RU2543339C2 - Oestrogen receptor ligands and methods of using them - Google Patents

Abstract

FIELD: medicine, pharmaceutics.

SUBSTANCE: invention refers to medicine, namely urology, and can be used for treating advanced prostate cancer in male individuals. That is ensured by administering a therapeutically effective amount of a compound of formula IV or its isomer, a pharmaceutically acceptable salt, a pharmaceutical product, a polymorph, a hydrate or any combination thereof

.

EFFECT: method provides treating effectively, inhibiting, reducing an incidence, relieving a severity or inhibiting advanced prostate cancer, as well as conducting the palliative treatment of advanced prostate cancer.

12 cl, 21 ex, 20 tbl, 23 dwg

Description

FIELD OF THE INVENTION

The present invention relates to methods for reducing testosterone levels by decreasing luteinizing hormone (LH) levels or regardless of LH levels in a male subject, and to methods for treating, suppressing, reducing the incidence, alleviating, or inhibiting the severity of advanced prostate cancer and palliative treatment of advanced prostate cancer glands.

BACKGROUND OF THE INVENTION

Estrogens belong to the group of endogenous and synthetic hormones that are important and are used to maintain tissues and bones. Estrogens are endocrine regulators of cellular processes involved in the development and maintenance of the reproductive system. The role of estrogen in reproductive biology, the prevention of postmenopausal hot flashes and the prevention of postmenopausal osteoporosis is widely recognized. The most important endogenous human estrogen is estradiol, and it is found in both women and men.

Two different subtypes of intracellular receptors, estrogen receptor alpha (ERα) and estrogen beta receptor (ERβ), are involved in the biological effects of estrogens and antiestrogens. Typically, endogenous estrogens are potent activators of both receptor subtypes. For example, estradiol acts as an ERα agonist in many tissues, including breast, bone, cardiovascular and central nervous systems. Typically, selective estrogen receptor modulators (SERMs) in different tissues act differently. For example, SERM may be an ERα antagonist in the mammary gland, but may be a partial ERα agonist in the uterus, bone and cardiovascular systems. Accordingly, compounds that act as estrogen receptor ligands are useful in treating a number of conditions and disorders.

Prostate cancer is one of the most commonly diagnosed types of non-skin cancer in men in the United States and the second most common cause of cancer death, and more than 180,000 new cases and nearly 29,000 deaths are expected this year. Patients diagnosed with advanced prostate cancer are subjected to antiandrogen therapy (ADT), usually either through luteinizing hormone releasing hormone releasing factor (LHRH) agonists, or through bilateral orchiectomy. As a result of antiandrogen therapy, not only testosterone levels decrease, but estrogen levels also decrease, since estrogen is obtained as a result of aromatization of testosterone, the levels of which are depleted as a result of ADT. Estrogen deficiency induced by antiandrogen therapy causes significant side effects that include hot flashes, gynecomastia and mastalgia, loss of bone mass, deterioration in bone quality and strength, osteoporosis and life-threatening fractures, adverse lipid changes and more frequent cases of cardiovascular disease and myocardial infarction, depression and other mood changes. It is believed that many side effects of ADT due to estrogen deficiency are mediated by ERα.

Leuprolide acetate (Lupron®) is a synthetic nonapeptide analogue of the natural gonadotropin releasing hormone (GnRH or LH-RH). Leuprolide acetate is an LH-RH superaggonator that ultimately inhibits LH secretion that occurs in the pituitary gland. Leuprolide acetate acts as a potent inhibitor of gonadotropin secretion, causing a suppression of ovarian and testicular steroidogenesis. The administration of leuprolide acetate to humans initially causes an increase in the levels of luteinizing hormone (LH) and follicle-stimulating hormone (FSH) in the bloodstream, leading to a temporary increase in the levels of gonadal steroids (testosterone and dihydrotestosterone in men and estrone and estradiol in women in the premenopausal period). However, prolonged administration of leuprolide acetate results in decreased levels of LH and FSH. In men, testosterone levels are reduced to levels corresponding to medical castration (below 50 ng / dl). In women in the premenopausal period, the estrogen content decreases to postmenopausal levels. Testosterone is a known stimulus for prostate cancer cells. Thus, inhibition of testosterone secretion or inhibition of the effects of testosterone is an essential component of prostate cancer therapy. Leiprolide acetate in the treatment of prostate cancer can be used to suppress LH, which is reflected in a decrease and decrease in serum testosterone levels to levels corresponding to medical castration.

Before administering LHRH agonists, testosterone levels corresponding to medical castration were achieved, increasing estrogenic activity in the pituitary gland through estrogens, primarily diethylstilbestrol (DES). Equally, DES was effective as an LHRH agonist in inhibiting testosterone to levels consistent with medical castration. Patients treated with DES did not have hot flashes or bone loss, but gynecomastia was observed to a greater extent than with ADT using LHRH agonists. Unfortunately, the use of highly effective pure estrogens like DES and estradiol is very often associated with a high risk of serious cardiovascular and thromboembolic complications that limit their clinical use.

The compounds of this invention reduce testosterone levels to levels consistent with medical castration, which may be useful for treating prostate cancer, while at the same time preventing an increased risk of thrombotic events and not causing loss of bone mass, hot flashes and / or gynecomastia.

SUMMARY OF THE INVENTION

In one embodiment, the invention provides a method for lowering total serum testosterone levels in a male subject, comprising administering a therapeutically effective amount of a compound of formula I-XII, as set forth herein below.

In one embodiment, the invention provides a method for lowering total serum testosterone levels in a male subject, comprising administering a therapeutically effective amount of a compound of formula I-XII as set forth herein below, such a decrease in total serum testosterone by reducing serum luteinizing hormone levels.

In one embodiment, the invention provides a method for lowering total serum testosterone levels in a male subject, comprising administering a therapeutically effective amount of a compound of formula I-XII as set forth herein below, wherein such a decrease in total serum testosterone is independent of the decrease serum luteinizing hormone levels

In one embodiment, the invention provides a method of lowering total serum testosterone levels in a male subject, comprising administering a therapeutically effective amount of a compound of formula I-XII as set forth herein below, wherein said administration of said compounds of formula I-XII prevents or leads to treatment of side effects associated with antiandrogen therapy (ADT), wherein said subject has prostate cancer.

In one embodiment, the invention provides a method of antiandrogenic therapy for a subject, comprising administering a therapeutically effective amount of a compound of formula I-XII, as set forth herein below. In another embodiment, said subject has prostate cancer.

In one embodiment, the invention provides a method of treating, suppressing, reducing the incidence, alleviating the severity or inhibiting advanced prostate cancer, comprising administering a therapeutically effective amount of a compound of formula I-XII, as set forth herein below.

In one embodiment, the invention provides a method for palliatively treating advanced prostate cancer, comprising administering a therapeutically effective amount of a compound of formula I-XII, as set forth herein below.

BRIEF DESCRIPTION OF GRAPHIC MATERIALS

The object recognized by the invention is specifically indicated and is stated clearly in the final part of this description. However, this invention, both in terms of organization and method of action, together with its objectives, features and advantages, can best be understood with reference to the following detailed description when considered in conjunction with the accompanying graphic materials, in which:

figure 1 shows the levels of testosterone (continuous line) and androgen in general (dashed line) in the blood serum of intact male macaques after daily oral (p / o) administration of Compound IV at a dose of 30 mg / kg (first dose in 0 day) (see Example 8);

figure 2 presents the levels of testosterone in intact rats treated with Compound IV (0.3; 1; 10; 30 mg / kg). ^I means P <0.05 compared with intact controls treated with diluent. BLOQ values (below the quantification limit) are presented graphically with a quantification limit of 0.08 ng / ml (see Example 9);

figure 3 presents the inhibitory effect of Compound IV on the activity of the enzyme 17β-HSD5 (see Example 12);

4 shows in vitro aggregation of human platelets in the presence of DES, 17β-estradiol (E2) and Compound IV. Platelet rich plasma (PRP) was incubated with diluent, E2, DES or Compound IV for 30 seconds, after which aggregation was induced using 0.3 units of thrombin. Aggregation monitoring was carried out for 5 minutes and expressed as a percentage of the diluent control (see Example 13);

Figure 5. A typical synthesis scheme for the preparation of Compounds II-XII (see Example 1);

6. The synthesis scheme for obtaining Compound IV (see Example 2);

7. A synthesis scheme for preparing Compound VI (see Example 3);

Fig. 8. The synthesis scheme for obtaining Compounds IX and X (see Example 5);

figure 9 presents the testosterone levels in intact rats treated with Compound IV at the doses of 3 mg / kg, 10 mg / kg and 300 mg / kg, after 24 hours, 72 hours and 168 hours (see Example 9);

10 shows LH levels (FIG. 10A), FSH levels (FIG. 10 B), testosterone levels (FIG. 10C), prostate mass levels (FIG. 10D), seminal vesicle mass levels (FIG. 10E) and levator mass (Fig. 10F) in intact and orchiectomized (ORX) rats treated with Compound IV at dosages of 0.3 mg / kg, 1 mg / kg, 3 mg / kg, 10 mg / kg and 30 mg / kg. ^I means P <0.05 compared with intact controls treated with diluent. ^O means P <0.05 compared with ORX controls treated with diluent. BLOQ values are presented graphically with a quantification limit of 0.08 ng / ml (see Example 9);

11 shows the size of the prostate in intact and ORX-treated rats after administration of Compound IV (Fig. 11A) and DES (Fig. 11B) in various dosages (see Example 15);

12 shows the differences between DES and Compound IV; DES gives a cross-reaction with the glucocorticoid receptor (GR), while Compound IV is not (Figa); DES cross-reacts with androgen receptor (AR). It weakly stimulates and weakly inhibits the action of AR (i.e., is a partial agonist / antagonist), while Compound IV is not (Fig. 12B); DES eliminates the transactivation of estrogen receptor related receptors (ERRs), while Compound IV is not (Fig. 12C) (see Example 15);

13 shows the effect of Compound IV at dosages of 5 mg / kg, 10 mg / kg, 15 mg / kg and 30 mg / kg on the weakening of hot flashes in the morphine withdrawal model. N = 7 animals per group. 17β-E2 was used at a dose of 5 mg / kg in 100% DMSO (dimethyl sulfoxide) (see Example 14);

on Fig presents a dose-dependent decrease in body weight (kg) of macaques (~ 20% at a dose of 100 mg / kg) after administration of Compound IV for 91 days. No signs of gynecomastia or hyperestrogenia were observed (see Example 16);

on Fig presents a dose-dependent decrease in serum testosterone (ng / ml) in macaques after daily oral administration of Compound IV in comparison with the positive control (LHRH agonist). The dashed line indicates the level of testosterone in chemically neutered patients, and the thick dashed line indicates the level of testosterone in surgically neutered macaques (see Example 16);

on Fig presents the dependence of the level of antigen of the prostate gland (PSA) (ng / ml) on the dose in macaques after administration of Compound IV at the initial time and on the 28th day. Treatment with Compound IV significantly reduced PSA levels (see Example 16);

on Fig presents the dependence of the volume of the prostate, measured using transrectal ultrasonography (TRUS), on the dose in macaques in comparison with the positive control (LHRH agonist) after administration of Compound IV, for the 6th week (see Example 16);

on Fig presents the dependence of the mass of organs (prostate, seminal vesicles and testes) on the dose in macaques after administration of Compound IV as a percentage of control on the 90th day (Figa). Macaque prostate mass at autopsy on the 13th day after daily oral administration of Compound IV (Fig. 18B) (see Example 16);

on Fig presents the dependence of the average levels of total testosterone (nmol / l) on the dose in people between 1 and 11 days after the administration of Compound IV (100 mg, 300 mg, 600 mg and 1000 mg) (see Example 17);

on Fig presents the dependence of the average levels of LH (ME (international units) / l) on the dose in humans between 1 and 10 days after the administration of Compound IV (100 mg, 300 mg, 600 mg and 1000 mg) (see Example 17);

on Fig presents the dependence of the average levels of free testosterone (pg / ml) on the dose in people between 1 and 10 days after the administration of Compound IV (100 mg, 300 mg, 600 mg and 1000 mg) (see Example 17);

on Fig presents the dependence of the average levels of PSA (μg / l) on the dose in humans between between 1 and 10 days after administration of Compound IV (100 mg, 300 mg, 600 mg and 1000 mg) (see Example 17);

on Fig presents the dependence of testosterone levels (ng / ml) on the dose in intact rats after 14 days of recovery after administration of Compound IV. ^I means P <0.05 compared with intact controls (see Example 10).

It should be borne in mind that for simplicity and clarity of illustration, the elements shown in the graphic materials do not have to be presented to scale. For example, the dimensions of some elements may be exaggerated compared to other elements for clarity. In addition, where appropriate, item numbers to indicate corresponding or similar elements may be repeated among the graphic materials.

DETAILED DESCRIPTION OF THE PRESENT INVENTION

In the following detailed description, numerous specific details are set forth in order to provide a thorough understanding of the invention. However, it will be understood by those skilled in the art that the present invention may be practiced without these characteristic details. In other instances, well-known methods, techniques, and components are not described in detail so as not to obscure the present invention.

In one embodiment, the compounds described herein and / or compositions containing these compounds can be used to lower total serum testosterone levels in a male subject.

In one embodiment, the compounds described herein and / or compositions containing these compounds can be used to lower serum total testosterone levels in a male subject, and such a decrease in serum total testosterone occurs by decreasing luteinizing levels hormone (LH) in serum.

In one embodiment, the compounds described herein and / or compositions containing these compounds can be used to lower total serum testosterone levels in a male subject, and such a decrease in serum total testosterone is independent of the decrease in levels serum luteinizing hormone.

In one embodiment, the invention provides a method of lowering total serum testosterone levels in a male subject, comprising administering a therapeutically effective amount of the compound or its isomer, pharmaceutically acceptable salt, pharmaceutical product, polymorph, hydrate, or any combination thereof represented by the structure of formula I :

Where

Y represents C (O) or CH ₂ ;

R ₁ , R ₂ independently represent hydrogen, halogen, hydroxyl, alkoxy, cyano, nitro, CF ₃ , N (R) ₂ , sulfonamide, SO ₂ R, alkyl, haloalkyl, aryl, O-Alk-NRaRe or O-Alk a heterocycle in which the heterocycle is a 3-7 membered substituted or unsubstituted heterocyclic ring, possibly aromatic;

R ₃ , R ₄ independently represent hydrogen, halogen, hydroxyalkyl, hydroxyl, alkoxy, cyano, nitro, CF ₃ , NHCOR, N (R) ₂ , sulfonamide, SO ₂ R, alkyl, haloalkyl, aryl or protected hydroxyl;

R represents alkyl, hydrogen, haloalkyl, dihaloalkyl, trihaloalkyl, CH ₂ F, CHF ₂ , CF ₃ , CF ₂ CF ₃ , aryl, phenyl, halogen, alkenyl, CN, NO ₂ or OH;

R ₅ and R ₆ independently represent hydrogen, phenyl, an alkyl group of 1-6 carbon atoms, a 3-7 membered cycloalkyl, a 3-7 membered heterocycle, a 5-7 membered aryl; or R ₅ and R ₆ form a 3-7 membered ring together with a nitrogen atom;

j and k are independently 1-4;

Alk is a linear alkyl of 1-7 carbon atoms, branched alkyl of 1-7 carbon atoms or cyclic alkyl of 3-8 carbon atoms.

In further embodiments of the methods described herein, a compound of formula I is represented by formula IA:

where R ₁ , R ₂ , R ₃ , R ₄ , j and k are as defined for formula I.

In one embodiment, the invention provides a method of lowering total serum testosterone levels in a male subject, comprising administering a therapeutically effective amount of a compound or its isomer, pharmaceutically acceptable salt, pharmaceutical product, polymorph, hydrate, or any combination thereof represented by a compound of formula II :

.

.

In one embodiment, the invention provides a method for lowering total serum testosterone levels in a male subject, comprising administering a therapeutically effective amount of a compound or its isomer, pharmaceutically acceptable salt, pharmaceutical product, polymorph, hydrate, or any combination thereof represented by a compound of formula III :

.

.

In one embodiment, the invention provides a method of lowering total serum testosterone levels in a male subject, comprising administering a therapeutically effective amount of a compound or its isomer, pharmaceutical product, pharmaceutically acceptable salt, polymorph, hydrate, or any combination thereof represented by a compound of formula IV :

.

.

In one embodiment, the invention provides a method of lowering total serum testosterone levels in a male subject, comprising administering a therapeutically effective amount of a compound or its isomer, pharmaceutically acceptable salt, pharmaceutical product, polymorph, hydrate, or any combination thereof represented by a compound of formula V :

.

.

In one embodiment, the invention provides a method of lowering total serum testosterone levels in a male subject, comprising administering a therapeutically effective amount of the compound or its isomer, pharmaceutically acceptable salt, pharmaceutical product, polymorph, hydrate, or any combination thereof represented by the compound of formula VI :

.

.

In one embodiment, the invention provides a method of lowering total serum testosterone levels in a male subject, comprising administering a therapeutically effective amount of a compound or its isomer, pharmaceutically acceptable salt, pharmaceutical product, polymorph, hydrate, or any combination thereof represented by a compound of formula VII :

.

.

In one embodiment, the invention provides a method of lowering total serum testosterone levels in a male subject, comprising administering a therapeutically effective amount of a compound or its isomer, pharmaceutically acceptable salt, pharmaceutical product, polymorph, hydrate, or any combination thereof represented by a compound of formula VIII :

.

.

In one embodiment, the invention provides a method for lowering total serum testosterone levels by decreasing luteinizing hormone (LH) levels in a male subject having prostate cancer, comprising administering a therapeutically effective amount of the compound or its isomer, pharmaceutically acceptable salt, pharmaceutical product , polymorph, hydrate, or any combination thereof, represented by a compound of formula IX:

.

.

In one embodiment, the invention provides a method of lowering total serum testosterone levels in a male subject, comprising administering a therapeutically effective amount of a compound or its isomer, pharmaceutically acceptable salt, pharmaceutical product, polymorph, hydrate, or any combination thereof represented by a compound of formula X :

.

.

In one embodiment, the invention provides a method of lowering total serum testosterone levels in a male subject, comprising administering a therapeutically effective amount of a compound or its isomer, pharmaceutically acceptable salt, pharmaceutical product, polymorph, hydrate, or any combination thereof represented by a compound of formula XI :

.

.

In one embodiment, the invention provides a method of lowering total serum testosterone levels in a male subject, comprising administering a therapeutically effective amount of a compound or its isomer, pharmaceutically acceptable salt, pharmaceutical product, polymorph, hydrate, or any combination thereof represented by a compound of formula XII :

In one embodiment, the invention provides a method for reducing total serum testosterone in a male subject, comprising administering a therapeutically effective amount of a compound of formula IA, I-XII or an isomer thereof, a pharmaceutically acceptable salt, pharmaceutical product, polymorph, hydrate, or any one thereof combinations. In another embodiment, the male subject has prostate cancer. In another embodiment, total serum testosterone is reduced to less than about 100 ng / dl. In another embodiment, total serum testosterone is reduced to less than about 50 ng / dl. In another embodiment, the concentration of total testosterone in serum is reduced to a level of less than about 25 ng / dl.

In one embodiment, the invention provides a method for reducing total serum testosterone in a male subject, comprising administering a therapeutically effective amount of a compound of formula IA, I-XII or an isomer thereof, a pharmaceutically acceptable salt, pharmaceutical product, polymorph, hydrate, or any one thereof combinations, and such a decrease in total serum testosterone occurs by decreasing serum luteinizing hormone (LH) levels. In another embodiment, the male subject has prostate cancer. In another embodiment, total serum testosterone is reduced to less than about 100 ng / dl. In another embodiment, total serum testosterone is reduced to less than about 50 ng / dl. In another embodiment, the concentration of total testosterone in serum is reduced to a level of less than about 25 ng / dl.

In one embodiment, the invention provides a method of lowering serum free testosterone levels in a male subject, comprising administering a therapeutically effective amount of a compound of formula IA, I-XII or an isomer thereof, a pharmaceutically acceptable salt, pharmaceutical product, polymorph, hydrate, or any one thereof combinations, and such a decrease in serum free testosterone occurs by decreasing serum luteinizing hormone (LH) levels. In another embodiment, the male subject has prostate cancer.

In one embodiment, the invention provides a method of lowering total serum testosterone levels in a male subject, comprising administering a therapeutically effective amount of a compound of formula IA, I-XII or an isomer thereof, a pharmaceutically acceptable salt, pharmaceutical product, polymorph, hydrate, or any one thereof combination, and such a decrease in total serum testosterone does not depend on a decrease in serum luteinizing hormone (LH) levels. In another embodiment, the male subject has prostate cancer. In another embodiment, total serum testosterone is reduced to less than about 100 ng / dl. In another embodiment, total serum testosterone is reduced to less than about 50 ng / dl. In another embodiment, the concentration of total testosterone in serum is reduced to a level of less than about 25 ng / dl.

In one embodiment, the invention provides a method of lowering serum free testosterone levels in a male subject, comprising administering a therapeutically effective amount of a compound of formula IA, I-XII or an isomer thereof, a pharmaceutically acceptable salt, pharmaceutical product, polymorph, hydrate, or any one thereof combinations, and such a decrease in serum free testosterone levels does not depend on a decrease in serum luteinizing hormone levels. In another embodiment, the male subject has prostate cancer.

In one embodiment, the invention provides methods for decreasing serum total testosterone levels or serum free testosterone levels in a male subject, said male subject having prostate cancer. In another embodiment, said subject has advanced prostate cancer.

In one embodiment, the decrease in serum testosterone concentrations is reversible and, after treatment with the compounds of the invention, a return to baseline levels is observed.

In another embodiment, serum testosterone concentrations are reversible after treatment with Compound IV, according to FIG. 23 and Example 10.

In one embodiment, the invention provides methods for reducing total serum testosterone levels in a male subject, comprising administering a therapeutically effective amount of a compound of formula IA, I-XII or an isomer thereof, a pharmaceutically acceptable salt, pharmaceutical product, polymorph, hydrate, or any one thereof combinations. In another embodiment, total serum testosterone is reduced to less than about 100 ng / dl. In another embodiment, total serum testosterone is reduced to less than about 50 ng / dl. In another embodiment, total serum testosterone is reduced to a level of less than about 25 ng / dl. In another embodiment, the total serum testosterone is reduced to less than about 75 ng / dl. In another embodiment, total serum testosterone is reduced to about a level in the range of 75 ng / dl to 100 ng / dl. In another embodiment, total serum testosterone is reduced to about a level in the range of 50 ng / dl to 75 ng / dl. In another embodiment, total serum testosterone is reduced to about a level in the range of 40 ng / dl to 50 ng / dl. In another embodiment, the concentration of total testosterone in serum is reduced to about a level in the range of 25 ng / dl to 50 ng / dl. In another embodiment, total serum testosterone is reduced to about a level in the range of 40 ng / dl to 60 ng / dl.

