters are metabolized to the respective compounds 4-OHA, 4-OHT and 4-OHBA, respectively.

[0071] Finally, the present invention also relates to a pharmaceutical composition for the use described herein, wherein the ERβ positive breast cancer is triple negative breast cancer (TNBC), and the pharmaceutical composition further comprises, in addition to the prodrug 4-OHA or an ester thereof, and / or in addition to a compound selected from 4-OHT, 4-OHBA and esters of 4-OHT and 4-OHBA, a further active ingredient different from the prodrug 4-OHA and different from a compound selected from 4-OHT, 4-OHBA and their esters, which binds to and thereby activates ERβ.

[0072] In a preferred embodiment, the further active ingredient is an anti-estrogen drug, and / or the further active ingredient is an anti-cancer indole compound.In a further preferred embodiment, the anti-estrogen drug is, for example, fulvestrant or tamoxifen.The anti-cancer indole compound is preferably an indole derivative that changes estrogen metabolism or is effective in stopping the growth of cancer cells.An example of an anti-cancer indole compound is indole-3-carbinol.

[0073] The following examples are illustrative of the present invention but are not intended to limit it. EXAMPLES

[0074] 1. Observations of breast tumor tissue shrinkage 1.1 A patient diagnosed (by mammography) with an ERβ-positive breast cancer tumor measuring 2 cm in maximum size was treated with a pharmaceutical composition comprising a prodrug of the invention: the prodrug was applied topically to the affected breast. Already after 3 weeks of treatment a reduction in the tumor size could be observed. 1.2 Application of 4 ml of cream containing 2.5% by weight of 4-OHA to breasts with ER-positive breast cancer (2 times per day). Already after 3-5 weeks, the size of the tumor was significantly reduced.

[0075] 2. Comparison of treatment success rates between anastrozole and letrozole and the prodrugs of the present invention The study was conducted on patients with ERβ-positive breast cancer. Initially, the patient was treated with the nonsteroidal aromatase inhibitors anastrozole or letrozole according to the respective current treatment guidelines, however, each treatment was unsuccessful. The patient was then treated with a pharmaceutical composition containing a prodrug according to the invention, ie, the prodrug (4-OHA) was applied topically to the affected breast. It was observed that stable disease for more than 6 months could be achieved in 50% of patients treated in this manner.

[0076] 3. Growth inhibition of MDA MB-231 cells upon incubation with 4-OHT The MDA-MB-231 cell line is an aggressive, highly invasive, and poorly differentiated epithelial human breast cancer cell line. It lacks expression of estrogen receptor alpha (ERα) and progesterone receptor (PR), as well as HER2 (human epidermal growth factor 2) amplification. The MDA-MB-231 cell line is ERβ positive. It is a model cell line for triple-negative breast cancer (TNBC), which is ERβ positive. Two different concentrations of MDA-MB-231 cells (1.0e3 and 2.5e3) were incubated with increasing concentrations of 4-OHT (K+; 0.5 μM; 1 μM; 5 μM; 10 μM; 20 μM). The cells were incubated for 6 days (d). The absorbance at 492 nm was then measured. The diagram in Figure 2 shows that significant inhibition of cell growth occurs starting from a final concentration of at least 5 μM / L 4-OHT. At 5 μM / L 4-OHT the inhibition of cell growth is approximately 50% (cell concentration 1.0e3), and at a 4-OHT concentration of 20 μM / L the inhibition of cell growth increases to approximately 75% to 80%.

[0077] 4. Observation of shrinkage of triple-negative breast tumor tissue (TNBC) Tumors from 16 breast cancer patients with ERα-positive and ERβ-positive breast cancer ("ER+") and 6 breast cancer patients with ERβ-positive and TNBC ("ER-": AR-negative; HER2neu-negative; ERα-negative) were treated with a cream containing 4-OHA as the active ingredient in a typical inactive cream base ("SAVE Cream"). Tumors were treated with 4-OHA for approximately 3 weeks (mean treatment duration: 21.2 days). The administered dose was 2 x 2 or 2 x 4 g of 2.0% by weight 4-OHT in cream base ("SAVE Cream") per breast per day. The mean tumor volume was assessed before and after treatment with the 4-OHT-containing cream. As can be seen, the tumor volume (mm 3) was reduced by 75%, and in ER- patients, the tumor volume was reduced by 14%. This is surprising and noteworthy, especially since TNBC is known to be a type of breast cancer that can be very aggressive. Since it does not express ERα, PR, and HER2 / neu, the effects of treatments known so far are not satisfactory. However, as can be seen, treatment with a cream containing 4-OHT was able to reach a reduction in tumor volume of 14%.

Claims

1. A compound of formula I as a prodrug for use in a method for treating a patient suffering from an estrogen receptor beta (ERβ) positive breast tumor. 【Chemistry 1】 An external preparation comprising the steroid compound 4-hydroxyandrost-4-ene-3,17-dione (4-OHA) or an ester thereof.

2. The ERβ-positive breast tumor is also androgen receptor (AR) positive, or The method according to claim 1, wherein the ERβ-positive breast tumor is an AR-negative breast tumor.

