WO2008122198A1

WO2008122198A1 - Non-steroidal androgen acceptor regulators and their medical use

Info

Publication number: WO2008122198A1
Application number: PCT/CN2008/000649
Authority: WO
Inventors: Mingwei Wang; Guohai Wu; Caihong Zhou; Qunyi Li
Original assignee: Shanghai Institute Of Materia Medica, Chinese Academy Of Sciences; The National Center For Drug Screening
Priority date: 2007-04-09
Filing date: 2008-03-31
Publication date: 2008-10-16
Also published as: CN101284795B; CN101284795A

Abstract

The invention discloses non-steroidal androgen acceptor regulators of formula I or pharmaceutical acceptable salts thereof, which are useful in preparation of non-steroidal medicaments for prophylaxis and/or treatment of partial androgen deficiency in the aging male, osteoporosis, sarcopenia, female sexual inadequacy, asitia, female hirsutism, severe androgenic alopecia, acne, cachexia, benign prostatic hyperplasia, prostatic cancer, breast cancer, Alzhermer's disease, Parkinson's disease and like. The invention also discloses pharmaceutical compositions containing the compounds of formula I or pharmaceutical acceptable salts thereof, and the use of the compounds of formula I or pharmaceutical acceptable salts thereof in preparation of medicaments for treatment of such conditions or diseases.

Description

A class of non-steroidal hormone receptor modulators and their medical uses

Technical field

The present invention relates to the use of a class of non-androgen receptor modulators and pharmaceutical compositions thereof for the prevention and/or treatment of symptoms or diseases caused by androgen and androgen receptor dysfunction, including but not limited to middle-aged men Androgen partial deficiency syndrome, osteoporosis, muscle consumption (Sarcopenia female sexual insufficiency, anorexia, female hirsutism, severe androgen-dependent alopecia, acne, dysentery, benign prostatic hyperplasia, prostate cancer, breast cancer, Alzheimer's disease (AD) and Parkinson's disease (PD).

Background technique

Androgen is an important steroidal sex hormone in the human body. It promotes cell differentiation and tissue growth by binding to the Androgen Receptor (AR) and participates in many key physiological functions such as male fetal reproductive organs (such as the prostate). , the formation of sputum, epididymis, etc.), the development and maintenance of secondary sexual characteristics, and sperm production. A certain proportion of androgens, such as Testosterone (T), Dihydrotestosterone (DHT), and Adrenal androsterone, are present in both men and women. In women, androsterone can be converted to estrogen in most target organs, but plays a normal physiological function in some tissues such as the brain. In the prostate and skin, androgen is converted to DHT by 5α-reductase, and DHT has a 3-5-fold higher affinity for AR than quinone.

AR is a member of the nuclear receptor superfamily, a ligand-activated transcription factor that is widely distributed in proliferating and non-proliferating tissues, including prostate and sperm, male and female genitalia, testes, ovaries, skin, edulis, sweat glands, Cartilage, myocardium, bone and smooth muscle, adrenal cortex, liver, pineal gland and multiple cerebral cortical and subcortical regions, including spinal motor neurons (Negro-Vilar, J Clinical Endocrinology and Metabolism, 1999, 84: 3459-3462 ). The AR protein has three domains: N-terminal structure (NTD), DNA-binding i or (DNA binding domain, DBD) binds to a ligand or (Ligand binding domain, LBD X He B, Kemppainen JA, 1999: J. Biol. Chem. 274: 37219-25). After the androgen and AR LBD form a complex, they bind to the Androgen response element (ARE) located in the promoter region of the target gene to activate or inhibit the expression of the purine gene, thereby regulating the physiological function of the target tissue. . Many AR-related diseases, either due to abnormal hormone levels or due to receptor dysfunction, disrupt normal interaction balance. In a diseased individual, the drug achieves the effect of a treatment-related disease by potentiating or inhibiting the level of receptor activation. Therefore, AR is an important target in the treatment of various diseases and symptoms associated with androgen action. For metabolic and endocrine diseases, AR modulators can be used to prevent/treat diseases associated with age and androgen metabolism, such as androgen deficiency in older men, androgen deficiency in women, osteoporosis, muscle loss, female sexual insufficiency , anorexia, hirsutism in women, severe androgen-dependent alopecia, acne, and several morbid diseases. In oncology, AR modulators can be used to treat benign prostatic hyperplasia, hormone-dependent prostate cancer, and breast cancer; for neurological diseases, AR modulators can be used to treat depression, Alzheimer's disease, and Parkinson's syndrome (Segal) S., Naraynan R. and Dalton J. Expert Opin. Investig. Drugs 2006, 15: 377-387).