Serum testosterone levels can be measured as “free” (i.e., bioavailable and unbound) or as “total” testosterone (including the percentage of protein bound and unavailable testosterone). Men over 40 who do not have prostate cancer show low testosterone levels, corresponding to a total testosterone level of less than 250 ng / dl (<8.7 nmol / l) or a free testosterone level of less than 0.75 ng / dl (<0, 03 nmol / l).

In one embodiment, among the methods of the present invention, there is provided a method for lowering total and / or free testosterone levels in blood serum regardless of decreasing levels of luteinizing hormone (LH) or by decreasing LH levels in a male subject having prostate cancer. In another embodiment, changes in testosterone levels will be a decrease relative to the level before treatment. In another embodiment, the level of total serum testosterone is reduced to less than 100 ng / dl. In another embodiment, total serum testosterone is reduced to less than 50 ng / dl. In another embodiment, total serum testosterone is reduced to less than 25 ng / dl. In another embodiment, the level of free testosterone is reduced to less than 2 ng / dl. In another embodiment, the level of free testosterone is reduced to less than 1 ng / dl. In another embodiment, the level of free testosterone is reduced to less than 0.5 ng / dl. In another embodiment, the level of free testosterone is reduced to less than 0.25 ng / dl.

Methods for determining serum free testosterone levels and total serum testosterone levels include monitoring testosterone levels throughout the treatment period by conducting a blood test. Total testosterone is a combination of testosterone circulating in the bloodstream associated with carrier proteins (albumin, SHBG (sex hormone-binding globulin), transcortin, transferrin), and free / unbound hormone. Testosterone can be influenced by a variety of factors, including the level of protein in the blood that the hormone transports in the body, the age, obesity, and interference associated with commonly used test methods.

Methods suitable for measuring free testosterone (FT) can be complex (equilibrium dialysis and calculation of free testosterone (CFT)) or simple (a commercially available FT Coat-A-Count determination kit) using a similar labeled substance. In another embodiment, measurements of total testosterone and free testosterone in blood serum can be achieved by simultaneously measuring total testosterone and SHBG (for example, using the immunoradiometric assay test kit) from DPC (Diagnostic Products Corporation)) and then calculating free testosterone (CFT). In another embodiment, the measurement of total testosterone and free testosterone is within the competence of a person skilled in the art.

In one embodiment, the invention provides a method of reducing total serum testosterone or serum free testosterone in a male subject, comprising administering a therapeutically effective amount of a combination of one or more other forms of ADT and a compound of formula IA, I-XII or an isomer thereof, a pharmaceutically acceptable salt, pharmaceutical product, polymorph, hydrate, or any combination thereof. In another embodiment, a decrease in total or free testosterone in serum occurs by decreasing the level of luteinizing hormone (LH) in serum. In another embodiment, a decrease in serum total or free testosterone levels is independent of a decrease in serum luteinizing hormone levels.

The methods of this invention include administering a combination of other forms of ADT and a compound of the invention. In one embodiment, other forms of ADT include the use of an LHRH agonist. In another embodiment, the LHRH agonist is leuprolide acetate (Lupron® (US 5480656; US 5575987; 5631020; 5643607; 5716640; 5814342; 6036976; all of which are incorporated herein by reference) or goserelin acetate (Zoladex®) (US 7118552; 7220247; 7220247; 7500964; all of which are incorporated herein by reference.) In one embodiment, other forms of ADT include the use of an LHRH antagonist. In another embodiment, the LHRH antagonist is a degarelix. In one embodiment, other forms of ADT include the use of antiandrogens. In another embodiment, the antiandrogens are bicalutamide, flutamide, finasteride, dutasteride, MDV3100, nilutamide, chlormadinone, or any combination thereof.

In one embodiment, the methods of the invention comprise administering a therapeutically effective amount of an antiandrogen and a compound of the invention. In one embodiment, the methods of this invention comprise administering a therapeutically effective amount of an LHRH agonist and a compound of the invention. In one embodiment, the methods of this invention comprise administering a therapeutically effective amount of an antiandrogen, an LHRH agonist, and a compound of the invention.

In one embodiment, the invention provides a method of lowering total testosterone levels and / or serum free testosterone levels by lowering luteinizing hormone (LH) levels or independently of decreasing luteinizing hormone levels in a male subject having prostate cancer, in order to antiandrogen therapy (ADT), comprising administering a therapeutically effective amount of a compound of formula IA, I-XII. In another embodiment, this compound is Compound IV.

In another embodiment, the invention provides a method of antiandrogen therapy (ADT) of a subject, comprising administering a therapeutically effective amount of a compound of formula IA, I-XII or an isomer thereof, a pharmaceutically acceptable salt, pharmaceutical product, polymorph, hydrate, or any combination thereof. In another embodiment, said subject has prostate cancer. In another embodiment, this compound is Compound IV.

In another embodiment, ADT is used to treat prostate cancer, to slow the progression of prostate cancer, or to prevent and / or treat recurrence of prostate cancer.

In one embodiment, the invention provides a method for treating prostate cancer, slowing the progression of prostate cancer, preventing and / or treating recurrence of prostate cancer, comprising administering a compound of the invention. In another embodiment, the administration of a compound of the invention in combination with LHRH analogs, reversible antiandrogens (such as bicalutamide or flutamide), antiestrogens, anti-cancer drugs, 5-alpha reductase inhibitors, aromatase inhibitors, progestins, selective androgen receptor modulators (SARMs) or agents acting through other receptors for nuclear hormones.

In one embodiment, the present invention provides a method for treating prostate cancer and decreasing serum total testosterone and / or serum free testosterone levels by decreasing LH levels or independently of decreasing LH levels, comprising administering a compound of formula IA, I-XII. In another embodiment, comprising the administration of Compound IV.

As a result of antiandrogen therapy, not only testosterone levels decrease, but estrogen levels also decrease, since estrogen is obtained as a result of aromatization of testosterone. Estrogen deficiency induced by antiandrogen therapy causes significant side effects, including flushing, gynecomastia, mastalgia, bone loss, deterioration of bone quality and strength, osteoporosis, osteopenia and life-threatening fractures, adverse lipid changes and more frequent cases of cardiovascular disease and myocardial infarction, loss of libido, impotence, loss of muscle mass (sarcopenia), fatigue, cognitive dysfunction and depression and other mood changes.

In other embodiments, the present invention provides a method of treating any disease, disorder or symptom associated with ADT. In other embodiments, the present invention provides a method of treating any disease, disorder or symptom associated with loss of testosterone. Each disease, disorder or symptom represents a separate embodiment of the present invention.

In one embodiment, the invention provides a method of lowering total serum testosterone levels in a male subject, comprising administering a therapeutically effective amount of a compound of formula IA, I-XII or an isomer thereof, a pharmaceutically acceptable salt, pharmaceutical product, polymorph, hydrate, or any one thereof combinations, where the specified introduction of the specified compounds of formula IA, I-XII or its isomer, pharmaceutically acceptable salt, pharmaceutical product, polymorph, hydrate, or any combination thereof, duprezhdaet occurrence or leads to the prevention, suppression, reduction in the frequency of occurrence, inhibition or treatment of side effects related to the antiandrogenic therapy (ADT), wherein said subject has prostate cancer. In another embodiment, such a decrease in serum total or free testosterone levels is accomplished by decreasing LH levels or independently of decreasing LH levels.

In one embodiment, administration of the compounds of the invention results in the suppression, reduction in incidence, inhibition or treatment of typical side effects associated with conventional antiandrogen therapy (ADT). In another embodiment, the subject has prostate cancer. Such prevention and / or reduction of side effects occurs relative to a placebo or control group. In one embodiment, typical side effects associated with conventional antiandrogen therapy (ADT) include hot flashes, gynecomastia, decreased bone mineral density and increased tendency to fracture. In another embodiment, the administration of the compounds of the invention prevents the occurrence of hot flashes that can be detected using conventional forms of antiandrogen therapy (ADT). In another embodiment, the administration of the compounds of the invention prevents gynecomastia, which can be detected using conventional forms of antiandrogen therapy (ADT). In another embodiment, the administration of the compounds of the invention prevents the appearance of reduced bone mineral density (BMD), which can be detected using conventional forms of antiandrogen therapy (ADT). In another embodiment, the administration of the compounds of the invention prevents an increased tendency to bone fracture, which can be detected using conventional forms of antiandrogen therapy (ADT). In another embodiment, an increased tendency to bone fracture is pathological fracture, non-traumatic fracture, spinal fracture, non-vertebral fracture, new morphometric fracture, clinical fracture, or a combination thereof.

In one embodiment, the term “conventional antiandrogen therapy” refers to orchiectomy (surgical castration) when the surgeon removes the testes. In another embodiment, the term “conventional antiandrogen therapy” refers to the administration of luteinizing hormone releasing hormone releasing factor (LHRH) analogues; these drugs lower the amount of testosterone produced by the testes. Examples of LHRH analogues commonly available in the United States include leuprolide (lupron, viadur (viadur), eligard), goserelin (zoladex), triptorelin (trelstar) and histrelin (vantas). In another embodiment, the term “conventional antiandrogen therapy” refers to the administration of antiandrogens; antiandrogens block the body’s ability to use any androgens. Even after orchiectomy or during treatment with LHRH analogues, a small amount of androgen is still produced by the adrenal glands. Examples of antiandrogenic drugs include flutamide (eulexin), bicalutamide (casodex) and nilutamide (nilandron). In another embodiment, the term “conventional antiandrogen therapy” refers to the administration of luteinizing hormone releasing hormone releasing factor (LHRH) antagonists such as abarelix (plenaxis); degarelix (firmagon) is a new LHRH antagonist, which in 2008 was approved by the FDA (Food and Drug Administration) for the treatment of advanced prostate cancer. In another embodiment, the term “conventional antiandrogen therapy” refers to the administration of 5α-reductase inhibitors such as finasteride (proscar) and dutasteride (avodart); 5α-reductase inhibitors block the body’s ability to convert testosterone to the more active androgen 5α-dihydrotestosterone (DHT). In another embodiment, the term “conventional antiandrogen therapy” refers to the administration of testosterone biosynthesis inhibitors such as ketoconazole (nizoral). In another embodiment, the term “conventional antiandrogen therapy” refers to the administration of estrogens such as diethylstilbestrol or 17β-estradiol.

In one embodiment, the term “hot flashes” refers to a sudden sensation of heat in the upper body or throughout the body, redness of the face and neck, red spots appearing on the chest, back and arms, intense sweating, cold shiver, etc.

In one embodiment, the term “gynecomastia” refers to a benign enlargement of the mammary glands in males resulting from the proliferation of the glandular component of the mammary gland, which may or may not be associated with pain. Clinically, gynecomastia is determined by the presence of an elastic or solid tumor, concentrically expanding from the nipples. A condition known as pseudogynecomastia or lipomastia is characterized by fat deposition without glandular proliferation. Although gynecomastia is usually bilateral, it can also be one-sided.

In one embodiment, the methods of this invention relate to treating men diagnosed with prostate cancer or advanced prostate cancer by lowering testosterone levels, which in addition does not cause bone loss and hot flashes. In another embodiment, Compounds IA, I-XII are used in the methods of this invention, and these compounds are capable of decreasing testosterone, the primary cause of prostate cancer, without also causing some side effects, such as bone loss and hot flashes, which are usually observed with modern antiandrogen therapies (ADT) for prostate cancer.

In another embodiment, Table 8 below (Example 11) shows a decrease in the amount of testosterone that also does not cause bone loss as a result of the administration of Compound IV.

In one embodiment, the methods of this invention relate to reducing testosterone levels, which further cures common prostate cancer by administering a compound of formula IA, I-XII. In one embodiment, the methods of this invention relate to decreasing testosterone levels, which further leads to the suppression, reduction in incidence, lessening of severity or inhibition of advanced prostate cancer by administering a compound of formula IA, I-XII. In one embodiment, the methods of this invention relate to decreasing testosterone levels, which further provides palliative treatment for advanced prostate cancer by administering a compound of formula IA, I-XII.

In one embodiment, the methods of this invention relate to the treatment of advanced prostate cancer. In one embodiment, the methods of this invention relate to suppressing, reducing the incidence, alleviating the severity, or inhibiting advanced prostate cancer. In one embodiment, the methods of this invention relate to palliative treatment of advanced prostate cancer. In another embodiment, Compounds IA, I-XII are used in the methods of this invention. In another embodiment, this invention relates to the treatment of advanced prostate cancer. In another embodiment, the invention relates to the suppression of advanced prostate cancer. In another embodiment, this invention relates to reducing the incidence of advanced prostate cancer. In another embodiment, this invention relates to alleviating the severity of advanced prostate cancer. In another embodiment, the invention relates to the inhibition of advanced prostate cancer.

The term "advanced prostate cancer" refers to metastatic cancer that occurs in the prostate gland and gives metastases far beyond the prostate gland, such as into surrounding tissues, covering the seminal vesicles, lymph nodes or bones of the pelvic region or other parts of the body. Pathological conditions of prostate cancer are classified according to the Gleason score from 1 to 5 in order of increasing malignancy. In another embodiment, this definition should include patients with a significant risk of progressive disease and / or death from prostate cancer, and any such patient with cancer that extends beyond the capsule of the prostate gland and is characterized by stages of the disease such as IIB or worse, obviously has "common" disease.

In one embodiment, the methods provided herein and / or using the compounds provided herein are effective in providing feedback on the hypothalamic-pituitary-testicular axis (NRT axis). Feedback refers to the ability of a substance produced in one organ or tissue to regulate the activity of another organ or tissue that affects its own activity. In one embodiment, the feedback in the hypothalamic-pituitary-testicular axis (NRT axis) leads to a decrease in LH levels. In one embodiment, the feedback in the hypothalamic-pituitary-testicular axis (NRT axis) leads to a decrease in total serum testosterone levels. In one embodiment, the feedback in the hypothalamic-pituitary-testicular axis (NRT axis) leads to a decrease in serum free testosterone levels. In one embodiment, the feedback in the hypothalamic-pituitary-testicular axis (NRT axis) leads to a decrease in androgen levels in blood serum, tissues or tumors.

The hypothalamic-pituitary-testicular (NRT) axis refers to the endocrine physiological system, which regulates hormone levels in the hypothalamus, pituitary, and testes. LHRH (luteinizing hormone releasing factor) is released by the hypothalamus and stimulates the pituitary gland for the synthesis and secretion of LH and FSH (gonadotropins). Further, LH and FSH act on the testes, stimulating the production of testosterone and sperm. Testosterone then has a direct negative feedback effect on the hypothalamic secretion of LHRH and an indirect negative feedback effect on the production of LH and FSH in the pituitary gland. Estrogens, androgens, and serum proteins (e.g., inhibin) also negatively affect LHRH secretion and LH and FSH secretion.

The pituitary gland is one of the glands that regulates the level of testosterone in the body. When testosterone levels drop, the pituitary gland releases luteinizing hormone (LH). This hormone induces the production of more testosterone in the testes. Testosterone levels increase during puberty. The highest level of testosterone is reached at the age of 20-40 years and then gradually decreases in elderly men. In women, the amount of testosterone is greatly reduced compared to men. However, testosterone plays an important role for the entire body of both men and women. It affects the brain, bone and muscle mass, the distribution of body fat, the vascular system, energy levels, tissue (external) genital organs and sexual function. Most of the testosterone in the blood is bound to a protein called sex hormone binding globulin (SHBG) or another whey protein called albumin. Testosterone that is not bound (or "free") can also be determined clinically.

In another embodiment, a decrease in serum total testosterone levels or serum free testosterone levels, independent of a decrease in serum luteinizing hormone levels, is due to an increase in sex hormone binding globulin (SHBG) levels. In another embodiment, a decrease in free testosterone levels, independent of a decrease in serum luteinizing hormone levels, is due to an increase in sex hormone binding globulin (SHBG) levels. In another embodiment, a decrease in serum total testosterone or serum free testosterone, independent of a decrease in serum luteinizing hormone (LH) levels, is due to inhibition of testicular production or secretion by Leydig cells in the testes. In another embodiment, a decrease in serum total testosterone or serum free testosterone, independent of a decrease in serum luteinizing hormone (LH) levels, is due to a decrease in adrenal steroidogenesis.

In one embodiment, the compounds described herein and / or compositions containing these compounds can be used to reduce luteinizing hormone (LH) levels. In another embodiment, the compounds and / or compositions of this invention can be used to reduce endogenous sex hormones.

Members of the hydroxysteroid dehydrogenase (HSD) family are involved in the conversion of steroids circulating in the bloodstream. 17β-HSD5 converts androstenedione to testosterone, and estrone to estradiol. In addition to this, she is also involved in the synthesis of prostaglandins. In one embodiment, the compounds of the invention inhibit HSD, specifically inhibit 17β-hydroxysteroid dehydrogenase 5 (17β-HSD5). Such inhibition can be useful in ADT, preventing peripheral / extragonas synthesis of testosterone, which can get out of the control of the NRT axis and lead to an insufficient decrease in total or free testosterone in the blood serum or make possible the presence of locally elevated intracellular testosterone levels, which in both cases may be adverse in ADT.

Antiandrogen therapy (ADT) provided by LHRH agonist therapy, i.e. the introduction of luteinizing hormone releasing hormone releasing factor agonists (LHRH) or their analogs, first stimulates the release of gonadotropin from the pituitary gland and produces testosterone from the testicles (the so-called “acute reaction” (from the English flare reaction)), then decreases the release of gonadotropin and decreases levels like testosterone and estrogen. This “acute reaction” caused by LHRH agonist therapy has a negative effect on the treatment of prostate cancer due to increased levels of androgen / testosterone. In addition, LHRH therapy is associated with an increased risk of diabetes and cardiovascular disease (Smith (2008) Current Prostate Reports. 6: 149-154).

To overcome the "acute" effects of LHRH therapy, antiandrogen monotherapy (bicalutamide, flutamide, chlormadinone), approaches with combined LHRH / antiandrogen therapy, and the use of LHRH antagonists (degarelix) have been proposed (Suzuki et al., (2008) Int. J. Clin. Oncol . 13; 401-410; Sharifi, N. et al., (2005) JAMA. 294 (2): 238-244). Antiandrogen monotherapy does not reduce subject's androgen levels. Anti-androgen monotherapy with bicalutamide has been shown to be less effective than ADT for patients with prostate cancer with bone metastases. In addition, the adverse effects observed with bicalutamide therapy include soreness and hypertrophy (gynecomastia and mastodynia) of the mammary glands. (Suzuki et al., Ibid.). Additional risks associated with antiandrogen therapy include elevated liver transaminases (Sharifi et al., Ibid.).

In one embodiment, the present invention provides a reduction in LH levels and thereby a decrease in serum total testosterone and / or serum free testosterone levels, without causing an “acute” effect and at the same time overcoming the adverse effects associated with estrogen deficiency, due to a decrease in testosterone levels using traditional ADT methods. Methods / applications of the compounds of the invention include the manifestation of tissue-selective estrogenic activities that maintain bone tissue (agonistic effect on bone tissue), reduction of coagulation potential and / or hot flashes and / or less or less strong or neutral effect on breast tissue compared to estradiol or diethylstilbestrol.

In one embodiment, Compound IV exhibits agonistic, but not antagonistic effects (Examples 6 and 7), i.e., the use of Compound IV will not lead to increased levels of gonadotropins and testosterone.

In one embodiment, Compound IV exhibits agonistic activity (Examples 8-11), demonstrating a sustained pharmacological response in the form of lower levels of serum hormones, testosterone and androgen in general.

In one embodiment, the methods provided herein employing the compounds and / or compositions provided herein are effective in reducing or eliminating resorptive effects in bone tissue caused by a decrease in LH levels using conventional forms of ADT. In one embodiment, the methods that are provided herein and / or which employ the compositions provided herein are effective in reducing or eliminating resorptive effects in bone tissue caused by lower testosterone levels using traditional forms of ADT. In one embodiment, the methods provided herein employing the compositions provided herein are effective in reducing or eliminating resorptive effects caused by a decrease in estrogen levels as a result of a decrease in LH level. In one embodiment, the methods provided herein using the compounds and / or compositions provided herein prevent bone resorptive effects associated with a decrease in LH levels using traditional forms of ADT. In one embodiment, the methods provided herein employing the compounds and / or compositions provided herein prevent bone loss associated with decreased levels of endogenous LH, testosterone and / or estradiol using conventional forms of ADT. In one embodiment, the methods provided herein using the compounds and / or compositions provided herein increase bone density (BMD) while providing a reduction in LH. In one embodiment, the methods provided herein using the compounds and / or compositions provided herein increase the percentage of bone volume while providing a reduction in the levels of endogenous LH, testosterone and / or estradiol.

In some embodiments, the invention provides a method for preventing and / or reducing thromboembolism by administering a compound of the invention or an isomer thereof, pharmaceutical product, polymorph, hydrate, or any combination thereof.

In one embodiment, the methods provided herein employ the compounds and / or compositions provided herein effective in breast tissue. In one embodiment, the methods provided herein using the compounds and / or compositions provided herein provide a reduction in LH while preventing gynecomastia associated with a decrease in LH resulting from conventional ADT.

In one embodiment, Example 13 presents the results of special toxicity studies, and in vitro studies on human platelets showed that Compound IV had a much lower procoagulating activity than DES. Thus, Compound IV, an ER-selective agonist, in relation to prostate cancer can provide a beneficial effect of DES with a lower risk of thrombotic events than DES, as well as providing a beneficial effect of an agonist or antagonist of LHRH without causing loss of bone mass, hot flashes or unfavorable lipid profiles.

Therapy with diethylstilbestrol alone (DES) or in conjunction with another ADT showed that the use of DES prevented bone resorption in patients diagnosed with prostate cancer. Although DES has been introduced as a therapy for prostate cancer, it is believed that the effects of DES on angiogenesis and malignancy are mediated by DES metabolites, and not by its action through an estrogen receptor. In addition, DES dosage levels for therapeutic use have numerous adverse side effects, including vascular disease, cardiovascular complications, thrombotic complications due to toxicity, gynecomastia, erectile dysfunction, and decreased libido (Scherrand Pitts, ibid., And Presti, JC Jr. (1996) JAMA. 275 (15): 1153-6).