3. The agent described in claim 1 or 2, wherein the ERβ-positive breast tumor is triple-negative breast cancer in that it is estrogen receptor alpha (ERα) negative, progesterone receptor (PR) negative, and HER2 receptor negative.

4. The method according to any one of claims 1 to 3, wherein the treatment is performed prior to planned surgical resection of breast tumor tissue and / or the treatment is for reducing breast tumor tissue.

5. The agent according to any one of claims 1 to 4, wherein the prodrug 4-OHA is metabolized to a pharmacologically active substance having hydroxyl groups at least at the C4 and C17 positions.

6. 6. The agent according to any one of claims 1 to 5, wherein the prodrug 4-OHA is metabolized into a pharmacologically active substance that has a higher binding affinity for estrogen receptor beta (ERβ) compared to its binding affinity for estrogen receptor alpha (ERα), and either both or only ERβ is expressed by breast tumor tissue, and wherein the treatment results in agonistic binding of ERβ.

7. The agent of claim 6, wherein the pharmacologically active substance exhibits a binding affinity for ERβ that is at least three times higher than its binding affinity for ERα.

8. The agent according to any one of claims 5 to 7, wherein the pharmacologically active substance is 4-hydroxy-5α-androstane-3β,17β-diol (4-OHBA) or 4-hydroxytestosterone (4-OHT).

9. A topical preparation comprising 4-hydroxytestosterone (4-OHT) or 4-hydroxy-5α-androstane-3β,17β-diol (4-OHBA), or a hydroxyester of 4-OHT or 4-OHBA, respectively, for use in a method of treating a patient suffering from an estrogen receptor β (ERβ) positive breast tumor.

10. An external preparation comprising 4-OHT for use in a method of treating a patient suffering from an estrogen receptor beta (ERβ) positive breast tumor, wherein the patient is suffering from triple negative breast cancer in that it is estrogen receptor alpha (ERα) negative, progesterone receptor (PR) negative, and HER2 receptor negative.

11. A compound of formula I as a prodrug for use in a method for treating a patient suffering from an estrogen receptor beta (ERβ) positive breast tumor. 【Chemistry 2】 A pharmaceutical composition for topical application comprising the steroid compound 4-hydroxyandrost-4-ene-3,17-dione (4-OHA) of the formula (I) or an ester thereof, and a pharma- ceutical acceptable carrier(s) and / or excipient(s).

12. The pharmaceutical composition of claim 11, wherein the treatment is a pre-operative treatment performed prior to planned surgical resection of breast tumor tissue.

13. The ERβ-positive breast tumor is also androgen receptor (AR) positive, or The pharmaceutical composition according to claim 11 or 12, wherein the ERβ-positive breast tumor is an AR-negative breast tumor.

14. The pharmaceutical composition described in any of claims 11 to 13, wherein the ERβ-positive breast tumor is a triple-negative breast cancer in that it is estrogen receptor alpha (ERα) negative, progesterone receptor (PR) negative, and HER2 receptor negative.

15. The pharmaceutical composition of any of claims 11 to 14, wherein the treatment is carried out for a period of time sufficient to achieve a reduction of the breast tumor tissue to a size that allows for breast-conserving surgical resection of the breast tumor tissue.

16. The pharmaceutical composition according to any one of claims 11 to 15, wherein the size of the tumor tissue as determined by mammography, X-ray or ultrasound is significantly reduced at the completion of treatment compared to the size of the breast tumor tissue at the start of treatment.

17. The pharmaceutical composition of claim 11, wherein the size of the tumor tissue as determined by mammography, x-ray or ultrasound is reduced by at least 14% by volume, or at least 20% by volume, at the completion of treatment compared to the size of the breast tumor tissue at the start of treatment.

18. The pharmaceutical composition of any of claims 11 to 17, wherein the treatment is pre-operative treatment, and the pre-operative treatment is initiated up to 6 months, or up to 5, 4, 3, or 2 months prior to the planned surgical procedure, i.e., breast-conserving surgical resection of the breast tumor tissue.

19. 19. The pharmaceutical composition of any of claims 11 to 18, wherein the prodrug 4-OHA or ester thereof is present in the pharmaceutical composition in an amount of about 0.01% to about 4% by weight, based on the weight of the pharmaceutical composition.

20. The pharmaceutical composition according to any one of claims 11 to 19, wherein the pharmaceutical composition is applied topically once or twice daily.

21. A pharmaceutical composition according to any one of claims 11 to 20, further comprising, in addition to the prodrug 4-OHA or an ester thereof, another active ingredient which binds to ERβ and thereby activates ERβ.

22. The pharmaceutical composition described in claim 21, wherein the ERβ-positive breast cancer is triple-negative breast cancer (TNBC).

23. 23. The pharmaceutical composition according to claim 21 or 22, wherein the other active ingredient is selected from antiestrogens and anticancer indole compounds.

24. The pharmaceutical composition described in claim 23, wherein the anti-estrogen drug is fulvestrant or tamoxifen.

25. The pharmaceutical composition of claim 23, wherein the anticancer indole compound is indole-3-carbinol.

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