Clinically used AR regulators are structurally classified into steroidal and non-steroidal. Steroidal androgens include, in addition to naturally occurring in the body, esters of androstenone such as cyclopentanoate, propionate, phenylpropionate, cyclopentylpropionate, heptanoate and decanoate. ), and other synthetic androgens (such as 7-methylnortestosterone and its acetate). Antiandrogen is represented by Cyproterone acetate (CPA). Non-steroidal AR modulators include Flutamide, Nilutamide, and Bicalutamide (Casodex), all of which are receptor antagonists. Some non-steroidal AR agonists have also been reported in the literature and patents, but they have not yet entered clinical application. Both the steroidal and non-steroidal AR modulators have certain side effects, including causing metabolic disorders of the androgen and causing male breast development and liver toxicity. Patients with benign prostatic hyperplasia/prostate cancer who have long-term use of flutamide or bicalutamide may have side effects such as gastrointestinal discomfort, nausea, vomiting, insomnia, fatigue, headache, anxiety, blurred vision, and loss of libido. After the androgen drug, there will be "anti-male" "Antiandrogen withdrawal syndrome" (AWS), which shows that the PSA level that was originally inhibited after a period of administration has risen rapidly and the tumor volume has increased. It can only stop taking the drug or switch to other anti-androgen drugs (Dicker AP, 2003, Lancet Oncol. 4: 30-36; Laufer M, Sinibaldi VJ, 1999, Urology 54: 745]. It has also been reported that AR modulators interact with the anticoagulant coumarin during use. There is an urgent need to research and develop non-steroidal AR modulators with new chemical structures.

The present inventors disclose a class of non-steroidal androgen receptor modulators, methods for their preparation, pharmaceutical compositions and uses in the patent application CN200510026252.2. Although the use of such compounds as androgen receptor modulators is disclosed in the use, only the use of the compounds as androgen receptor antagonists is disclosed, and no use is made for their use as androgen receptor agonists. set forth.

Summary of the invention

In view of the defects or deficiencies of the above prior art, the present invention prepares and optimizes a novel class of non-body small molecule compounds through chemical synthesis and structure-activity relationship studies. The androgen receptor competition experiments showed that these compounds have an affinity for the receptor of less than 10 ΉΜ, which is equivalent to DHT; luciferase 4 gene co-transfection experiments and prostate cancer cell line LNCaP proliferation experiments show that they have AR Partial agonism and antagonistic activity represent the potential for further development as a novel androgen receptor modulator.

Accordingly, it is an object of the present invention to provide a non-steroidal androgen receptor modulator compound having the structure of the following formula I or a pharmaceutically acceptable salt thereof.

A further object of the present invention is to provide a pharmaceutical composition comprising a compound of the formula I or a pharmaceutically acceptable salt thereof.

A further object of the present invention is to provide a compound of the formula I or a pharmaceutically acceptable salt thereof as a non-steroidal androgen receptor modulator for the prevention or/and treatment of androgen and androgen receptor dysfunction The cause of the symptoms or diseases caused. The compound of the present invention having a structure of the formula I or a pharmaceutically acceptable salt thereof is used as a non-steroidal androgen receptor agonist for the preparation of a prophylactic or/and therapeutic symptom caused by androgen and androgen receptor dysfunction or Use in disease.