In one embodiment, the present invention overcomes the negative side effects of therapy with LHRH agonists or antagonists, used alone or in combination with antiandrogens or DES. In another embodiment, the methods of the subject invention provide antiandrogen therapy without adverse side effects associated with loss of estrogen, such as adverse conditions associated with bone tissue, and without adverse side effects associated with stimulation of estrogen such as gynecomastia. In another embodiment, the methods of the present invention provide for lowering LH levels and thereby lowering serum total and / or free testosterone levels without causing an “acute” effect while overcoming the adverse effects associated with estrogen deficiency caused by a decrease in LH and overcoming the adverse effects associated with a general increase in estrogen agonist levels observed with DES therapy. Methods / applications of the compound of the invention provide for the manifestation of tissue-selective estrogenic activities, thereby maintaining bone tissue (agonistic effect on bone tissue), reducing coagulation potential and neutralizing breast tissue.

The antiestrogenic effects of traditional selective estrogen receptor modulators (SERMs), such as tamoxifen, toremifene, and raloxifene, at the hypothalamic level are expressed in increased levels of gonadotropins or increased levels of LH in men and thus potentially lead to increased serum testosterone levels (Tsouri et al. 2008 Fertility and Sterility doi: 10.1016). Conversely, methods of the subject invention, comprising administering a Compound of Formula IA, I-XII, comprise reducing a LH level in a male subject.

Additional Embodiments Compounds of Formula I

In one embodiment of the methods of this invention, Y in a Compound of Formula I is C (O). In another embodiment, Y represents CH ₂ . In another embodiment, R ₁ and R ₂ in the Compound of Formula I or IA independently are O-Alk-NR ₅ R ₆ or an O-Alk heterocycle. In another embodiment, Alk in these groups O-Alk-heterocycle, O-Alk-NR ₅ R ₆ , -Alk-heterocycle and Alk-NR ₅ R ₆ , which are described above, is a linear alkyl of 1-7 atoms carbon, branched alkyl of 1-7 carbon atoms or cyclic alkyl of 3-8 carbon atoms. In another embodiment, the alkyl is ethylene (—CH ₂ CH ₂ -). In another embodiment, Alk is methylene (—CH ₂ -). In another embodiment, Alk is propylene (—CH ₂ CH ₂ CH ₂ -). In another embodiment, Alk is 2-methylpropylene (—CH ₂ CH (CH ₃ ) CH ₂ -).

. В другом воплощении способов R₁ в Соединении формулы I или IA представляет собой гидроксил. В другом воплощении способов R₁ в Соединении формулы I или IA представляет собой алкокси. В другом воплощении способов R₁ и R₂ независимо представляют собой водород, галоген, гидроксил, алкокси, циано, нитро, CF₃, N(R)₂, сульфонамид, SO₂R, алкил, галогеноалкил, арил, O-Alk-NR₅R₆ или O-Alk-гетероцикл, где гетероцикл представляет собой 3-7-членное замещенное или незамещенное гетероциклическое кольцо, возможно ароматическое. В другом воплощении способов R₁ и R₂ в Соединении формулы I или IA независимо представляют собой галоген, гидроксил, алкокси, циано, нитро, CF₃, N(R)₂, сульфонамид, SO₂R, алкил, галогеноалкил, арил, O-Alk-NR₅R₆ или O-Alk-гетероцикл, где гетероцикл представляет собой 3-7-членное замещенное или незамещенное гетероциклическое кольцо, возможно ароматическое. В другом воплощении способов R₂ в Соединении формулы I или IA представляет собой галоген. В другом воплощении способов R₂ в Соединении формулы I или IA представляет собой F. В другом воплощении способов R₂ в Соединении формулы I представляет собой Cl. В другом воплощении способов R₂ в Соединении формулы I или IA представляет собой Br. В другом воплощении способов R₂ в Соединении формулы I или IA представляет собой I. В другом воплощении способов R₂ в Соединении формулы I или IA представляет собой гидроксил. В другом воплощении способов R₁ и/или R₂ представляет собой CF₃. В другом воплощении R₁ и/или R₂ представляет собой СН₃. В другом воплощении R₁ и/или R₂ представляет собой галоген. В другом воплощении R₁ и/или R₂ представляет собой F. В другом воплощении R₁ и/или R₂ представляет собой Cl. В другом воплощении R₁ и/или R₂ представляет собой Br. В другом воплощении R₁ и/или R₂ представляет собой I. В другом воплощении R₂ в Соединении формулы I находится в пара-положении.In one embodiment of the methods of this invention, R ₁ in the Compound of Formula I or IA is in the para position. In one embodiment of the methods of this invention, R ₁ and R ₂ in the Compound of Formula I or IA are different. In another embodiment of the methods of this invention, R ₁ and R ₂ in the Compound of formula I or IA are the same. In another embodiment of the methods of this invention, R ₁ in the Compound of formula I or IA is

. In another embodiment of the methods, R ₁ in the Compound of Formula I or IA is hydroxyl. In another embodiment of the methods, R ₁ in the Compound of Formula I or IA is alkoxy. In another embodiment of the methods, R ₁ and R ₂ independently represent hydrogen, halogen, hydroxyl, alkoxy, cyano, nitro, CF ₃ , N (R) ₂ , sulfonamide, SO ₂ R, alkyl, haloalkyl, aryl, O-Alk-NR ₅ R ₆ or an O-Alk heterocycle, wherein the heterocycle is a 3-7 membered substituted or unsubstituted heterocyclic ring, optionally aromatic. In another embodiment of the methods, R ₁ and R ₂ in the Compound of Formula I or IA independently represent halogen, hydroxyl, alkoxy, cyano, nitro, CF ₃ , N (R) ₂ , sulfonamide, SO ₂ R, alkyl, haloalkyl, aryl, O -Alk-NR ₅ R ₆ or O-Alk-heterocycle, where the heterocycle is a 3-7 membered substituted or unsubstituted heterocyclic ring, possibly aromatic. In another embodiment of the methods, R ₂ in the Compound of formula I or IA is halogen. In another embodiment of the methods, R ₂ in the Compound of Formula I or IA is F. In another embodiment of the methods R ₂ in the Compound of Formula I is Cl. In another embodiment of the methods, R ₂ in the Compound of formula I or IA is Br. In another embodiment of the methods, R ₂ in the Compound of Formula I or IA is I. In another embodiment of the methods R ₂ in the Compound of Formula I or IA is hydroxyl. In another embodiment of the methods, R ₁ and / or R ₂ is CF ₃ . In another embodiment, R ₁ and / or R ₂ represents CH ₃ . In another embodiment, R ₁ and / or R ₂ is halogen. In another embodiment, R ₁ and / or R ₂ is F. In another embodiment, R ₁ and / or R ₂ is Cl. In another embodiment, R ₁ and / or R ₂ represents Br. In another embodiment, R ₁ and / or R ₂ is I. In another embodiment, R ₂ in the Compound of Formula I is in the para position.

In one embodiment of the methods of this invention, R ₃ and R ₄ in the Compound of Formula I or IA are the same. In another embodiment of the methods of this invention, R ₃ and R ₄ in the Compound of formula I or IA are different. In another embodiment of the methods j and k in the Compound of Formula I or IA independently represent 1. In another embodiment of the methods R ₃ and R ₄ in the Compound of Formula I or IA independently represent halogen, haloalkyl, hydroxyl or alkyl. In another embodiment of the methods R ₃ and R ₄ in the Compound of Formula I or IA independently represent F. In another embodiment of the methods R ₃ and R ₄ in the Compound of Formula I or IA independently represent Br. In another embodiment of the methods, R ₃ and R ₄ in the Compound of Formula I or IA are independently Cl. In another embodiment, R ₄ is in the para position. In another embodiment, R ₃ is in the ortho position. In another embodiment, R ₃ is in the meta position. In another embodiment, R ₃ and / or R ₄ is CF ₃ . In another embodiment, R ₃ and / or R ₄ represents CH ₃ .

In one embodiment of the methods of this invention, R ₅ and R ₆ in the Compound of Formula I or IA form a 3-7 membered ring together with a nitrogen atom. In another embodiment, the ring is a saturated or unsaturated ring. In another embodiment, the ring is a substituted or unsubstituted ring. In another embodiment of the methods of this invention, R ₅ and R ₆ in the Compound of Formula I or IA form a piperidine ring together with nitrogen. In another embodiment of the methods, R ₅ and R ₆ in the Compound of Formula I or IA form a pyrazine ring together with nitrogen. In another embodiment of the methods, R ₅ and R ₆ in the Compound of Formula I or IA form a piperazine ring together with nitrogen. In another embodiment of the methods, R ₅ and R ₆ in the Compound of Formula I or IA form a morpholine ring together with nitrogen. In another embodiment of the methods, R ₅ and R ₆ in the Compound of Formula I or IA form a pyrrole ring together with nitrogen. In another embodiment of the methods, R ₅ and R ₆ in the Compound of Formula I or IA form pyrrolidine. In another embodiment of the methods, R ₅ and R ₆ in the Compound of Formula I or IA form a pyridine ring together with nitrogen. In another embodiment, this ring is substituted with halogen, alkyl, alkoxy, alkylene, hydroxyl, cyano, nitro, amino, amide, COOH or aldehyde.

In another embodiment of the methods of the invention, R ₁ in the Compound of Formula I or IA and R ₂ in the Compound of Formula I or IA are independently an O-Alk heterocycle or an OCH ₂ CH ₂ heterocycle. In another embodiment, the term “heterocyclic” group refers, in one embodiment, to a ring structure containing, in addition to carbon atoms, sulfur, oxygen, nitrogen atoms, or any combination thereof, as part of a ring. In another embodiment, the heterocycle is a 3-12 membered ring. In another embodiment, the heterocycle is a 6 membered ring. In another embodiment, the heterocycle is a 5-7 membered ring. In another embodiment, the heterocycle is a 4-8 membered ring. In another embodiment, the heterocyclic group may be unsubstituted or may be substituted with halogen, haloalkyl, hydroxyl, alkoxy, carbonyl, amido, alkylamido, dialkylamido, cyano, nitro, CO ₂ H, amino, alkylamino, dialkylamino, carboxyl, thio and / or thioalkyl. In another embodiment, the heterocyclic ring may be fused to another saturated or unsaturated cycloalkyl or heterocyclic 3-8 membered ring. In another embodiment, the heterocyclic ring is a saturated ring. In another embodiment, the heterocyclic ring is an unsaturated ring. In another embodiment, the heterocycle is piperidine. In another embodiment, the heterocycle is pyridine. In another embodiment, the heterocycle is piperidine, pyridine, furan, thiophene, pyrrole, pyrrolidine, pyrazine, piperazine or pyrimidine.

The term “cycloalkyl” refers to a non-aromatic monocyclic or polycyclic ring containing carbon and hydrogen atoms. The cycloalkyl group may contain one or more than one carbon-carbon double bond in the ring, provided that the ring does not become aromatic due to their presence. Examples of cycloalkyl groups include, but are not limited to, (C3-C7) cycloalkyl groups such as cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl and cycloheptyl and saturated cyclic and bicyclic terpenes, and (C3-C7) cycloalkenyl groups such as cyclopropenyl, cyclobutenyl , cyclopentenyl, cyclohexenyl and cycloheptenyl and unsaturated cyclic and bicyclic terpenes. The cycloalkyl group may be unsubstituted or may be substituted with one or two substituents. Preferably, the cycloalkyl group is a monocyclic ring or a bicyclic ring.

The term “alkyl” refers, in one embodiment, to a saturated aliphatic hydrocarbon, including straight chain, branched chain, and cyclic alkyl groups. In one embodiment, the alkyl group contains 1-12 carbon atoms. In another embodiment, the alkyl group contains 1-7 carbon atoms. In another embodiment, the alkyl group contains 1-6 carbon atoms. In another embodiment, the alkyl group contains 1-4 carbon atoms. In another embodiment, the cyclic alkyl group contains 3-8 carbon atoms. In another embodiment, the cyclic alkyl group contains 3-12 carbon atoms. In another embodiment, branched alkyl is alkyl substituted with alkyl side chains containing 1-5 carbon atoms. In another embodiment, branched alkyl is alkyl substituted with haloalkyl side chains containing 1-5 carbon atoms. The alkyl group may be unsubstituted or may be substituted with halogen, haloalkyl, hydroxyl, alkoxy, carbonyl, amido, alkylamido, dialkylamido, nitro, amino, alkylamino, dialkylamino, carboxyl, thio and / or thioalkyl.

The term “alkenyl” group refers, in another embodiment, to an unsaturated hydrocarbon, including straight chain, branched chain and cyclic alkyl groups containing one or more double bonds. An alkenyl group may have one double bond, two double bonds, three double bonds, etc. In another embodiment, the alkenyl group contains 2-12 carbon atoms. In another embodiment, the alkenyl group contains 2-6 carbon atoms. In another embodiment, the alkenyl group contains 2-4 carbon atoms. Examples of alkenyl groups are ethenyl, propenyl, butenyl, cyclohexenyl, etc. The alkenyl group may be unsubstituted or may be substituted with halogen, hydroxy, alkoxy, carbonyl, amido, alkylamido, dialkylamido, nitro, amino, alkylamino, dialkylamino, carboxyl, thio and / or thioalkyl.

The term “aryl” group refers to an aromatic group containing at least one carbocyclic aromatic group or heterocyclic aromatic group, which may be unsubstituted or may be substituted by one or more groups selected from halogen, haloalkyl, hydroxy, alkoxy, carbonyl, amido, alkylamido, dialkylamido, nitro, amino, alkylamino, dialkylamino, carboxy or thio or thioalkyl. Non-limiting examples of aryl rings are phenyl, naphthyl, pyranyl, pyrrolyl, pyrazinyl, pyrimidinyl, pyrazolyl, pyridinyl, furanyl, thiophenyl, thiazolyl, imidazolyl, isoxazolyl and the like. In one embodiment, the aryl group is a 4-8 membered ring. In another embodiment, the aryl group is a 4-12 membered ring (a). In another embodiment, the aryl group is a 6 membered ring. In another embodiment, the aryl group is a 5 membered ring. In another embodiment, the aryl group is a system of 2-4 fused rings.

The term “aldehyde” group refers, in one embodiment, to an alkyl or alkenyl substituted with a formyl group, wherein the alkyl or alkenyl is as defined above. In another embodiment, the aldehyde group is an aryl or phenyl group substituted with a formyl group, wherein aryl is as defined above. Examples of aldehydes are formyl, acetal, propanal, butanal, pentanal, benzaldehyde. In another embodiment, the aldehyde group is a formyl group.

The term “haloalkyl” group refers, in another embodiment, to an alkyl group as defined above that is substituted by one or more than one halogen atom, for example F, Cl, Br or I.

The term “hydroxyl” group refers, in another embodiment, to an OH group. It will be apparent to those skilled in the art that when R ₁ , R _2, or R ₃ in the compounds of the present invention are OR, then R is not OH.

In one embodiment, the term “halogen” or “halogen” refers to a halogen such as F, Cl, Br or I.

In another embodiment, the word “phenol” refers to any benzene derivative having an alcohol group (OH).

Reference to protected hydroxyl made in some embodiments includes the introduction of a substituent linked to the oxygen moiety of the benzene ring, and this substituent can be easily removed. In some embodiments, phenol protecting groups may include groups: methyl ether (methoxy), alkyl ether (alkoxy), benzyl ether (Bn), methoxymethyl (MOM) ether, benzoyloxymethyl (BOM) ether, benzyl, carbobenzoxy, methoxyethoxymethyl (MEM) ether, 2- (trimethylsilyl) ethoxy methyl (SEM) ether, methylthiomethyl (MTM) ether, phenylthiomethyl (RTM) ether, azidomethyl ether, cyanomethyl ether, 2,2-dichloro-1 simple , 1-difluoroethyl ether, 2-chloroethyl ether, 2-bromoethyl ether, tetrahydropyranyl (THP) ether, 1-ethoxyethyl (EE) ether, phenacyl ether, 4-bromophenacyl ether, cyclopropylmethyl methyl ether, ether, propargyl ether, isopropyl ether, cyclohexyl ether, tert-butyl ether, 2,6-dimethylbenzyl ether, 4-methoxybenzyl ether, o-nitrobenzyl ether, simple 2,6-dichlorobenzyl ether, 3,4-dichlorobenzyl ether, 4- (dimethylamino) carbonylbenzyl ether, 4-methylsulphinyl-benzyl ether, 4-anthrilmethyl ether, 4-picolyl ether, heptafluoro-p-tolyl tetrafluoro-4-pyridyl ether, trimethylsilyl ether (TMS) ether, tert-butyldimethylsilyl ether (TBDMS) ether, tert-butyl diphenylsilyl ether (TBDPS) ether, triisopropylsilyl ether (aryl formate, aryl formate, aryl aryl aryl aryl aryl aryl aryl aryl aryl aryl aryl aryl aryl aryl aryl aryl aryl aryl , ar l-9-fluorenecarboxylate, arylmethyl carbonate, 1-adamantyl carbonate, tert-butyl carbonate, 4-methylsulfinyl benzyl carbonate, 2,4-dimethylpent-3-yl carbonate, aryl-2,2,2-trichloroethyl carbonate, arylbenzyl carbonate, aryl, aryl Dmp-OAr), dimethylphosphinothionyl ester (Mpt-OAr), diphenylphosphinothionyl ester (Dpt-OAr), arylmethanesulfonate, aryltoluenesulfonate or aryl-2-formylbenzenesulfonate.

In one embodiment, the methods of this invention use N, N-bis (4-hydroxyphenyl) -4-propylbenzamide (II) or an isomer thereof, a pharmaceutically acceptable salt, pharmaceutical product, polymorph, hydrate, or any combination thereof. In another embodiment, the methods of this invention use 4,4 '- (2,3-dimethyl-benzylazanediyl) diphenol (III) or an isomer thereof, a pharmaceutically acceptable salt, pharmaceutical product, polymorph, hydrate, or any combination thereof. In another embodiment, the methods of this invention use 3-fluoro-N- (4-fluorophenyl) -4-hydroxy-N- (4-hydroxyphenyl) benzamide (IV) or an isomer thereof, a pharmaceutically acceptable salt, pharmaceutical product, polymorph, hydrate or any combination thereof. In another embodiment, the methods of this invention use N, N-bis (4-hydroxyphenyl) -2,3-dimethylbenzamide (V) or an isomer thereof, a pharmaceutically acceptable salt, pharmaceutical product, polymorph, hydrate, or any combination thereof. In another embodiment, the methods of this invention use N, N-bis (4-hydroxyphenyl) -2-naphthylamide (VI) or an isomer thereof, a pharmaceutically acceptable salt, pharmaceutical product, polymorph, hydrate, or any combination thereof. In another embodiment, the methods of this invention use 3-fluoro-4-hydroxy-N, N-bis (4-hydroxyphenyl) benzamide (VII) or an isomer thereof, a pharmaceutically acceptable salt, pharmaceutical product, polymorph, hydrate, or any combination thereof . In another embodiment, the methods of this invention use 4 - ((4-fluorophenyl) (4-hydroxybenzyl) amino) phenol (VIII) or an isomer thereof, a pharmaceutically acceptable salt, pharmaceutical product, polymorph, hydrate, or any combination thereof. In another embodiment, the methods of this invention use 4-fluoro-N- (4-hydroxyphenyl) -N- [4- (2-piperidin-1-yl-ethoxy) phenyl] -2-trifluoromethylbenzamide (IX ) or an isomer thereof, a pharmaceutically acceptable salt, pharmaceutical product, polymorph, hydrate, or any combination thereof. In another embodiment, the methods of this invention use a hydrochloride salt of compound IX (HCl salt of compound IX) or 4-fluoro-N- (4-hydroxy-phenyl) -N- [4- (2-piperidin-1-yl-ethoxy) hydrochloride ) -phenyl] -2-trifluoromethyl-benzamide (X) or an isomer thereof, a pharmaceutically acceptable salt, pharmaceutical product, polymorph, hydrate, or any combination thereof. In another embodiment, the methods of this invention use 3-fluoro-4-hydroxy-N- (4-hydroxyphenyl) -N-phenylbenzamide (XI) or an isomer thereof, a pharmaceutically acceptable salt, pharmaceutical product, polymorph, hydrate, or any combination thereof. In another embodiment, the methods of this invention use 3-fluoro-N, N-bis- (4-hydroxy-phenyl) -2-methyl-benzamide (XII) or an isomer thereof, a pharmaceutically acceptable salt, pharmaceutical product, polymorph, hydrate or any combination of them.

In one embodiment, “pharmaceutically acceptable salts” of a compound that can be prepared by reacting a compound of the invention with an acid or base are used in the methods of the invention.

Suitable pharmaceutically acceptable salts of the amines of the compounds according to the methods of this invention can be obtained from an inorganic acid or from an organic acid. In one embodiment, examples of inorganic salts of amines are bisulfates, borates, bromides, chlorides, hemisulphates, hydrobromates, hydrochlorides, 2-hydroxyethylsulfonates (hydroxyethanesulfonates), iodates, iodides, isothionates, nitrates, persulfates, phosphates, sulfates, sulfates ( sulfonylates, sulfonic acid salts (alkyl sulfonates, aryl sulfonates, halogen-substituted alkyl sulfonates, halogen-substituted aryl sulfonates), sulfonates and thiocyanates.

In one embodiment, examples of organic amine salts can be selected from the classes of aliphatic, cycloaliphatic, aromatic, araliphatic, heterocyclic, carboxylic and sulfonic organic acids, examples of which are acetates, arginines, aspartates, ascorbates, adipates, anthranilates, algenates, alkanes -carboxylates, substituted alkane-carboxylates, alginates, benzenesulfonates, benzoates, bisulfates, butyrates, bicarbonates, bitartrates, carboxylates, citrates, camphorites, camphorsulfonates, cyclohexyl sulfamates, c clopentane propionates, calcium edetates, camsylates, carbonates, clavulanates, cinnamates, dicarboxylates, digluconates, dodecyl sulfonates, dihydrochlorides, decanoates, enanthates, ethanesulfonates, edetates, edisylates, glyatates, glucurate gum, fumate, fumate, gum, fuma , glucorates, glucoheptanoates, glycerophosphates, gluceptates, glycolyl arsenylates, glutarates, glutamate, heptanoates, hexanoates, hydroxy maleates, salts of hydroxycarboxylic acids, hexyl resorcinates, hydroxybenzoates, hydroxy naphthoates, hydrofluorates, la tats, lactobionates, laurates, malates, maleates, methylenebis (beta-oxine naphthoate), malonates, mandelates, mesylates, methanesulfonates, methyl bromides, methyl nitrates, methyl sulfonates, monopotassium maleates, mucates, monocarboxylates, nitrates, naphthalenesulfonates, nickelsulfonates, niconates -methylglucamines, oxalates, octanoates, oleates, pamoates, phenylacetates, picrates, phenylbenzoates, pivalates, propionates, phthalates, pectinates, phenylpropionates, palmitates, pantothenates, polygalacturates, pyruvates, quinic acid salts, salicylates, salicylates, sulfonylates, subacetates, tartrates, theophylline acetate, p-toluenesulfonates (tosylates), trifluoroacetates, terephthalates, tannates, theoclates, trihaloacetates, triethiodide, tricarboxylates, undecanoates and valerates.

In one embodiment, examples of inorganic salts of carboxylic acids or phenols may be selected from ammonium, alkali metal salts including lithium, sodium, potassium, cesium; alkaline earth metals, including calcium, magnesium, aluminum; zinc, barium, choline, quaternary ammonium.

In another embodiment, examples of organic salts of carboxylic acids or phenols may be selected from salts of arginine, organic amines, including aliphatic organic amines, alicyclic organic amines, aromatic organic amines, benzatines, tert-butylamines, benethamines (N-benzylphenethylamine), dicyclohexylamines, dimethyl diethanolamines, ethanolamines, ethylenediamines, hydrabamines, imidazoles, lysines, methylamines, meglumines, N-methyl-D-glucamines, N, N'-dibenzylethylenediamines, nicotinamides, organic amines, ornithines, pyridines, picolines, piperazines, procaine, tris (hydroxymethyl) methylamines, triethylamines, triethanolamines, trimethylamines, tromethamines and urea.

In one embodiment, such salts can be formed by conventional methods, for example, by reacting a product in the form of a free base or free acid with one or more equivalents of the corresponding acid or base in a solvent or medium in which the salt is insoluble or in a solvent such as water, which is removed in vacuo, or by freeze-drying, or by exchanging ions of a salt available for another ion on a suitable ion-exchange resin.

In one embodiment, a pharmaceutically acceptable salt of the compounds of this invention is used in the methods of this invention. In one embodiment, a pharmaceutically acceptable salt of the Compounds of Formula IA, I-XII is used in the methods of this invention. In one embodiment, the amine salt of the Compounds of Formula IA, I-XII of the present invention is used in the methods of this invention. In one embodiment, the phenol salt of the Compounds of Formula IA, I-XII of the present invention is used in the methods of this invention.

In one embodiment, the methods of this invention employ a compound of formula IA, I-XII and / or an isomer thereof, a pharmaceutical product, a hydrate, a polymorph, or a combination thereof, in the form of a free base, free acid, in an uncharged form or in a non-complexing form .

In some embodiments of the invention, the compounds of the invention contain three phenyl groups that are held together by an amide bond. In one embodiment, the compounds of the invention are uncharged structures. In another embodiment, the compounds of the invention are free base structures. In another embodiment, the compounds of the invention are free acid structures. In another embodiment, the compounds of the invention are non-complexing structures. In another embodiment, the compounds of the invention are non-ionized structures. In another embodiment, the compounds of the invention are pharmaceutically acceptable salts. In another embodiment, some of the compounds of the invention are hydrochloride (HCl) salts.

In one embodiment, the isomer of the Compound of Formula IA, I-XII is used in the methods of this invention. In one embodiment, a pharmaceutical product of a Compound of Formula IA, I-XII is used in the methods of this invention. In one embodiment, the hydrate of the Compound of Formula IA, I-XII is used in the methods of this invention. In one embodiment, a polymorph of Compounds of formula IA, I-XII is used in the methods of this invention. In one embodiment, a metabolite of the Compound of Formula IA, I-XII is used in the methods of this invention. In another embodiment, the methods of this invention use a composition comprising a Compound of formula IA, I-XII, as described herein, or, in another embodiment, a combination of an isomer, metabolite, pharmaceutical product, hydrate, polymorph of a Compound of formula IA, I-XII .

In one embodiment, the term "isomer" includes, but is not limited to, optical isomers and analogs, structural isomers and analogs, conformational isomers and analogs, and the like.

In one embodiment, the term "isomer" is intended to include the optical isomers of a compound. In one embodiment, the term "isomer" is intended to include stereoisomers of a compound. In the compounds of the invention, there is an amide bond which can be characterized by its cis or trans isomerization. It should be understood that the present invention covers any optically active or stereoisomeric forms or mixtures thereof, and it is believed that their use for any application is within the scope of this invention.

In another embodiment, the invention also includes hydrates of the compound. In one embodiment, the term "hydrate" refers to hemihydrate, monohydrate, dihydrate, trihydrate, or other forms that are known in the art.

Synthesis Methods

Compounds of formula I or IA can easily be prepared, for example, by reacting substituted diphenylamine with benzoic acid or benzoyl halide in the presence of a base to give benzamide. In one embodiment, the base is pyridine. In another embodiment, the benzoyl halide is benzoyl chloride. In another embodiment, the hydroxyl substituent is protected during the reaction between diphenylamine and benzoic acid or benzoyl halide. In another embodiment, the hydroxyl protecting group is optionally removed in the last step. See also U.S. Publication No. 2009/00624231, which is incorporated herein by reference in its entirety.

For example, the Compound of formula IA:

,

,

where R ₁ , R ₂ , R ₃ and R ₄ , j and k are as described above,

can be obtained by a method including bringing into interaction

from

with receipt

; и

; and

reacting diphenylamine (3) with

in the presence of a base to obtain

,

,

where if R ₁ , R ₂ , R ₃ and R ₄ independently represent OH, O-Alk-R ₅ R ₆ or O-Alk heterocycle, then R ₁ ', R ₂ ', R ₃ ', R ₄ ' represent a protected hydroxyl group, the protective group being removed to give free hydroxyl, or possibly followed by reaction with a Cl-Alk heterocycle or Cl-Alk-NR ₅ R ₆ to give a Compound of formula IA:

where if R ₁ , R ₂ , R ₃ and R _{4 are} independently not OH, O-Alk-NR ₅ R _6, or O-Alk heterocycle, then R ₁ ', R ₂ ', R ₃ 'and R ₄ ' are R ₁ , R ₂ , R ₃ and R _4, respectively.

Another example of a process for preparing a compound of formula IA:

,

,

where R ₁ , R ₂ , R ₃ and R ₄ are as described above, includes bringing into interaction

from

in the presence of a base to obtain

,

,

where if R ₁ , R ₂ , R ₃ and R ₄ independently represent OH, O-Alk-R ₅ R ₆ or O-Alk heterocycle, then R ₁ ', R ₂ ', R ₃ 'and R ₄ ' represent a protected hydroxyl group, where the protective group is removed to give free hydroxyl, or possibly followed by reaction with a Cl-Alk heterocycle or Cl-Alk-NR ₅ R ₆ to give a compound of formula IA:

,

,

where if R ₁ , R ₂ , R ₃ and R _{4 are} independently not OH, O-Alk-NR ₅ R _6, or O-Alk heterocycle, then R ₁ ', R ₂ ', R ₃ 'and R ₄ ' are R ₁ , R ₂ , R ₃ and R _4, respectively.

In one example, Compound II is prepared according to Example 1 and Figure 5.

In another example, Compound III was prepared according to Example 1 and FIG. 5.

In the following example, a compound of formula IV:

can be obtained as a result of interaction

from

in the presence of a base to obtain

followed by deprotection of the protected groups to give Compound IV:

,

,

where P and P 'are the same or different protecting groups. In one example, Compound IV is prepared according to Example 2 and FIG. 6.

In another example, Compound V is prepared according to Example 1 and FIG. 5.

In the following example, Compound VI was prepared according to Example 3 and FIG. 7.

In another example, Compound VII was prepared according to Example 1 and FIG. 5.

In another example, Compound VIII was prepared in accordance with Example 4 and FIG. 5.

In another example, Compound IX was prepared according to Example 5 and FIG. 8.

In another example, Compound X hydrochloride is prepared in accordance with Example 5 and FIG. 8.

In another example, Compound XI is prepared according to Example 1 and FIG. 5.

In another example, Compound XII is prepared in accordance with Example 1 and FIG. 5.

Suitable hydroxyl protecting groups include, for example, methyl ether (methoxy), benzyl ether (benzyloxy), methoxymethyl (MOM) ether, benzoyloxymethyl (BOM) ether, benzyl, carbobenzoxy, methoxyethoxymethyl (MEM) ether, 2- (trimethylsilyl) ethoxy methyl (SEM) ether, methylthiomethyl (MTM) ether, phenylthiomethyl (RTM) ether, azidomethyl ether, cyanomethyl ether, 2,2-dichloro-1,1-difluoroethyl ether, simple o 2-chloroethyl ether, 2-bromoethyl ether, tetrahydropyranyl ether (THP) ether, 1-ethoxyethyl (EE) ether, phenacyl ether, 4-bromophenacyl ether, cyclopropylmethyl ether, allyl ether, proparyl ether isopropyl ether, cyclohexyl ether, tert-butyl ether, benzyl ether, 2,6-dimethylbenzyl ether, 4-methoxybenzyl ether, o-nitrobenzyl ether, simply o 2,6-dichlorobenzyl ether, 3,4-dichlorobenzyl ether, 4- (dimethylamino) carbonylbenzyl ether, 4-methylsulfinylbenzyl ether, 4-anthrylmethyl ether, 4-picolyl ether, heptafluoro-p-tolyl tetrafluoro-4-pyridyl ether, trimethylsilyl ether (TMS) ether, tert-butyldimethylsilyl ether (TBDMS) ether, tert-butyl diphenyl silyl ether (TBDPS) ether, triisopropylsilyl ether (aryl formate, aryl formate, aryl aryl aryl aryl aryl aryl aryl aryl aryl aryl aryl aryl aryl aryl aryl aryl aryl aryl aryl aryl aryl aryl-9- luorenecarboxylate, arylmethyl carbonate, 1-adamantyl carbonate, tert-butyl carbonate, 4-methylsulfinyl benzyl carbonate, 2,4-dimethyl pent-3-yl carbonate, aryl-2,2,2-trichloroethyl carbonate, aryl benzyl carbonate, aryl carbamide ether dimethylphosphinothionyl ester (Mpt-OAr), diphenylphosphinothionyl ester (Dpt-OAr), arylmethanesulfonate, aryltoluenesulfonate or aryl-2-formylbenzenesulfonate.

The methods of this invention include the use of compounds IA, I-XII, the process for preparing the compounds of the invention involves reacting diphenylamine with benzoyl chloride in the presence of a base. Suitable bases include, for example, pyridine, triethylamine, K ₂ CO ₃ , Cs ₂ CO ₃ , Na ₂ CO ₃ , methylamine, imidazole, benzimidazole, histidine, tributylamine, or any combination thereof. In one embodiment, the base is pyridine.

The methods of this invention include the use of Compounds IA, I-XII, wherein the process for preparing the compounds of the invention involves deprotecting the protected hydroxyl. In another embodiment, the deprotection conditions are dependent on the protecting group. In some embodiments, the deprotection step involves hydrogenation in the presence of Pd / C. In another embodiment, deprotection involves interaction with BBr ₃ . In another embodiment, the deprotection step comprises reacting with an acid.

In the following examples, Compounds IA, I-XII are protected in accordance with FIGS. 5-8 and Examples 1-5.

Pharmaceutical Compositions

In some embodiments, the present invention provides methods of use that include administering a composition containing the described compounds. As used herein, “pharmaceutical composition” means a “therapeutically effective amount” of an active ingredient, i.e. the compounds of the invention, together with a pharmaceutically acceptable carrier or diluent. The term "therapeutically effective amount", as used herein, refers to the amount that provides a therapeutic effect for a given state and mode of administration.

As used herein, the term “administration” refers to bringing a subject into contact with a compound of the present invention. As used herein, administration can be carried out in vitro, i.e. in vitro, or in vivo, i.e. in the cells or tissues of living organisms, such as humans. In one embodiment, the present invention encompasses the administration of the compounds of the present invention to a male subject.

The present invention provides, in other embodiments, pharmaceutical products of the compounds described herein. The term "pharmaceutical product" refers, in other embodiments, to a composition suitable for pharmaceutical use (pharmaceutical composition), for example, as set forth herein.

The compounds of the invention may be administered on their own or as an active ingredient in the composition. Thus, the present invention also includes pharmaceutical compositions of Compounds of Formula I, containing, for example, one or more pharmaceutically acceptable carriers.

A huge number of standard references are available that describe techniques for preparing various compositions suitable for administering the compounds of the invention. Examples of possible compositions and preparations are contained, for example, in the Handbook of Pharmaceutical Excipients (American Pharmaceutical Association, current edition); Pharmaceutical Dosage Forms: Tablets (editors of Lieberman, Lachman and Schwartz, current edition, published by Marcel Dekker, Inc.), and Remington's Pharmaceutical Sciences (editor of Arthur Osol), 1553-1593 (current edition).

The route of administration and dosage forms are closely related to therapeutic amounts of compounds or compositions that are desirable and effective for a given therapeutic use.

Suitable dosage forms include those for oral, rectal, sublingual, mucosal, nasal, ophthalmic, subcutaneous, intramuscular, intravenous, transdermal, spinal, intrathecal, intraarticular, intraarterial, subarachnoid, intrabronchial, lymphatic and intrauterine delivery and other intrauterine forms ingredients, but are not limited to. Formulations suitable for oral administration are preferred.

In order to prepare such pharmaceutical dosage forms, the active ingredient can be mixed with a pharmaceutical carrier in accordance with traditional pharmaceutical blending methods. The carrier may be in a wide variety of forms, depending on the form of the drug required for administration.

In the manufacture of compositions in the composition of the oral dosage form, you can use any of the usual pharmaceutical environments. Thus, for liquid oral preparations, such as, for example, suspensions, elixirs and solutions, suitable carriers and additives include water, glycols, oils, alcohols, flavoring agents, preservatives, colorants and the like. For solid oral preparations, such as, for example, powders, capsules and tablets, suitable carriers and additives include starches, sugars, diluents, granulating agents, lubricants, binders, disintegrants and the like. Because of their ease in administration, tablets and capsules represent the most advantageous oral dosage unit form. If desired, sugar or enteric coating may be applied to the tablets using standard methods.

For parenteral compositions, the carrier will usually contain sterile water, although other ingredients, for example, solubility or preservation promoting ingredients, may be included. Injectable solutions may also be prepared, in which case appropriate stabilizers may be used.

In some cases, it may be advantageous to use the active agent in a “vectorized” form, such as by encapsulating the active agent in a liposome or other encapsulating medium or by “fixing” the active agent, for example, by covalent binding, chelation or the formation of associative coordination bonds, on a suitable biological molecule, such as molecules selected from proteins, lipoproteins, glycoproteins and polysaccharides.

The methods of treatment of the present invention using compositions suitable for oral administration can be represented by methods using separate units, such as capsules, cachets, tablets or lozenges, each of which contains a predetermined amount of the active ingredient in the form of, for example, powder or granules . It is possible that a suspension in an aqueous solution or non-aqueous liquid, such as a syrup, elixir, emulsion or extract, may be used.

A tablet may be made by compression or molding or by wet granulation, optionally together with one or more auxiliary ingredients. Compressed tablets can be made by compression in a machine suitable for this, wherein the active compound is in a free-flowing form, such as powder or granules, which may be mixed, for example, with a binder, disintegrant, lubricant, inert diluent, surface-active substance or brightening agent. Molded tablets consisting of a mixture of the powdered active compound with a suitable carrier can be made by molding in a suitable machine.

A syrup can be made by adding the active compound to a concentrated aqueous solution of sugar, for example sucrose, to which any auxiliary ingredient (s) can also be added. Such auxiliary ingredient (s) may include flavoring agents, a suitable preservative, agents to slow the crystallization of sugar, and agents to increase the solubility of any other ingredient, such as a polyhydric alcohol, for example glycerol or sorbitol.

Formulations suitable for parenteral administration may contain a sterile aqueous composition of the active compound, which is preferably isotonic with the blood of the recipient (e.g., saline). Such formulations may include suspending agents and thickening agents and liposomes or other microparticle systems that are designed to provide targeted delivery of the compound to blood components or to one or more organs. Such preparations may be presented in a single dosage form or in a dosage form for repeated administration.

Parenteral administration may refer to any suitable form of systemic delivery. Administration may be, for example, intravenous, intraarterial, intrathecal, intramuscular, subcutaneous, intraabdominal (e.g., intraperitoneal), etc., and may be carried out by means of infusion pumps (external or implantable) or any other suitable means corresponding to the desired route of administration.

Nasal compositions and other compositions for administration by spraying onto mucous membranes (for example, inhaled forms) may contain purified aqueous solutions of the active compounds with preservatives and isotonic agents. Preferably, the pH and isotonic state of such compositions is adjusted to values compatible with the nasal or other mucous membranes. Alternatively, they may be in the form of finely divided solid powders suspended in a gaseous carrier. Such compositions can be delivered using any suitable means or method, for example, by means of a nebulizer, atomizer, metered dose inhaler or the like.

Formulations for rectal administration may be presented as a suppository with a suitable carrier, such as cocoa butter, hydrogenated fats or hydrogenated fatty carboxylic acids.

Transdermal compositions can be prepared by introducing the active agent into a thixotropic or gel-like carrier, such as a cellulosic medium, for example methyl cellulose or hydroxyethyl cellulose, the resulting composition being then placed in a transdermal device adapted to ensure dermal contact with the skin of the user.

In addition to the aforementioned ingredients, the compositions of this invention may further include one or more accessory ingredient (s) selected from, for example, diluents, buffers, flavoring agents, binders, disintegrants, surfactants, thickeners, lubricants, preservatives (including antioxidants) and the like.

The compositions of the present invention may have an immediate release profile, continuous release, delayed onset release profile, or any other release profile known to one skilled in the art.

In one embodiment, the invention provides methods of: a) reducing serum total testosterone levels; b) lowering serum free testosterone levels by decreasing luteinizing hormone (LH) or regardless of decreasing LH hormone in a male subject having prostate cancer, comprising administering an oral composition comprising a Compound of Formula IA, I-XII. In further embodiments, the methods of this invention use an oral composition comprising a Compound of Formula II, Formula III, Formula IV, Formula V, Formula VI, Formula VII, Formula VIII, Formula IX, Formula X, Formula XI, or Formula XII.

In one embodiment, the invention provides a method for treating prostate cancer by lowering LH levels or regardless of decreasing LH levels in a male subject having prostate cancer, comprising administering an oral composition comprising a Compound of formula IA, I-XII. In further embodiments the present invention provides methods for treating prostate cancer by decreasing LH levels or regardless of decreasing LH levels in a male subject having cancer of the present Yelnia cancer, comprising administering an oral composition containing a compound of Formula II, Formula III, Formula IV, Formula V, Formula VI, Formula VII, Formula VIII, Formula IX, Formula X, Formula XI or Formula XII.

It should be understood that this invention encompasses any embodiment of a compound that is described herein and which in some embodiments is referred to as a “compound of the invention.”

In one embodiment, the methods of this invention may include administering the compounds of the invention in various dosages. In one embodiment, the compound of the invention is administered at a dosage of 1-1500 mg per day. In further embodiments, the compound of the invention is administered at a dose of 1-10 mg per day, 3-26 mg per day, 3-60 mg per day, 3-16 mg per day, 3-30 mg per day, 10-26 mg per day , 15-60 mg per day, 50-100 mg per day, 50-200 mg per day, 150-300 mg per day, 20-50 mg per day, 5-50 mg per day, 200-500 mg per day, 150-500 mg per day, 200-1000 mg per day, 300-1500 mg per day or 100-1000 mg per day.

In one embodiment, the methods of this invention may include administering the compounds of the invention in various dosages. In one embodiment, the compound of the invention is administered in a dosage of 3 mg. In further embodiments, the compound of the invention is administered in a dosage of 10 mg, 30 mg, 50 mg, 100 mg, 200 mg, 300 mg, 450 mg, 500 mg, 600 mg, 900 mg, 1000 mg or 1500 mg.

In one embodiment, the methods of this invention may include administering the compounds of the invention in various dosages. In one embodiment, the compound of the invention is administered at a dosage of 0.1 mg / kg / day. In further embodiments, the compound of the invention is administered in a dosage of 0.2 to 30 mg / kg / day or 0.2 mg / kg / day, 0.3 mg / kg / day, 1 mg / kg / day, 3 mg / kg / day, 5 mg / kg / day, 10 mg / kg / day, 20 mg / kg / day or 30 mg / kg / day.

In one embodiment, the methods of this invention are provided for the use of a pharmaceutical composition comprising a Compound of formula IA, I-XII. In further embodiments, the methods of this invention are provided for using a pharmaceutical composition comprising a Compound of Formula II, Formula III, Formula IV, Formula V, Formula VI, Formula VII, Formula VIII, Formula IX, Formula X, Formula XI, or Formula XII.

In some embodiment, the pharmaceutical composition is a solid dosage form. In another embodiment, the pharmaceutical composition is a tablet. In another embodiment, the pharmaceutical composition is a capsule. In another embodiment, the pharmaceutical composition is a solution. In another embodiment, the pharmaceutical composition is a transdermal patch.

In one embodiment, the use of a compound of the invention or a composition containing the compound will be used to inhibit, suppress, enhance or stimulate a desired response in a subject, which is apparent to one skilled in the art. In another embodiment, the compositions may also contain additional active ingredients whose activity is useful for the particular application for which the compound of the invention is administered.

When administered to mammals and in particular humans, it is expected that the physician will determine the actual dosage and duration of treatment, which will be most suitable for the individual and may vary depending on the age, weight, heredity and / or response of the individual.

In some embodiments, any of the compositions of this invention will comprise a compound of the invention in any form or any embodiment as set forth herein. In some embodiments, any of the compositions of this invention will comprise a compound of the invention in any form or any embodiment as set forth herein. In some embodiments, the compositions of this invention will consist essentially of the compounds of the invention in any form or any embodiment as set forth herein. In some embodiments, the term “comprise” refers to the inclusion of said active agent, such as a compound of the invention, as well as the inclusion of other active agents and pharmaceutically acceptable carriers, excipients, emollients, stabilizers, etc., such as are known in the pharmaceutical industry . In some embodiments, the term “consisting essentially of” refers to a composition whose active ingredient is only the specified active ingredient, but other compounds intended for stabilization, preservation, etc., may be included. this composition, but not directly involved in the therapeutic effect of the specified active ingredient. In some embodiments, the term “consisting essentially of” may refer to components that contribute to the release of the active ingredient. In some embodiments, the term "consisting" refers to a composition that contains an active ingredient and a pharmaceutically acceptable carrier or excipient,

It should be understood that any use of any of the Compounds that are described in this description can be used in the treatment of any disease, disorder or condition that is described in this description, and it is an embodiment of the present invention. In one embodiment, the compounds are in the form of a free base, a free acid, in an uncharged form or in a non-complexing form.