The compound of the present invention having a structure of the formula I or a pharmaceutically acceptable salt thereof is used as a non-steroidal androgen receptor antagonist in the preparation of a prophylactic or/and therapeutic agent for symptoms caused by androgen and androgen receptor dysfunction or Use in disease.

The non-androgen receptor modulator or a pharmaceutically acceptable salt thereof provided by the present invention has the structure of the following formula I:

Where: 1 is? , Cl, Br or N0 ₂ ; specifically: compound 3-phenyl-3-(4-nitroanilino)-1-(4-chlorophenyl)-1-propanone (MWW6003 3-phenyl-3- (4-nitroanilino)-1-(4-nitrophenyl)-1-propanone (MWW6009), 3-phenyl-3-(4-nitroanilino)-1-(4-bromobenzene Base)-1-propanone (MWW6015) and 3-phenyl-3-(4-nitroanilino)-1-(4-fluorophenyl)-1-propanone (MWW6040), the structural formula is as follows:

MWW6003 MWW6015

MWW6009 MWW6040 For the compound of the formula I of the present invention or a pharmaceutically acceptable salt thereof, when a chiral carbon is present in the molecule, it is a racemate or an optically active substance. The method obtains a pharmaceutically acceptable salt thereof, for example, the acid is a mineral acid such as hydrochloric acid, hydrobromic acid, nitric acid, sulfuric acid, phosphoric acid, etc.; an organic acid such as formic acid, acetic acid, propionic acid, benzoic acid, horse Acid, fumaric acid, succinic acid, tartaric acid, citric acid, etc.; alkylsulfonic acid, such as methanesulfonic acid, ethylsulfonic acid, etc.; arylsulfonic acid, such as benzenesulfonic acid, p-toluenesulfonic acid, etc. use.

The pharmaceutical compositions provided herein comprise a therapeutically effective amount of one or more compounds of the formula I, or a pharmaceutically acceptable salt thereof, which may further comprise one or more pharmaceutically acceptable carriers Or an excipient.

A desirable ratio of the pharmaceutical composition provided by the present invention is that the compound of the formula I or a pharmaceutically acceptable salt thereof has an active ingredient in an amount of from 50% to 99.5% by weight based on the total weight ratio, and the remainder is less than 50% by weight based on the total weight. A further object of the present invention is to provide prevention by administering an androgen receptor modulator or a pharmaceutically acceptable salt thereof (including a pharmaceutical composition thereof) as described herein, as an androgen receptor agonist and/or an antagonist. And/or methods for treating various symptoms or diseases caused by androgen and androgen receptor dysfunction, including but not limited to osteoporosis, muscle wasting, female sexual insufficiency, anorexia, female hirsutism, severe androgen Dependent hair loss, acne, seborrhea, dysentery (including but not limited to AIDS), benign prostatic hyperplasia, prostate cancer, androgenetic alopecia, benign or malignant tumors that express androgen receptors (eg, in the breast, brain, Cancer of tissues such as skin, ovary, bladder, lymph, liver, kidney and pancreas), Endometriosis, ovarian syndrome, Alzheimer's disease, Parkinson's disease, androgen-dependent/age-related diseases (eg, androgen partial deficiency syndrome in middle-aged men, male or female) Dysfunction and muscle atrophy in ambulatory patients, etc.). It is particularly preferably used for the treatment/prevention of androgen-dependent tumors, particularly prostate cancer, which reduces the incidence of prostate cancer, prevents the occurrence of prostate cancer or causes regression of prostate cancer.

DRAWINGS

Figure 1 is a competitive binding activity test of a compound with DHT for AR. Compound MWW6003 and MWW6015 rather, IC ₅ and DHT AR affinity for both. Values were 2.23 ΉΜ and 3.48 ΉΜ; compound MWW6009 and MWW6040 relatively weak affinity for AR, IC ₅₀ values were 21.60 nM and 14.40 ΉΜ, under the same conditions DHT IC ₅₀ of 7.75 ΉΜ.