The following examples are presented to more fully illustrate preferred embodiments of the invention. They should in no way be construed as limiting the broad scope of the invention.

EXAMPLES

EXAMPLE 1

General Methods for the Synthesis of Compounds of Formulas II-XII and Synthetic Intermediates

Organic solvents, surfactants, antioxidants, etc. that can be used in the compositions described herein are usually available from commercial sources. For example, PEG-300, Polysorbate 80, Captex ™ 200, Capmul ™ MCM C8 can be obtained, for example, from the Dow Chemical Company (Midland, Ml), ICI Americas, Inc. (Wilmington, DE) or Abitec Corporation (Janesville, WI).

The estrogen receptor ligands described herein can be prepared by a variety of methods well known to those skilled in the art. For example, the estrogen receptor ligands described herein can be prepared by the synthetic methods described in US Patent Application Serial No. 2009/0062341; thereby the description of each of them is incorporated in its entirety by reference.

General synthesis of derivatives of N, N-bis-arylbenzamides

General synthesis of diarylanilins (Figure 5). Arylamine (1.5 equivalents), aryl iodide (1 equivalent), K ₂ CO ₃ (2 equivalents), CuI (0.1 equivalents) and L-proline (0.2 equivalents) were mixed together and dissolved in anhydrous DMSO at room temperature . Then the reaction mixture was stirred and heated to 90 ° C for 28 hours. The mixture was cooled to room temperature and hydrolyzed with water. To carry out the separation of the solution, EtOAc was added. The EtOAc-containing layer was separated, washed with brine and dried over anhydrous MgSO ₄ . The solvent was removed under reduced pressure. The solid residue was purified by flash column chromatography (silica gel) using a 5% EtOAc / hexanes mixture as eluent, which afforded the corresponding diarylaniline.

Bis- (4-methoxyphenyl) amine (1a): pale yellow solid, 73% yield. Mp 98.6-99.0 ° C. ¹ H NMR (CDCl ₃ , 300 MHz) δ 6.93-6.81 (m, 8H), 5.37 (s, br, 1H), 3.78 (s, 6H). MS m / z 228.4 (MH) ⁺ .

N- (4-Methoxyphenyl) phenylamine (1b): pale yellow solid, 70% yield. Mp 106.3-106.5 ° C. ¹ H NMR (CDCl ₃ , 300 MHz) δ 7.24-7.18 (m, 3H), 7.08-7.06 (m, 2H), 6.92-6.84 (m, 4H), 5.61 (s, br, 1H), 3.79 (s , 3H). MS m / z 200.1 (M + H) ⁺ .

N- (4-Fluorophenyl) -N-4-methoxyphenylamine (1c): pale yellow solid, 54% yield. Mp 60.6-61.0 ° C. ¹ H NMR (CDCl ₃ , 300 MHz) δ 7.01-6.83 (m, 8H), 3.78 (s, 3H). MS m / z 217 (M) ⁺ .

N- (4-Benzyloxyphenyl) -N-4-methoxyphenylamine (1d): pale yellow solid, 54% yield. Mp 108.0-108.4 ° C. ¹ H NMR (CDCl ₃ , 300 MHz) δ 7.34-7.08 (m, 5H), 6.90-6.81 (s, 3H), 3.78 (s, 3H). MS m / z 306 (M + H) ⁺ .

General synthesis of benzamides. Arylaniline (1 equivalent), benzoyl chloride (1.3 equivalents) and pyridine (6 equivalents) were mixed together and dissolved in anhydrous THF (tetrahydrofuran) at room temperature. The mixture was stirred and refluxed for 24 hours. The reaction solution was cooled to room temperature and hydrolyzed by adding 2 N. HCl solution. The solution was extracted with ethyl acetate. The organic layer was washed with saturated aqueous NaHCO ₃ to remove excess acid, dried over anhydrous MgSO ₄ , filtered, and concentrated under reduced pressure. The residue was purified by flash column chromatography using EtOAc / hexanes (3/7 v / v), which afforded the compounds of the corresponding benzamides.

3-Fluoro-N- (4-fluorophenyl) -4-methoxy-N- (4-methoxyphenyl) benzamide (2a): yellow solid. Mp 54-56 ° C. ¹ H NMR (CDCl ₃ / TMS (tetramethylsilane)) δ 7.24-7.11 (m, 4H), 7.05-6.97 (m, 4H), 6.85-6.78 (m, 3H), 3.86 (s, 3H), 3.79 (s , 3H). MS (ESI (Electrospray Ionization)) m / z 370.1 [M + H] ⁺ .

4-Fluoro-N, N-bis (4-methoxyphenyl) -2- (trifluoromethyl) benzamide (2b): colorless oil, yield 84.2%. ¹ H NMR (CDCl ₃ , 300 MHz) δ 7.34-7.26 (m, 4H), 7.09-7.01 (m, 3H), 6.91 (d, 2H, J = 8.7 Hz), 6.87 (d, 2H, J = 8.7 Hz), 3.80 (s, 3H), 3.71 (s, 3H). MS m / z 442.1 (M + Na) ⁺ .

4-Methoxy-N- (4-methoxyphenyl) -N- (4-fluorophenyl) benzamide (2c): white solid, 97% yield. Mp 133.5-134.5 ° C. ¹ H NMR (CDCl ₃ , 300 MHz) 6 8.11-6.66 (m, 15H), 3.74 (s, 3H), 3.73 (s, 3H). MS m / z 384 (M + H) ⁺ .

N- (4-PLEToxyphenyl) -N- (4-benzyloxyphenyl) -2-naphthylamide (2d): white solid, 58% yield. Mp 174.9-175.5 ° C. ¹ H NMR (CDCl ₃ , 300 MHz) δ 8.04 (s, 1H), 7.77-7.74 (m, 2H), 7.64-7.61 (m, 1H), 7.51-7.43 (m, 4H), 7.40-7.31 (m , 4H), 7.13-7.10 m, 4H), 6.88-6.78 (m, 4H), 4.99 (s, 2H), 3.74 (s, 3H). MS m / z 460 (M + H) ⁺ .

4-Fluoro-N, N-bis (4-methoxyphenyl) -2- (trifluoromethyl) benzamide (2e): colorless oil, yield 84.2%. ¹ H NMR (CDCl ₃ , 300 MHz) δ 7.34-7.26 (m, 4H), 7.09-7.01 (m, 3H), 6.91 (d, 2H, J = 8.7 Hz), 6.87 (d, 2H, J = 8.7 Hz), 3.80 (s, 3H), 3.71 (s, 3H). MS m / z 442.1 (M + Na) ⁺ .

General procedure for the demethylation of benzamide derivatives using BBr ₃ . The methoxybenzamide compound was dissolved in anhydrous CH ₂ Cl ₂ . BBr ₃ (1.0 M solution in CH ₂ Cl ₂ ) was added dropwise at 0 ° C. The reaction solution was slowly warmed to room temperature and allowed to stir overnight at room temperature. The mixture was cooled to 0 ° C in an ice bath and hydrolyzed by adding water. To carry out the separation of the solution, EtOAc was added. The organic layer was separated; the aqueous layer was extracted with EtOAc. The organic layer was washed with brine and dried over anhydrous MgSO ₄ . The solvent was removed under reduced pressure. The residue was purified by flash column chromatography using CH ₃ OH / CH ₂ Cl ₂ (1/9 v / v), which allowed the compounds of the corresponding phenols to be obtained.

4-Fluoro-N, N-bis (4-hydroxyphenyl) -2- (trifluoromethyl) benzamide (3a): white solid, yield 92.5%. ¹ H NMR (DMSO-d ₆ , 300 MHz) δ 9.55 (s, 1H), 9.53 (s, 1H), 7.69-7.58 (m, 2H), 7.46-7.39 (m, 1H), 7.18 (d, 2H , J = 8.7 Hz), 6.93 (d, 4H, J = 8.7 Hz), 7.03 (d, 2H, J = 8.4 Hz), 6.78 (d, 2H, J = 8.7 Hz) 6.57 (d, 2H, J = 8.7 Hz). MS m / z 392.1 (M + H) ⁺ .

The following compounds were synthesized as described above, characteristics were determined, and the results were summarized in Table 1: N, N-bis (4-hydroxyphenyl) -4-propylbenzamide (II); 3-fluoro-N- (4-fluorophenyl) -4-hydroxy-N- (4-hydroxyphenyl) benzamide (IV); N, N-bis (4-hydroxyphenyl) -2,3-dimethylbenzamide (V); 3-fluoro-4-hydroxy-N, N-bis (4-hydroxyphenyl) benzamide (VII); 3-fluoro-4-hydroxy-N- (4-hydroxyphenyl) -N-phenylbenzamide (XI); and 3-fluoro-N, N-bis (4-hydroxyphenyl) -2-methylbenzamide (XII).

General methods for the debenzylation of benzyloxyphenylbenzamides. The compound was dissolved in EtOH in a 250 ml hydrogenation vessel. Pd / C powder (5 mol%) was added to the solution. The reaction vessel was installed in a hydrogenation apparatus under a pressure of gaseous hydrogen of 20 psi. inch (138 kPa). The progress of the reaction was monitored using TLC (thin layer chromatography) until the starting material disappeared. Then, the solvent was removed under reduced pressure. The residue was purified by flash column chromatography using hexanes / EtOAc (3/2 v / v), which afforded the desired product.

The following compounds were synthesized as described above, characteristics were determined, and the results were summarized in Table 1: N, N-bis (4-hydroxyphenyl) -2-naphthylamide (VI).

General methods for the recovery of benzamides with deprotected. Benzamide compounds were dissolved in 20 ml of anhydrous THF at room temperature. H ₃ B (SMe ₂ ) was added using a syringe at room temperature in an argon atmosphere. The reaction solution was stirred and heated to reflux for 6 hours. Then the reaction was quenched by adding 10 ml of MeOH at 0 ° C. The solvent was removed under reduced pressure. The residue was subjected to flash column chromatography (silica gel, CH ₂ Cl ₂ / MeOH = 9/1 vol./about.), Which allowed to obtain the desired product.

The following compounds were synthesized as described above, characteristics were determined, and the results were summarized in Table 1: 4.4 '- (2,3-dimethylbenzylazanediyl) diphenol (III); 4 - ((4-fluorophenyl) (4-hydroxybenzyl) amino) phenol (VIII).

General synthesis of O- (2-piperidin-1-ylethoxy) benzamides and analogues. To a solution of a hydroxyphenyl-containing benzamide analog (1 equivalent) in acetone was added K ₂ CO ₃ (3 equivalents) and a salt of N-chloroethylpiperidine hydrochloride (1.2 equivalents). The solution was heated to reflux for 6 hours. The solution was evaporated to dryness. The residue was hydrolyzed by adding water, and then extracted with ethyl acetate. The organic layers were separated and dried over anhydrous MgSO ₄ . The solvent was removed under reduced pressure. The residue was purified by flash chromatography using methylene chloride / methanol (9/1 v / v) to give the desired compound. Hydrochloride salts were prepared by adding HCl in Et ₂ O to a methanol solution of the compounds, followed by evaporation of the solvents.

The following compounds were synthesized as described above, characteristics were determined and the results were summarized in Table 1: 4-fluoro-N- (4-hydroxyphenyl) -N- (4- (2- (piperidin-1-yl) ethoxy) phenyl) -2- (trifluoromethyl) benzamide (IX); and 4-fluoro-N- (4-hydroxyphenyl) -N- (4- (2- (piperidin-1-yl) ethoxy) phenyl) -2- (trifluoromethyl) benzamide hydrochloride (X), which is an HCl salt of the compound IX.

TABLE 1. Physical characteristics of the compounds of formulas II-XII Connection No Structure physical characteristics II

¹ H NMR (DMSO-d ₆ , 300 MHz) δ 9.46 (s, 2H, 2 × OH), 7.27-7.26 (m, 2H, ArH), 7.06-7.04 (m, 2H, ArH), 6.99-6.97 ( m, 4H, ArH), 6.66-6.65 (m, 4H, ArH), 2.50 (s, 2H, CH ₂ overlapping with the DMSO peak), 1.53-1.52 (m, 2H, CH ₂ ), 0.82 (t , J = 7.33 Hz, 3H, CH ₃ ). m / z 346.0 (M-H) ^- . III

Reddish-brown foam, yield 41%. Mp 147-150 ° C. ¹ H NMR (DMSO-d ₆ , 300 MHz) δ 8.92 (s, 2H), 7.07 (d, J = 7.33 Hz, 1 H), 7.00-6.94 (m, 2H), 6.76-6.72 (m, 4H), 6.63-6.59 (m, 4H), 4.72 (s, 2H), 2.23 (s, 3H), 2.16 (s, 3H). m / z 320.2 (M + H) ⁺ . IV

A tan solid, yield 92%. Mp 110-112 ° C. ¹ H NMR (DMSO-d ₆ , 300 MHz) δ 10.14 (bs, 1H), 9.71 (bs, 1H), 7.26-7.11 (m, 5H), 7.05-6.99 (m, 3H), 6.78 (t, J = 8.61 Hz, 2H); 6.68 (d, J = 8.68 Hz, 2H). m / z 364.1 (M + Na) ⁺ . V

¹ H NMR (DMSO-d ₆ , 300 MHz) δ 9.47 (bs, 2H, 2 × OH), 7.18 (d, J = 8.30 Hz, 2H, ArH), 7.06 (d, J = 7.08 Hz , 1H, ArH), 7.00-6.92 (m, 4H, ArH), 6.78 (d, J = 8.30 Hz, 2H, ArH), 6.51 (d, J = 8.06 Hz, 2H, ArH), 2.22 (s, 3H, CH ₃ ), 2.15 (s, 3H, CH ₃ ). m / z 334.3 (M + H) ⁺ . VI

White solid, 70% yield. Mp 264.3-265.2 ° C (with decomposition). ¹ H NMR (DMSO-d ₆ , 500 MHz) δ 9.46 (s, 2H), 7.98 (s, 1H), 7.85-7.75 (m, 2H), 7.75-7.73 (m, 2H), 7.54-7.48 (m , 2H), 7.45-7.43 (m, 1H), 7.05 (s, 4H), 6.66 (s, 4H). m / z 356 (M + H) ⁺ . VII

¹ H NMR (DMSO-d ₆ , 300 MHz) δ 10.25 (bs, 1H, OH), 9.48 (bs, 2H, 2 × OH), 7.12-6.95 (m, 6H, ArH), 6.80-6.65 (m, 5H, ArH). m / z 338.0 (M-H) ^- . VIII

Yellow oil, yield 92%. ¹ H NMR (DMSO-d ₆ , 500 MHz) δ 9.29 (s, 1H), 9.24 (s, 1H), 7.09 (d, 2H, J = 8.3 Hz), 6.98 (d, 2H, J = 9 , 0 Hz), 6.94-6.91 (m, 2H), 6.73 (d, 2H, J = 9.0 Hz), 6.68-6.64 (m, 4H), 4.70 (s, 2H). m / z 307.8 (M-H) ^- . IX and X (HCl salt of compound IX)

White solid, 57.7% yield. ¹ H NMR (DMSO-d ₆ , 300 MHz) δ 9.57 (s, 1H), 7.71-7.68 (m, 2H), 7.47-7.44 (m, 1 H), 7.28 (d, 1H, J = 9.0 Hz), 7.18 (d, 1H, J = 8.7 Hz), 7.13 (d, 1H, J = 8.7 Hz), 7.05 (d, 1H, J = 8.4 Hz), 6.97 (d, 1H , J = 9.0 Hz), 6.80-6.76 (m, 2H), 6.57 (d, 1H, J = 8.7 Hz), 4.06 (t, 1 H, J = 6.0 Hz), 3.93 (t , 1H, J = 6.0 Hz), 2.66 (t, 1 H, J = 5.7 Hz), 2.55 (t, 1H, J = 5.4 Hz), 2.44 (s, 2H), 2.36 (s , 2H), 1.49-1.37 (m, 6H). m / z 501.0 (M-H) ^- . Xi

¹ H NMR (DMSO-d ₆ , 300 MHz) δ 9.95 (bs, 1H, OH), 9.47 (bs, 2H, 2 × OH), 7.02-6.95 (m, 6H, ArH), 6.75-6.72 (m, 1H, ArH), 6.68-6.66 (m, 4H, ArH). m / z 324.0 (M + H) ⁺ . XII

Pale red solid. Yield 72.0%. Mp > 240 ° C. ¹ H NMR (DMSO-d ₆ , 300 MHz) δ 9.50 (bs, 2H), 7.19-6.79 (m, 7H), 6.61 (d, J = 8.93 Hz, 2H), 6.53 (d, J = 7 , 79 Hz, 2H); 2.23 (s, 3H). m / z 336.0 (MH) ^- .

EXAMPLE 2

Synthesis of Compounds of Formula IV (FIG. 6)

Stage 1: synthesis of 4-fluoro-N- (4-methoxyphenyl) aniline (1C)

4-Fluoroaniline (78.63 g; 0.708 mol), 4-iodoanisole (138.00 g; 0.590 mol), anhydrous K ₂ CO ₃ (122.23 g; 0.884 mol), CuI (11.23 g; 58, 96 mmol) and L-proline (13.58 g; 0.118 mol) were mixed together in a 1 L dried three-necked round bottom flask equipped with a stirring bar, reflux condenser and argon inlet. Anhydrous DMSO (300 ml) was added at room temperature. The reaction mixture was stirred and heated to 90 ° C. for 20 hours under argon. Then the mixture was cooled to room temperature and hydrolyzed with water (300 ml). EtOAc (200 ml) was added to effect solution separation. The EtOAc-containing layer was separated. The aqueous layer was extracted with 100 ml EtOAc. EtOAc-containing layers were combined, washed with brine (2 × 100 ml) and dried over anhydrous MgSO ₄ (50 g). The solvent was removed under reduced pressure. The brown oil residue was purified by flash column chromatography (silica gel, hexanes / EtOAc = 9/1 v / v), which afforded the product 4-fluoro-N- (4-methoxyphenyl) aniline (1c) as a yellow solid substances (99.70 g); yield 77.8%. Mp 46-48 ° C. MS (ESI) m / z 218.1 [M + H] ⁺ . ¹ H NMR (DMSO-d ₆ , 300 MHz) δ 7.77 (bs, 1H), 7.03-6.98 (m, 4H), 6.93-6.82 (m, 4H), 3.70 (s, 3H).

Stage 2: synthesis of 3-fluoro-N- (4-fluorophenyl) -4-methoxy-N- (4-methoxyphenyl) benzamide (2a)

4-Fluoro-N- (4-methoxyphenyl) aniline (1c) (90.78 g; 0.418 mol) and 3-fluoro-4-methoxybenzoyl chloride (94.55 g; 0.501 mol) were mixed together and dissolved in anhydrous THF (200 ml) in a dried three-necked round-bottomed flask with a capacity of 1 l, equipped with a stirring rod, a reflux condenser and an argon inlet. Anhydrous pyridine (132.22 g; 1.672 mol) was added using a syringe at room temperature in an argon atmosphere. The reaction mixture was stirred and heated to reflux overnight. Then the reaction mixture was cooled to room temperature and filtered to remove the pyridine salt. The solution was concentrated to remove the THF solvent. The oil residue was washed with 200 ml of 2 N. HCl solution and was extracted with ethyl acetate (2 × 200 ml). The combined organic layer was washed with saturated aqueous Na ₂ CO ₃ (150 ml) to remove excess benzoyl chloride and acid, dried over MgSO ₄ (50 g), filtered and concentrated under reduced pressure to give an oil. The residue was purified by flash column chromatography using silica gel and CH ₂ Cl ₂ / acetone (50/1 v / v), which afforded the pure compound of the corresponding benzamide as a yellow solid. Mp 54-56 ° C. MS (ESI) m / z 370.1 [M + H] ⁺ . ¹ H NMR (CDCl ₃ / TMS) δ 7.24-7.11 (m, 4H), 7.05-6.97 (m, 4H), 6.85-6.78 (m, 3H), 3.86 (s, 3H), 3.79 (s, 3H) .

Stage 3: synthesis of 3-fluoro-N- (4-fluorophenyl) -4-hydroxy-N- (4-hydroxyphenyl) benzamide (IV)

The compound 3-fluoro-N- (4-fluorophenyl) -4-methoxy-N- (4-methoxyphenyl) benzamide (2a) (138.0 g; 0.374 mol) was dissolved in anhydrous CH ₂ Cl ₂ (600 ml) at room temperature (CT) in an argon atmosphere. BBr ₃ (374.75 g; 1.496 mol) was added dropwise using a syringe with stirring at an ice bath in an argon atmosphere. The reaction solution was allowed to stir at room temperature overnight. Then the solution was poured into 1 liter of ice water with stirring. This suspension was stirred at room temperature for 2 hours. The white precipitate was filtered off, washed with water (2 × 100 ml) and dried under vacuum. The CH ₂ Cl ₂ layer was separated, dried over anhydrous MgSO ₄ (50 g), filtered and concentrated under reduced pressure to dryness. The white precipitate and the residue from the CH ₂ Cl ₂ solution were combined and purified by flash column chromatography (silica gel, CH ₂ Cl ₂ / acetone / MeOH = 90/7/3 v / v / v) to give a light tan a solid which was recrystallized twice from a hot EtOAc / hexanes solution to give a white crystalline solid (104.0 g); yield 81.6%. Mp 110-112 ° C. MS (ESI) m / z 364.1 [M + Na] ⁺ . ¹ H NMR (DMSO-d ₆ ) δ 10.14 (bs, 1H), 9.71 (bs, 1H), 7.25-7.11 (m, 5H), 7.05-6.99 (m, 3H), 6.78 (t, J = 8, 6 Hz, 1H); 6.68 (d, J = 8.7 Hz, 2H).

EXAMPLE 3

The synthesis of the compounds of formula VI (Fig.7)

Synthesis of 4- (benzyloxy) -N- (4-methoxyphenyl) aniline (1d) 4-Benzyloxyaniline (16.6 g; 83.31 mmol), 4-iodoanisole (15.0 g; 64.09 mmol), K ₂ CO ₃ (17.72 g; 128.18 mmol), CuI (1.22 g; 6.41 mmol) and L-proline (1.48 g; 12.82 mmol) were mixed together and dissolved in anhydrous DMSO (120 ml) at room temperature. Then the reaction mixture was stirred and heated to 90 ° C for 48 hours. The mixture was cooled to room temperature and hydrolyzed with water. To carry out the separation of the solution, EtOAc was added. The EtOAc-containing layer was separated, washed with brine, dried over anhydrous MgSO ₄ . The solvent was removed under reduced pressure. The solid residue was purified by flash column chromatography (silica gel) using EtOAc / hexanes (1/9 v / v), which afforded the corresponding diarylaniline as a yellow solid (9.8 g); yield 50%. Mp 108.0-108.4 ° C. ¹ H NMR (CDCl ₃ , 300 MHz) δ 7.34-7.25 (m, 5H), 6.90-6.81 (m, 8H), 5.02 (s, 2H), 3.78 (s, 3H). MS m / z 306 (M + H) ⁺ .

Synthesis of N- (4-benzyloxyphenyl) -N- (4-methoxyphenyl) -2-naphthamide (2d) One equivalent of 4- (benzyloxy) -N- (4-methoxyphenyl) aniline (0.80 g; 2.62 mmol) mixed with 1.5 equivalents of 2-naphthoyl chloride (0.75 g; 3.93 mmol) and 4 equivalents of pyridine (Py) (0.83 g; 10.48 mmol) in a dried three-necked round-bottom flask equipped with a magnetic stir bar and reflux condenser. The mixture was dissolved in anhydrous THF (30 ml) and heated to reflux for 20 hours. The reaction solution was cooled to room temperature and filtered. The solvent was removed under reduced pressure. The residue was purified by flash column chromatography using silica gel and EtOAc / hexanes (3/7 v / v), which afforded the pure compound of the corresponding naphthamide as a white solid (0.70 g); yield 58%. Mp 174.9-175.5 ° C. ¹ H NMR (CDCl ₃ , 300 MHz) δ 8.04 (s, 1H), 7.77-7.74 (m, 2H), 7.64-7.61 (m, 1H), 7.51-7.43 (m, 4H), 7.40-7.31 (m , 4H), 7.13-7.10 (m, 4H), 6.88-6.78 (m, 4H), 4.99 (s, 2H), 3.74 (s, 3H). MS m / z460 (M + H) ⁺ .

Synthesis of N, N-bis (4-hydroxyphenyl) -2-naphthylamide (VI)

The compound N- (4-benzyloxyphenyl) -N- (4-methoxyphenyl) -2-naphthamide (2d) (0.50 g; 1.09 mmol) was dissolved in anhydrous CH ₂ Cl ₂ (30 ml) at room temperature. BBr ₃ (3.26 ml of a 1.0 M solution in CH ₂ Cl ₂ ; 3.26 mmol) was added dropwise using a syringe with stirring at room temperature. The reaction solution was allowed to stir overnight at room temperature. The mixture was cooled to 0 ° C in an ice bath and hydrolyzed by adding water. To carry out the separation of the solution, EtOAc was added. The organic layer was separated; the aqueous layer was extracted twice with EtOAc. The organic layers were combined, washed with brine and dried over anhydrous MgSO ₄ . The solvent was removed in vacuo. The residue was purified by flash column chromatography using silica gel and CH ₃ OH / CH ₂ CH ₂ (1/9 v / v), which afforded the desired pure phenol compound as a white solid (0.27 g); yield 70%. Mp 264.3-265.2 ° C (with decomposition). ¹ H NMR (DMSO-d ₆ , 500 MHz) δ 9.46 (s, 2H), 7.98 (s, 1H), 7.85-7.75 (m, 2H), 7.75-7.73 (m, 2H), 7.54-7.48 (m , 2H), 7.45-7.43 (m, 1H), 7.05 (s, 4H), 6.66 (s, 4H). MS m / z356 (M + H) ⁺ .

EXAMPLE 4

Synthesis of Compounds of Formula VIII

Synthesis of 4 - ((4-fluorophenyl) (4-hydroxybenzyl) amino) phenol (VIII)

The compound N- (4-fluorophenyl) -4-hydroxy-N- (hydroxyphenyl) benzamide (0.30 g; 0.93 mmol) was dissolved in 20 ml of anhydrous THF at room temperature. H ₃ B (SMe ₂ ) (1.86 ml of a 2 M solution in THF; 3.71 mmol) was added via syringe at room temperature under argon. The reaction solution was stirred and heated to reflux for 6 hours. Then the reaction was quenched by adding 10 ml of MeOH at 0 ° C. The solvent was removed under reduced pressure. The residue was flash column chromatographed (silica gel, CH ₂ Cl ₂ / MeOH = 9/1 v / v) to give a yellow oil (0.26 g); yield 92%. ¹ H NMR (DMSO-d ₆ , 500 MHz) δ 9.29 (s, 1H), 9.24 (s, 1H), 7.09 (d, 2H, J = 8.3 Hz), 6.98 (d, 2H, J = 9 , 0 Hz), 6.94-6.91 (m, 2H), 6.73 (d, 2H, J = 9.0 Hz), 6.68-6.64 (m, 4H), 4.70 (s, 2H). MS m / z 307.8 (MH) ^- .

EXAMPLE 5 Synthesis of Compounds of Formulas IX and X (FIG. 8)

Synthesis of diarylanilines. Arylamine (1.5 equivalents), aryl iodide (1 equivalent), K ₂ CO ₃ (2 equivalents), CuI (0.1 equivalents) and L-proline (0.2 equivalents) were mixed together and dissolved in anhydrous DMSO at room temperature . Then the reaction mixture was stirred and heated to 90 ° C for 28 hours. The mixture was cooled to room temperature and hydrolyzed with water. To carry out the separation of the solution, EtOAc was added. The EtOAc-containing layer was separated, washed with brine, dried over anhydrous MgSO ₄ . The solvent was removed under reduced pressure. The solid residue was purified by flash column chromatography (silica gel) using EtOAc / hexanes (3/7 v / v) as solvent, which afforded the corresponding diarylaniline. Bis- (4-methoxyphenyl) amine (1a): pale yellow solid; yield 73%. ¹ H NMR (CDCl _3, 300 MHz) δ 6.93-6.81 (m, 8H) , 5.37 (s, br, 1H), 3.78 (s, 6H). MS m / z 228.4 (MH) ⁺ .

Synthesis of 4-fluoro-N, N-bis (4-methoxyphenyl) -2- (trifluoromethyl) benzamide (2e)

1 equivalent of bis- (4-methoxyphenyl) amine (1a) (0.73 g; 3.18 mmol) was mixed with 1.2 equivalents of 4-fluoro-2-trifluoromethylbenzoyl chloride (0.87 g; 3.82 mmol) and 6 pyridine equivalents (1.51 g; 19.08 mmol) in a dried three-necked round-bottom flask equipped with a magnetic stir bar and reflux condenser. The mixture was dissolved in anhydrous THF (20 ml) and heated to 90 ° C. for 20 hours. The reaction solution was cooled to room temperature and filtered. The solvent was removed under reduced pressure. The residue was purified by flash column chromatography using silica gel and EtOAc / hexanes (3/7 v / v), which afforded the pure compound of the corresponding benzamide as a colorless oil (1.12 g); yield 84.2%. ¹ H NMR (CDCl ₃ , 300 MHz) δ 7.34-7.26 (m, 4H), 7.09-7.01 (m, 3H), 6.91 (d, 2H, J = 8.7 Hz), 6.87 (d, 2H, J = 8.7 Hz), 3.80 (s, 3H), 3.71 (s, 3H). MS m / z 442.1 (M + Na) ⁺ .

Synthesis of 4-fluoro-N, N-bis (4-hydroxyphenyl) -2- (trifluoromethyl) benzamide (3a)

The compound 4-fluoro-N, N-bis (4-methoxyphenyl) -2- (trifluoromethyl) benzamide (2e) (1.00 g; 2.38 mmol) was dissolved in anhydrous CH ₂ Cl ₂ (30 ml) at room temperature . BBr ₃ (10 ml of a 1.0 M solution in CH ₂ Cl ₂ ; 10.0 mmol) was added dropwise with a syringe at room temperature. The reaction solution was allowed to stir overnight at room temperature. The mixture was cooled to 0 ° C in an ice bath and hydrolyzed by adding water. To carry out the separation of the solution, EtOAc was added. The organic layer was separated; the aqueous layer was extracted twice with EtOAc. The organic layers were combined, washed with brine and dried over anhydrous MgSO ₄ . The solvent was removed in vacuo. The residue was purified by flash column chromatography using silica gel and CH ₃ OH / CH ₂ Cl ₂ (1/9 v / v), which afforded the desired pure phenol compound as a white solid (0.86 g); yield 92.5%. ¹ H NMR (DMSO-d ₆ , 300 MHz) 59.55 (s, 1H), 9.53 (s, 1H), 7.69-7.58 (m, 2H), 7.46-7.39 (m, 1H), 7.18 (d, 2H, J = 8.7 Hz), 6.93 (d, 4H, J = 8.7 Hz), 7.03 (d, 2H, J = 8.4 Hz), 6.78 (d, 2H, J = 8.7 Hz), 6.57 (d, 2H, J = 8.7 Hz). MS m / z 392.1 (M + H) ⁺ .

Synthesis of 4-fluoro-N- (4-hydroxyphenyl) -N- [4- (2-piperidin-1-yl) ethoxy) phenyl] -2- (trifluoromethyl) benzamide (IX)

To a solution of 4-fluoro-N, N-bis (4-hydroxyphenyl) -2- (trifluoromethyl) benzamide (3a) (0.61 g; 1.56 mmol) in acetone was added K ₂ CO ₃ (1.29 g; 9.36 mmol) and N-chloroethylpiperidine hydrochloride salt (0.34 g; 1.87 mmol). The solution was heated to reflux for 20 hours. The solution was evaporated to dryness. The residue was purified by flash chromatography (silica gel; methylene chloride / methanol = 9/1 v / v) to obtain the desired compound as a white solid (0.45 g); yield 57.7%. ¹ H NMR (DMSO-d ₆ , 300 MHz) δ 9.57 (s, 1H), 7.71-7.68 (m, 2H), 7.47-7.44 (m, 1H), 7.28 (d, 1H, J = 9.0 Hz ), 7.18 (d, 1H, J = 8.7 Hz), 7.13 (d, 1H, J = 8.7 Hz), 7.05 (d, 1H, J = 8.4 Hz), 6.97 (d, 1H, J = 9.0 Hz), 6.80-6.76 (m, 2H), 6.57 (d, 1H, J = 8.7 Hz), 4.06 (t, 1H, J = 6.0 Hz), 3.93 (t, 1H , J = 6.0 Hz), 2.66 (t, 1H, J = 5.7 Hz), 2.55 (t, 1H, J = 5.4 Hz), 2.44 (s, 2H), 2.36 (s, 2H) 1.49-1.37 (m, 6H). MS m / z 501.0 (M-H) ^- .

The hydrochloride salt of compound (X) was obtained by adding HCl in Et ₂ O to a methanol solution of the compounds, followed by evaporation of the solvents.

EXAMPLE 6

Estrogen receptor binding affinities, agonistic and antagonistic activity

The binding affinity of the compounds with ER was determined using a competition binding assay of the radioactive ligand in vitro using natural ligand having a high affinity ER [2,4,6,7- ³ H (N)] - estradiol ^([3 H] E ₂₎ and bacterially an expressed, GST fused protein containing a ligand binding domain (LBD) of an ER-α or ER-β receptor.

Method

Recombinant ER-α or ER-β was combined with [ ³ H] E ₂ to determine the equilibrium dissociation constant (K _d ) for [ ³ H] E ₂ . Protein was incubated with [ ³ H] E ₂ in increasing concentrations in the presence and absence of unlabeled E ₂ in high concentration at 4 ° C for 18 h to determine total and non-specific binding. Non-specific binding data were subtracted and K _d for E ₂ (ERα: 0.71 nM; ERβ: 1.13 nM) was determined using non-linear regression. In addition, the concentration [ ³ H] E ₂ necessary for saturation of ER-α and ER-β-LBD was determined, which turned out to be 4-6 nM.

Compounds in increasing concentrations (range: 10 ^-11 -10 ^-6 M) were incubated with [ ³ H] E ₂ (5.7 nM) and ER-LBD under the conditions described above. After incubation, the material from the plates was collected using GF / B filters using a Unifilter-96 harvester (PerkinElmer) and washed three times with ice-cold buffer B (50 mM Tris; pH 7.2). The filter plates were dried at room temperature, then 35 μl Microscint-O cocktail was added to each well, and the filter plates were tightly closed using TopSeal-A. Radioactivity was counted on a TopCount® NXT microplate scintillation counter using ³ H settings in a Microscint cocktail (PerkinElmer).

Specific binding of [ ³ H] E ₂ for each concentration of compounds was determined by subtracting data for non-specific binding of [ ³ H] E ₂ (determined by incubation with unlabeled E ₂ at a concentration of 10 ^-6 M) and expressing it as a percentage of specific binding in lack of test compound. The concentration of compounds was determined, which leads to a decrease in specific binding of [ ³ H] E ₂ by 50% (IC ₅₀ ). Next, the equilibrium binding constant (K _i ) was calculated for the compounds: K _i = K _d × IC ₅₀ / (K _d + L), where K _d represents the equilibrium dissociation constant [ ³ H] E ₂ (ER-α = 0.71 nM; ER-β = 1.13 nM), and L represents the concentration of [ ³ H] E ₂ (ER-a: 5.7 nM; ER-β: 5.7 nM).

results

Binding assays confirmed that ligands bind to ER-α and ER-β in various concentrations ranging from 3.75 nM to more than 1000 nM, and selectivity varies from a compound that is isoform selective to non-isoform selective. The results for representative compounds are listed in Table 2.

Table 2. Binding Results for Individual Compounds COMPOUNDS K _i (nM) for ER-α K _i (nM) for ER-β

3.75 81.6

3.81 6.44

21.7 15,2

7.13 35.9

9 72

6.06 76.92

13 19

14.79 646.32

fifteen 57

15.12 25.02

Compound IV binds to ERα and ERβ. The binding affinity of Compounds IV to ER was determined using a competitive binding in vitro radioligand assay using natural ER having a high affinity ligand - [2,4,6,7- ³ H (N)] - estradiol ^([3 H] E ₂₎ and a bacterially expressed, GST fused protein comprising a ligand binding domain (LBD) of an ER-α or ER-β receptor. According to this binding affinity analysis (K _i values) of Compound IV with ERα and ERβ were 21.7 ± 1.7 nM (n = 3) and 15.2 ± 4.1 nM (n = 3), respectively. After binding to ER, Compound IV initiates a complex series of molecular events leading to tissue-selective expression or repression of target genes associated with the pharmacological response. In transient transfection assays, Compound IV behaves as an ERα and ERβ agonist, demonstrating higher efficacy in stimulating ERα-mediated transcriptional activation compared to ERβ. While estradiol activates ERα and ERβ, exhibiting 5.1 times greater selectivity for ERα, Compound IV shows 49.0 times greater selectivity for ERα. Thus, the relative transactivation efficiency (normalized to the values for estradiol) of Compound IV is characterized by a 9.7-fold higher selectivity for ERα compared to ERβ. In addition, under the action of Compound IV at concentrations up to 10 μM inclusive, no antagonistic effects were observed with respect to estradiol-stimulated (1 nM) transcriptional activation. Although many steroidal ligands cross-react with other nuclear hormone receptors, the effect of Compound IV is specific for ERα and ERβ. In transcriptional activation assays, cross-reactivity was screened for both agonistic and antagonistic activity of Compound IV against glucocorticoid receptor (GR) isoforms, mineralocorticoid receptor (MR), progesterone receptor (PR), rat androgen receptor (AR) and farnesoid isoforms X receptor (FXR), hepatic X receptor (LXR), peroxisome proliferator activated receptors (PPAR-α and PPAR-γ) and human retinoid X receptor (RXR-α). Compound IV did not show any agonistic or antagonistic activity in any of these assays, which confirms the conclusion that Compound IV does not have functional cross-reactivity against members of this superfamily of nuclear hormone receptors.

EXAMPLE 7

Transactivation by individual compounds

In order to determine whether the compound is an agonist, antagonist, or partial agonist or antagonist, transactivation analyzes were performed for an agonistic and antagonistic mode of action.

Method

Rat estrogen receptors (ER-α and ER-β) were cloned from rat ovary cDNA into the backbone of plasmid vector PCR3.1. To establish the absence of any mutations, sequencing was performed. HEK-293 cells (kidneys of the human embryo) were scattered at 100,000 cells per well in a 24-well plate in Dulbecco's minimum maintenance medium (DMEM) supplemented with 5% purified on activated carbon fetal calf serum (csFBS). Cells were transfected using lipofectamine (Invitrogen, Carlsbad, Calif.) Along with 0.25 μg ERE-LUC (estrogen response element and luciferase constructs; from the English estrogen response element-luciferase), 0.02 μg CMV-LUC (promoter constructs ) cytomegalovirus and luciferase) (Renilla coral luciferase) and 12.5 ng rat ER-α or 25 ng rat ER-β. 24 hours after transfection, cells were treated with compounds at various concentrations or with a combination of compounds and estradiol to determine antagonistic activity. Luciferase assays were performed 48 hours after transfection.

results

Screening of the compounds of this invention in a system for transactivation analysis revealed that the compounds belonged to all three classes, i.e., agonists, antagonists, and partial agonists. An example of an agonist and antagonist is shown in Table 3. The results of transactivation are very well consistent with the results of binding for selectivity towards isoforms.

Table 3 shows the EC ₅₀ and IC ₅₀ values obtained in the transactivation analysis for some of the individual compounds of this invention.

Table 3. The results of the analysis of transactivation (both for agonist and antagonist) under the action of individual compounds according to the invention COMPOUND EC ₅₀ (nM) for ER-α EC ₅₀ (nM) for ER-β IC ₅₀ (nM) for ER-α IC ₅₀ (HM) for ER-β

0.65 40,4 > 1000 > 1000

> 1000 > 1000 2,207 145

EXAMPLE 8

Javanese Macaque Testosterone Suppression

Two-year-old male sexually invasive male Javanese macaques (n = 2) were kept in accordance with USDA (United States Department of Agriculture) guidelines with free access to primate feed and water (with the exception of fasting before an oral dose). Animals once a day for 7 consecutive days orally through a gastric tube received a dose of the Compound of formula IV (30 mg / kg) as a microemulsion in a diluent Tween 80 / deionized water. Blood samples for serum were taken by venipuncture before oral administration of the dose on the 0th (starting point), 2nd, 3rd, 4th, 5th and 6th day. Testosterone and androgen levels were generally quantified using an enzyme-linked immunosorbent assay (EIA), whether or not combined with HPLC (high performance liquid chromatography), respectively. After treatment with a Compound of Formula IV for 6 days, a time-dependent decrease in testosterone and androgen levels in general (testosterone / dihydrotestosterone) became apparent. The compound of formula IV inhibited testosterone levels by 58% and 64% in animal No. 1 and animal No. 3, respectively, relative to the baseline values (see solid lines in FIG. 1, Table 4). Similarly, the level of androgens was generally suppressed by 56% in both animals No. 1 and No. 3 (see dotted lines in Figure 1, Table 4) compared to the initial values.

Consistent with the presence of estrogen feedback in the pituitary-testicular axis in males, these results demonstrate a reliable pharmacological response to the suppression of serum hormones (testosterone and androgens in general) in intact non-human primates (cynomolgus monkeys) after repeated administration oral doses (30 mg / kg) Compounds of formula IV.

Table 4. Levels of testosterone and androgens in general in the blood serum of intact male macaques with daily oral administration of the Compound of Formula IV (30 mg / kg; first dose on day 0) Testosterone Androgens in general Serum Level (pg / ml) Serum Level (pg / ml) Day Animal 1 Animal 3 Animal 1 Animal 3 0 (starting point) 1120 617 1868 1643 2 937 479 1178 847 3 784 437 1078 786 four 552 415 988 924 5 403 276 966 664 6 474 221 819 726 The percentage reduction from the starting point The percentage reduction from the starting point Day Animal 1 Animal 3 Animal 1 Animal 3 0 (starting point) one hundred one hundred one hundred one hundred 2 16 22 37 48 3 thirty 29th 42 52 four 51 33 47 44 5 64 55 48 60 6 58 64 56 56

EXAMPLE 9

Suppression of LH and testosterone hormone levels in rats

To evaluate the effect of Compound IV on LH suppression in intact and orchiectomized (ORX) male rats, an in vivo dose-response study was conducted. In intact and ORX-treated animals, Compound IV at doses of S10 mg / kg / day significantly inhibited LH levels compared to the corresponding controls. (A similar pattern of suppression was observed for FSH levels.) LH suppression led to a steady decrease in testosterone levels below the limit of quantitation (BLOQ), which is 0.08 ng / ml, and a decrease in the masses of the prostate, seminal vesicles and muscle mass of the levator, as they are highly androgen-dependent organs. In intact animals, a dose-dependent decrease in the mass of these target organs was noted, while the mass of seminal vesicles and muscle mass of the levator reached the level of controls corresponding to medical castration. Although the masses of the prostate gland in intact animals were significantly reduced, these values did not reach the level of controls corresponding to medical castration. The results are summarized in Table 6 below.

Materials and methods

Male Sprague-Dawley rats weighing approximately 200 g were maintained in a 12 h light / dark cycle regimen with unlimited access to food (rodent food with a protein content of 16% Teklad Global 2016, Harlan, Madison, WI) and water. The animal protocol has been reviewed and approved by the University of Tennessee's Bioethics Committee.

The product tested in this study was weighed and dissolved in 10% DMSO (Fisher) diluted with PEG300 (Acros Organics, NJ) to obtain appropriate dose formulations. For this study, sixty (60) male Sprague-Dawley rats were randomly weighted and assigned to one of twelve treatment groups (n = 5 animals / group). The groups to be treated are listed in Table 5. Animals were kept in groups of 2-3 animals per cage. The control groups (intact and orchiectomized (ORX)) were diluted daily. Compound IV was administered by subcutaneous injection (200 μl) at doses of 0.3, 1, 3, 10 and 30 mg / kg / day for animals of both intact and ORX-treated groups.

After the administration regimen was administered for 14 days, the animals were killed under anesthesia (ketamine / xylazine, 87:13; mg / kg) and body weight was recorded. In addition, the ventral lobe of the prostate, seminal vesicles and levator were removed, cleaned of foreign tissue and weighed separately. Organ mass was normalized to body weight and expressed as a percentage of intact control. Under isoflurane anesthesia, blood was drawn from the abdominal aorta and left to coagulate. Blood serum was separated by centrifugation and stored at -80 ° C until hormone levels were determined in it. Serum luteinizing hormone (LH) and follicle-stimulating hormone (FSH) concentrations were determined using a Luminex assay to determine rat pituitary hormones (Rat Pituitary Luminex Assay) (Millipore, Billerica, MA) according to the manufacturer's instructions. The lower limit of quantitation (LLOQ) for this assay was 3.2 pg / ml for LH and 32 pg / ml for FSH. Testosterone was measured using an EIA to determine testosterone (Alpco Diagnostics, Salem, NH) with an LLOQ of 0.08 ng / ml. Serum hormone levels below the lower limit of quantification (BLOQ) were discarded in the analyzes of the mean values within the group. In view of this, the reported value for LH and T (testosterone) in groups with BLOQ samples will be higher than the true value. This assay method provides a deliberately underestimated estimate of LH and T suppression. To compare groups that received individual doses with intact groups and ORX-treated control groups that received diluent, the Fisher test was used to determine statistically significant differences. Statistical significance was determined a priori as a value of P <0.05.

Table 5. Processed Groups Group Gonadal status Dose (mg / kg / day) Product under test one Intact - Diluent 2 ORX - Diluent 3 Intact 0.3 Compound IV four Intact one Compound IV 5 Intact 3 Compound IV 6 Intact 10 Compound IV 7 Intact thirty Compound IV 8 ORX 0.3 Compound IV 9 ORX one Compound IV 10 ORX 3 Compound IV eleven ORX 10 Compound IV 12 ORX thirty Compound IV

Luteinizing hormone levels in intact and ORX-treated rats (Table 6)

LH levels (mean ± SD (standard deviation)) in the intact and ORX-treated diluent control groups were 1.46 ± 0.64 and 11.1 ± 3.9 ng / ml, respectively. Compound IV dose-dependently reduced LH levels in intact animals with statistically significant decreases when daily doses of ≥ 3 mg / kg were used. LH levels in intact Compound IV-treated animals were 0.863 ± 0.384, 0.704 ± 0.530, 0.395 ± 0.302, 0.226 ± 0.165 and 0.236 ± 0.176 ng / ml after doses of 0.3, 1, 3, 10 and 30 mg / kg / day respectively. LH levels in ORX-treated males also decreased significantly as a result of treatment with Compound IV. In ORX-treated animals, LH levels were 15.4 ± 2.9, 13.5 ± 2.2, 6.5 ± 5.6, 0.425 ± 0.135 and 0.368 ± 0.119 ng / ml after doses of 0.3, 1, 3, 10 and 30 mg / kg / day, respectively. The results are presented graphically in FIG. 10A.

Follicle-stimulating hormone levels in intact and ORX-treated rats (Table 6)

Serum FSH levels in the intact and ORX-treated diluent control groups were 20.9 ± 8.5 and 93.5 ± 13.8 ng / ml, respectively. In intact animals, Compound IV dose-dependently reduced FSH levels to obtain statistically significant decreases observed at doses of ≥ 10 mg / kg / day. FSH levels in intact Compound IV-treated animals were 17.3 ± 6.4; 15.7 ± 7.3; 18.4 ± 7.7; 9.2 ± 4.0 and 6.3 ± 1.8 ng / ml after doses of 0.3, 1, 3, 10 and 30 mg / kg / day, respectively. In ORX-treated animals, LH levels were 115 ± 17, 114 ± 22, 65.2 ± 31.9, 27.6 ± 8.2 and 15.1 ± 4.1 ng / ml after doses of 0.3, 1, 3, 10 and 30 mg / kg / day, respectively. The results are presented graphically in FIG. 10B.

Testosterone levels in intact and ORX rats

Serum testosterone levels in intact diluent-treated control groups were 2.4 ± 1.1 ng / ml. The lower limit of quantification for T was 0.08 ng / ml. Values less than 0.08 ng / ml are considered as values below the limit of quantification (BLOQ). In intact animals, the Compound of Formula IV dose-dependently reduced T levels with a significant decrease observed at doses of ≥ 3 mg / kg per day. Testosterone levels in intact animals treated with the Compound of Formula IV were 2.6 ± 1.7, 1.6 ± 1.0, 0.7 ± 0.4, BLOQ and BLOQ ng / ml after doses of 0.3, 1 , 3, 10 and 30 mg / kg per day, respectively. In ORX-treated animals, T levels were BLOQ for all groups treated with Compound IV and the group treated with diluent. Results for intact animals are shown graphically in FIG. 10C (and FIG. 2) (for graphical purposes, BLOQ values are represented by the limit of quantification).

The fast and effective inhibition of serum testosterone in intact male rats, measured after administration of Compound IV at doses of 3 mg / kg, 10 mg / kg and 300 mg / kg after 24 hours, 72 hours and 168 hours, is shown in FIG. 9.

Organ masses (Table 6)

The masses of the prostate gland, seminal vesicles and the muscle mass of the levator were measured to confirm the suppression of T. Organ masses (mean ± SD) are shown in Figures 10D, 10E and 10F, respectively. Dose-dependent reductions in the mass of the prostate, seminal vesicles and muscle mass of the levator were observed in intact animals treated with Compound IV. Prostate masses in intact animals were 84.0 ± 19.2, 75.2 ± 20.7, 68.2 ± 8.1, 45.1 ± 20.0 and 43.6 ± 8.8% after dosing 0.3, 1, 3, 10 and 30 mg / kg / day, respectively. Prostate masses in ORX-treated animals were 19.0 ± 4.2, 17.4 ± 3.4, 19.6 ± 6.7, 22.9 ± 5.4 and 20.6 ± 2.1% after administration doses of 0.3, 1, 3, 10 and 30 mg / kg / day, respectively. The mass of seminal vesicles in intact animals was 76.2 ± 7.8, 66.3 ± 27.2, 51.8 ± 28.5, 19.1 ± 7.0 and 17.9 ± 3.3% after dose administration 0.3, 1, 3, 10 and 30 mg / kg / day, respectively. The masses of seminal vesicles in animals subjected to ORX were 12.2 ± 1.3, 16.6 ± 5.4, 16.5 ± 4.8, 13.3 ± 1.9 and 12.9 ± 2.1% after administration doses of 0.3, 1, 3, 10 and 30 mg / kg / day, respectively. The levator masses in intact animals were 86.9 ± 10.0, 82.1 ± 12.1, 65.2 ± 4.4, 57.8 ± 11.2 and 58.1 ± 4.7% after doses of 0 , 3, 1,3, 10 and 30 mg / kg / day, respectively. The levator masses in animals subjected to ORX were 54.5 ± 6.6, 49.6 ± 7.0, 53.6 ± 10.0, 51.1 ± 4.9 and 49.2 ± 4.2% after dosing 0.3, 1, 3, 10 and 30 mg / kg / day, respectively.

Data on LH suppression and organ mass reduction are summarized in Table 6.

Table 6. The effect of in vivo Compounds of formula IV on serum hormones and organ mass Gonadal status Compound Dose (mg / kg per day) LH (ng / ml) FSH (ng / ml) Predstat. iron (% of intact.) Seminal vesicles (% of intact.) Levator (% of intact.) Intact Diluent - SD average 1.46 ^b
0.642 20.9 ^b
8.49 100.0 ^b
28.6 100.0 ^b
13,4 100.0 ^b
4.97 ORX Diluent - SD average 11.1 ^a
3.87 93.5 ^a
13.8 13.7 ^a
2,56 14.0 ^a
2.93 58.8 ^a
6.62 Intact Compound IV 0.3 SD average 0.863 ^b
0.384 17.3 ^a
6.44 84.0 ^b
19.2 76.2 ^{a, b}
7.83 86.9 ^{a, b}
10 Intact Compound IV one SD average 0.704 ^b
0.53 15.7 ^b
7.26 75.2 ^b
20.7 66.3 ^{a, b}
27,2 82.1 ^{a, b}
12.1 Intact Compound IV 3 SD average 0.395 ^{a, b}
0.302 18.4 ^b
7.72 68,2 ^{a, b}
8.12 51.8 ^{a, b}
28.5 65.2 ^a
4.35 Intact Compound IV 10 SD average 0.226 ^{a, b}
0.165 9.25 ^{a, b}
3.97 45.1 ^{a, b}
twenty 19.1 ^a
6.98 57.8 ^a
11,2 Intact Compound IV thirty SD average 0.236 ^{a, b}
0.176 6.25 ^{a, b}
1.82 43.6 ^{a, b}
8.75 17.9 ^a
3.33 58.1 ^a
4.71 ORX Compound IV 0.3 SD average 15.4 ^a
2.94 116 ^a
17,2 19.0 ^{a, b}
4.19 12.2 ^a
1.31 54.5 ^a
6.56 Gonadal status Compound Dose (mg / kg per day) LH (ng / ml) FSH (ng / ml) Predstat. iron (% of intact.) Seminal vesicles (% of intact.) Levator (% of intact.) ORX Compound IV one SD average 13.5 ^a
2.18 114 ^a
22.3 17.4 ^a
3.4 16.6 ^a
5.36 49.6 ^a
7.04 ORX Compound IV 3 SD average 6.5
5.63 65.2 ^a
31.9 19.6 ^a
6.67 16.5 ^a
4.82 53.6 ^a
10 ORX Compound IV 10 SD average 0.425 ^{a, b}
0.135 27.6 ^b
8.16 22.9 ^{a, b}
5.44 13.3 ^a
1.91 51.1 ^a
4.88 ORX Compound IV thirty SD average 0.368 ^{a, b}
0.119 15.1 ^b
4.11 20.6 ^{a, b} 2.08 12.9 ^a
2.14 49.2 ^{a, b}
4.21 ^and P <0.05 compared with intact, receiving a diluent. ^b P <0.05 compared with subjected to ORX treated with diluent.

EXAMPLE 10

Recovery of testosterone levels after suppression by Compound IV in rats and macaques

The reversibility of chemical castration by Compound IV was investigated.

Materials and methods

Thirty-five (35) male Sprague-Dawley rats weighing approximately 200 g were maintained on a 12-hour light / dark cycle with unlimited access to food (rodent food with 16% protein Teklad Global 2016, Harlan, Madison, WI) and water . The animal protocol has been reviewed and approved by the University of Tennessee's Bioethics Committee.

The product tested in this study was weighed and dissolved in PEG300 (100%) (Acros Organics, NJ) to obtain appropriate dose formulations. Animals were randomly assigned to one of ten treatment groups (n = 5 animals / group). The groups to be treated are listed in Table 7. Animals were kept in groups of 2-3 animals per cage. Group 1 animals were euthanized at the start of the study (day 1) to determine baseline testosterone levels in intact animals. Animals of groups 2–7 received daily doses of 1, 3, or 30 mg / kg (~ 200 μl) orally through a gastric tube for three days. Animal groups 2, 3, and 4 were euthanized on day 4 to measure maximum testosterone suppression. Animals of groups 5, 6 and 7 were left to recover during the washout period (14 days) without drug administration.

Table 7. Processed Groups Group Compound IV, p / o dose Treatment Group 1 - Starting point Group 2 1 mg / kg for 3 days No recovery Group 3 3 mg / kg for 3 days No recovery Group 4 30 mg / kg for 3 days No recovery Group 5 1 mg / kg for 3 days 14 day recovery Group 6 3 mg / kg for 3 days 14 day recovery Group 7 30 mg / kg for 3 days 14 day recovery

results

Serum testosterone levels in intact rats were 6.4 ± 3.1 ng / ml (mean ± SD) at baseline. Compound IV, administered at doses of 3 and 30 mg / kg for three days, significantly inhibited serum testosterone levels to 1.47 ± 0.26 and 1.62 ± 0.49 ng / ml, respectively. No significant suppression was observed in animals that received Compound IV at a dose of 1 mg / kg for three days. Most importantly, serum testosterone levels were 3.3 ± 1.92, 3.00 ± 1.06 and 3.8 ± 1.72 ng / ml in animals that received Compound IV at a dose of 1 for three days , 3 or 30 mg / kg, respectively, according to the results of measurements after 14 days of the recovery period, and there were no statistically significant differences from the concentrations of testosterone in serum at the starting point in intact rats, as shown in Fig.23.

This study confirms previous results showing that Compound IV rapidly suppresses serum testosterone levels in intact male rats. The inventors observed suppression of serum testosterone levels in groups receiving a dose of ≥ 3 mg / kg / day for 3 days. A significant decrease in serum testosterone levels was not observed for the group with a dose of 1 mg / kg. However, after 14 days of recovery, serum testosterone levels returned to the level of intact controls. This study shows that pharmacological castration in rats under the action of Compound IV is reversible.

The effect of Compound IV on the suppression and restoration of testosterone levels in intact male macaques was evaluated in combination with an oral pharmacokinetic study. Three processed, previously untreated male Cynomolgus monkeys (aged 2–3 years) were administered Compound IV daily at a dose of 30 mg / kg orally through a gastric tube for the next 7 days. Blood samples were collected and separated to obtain serum and plasma for quantitative measurements of testosterone and Compound IV, respectively. The results show that daily administration of oral doses of Compound IV significantly reduced the levels of circulating androgens (mainly testosterone and dihydrotestosterone) in all three macaque males by up to 47% inclusive compared to baseline levels (levels of 1591 ± 72.5, 997 ± 104 and 852 ± 136 ng / ml, respectively, for the starting point, on the 2nd day and 6th day of treatment (mean ± SEM (standard error of the mean))). After an 18-day recovery period without drug administration, androgen levels returned to normal and did not significantly differ from baseline levels before treatment (1757.7 ± 369.5 ng / ml after recovery).

EXAMPLE 11

Preservation of bone mass despite a decrease in LH and testosterone levels in rats (Table 8)

The effect on bone caused by treatment with a Compound of Formula IV was investigated. An orally administered Compound of Formula IV completely prevented bone loss associated with suppression of LH in intact male rats. A significant decrease in LH was induced by the Compound of Formula IV in intact animals at dose levels of ≥ 10 mg / kg per day. Although at a dose of 1 mg / kg per day, the Compound of Formula IV did not significantly reduce LH, a significant decrease in the mass of the prostate gland, seminal vesicles and muscle mass of the levator was evident at this dose, indicating that the decrease in the level of circulating testosterone was physiologically relevant to to these androgen-sensitive organs. However, administration of a Compound of Formula IV at a dose of 1 mg / kg per day maintained trabecular bone volume (measured in the distal femur) at the level of intact controls. When administered at doses of 10 and 30 mg / kg per day, the Compound of Formula IV increased bone volume in the distal femur significantly more than in intact controls. These data show that Compound IV increases the mineral density of the trabecular bone (BMD) and the percentage of bone volume at dose levels that lower LH levels in intact rats. The data from this study are presented in Table 8.

Table 8. The in vivo effect of the Compounds of formula IV on bones, organ mass and hormonal parameters of blood serum in rats Gonadal status Compound Dose (mg / kg per day) Mineral. bone density (g / cm ³ ) Percentage, bone volume (BV / TV) (%) Predstat. iron (% of intact.) Semenn. vesicles (% of intact.) Levator (% of intact.) FSH (ng / ml) LH (ng / ml) Intact Diluent - Avg SD 0.274 ^b
0,033 20,2 ^b
3.57 100.0 ^b
11.1 100.0 ^b
15.9 100.0 ^b
11.6 9.93 ^b
2.94 0.781 ^b
0.263 ORX Diluent - Avg SD 0.224 ^a
0,025 15.4 ^a
2.6 14.8 ^a
4.08 10.3 ^a
0.767 59.4 ^a
7.26 117 ^a
40,2 22.0 ^a
5.81 Intact Compound IV one Avg SD 0.273 ^b
0.04 20,0 ^b
4.08 69.2 ^{a, b}
13.5 44.6 ^{a, b}
15.7 80.0 ^{a, b}
6.69 14.1 ^{a, b}
4.07 0.820 ^b
0.392 Intact Compound IV 10 Avg SD 0.326 ^{a, b}
0,048 25.9 ^{a, b}
4.76 30.7 ^{a, b}
12,4 12.8 ^{a, b}
0.886 58.1 ^a
9.68 5.48 ^{a, b}
1.97 0,060 ^{a, b}
0,092 Intact Compound IV thirty Avg SD 0.326 ^{a, b}
0,046 25.5 ^{a, b}
4.49 30.1 ^{a, b}
17.4 14.4 ^{a, b}
1.45 56.1 ^a
4.67 6.32 ^{a, b}
3.4 0,078 ^{a, b}
0.114 ^and P <0.05 compared with intact, receiving a diluent. ^b P <0.05 compared with subjected to ORX treated with diluent.
BV / TV = the ratio of the volume of the (trabecular) bone to the total volume (bone) (from the English bone volume / total volume).

EXAMPLE 12

Effect on the activity of the enzyme 17β-hydroxysteroid-dehydrogenase type 5 (17β-HSD5)

Members of the HSD family are involved in the conversion of circulating steroids. 17β-HSD5 converts androstenedione to testosterone, and estrone to estradiol. In addition, she is also involved in the synthesis of prostaglandins. Here, the ability of certain individual compounds of the invention to inhibit the activity of 17β-HSD5 has been demonstrated.

Method

Human 17β-HSD5 was cloned into pGEX 4t1 vector and purified protein was obtained. The purified protein was incubated together with a representative compound of the invention, ¹⁴ C-androstenedione and NADPH (reduced form of nicotinamide adenine dinucleotide phosphate) in an appropriate buffer. The synthesized testosterone was extracted using ethyl acetate, dried in air, applied to a thin layer chromatography plate (TLC) and separated. The TLC plate was exposed on a phospho-imager screen and the intensity of the testosterone band was quantified. Indomethacin was used as a positive control (LHRH agonist).

results

The tested Compound IV had a partial inhibitory effect on the activity of the 17β-HSD5 enzyme. Positive control - indomethacin (LHRH agonist), as expected, showed strong inhibition of this enzyme, as shown in Figure 3.

EXAMPLE 13

Toxicity studies

To compare the coagulation potential of Compound IV and diethylstilbestrol (DES, positive control), a study was performed using an in vitro analysis of human platelet aggregation. In this study, the blood of healthy male donors was used, since men are the contingent for which Compound IV treatment (LH suppression) is intended. Platelet-rich blood plasma was preincubated with each of the following: estradiol (E2), DES, Compound IV or diluent for 30 seconds and then thrombin (0.3 units) was added to induce platelet aggregation. The results of the study show that pre-incubation with DES increases thrombin-induced platelet aggregation by approximately 10 times. However, Compound IV and estradiol reduce aggregation in platelet-rich blood plasma. These data demonstrate that Compound IV reduces the in vitro reactivity of human platelets compared with DES, and suggests that Compound IV may have lower thromboembolic potential than DES (Figure 4).

EXAMPLE 14

The effect of Compound IV on the "tides"

A study was made of the effect of Compound IV on hot flashes using the morphine-dependent rat model (MD model) developed by Simpkins et al. (1983), for which a number of similarities with menopausal hot flashes have been shown. In addition to resembling human nature, this experimental animal model has a short lead time, making it a useful tool for high throughput screening using tail skin temperature (TST) measurements to identify compounds that can alleviate vasomotor symptoms. TST-sensors TA-40 (Data Sciences International, MN) were attached with adhesive tape to the base of the tails and the initial temperature was recorded for 15 minutes. After 15 minutes, the animals were treated with naloxone (1 mg / kg, SQ) to reverse the effects of morphine. During the experiment, tail skin temperature (TST) was measured for one hour after treatment with naloxone with a measurement frequency of 5 s. After collecting the data, a moving average of the temperature recorded for every 60 seconds for each animal was calculated and subjected to subsequent analysis. The initial temperature was calculated as the average of 15 minutes prior to the introduction of naloxone. The area under the curve (AUC) was calculated by subtracting all values after administration of naloxone from the initial value using the linear trapezoidal method.

Compound IV attenuated hot flashes in the morphine withdrawal model (see FIG. 13) with best results for 10 mg of Compound IV. 17β-E2 was used at a concentration of 5 mg / kg in 100% DMSO.

EXAMPLE 15

Comparison of Compound IV and DES in Rat

Prior to the administration of LHRH agonists, testosterone levels corresponding to medical castration were achieved, increasing estrogenic activity in the pituitary gland by means of estrogens, primarily diethylstilbestrol (DES). DES was as effective as LHRH agonists in inhibiting testosterone to levels consistent with medical castration. Patients treated with DES did not have hot flashes or bone loss, but gynecomastia was observed to a greater extent than with ADT using LHRH agonists. Unfortunately, the use of highly effective pure estrogens like DES and estradiol is very often associated with a high risk of serious cardiovascular and thromboembolic complications that limit their clinical use.

It was hypothesized, but not proven, that the increased risk of developing venous thromboembolic complications as a result of DES is associated with its cross-reactivity with receptors of other hormones. In vitro studies using human platelets have shown that Compound IV has much less procoagulating activity than DES. Thus, Compound IV, a selective ER-alpha agonist, against prostate cancer can provide the beneficial effects of DES and also provide the beneficial effects of the LHRH agonist without causing osteoporosis or unfavorable lipid profiles.

Compound IV is as effective as DES in reducing the size of the prostate in rats (FIG. 11A) and gives a moderate increase in the size of the prostate in subjected to ORX rats (FIG. 11B).

The differences between DES and Compound IV are shown in FIGS. 12A-12C, where DES is seen to cross-react with a glucocorticoid receptor (GR) (FIG. 12A) and androgen receptor (AR) (FIG. 12B), while Compound IV is not . In addition, DES antagonizes the transactivation of estrogen receptor related receptors (ERRs), while Compound IV is not. Compound IV showed no cross-reaction with any of the three ERR isoforms (ERR-α, ERR-β and ERR-γ), as shown in Fig. 12C.

EXAMPLE 16

Macaque toxicity study - 90 days

Javanese macaques living in families were obtained from Mauritius. A prospective study was planned to obtain a pharmacological and toxicological evaluation of the results of the oral administration of Compound IV and a positive control (LHRH agonist) for 39 weeks on male cynomolgus monkeys with an intermediate period of 13 weeks. A total of 39 sexually mature male macaques aged 5–8 years were randomly assigned to five groups before treatment. The groups included: 1) diluent as a control, 2) Compound IV at a dose of 1 mg / kg, 3) Compound IV at a dose of 10 mg / kg, 4) Compound IV at a dose of 100 mg / kg and 5) positive control (LHRH agonist ) The drug was delivered orally using a cage-side administration once daily for 39 weeks, with diluent as a control (Tween 80 / PRANG ™) for Groups 1 and 5 or Compound IV in a diluent for Groups 2 , 3 and 4. Dose levels of Compound IV were 1, 10 and 100 mg / kg / day for Groups 2, 3 and 4, respectively. Oral doses were delivered in a dose volume of 10 ml / kg calculated on the basis of the most recent measurement of body weight for each animal (Fig. 14). Animals in Group 5, in addition, received a once-daily subcutaneous injection of a positive control (LHRH agonist) (constantly in a volume of 0.02 ml) during the study period of 39 weeks. The general appearance and clinical symptoms were noted and recorded daily. Conventional assessments and some other studies were performed as indicated in the study protocol. Sample parameters include, but are not limited to, testosterone, prostate antigen (PSA), and volume and mass of the prostate.

Testosterone levels and total PSA levels were quantified in serum samples (following the standard procedure) using enzyme-linked immunosorbent assay (EIA) and chemiluminescent immunoassay (LIA, ALPCO Diagnostics, Salem, NH), respectively. Blood samples for assessing the level of testosterone were taken from all animals (on an empty stomach) at the initial moment (i.e., before treatment) and on the 1st, 3rd, 7th, 14th, 28th, 64th and the 90th day. Blood samples for determining PSA were taken from all animals (on an empty stomach) at baseline and during the 6th week. To conduct the discussion, the results for samples with concentrations below the limit of quantitation (BLOQ) in the testosterone and PSA analyzes were taken into account as ½ of the lower limit of quantification (LLOQ) in the analysis and were considered as “estimated final concentrations”. The data shown in Tables 9-16 are presented as “only quantifiable concentrations” (that is, BLOQ values are excluded) in addition to “estimated final concentrations” (that is, BLOQ results are included as ½ LLOQ analysis). Prostate volume was measured on live animals under anesthesia using a transrectal ultrasonography (TRUS) procedure at baseline and at week 6. The width and height of the prostate were recorded. Prostate volume was calculated as (width × width × height × π / 6) and normalized to body weight. The wet mass of the prostate was recorded at autopsy after cleansing the tissue from fat and foreign tissue.

Results and discussion

Serum testosterone levels are shown in FIG. 15 and Tables 9-12. At the starting point, testosterone levels for all macaques in this study were in the range typical for mature adult male Javanese macaques. However, testosterone levels decreased significantly in macaques receiving Compound IV at a dose of 100 mg / kg / day, and in macaques treated with a positive control (LHRH agonist). A biphasic change in testosterone levels in the positive control group (LHRH agonist) was demonstrated, initially with a significant increase (ie, “acute” reaction) of 47.4% and 547% (p <0.01) by the 1st and 3rd day, respectively, then with a decrease of 3.6%, 67%, 73%, 83% and 85% on the 7th, 14th, 28th, 64th and 90th days (see Fig. 15 and Tables 9-12). A similar "acute" reaction was not observed for any animal treated with Compound IV even with the highest dose level (i.e., 100 mg / kg / day). The dose and duration of treatment were important for the pharmacological action of Compound IV, while doses of 100 mg / kg / day suppressed serum testosterone levels by 60%, 51%, 42%, 79% and 92% on the 3rd, 7th , 14th, 28th and 64th days, respectively, relative to the value of the starting point (See Fig. 15 and Tables 9 and 10). After 90 days of treatment with Compound IV (100 mg / kg / day), testosterone levels in 6 out of 10 Group 4 macaques decreased to concentrations below the quantification limit of this assay (reference to Table 11). The average serum testosterone level of Group 4 macaques was reduced by 96% compared to the corresponding initial values ("estimated final concentrations", i.e. testosterone levels for 6/10 macaques with BLOQ values are calculated as 50% of the LLOQ concentration, cm .Table 10). It is important to note that on the 90th day, Compound IV at a dose of 100 mg / kg / day reduced serum testosterone levels to significantly lower levels than the positive control (LHRH agonist) (p = 0.013).

Table 9. Average serum testosterone levels (ng / ml) in intact male macaques after daily oral administration of Compound IV; ^@ estimated final concentrations Day Control - thinner Compound IV 1 mg / kg Compound IV 10 mg / kg Compound IV 100 mg / kg Positive control (LHRH agonist) Average Sem N Average Sem N Average Sem N Average Sem N Average Sem N 0 (original dot) 6.1 1,2 10 7.3 1,0 6 4.9 0.6 6 4.4 0.6 10 4.9 0.9 7 one 8.0 1.7 10 eleven 1,6 6 7.6 1,1 6 8.0 2.2 10 7.2 0.8 7 3 8.2 2,3 10 7.4 1,2 6 5.1 1,1 6 1.8 * 0.5 10 32 ^{# $} 3.8 7 7 5.9 1,2 10 6.7 0.8 6 7.7 1.9 6 2.2 * 0.7 9 4.7 2.6 7 fourteen 3.4 0.5 10 3.8 0.4 6 7.1 1,6 6 2.6 0.9 9 1.6 ^# 0.2 7 28 3.8 0.6 10 4.7 0.9 6 9,4 2.1 6 0.9 * 0.2 10 1.3 ^# 0.2 7 64 5.1 1,1 10 4.3 0.6 6 5,4 1,5 6 0.3 * 0.1 9 0.8 ^{# $} 0.2 7 90 3.6 0.6 9 4.2 0.6 four 4.6 1,0 5 0.2 * 0,0 10 0.8 ^{# $} 0.2 7 LLOQ in testosterone analysis = 0.246 ng / ml; ^@ BLOQ values calculated as 0.123 ng / ml, half of LLOQ.
*: Statistically significant (p <0.05) results for Compound IV (100 mg / kg) compared with the diluent control.
#: Statistically significant (p <0.05) results for the positive control (LHRH agonist) compared to the diluent control.
$: Statistically significant (p <0.05) results for a positive control (LHRH agonist) compared to Compound IV (100 mg / kg).

Table 10. Percentage change (%) in average serum testosterone levels compared to baseline; ^@ estimated final concentrations Day Control - thinner Compound IV 1 mg / kg Compound IV 10 mg / kg Compound IV 100 mg / kg Will put. the control one 31 44 54 82 47 3 35 1.8 3,5 -60 547 7 -3.2 -8.1 57 -51 -3.6 fourteen -44 -48 45 -42 -67 28 -38 -35 92 -79 -73 64 -16 -41 eleven -92 -83 90 -42 -42 -5.5 -95 -85 LLOQ in testosterone analysis = 0.246 ng / ml; ^@ BLOQ values calculated as 0.123 ng / ml, half of LLOQ.

Table 11. Average serum testosterone levels (ng / ml) in intact male macaques after daily oral administration of Compound IV; ^λ only quantifiable concentrations Day Control - thinner Compound IV 1 mg / kg Compound IV 10 mg / kg Compound IV 100 mg / kg Positive control (LHRH agonist) Average Sem N Average Sem N Average Sem N Average Sem N Average Sem N 0 (original dot) 6.1 1,2 10 7.3 1,0 6 4.9 0.6 6 4.4 0.6 10 4.9 0.9 7 one 8.0 1.7 10 eleven 1,6 6 7.6 1,1 6 8.0 2.2 10 7.2 0.8 7 3 8.2 2,3 10 7.4 1,2 6 5.1 1,1 6 1.8 0.5 10 32 3.8 7 7 5.9 1,2 10 6.7 0.8 6 7.7 1.9 6 2.2 0.7 9 4.7 2.6 7 fourteen 3.4 0.5 10 3.8 0.4 6 7.1 1,6 6 2.6 0.9 9 1,6 0.2 7 28 3.8 0.6 10 4.7 0.9 6 9,4 2.1 6 0.9 0.2 10 1.3 0.2 7 64 5.1 1,1 10 4.3 0.6 6 5,4 1,5 6 0.3 0.1 9 0.8 0.2 7 90 3.6 0.6 9 4.2 0.6 four 4.6 1,0 5 0.2 0,0 four 0.8 0.2 7 LLOQ in testosterone analysis = 0.246 ng / ml; ^λ BLOQ values are excluded.

Table 12. Percentage change (%) of average testosterone levels compared to the starting point; ^λ only quantifiable concentrations Day Control - thinner Compound IV 1 mg / kg Compound IV 10 mg / kg Compound IV 100 mg / kg Will put. the control one 31 44 54 82 47 3 35 1.8 3,5 -60 547 7 -3.2 -8.1 57 -51 -3.6 fourteen -44 -48 45 -42 -67 28 -38 -35 92 -79 -73 64 -16 -41 eleven -92 -83 90 -42 -42 -5.5 -95 -85 LLOQ in testosterone analysis = 0.246 ng / ml; ^λ BLOQ values are excluded.

Serum PSA levels were also significantly suppressed by Compound IV four weeks after the start of treatment. A decrease in PSA levels of 69% and 87% (on average) was observed for macaques receiving Compound IV at a dose of 10 mg / kg and 100 mg / kg for 4 weeks, while in the positive control group (LHRH agonist) PSA decreased by 60% (Fig. 16 and Tables 13-16).

Table 13. Mean serum PSA levels (ng / ml) in intact male macaques after daily oral administration of Compound IV; ^@ estimated final concentrations Control - thinner Compound IV 1 mg / kg Compound IV 10 mg / kg Compound IV 100 mg / kg Positive control (LHRH agonist) Average Sem N Average Sem N Average Sem N Average Sem N Average Sem N Before processing. 1,1 0.2 10 1,0 0.2 6 0.8 0.1 6 1,0 0.1 10 1,0 0.1 7 4th week 1,0 0.2 10 0.9 0.2 6 0.3 ^* 0.1 6 0.1 ^& 0.1 10 0.4 ^{# $} 0.1 7 LLOQ in the analysis of PSA = 0.0575 ng / ml; ^@ BLOQ values calculated as 0.02875 ng / ml, half of LLOQ.
*: Statistically significant (p <0.05) results for Compound IV (10 mg / kg) compared with the diluent control.
&: Statistically significant (p <0.05) results for Compound IV (100 mg / kg) compared with the diluent control.
#: Statistically significant (p <0.05) results for the positive control (LHRH agonist) compared to the diluent control.
$: Statistically significant (p <0.05) results for a positive control (LHRH agonist) compared to Compound IV (100 mg / kg).

Table 14. Percentage change (%) in average PSA levels from baseline; ^@ estimated final concentrations The control Compound IV 1 mg / kg Compound IV 10 mg / kg Compound IV 100 mg / kg Positive control (LHRH agonist) 4th week -7.1 -eleven -69 -87 -60 LLOQ in the analysis of PSA = 0.0575 ng / ml; ^@ BLOQ values calculated as 0.02875 ng / ml, half of LLOQ.

Table 15. Mean serum PSA levels (ng / ml) in intact male macaques after daily oral administration of Compound IV; ^λ only quantifiable concentrations Control - thinner Compound IV 1 mg / kg Compound IV 10 mg / kg Compound IV 100 mg / kg Positive control (LHRH agonist) Average Sem N Average Sem N Average Sem N Average Sem N Average Sem N Before processing. 1,2 0.2 9 1,0 0.2 6 0.8 0.1 6 1,0 0.1 10 1,0 0.1 7 4th week 1,1 0.1 9 0.9 0.2 6 0.3 0.1 5 0.3 0.1 four 0.4 0.1 7 LLOQ in the analysis of PSA = 0.0575 ng / ml; ^λ BLOQ values are excluded in this table.

Table 16. Percentage change (%) in average PSA levels from baseline; ^λ only quantifiable concentrations The control Compound IV 1 mg / kg Compound IV 10 mg / kg Compound IV 100 mg / kg Positive control 4th week -7.1 -eleven -64 -72 -60 LLOQ in the analysis of PSA = 0.0575 ng / ml; ^λ BLOQ values are excluded in this table.

Prostate volumes were measured periodically using TRUS throughout the study. Results obtained after six weeks of treatment demonstrate the potent effect of Compound IV and a positive control (LHRH agonist) on the macaque prostate. Compound IV significantly, by 25% and 45%, suppressed prostate volumes at dose levels of 10 mg / kg and 100 mg / kg, respectively, while in the positive control group (LHRH agonist), prostate volumes decreased by 28% (FIG. 17 and Tables 17 and 18).

Table 17. Average prostate volumes (in relative units) in male macaques after daily oral administration of Compound IV Control Thinner Compound IV 1 mg / kg Compound IV 10 mg / kg Compound IV 100 mg / kg Positive control (LHRH agonist) Average Sem N Average Sem N Average Sem N Average Sem N Average Sem N 6th week 438 78 10 468 78 6 327 33 6 242 28 10 315 47

Table 18. Percentage change (%) in average volumes of the prostate compared to the starting point The control Compound IV 1 mg / kg Compound IV 10 mg / kg Compound IV 100 mg / kg Positive control 6th week 0 6.8 -25 -45 -28

A reduction in prostate volume associated with Compound IV was confirmed by an autopsy evaluation of prostate mass. After thirteen weeks of treatment with Compound IV, significantly, by 24% and 21%, the average masses of the prostate gland in animals receiving 10 and 100 mg / kg / day decreased, respectively (Fig. 18B and Tables 19 and 20).

Table 19. The average mass of the prostate (in grams) at dissection in macaques with daily oral administration of Compound IV Control - thinner Compound IV 1 mg / kg Compound IV 10 mg / kg Compound IV 100 mg / kg Average Sem N Average Sem N Average Sem N Average Sem N 13th week 1.8 0.2 3 1.8 0.4 3 1.3 0.1 3 1.4 0.1 3

Table 20. Percentage change (%) of average prostate mass compared to baseline The control Compound IV 1 mg / kg Compound IV 10 mg / kg Compound IV 100 mg / kg 13th week 0 1.7 -24 -21

No pronounced effects on platelet aggregation, prothrombin time (RT) or activated partial thromboplastin time (ARTT) were observed.

EXAMPLE 17

Compound IV Studies in Humans

A study was conducted to determine the effect of Compound IV on men. Studies were performed on 12 subjects per group using dosages of Compounds IV of 100, 300, 600 and 1000 mg. Table 21 presents the average changes in the levels of LH, PSA, free testosterone and total testosterone in the blood serum of men after administration of Compound IV in dosages of 100, 300, 600 and 1000 mg. Dose-dependent mean levels of total testosterone (nmol / L) in humans were measured between 1 and 11 days (Figure 19). The level of total testosterone decreased by 51.9% and 47.9% for dosages of 600 mg and 1000 mg, respectively.

Dose-dependent mean levels of LH (IU / L) in humans were measured between 1 and 10 days (Figure 20). LH levels increased by 20.7%, 46.9%, 27.6% and 29.2% for dosages of 100 mg, 300 mg, 600 mg and 1000 mg, respectively.

Dose-dependent mean levels of free testosterone (pg / ml) in humans were measured between 1 and 10 days (Figure 21). Free testosterone levels decreased by 17.0%, 18.5%, 72.7% and 53.2% for dosages of 100 mg, 300 mg, 600 mg and 1000 mg, respectively.

Dose-dependent mean PSA levels (μg / L) in humans were measured between 1 and 10 days (Figure 22). PSA levels decreased by 9.2%, 24.4%, 27.5% and 29.9% for dosages of 100 mg, 300 mg, 600 mg and 1000 mg, respectively. No changes were noted for doses of 10 and 30 mg.

Table 21. Average change from baseline 100 mg 300 mg 600 mg 1000 mg Serum PSA -9.2% -24.4% -27.5% -29.9% Lh 20.7% 46.9% 27.6% 29.2% Free testosterone -17.0% -18.5% -72.7% -53.2% Total testosterone 3.9% 7.3% -51.9% -47.9%

EXAMPLE 18

Bioavailability of Compound IV

Compound IV was rapidly absorbed after oral administration to rats, dogs, and macaques. The oral bioavailability of Compound IV for rats ranged from 6% to 25% depending on the composition in which the dose was administered. Compositions prepared using polyethylene glycol 300 (PEG300), as a rule, had a greater effect than microemulsions prepared on Tween 80 diluted with deionized water. Visual control of blood plasma concentration-time profiles in dogs suggested that Compound IV undergoes enterohepatic recirculation, as indicated by the second peak in the final phase. It is important to note that the effect on dogs in the group of males receiving an oral dose of 30 mg / kg in PEG300 exceeded the effect necessary to maximize the effect on prostate reduction in the rat model of LH suppression. Preliminary pharmacokinetic studies in macaques suggested that the oral bioavailability in this species is approximately equal to or greater than that in dogs, which was confirmed by measuring plasma concentrations of Compound IV and suppressing serum testosterone levels for seven days. In general, these data indicate that a sufficient effect with oral administration can be achieved in two species of animals that are not rodents to obtain the desired pharmacological effect (based on AUC data). Further, data on the endocrine system in rats and macaques suggest that the pharmacological effects of Compound IV are reversible (i.e., serum testosterone concentrations return to baseline or normal levels after discontinuation of Compound IV treatment).

EXAMPLE 19

Pharmacokinetics of Compound IV

Preliminary studies of in vitro metabolism (mouse, rat, dog, macaque and humans) and in vivo (rat) suggested that the combined participation of Compound IV, its hydroxylated metabolite (s) and its N-dealkylated metabolite contributes to the overall distribution Compounds IV in animals and humans. The results of interspecific comparisons, although they are only qualitative, show that the general metabolic profiles of species not participating in clinical studies adequately reflect the profile generated in human liver microsomes. According to these results, rats and dogs are species of rodents and animals that are not rodents, respectively, suitable for pharmacological and toxicological evaluations. In vitro studies show that Compound IV does not induce relevant isoforms of CYP450 (cytochrome P450-dependent monooxygenase) (CYP1A2, CYP2B6 or CYP3A4) and does not inhibit CYP1A2, CYP2C19, CYP2D6 or CYP3A4 / 5 at concentrations <30 μM. CYP2C9 is inhibited by Compound IV, but only at high concentrations (K _i = 8 μM) and possible pharmacokinetic drug-drug interactions are considered unlikely.

EXAMPLE 20

The biological activity of Compound IV

Compound IV has a slight inhibitory effect or no in vitro inhibitory effect (IC ₅₀ ≥ 300 μM) on the hERG channel (the product of the specific human heart potassium channel gene, from the human ether a-go-go-related gene). The compound at a concentration of 10 and 100 μM in a dose-dependent manner reduces APD ₅₀ and APD ₉₀ (duration of action potential at 50% and 90% repolarization) in isolated Purkinje fibers of a dog in vitro. However, Compound IV does not affect hemodynamic or cardiac function (blood pressure, heart rate, electrocardiogram structure or QT intervals (cardiac output)) when measured by telemetry in dogs at any dose (up to 300 mg / kg inclusive). No neuropharmacological or pulmonary effects were observed. No significant effects on renal function were noted with a single oral dose of Compound IV up to and including 30 mg / kg. Only increased excretion of urine and excretion of potassium and chlorine ions into the urine were observed with the introduction of the highest tested dose (100 mg / kg). Oral administration of Compound IV at doses of 30-300 mg / kg in rats caused a significant increase in peristalsis, and oral administration of Compound IV at a dose of 30 mg / kg in rats caused a significant increase in gastrointestinal motility and gastric acidity (in all probability not due to smooth muscle effects).

Compound IV at concentrations up to 200 μM, inclusive, was not mutagenic and did not cause structural or quantitative chromosomal aberrations in human peripheral blood lymphocytes in vitro. Compound IV was well tolerated by rats and dogs after single and repeated oral administration (up to and including 28 days). No pathological changes were observed in the kidneys, liver, heart and other organs that are not targets. There were no serious physical symptoms, effects on body weight, clinical pathological changes, ophthalmic, electrocardiographic or histopathological changes associated with the oral administration of Compound IV to male or female dogs for up to 28 days, inclusive.

Along with the fact that some features of the invention are illustrated and set forth herein, those skilled in the art will further find numerous modifications, substitutions, changes and equivalents. Therefore, it should be understood that the claims are intended to include all such modifications and changes that fall within the concept of the real essence of the invention.

Claims (12)

1. A method of treating advanced prostate cancer in a male subject, essentially consisting of administering a therapeutically effective amount of a compound of formula IV or an isomer thereof, a pharmaceutically acceptable salt, pharmaceutical product, polymorph, hydrate, or any combination thereof:

2. The method according to claim 1, where the specified treatment of advanced prostate cancer is carried out by lowering the levels of total testosterone in blood serum or free testosterone in the blood serum, and where the specified decrease in the levels of total testosterone in blood serum or free testosterone in the blood serum serum luteinizing hormone levels. 3. The method according to claim 1, where the specified treatment of advanced prostate cancer is carried out by reducing the levels of total testosterone in serum or free testosterone in the blood serum, and where the specified decrease in levels of total testosterone in the blood serum or free testosterone in the blood serum does not depend on reduce serum luteinizing hormone levels. 4. The method of claim 3, wherein said decrease in total serum testosterone or free serum testosterone levels occurs due to an increase in sex hormone binding globulin (SHBG) levels, inhibition of testosterone production or secretion by Leydig cells in the testes, or a decrease in adrenal steroidogenesis . 5. The method according to claim 4, where the specified total testosterone in serum contains free testosterone and associated testosterone, where a decrease in these levels of free testosterone, independent of a decrease in serum luteinizing hormone levels, occurs due to an increase in SHBG levels. 6. The method according to claim 2 or 3, where the total serum testosterone or free serum testosterone is reduced to a level of less than about 100 ng / dl, 50 ng / dl or 25 ng / dl. 7. The method according to claim 1, where the introduction of the specified compound produces a decrease in the levels of total testosterone in serum or free testosterone in blood serum, without causing side effects associated with antiandrogen therapy (ADT). 8. The method according to claim 1, where the introduction of the specified compound produces a decrease in the levels of total testosterone in blood serum or free testosterone in blood serum without causing flushing, gynecomastia, reduced bone mineral density or an increased tendency to bone fracture. 9. The method of claim 8, wherein said decrease in serum total testosterone or serum free testosterone levels occurs by decreasing serum luteinizing hormone levels. 10. The method of claim 8, wherein said decrease in serum total testosterone or serum free testosterone does not depend on a decrease in serum luteinizing hormone levels. 11. The method according to claim 1, where the specified method also suppresses, reduces the incidence, reduces the severity or inhibits advanced prostate cancer. 12. The method according to claim 1, where the specified method also provides palliative treatment of advanced prostate cancer.

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