Figure 2 is a cytotoxicity test of compounds in HeLa cells. Compounds MWW6003 and MWW6015 were incubated with HeLa cells for three days and their effects on cell growth were determined by MTT assay, both showing only low cytotoxicity, with IC _{5 of} compound MWW6003. The value is 2.8 μΜ and the IC ₅₀ of the MWW6015 is between 10 and 40 μΜ.

Figure 3 is a pharmacological activity test of a compound in a CV-1 cell reporter transfection experiment. Dihydrotestosterone (DHT) induces the expression of luciferase in cells to increase chemiluminescence readings, and the antagonism of compounds on androgen receptors appears to inhibit DHT-induced chemiluminescence. MWW6003 and MWW6015 showed AR agonistic activity in agonistic mode, with potency of 122.7% and 107.0%, respectively, and EC ₅₀ of 70.8 ΉΜ and 122.1 nM, respectively; Compound MWW6009 showed partial agonistic activity, EC ₅₀ was 1206 ΉΜ, and the potency was DHT. of 26.2%, under the same conditions of DHT EC ₅₀ of 0.4 ΉΜ; in antagonist mode, the compound MWW6009 displays some AR antagonistic activity, an IC ₅₀ of 5989 nM, under conditions similar bicalutamide (bicalutamide, Casodex) of The IC ₅₀ is 97.7. Figure 4 is a pharmacological activity test of the compound in the MDA-MB-453 cell reporter transfection experiment. MWW6003, MWW6009 and MWW6015 show partial AR agonistic activity in the agonistic mode, The potency was 19.5%, 9.2%, and 20.4%, respectively, and EC ₅ o was 239.3 ΉΜ, 561.7 ΉΜ, and 102.1 分别, respectively. Under the same conditions, the EC ₅₀ of DHT was 0.6 ΉΜ; in the antagonistic mode, the compounds MWW6003 and MWW6015 showed A certain AR antagonistic activity, IC ₅ o was 812.6 ΉΜ and 84.8 分别, respectively, the inhibition rates were 99.1% and 23.9%, respectively, while the IC ₅₀ of Casodex was 400.4 同样 under the same conditions.

Figure 5 is a pharmacological activity test of the compound in the LNCaP cell proliferation assay. LNCaP is a hormone-dependent prostate cancer cell, and both DHT and AR agonists induce proliferation. AR antagonists can inhibit DHT-induced cell proliferation. The results showed that the compounds MWW6003 and MWW6015 showed some activity trends in both the agonistic and antagonistic modes.

detailed description

In order to clarify the invention and not to be limited thereto, the invention is described in detail in the following subsections.

Definition

The technical and scientific terms used in the present invention have the same meaning as the general understanding of the general art in the field to which the invention pertains, unless otherwise defined. All patents, applications, published applications and other publications and sequences derived from gene banks and other databases referred to herein are incorporated by reference in their entirety. If the definitions set forth in this section are in contrast to the definitions of income and citations of all patents, applications, published applications, and other publications and sequences derived from the Gene Bank and other databases referenced in this patent, or inconsistent, The definition of clarification shall prevail.

As used herein, "a" or "an" refers to "at least one" or "one or more."

As used herein, "modulator" refers to a class of compounds that enhance (eg, agonists, have agonistic activity) or inhibit (eg, antagonists, have antagonistic activity) the biological activity or function of a prion protein (including enzymes, receptors, etc.), Moreover, its enhancement and inhibition only temporarily act on the occurrence of a specific event in a specific cell or tissue, such as signal transduction, transcriptional activation and the like.

As used herein, "middle-aged male androgen partial deficiency syndrome" (Partial androgen) Deficiency of the aging male , PAD AM ) refers to the progressive reduction of male androgen production after middle age. Clinical manifestations include fatigue, depression, decreased libido, sexual dysfunction, erectile dysfunction, hypogonadism, osteoporosis, alopecia, obesity , senile muscle atrophy, osteopenia, benign prostatic hyperplasia, anemia, mood and cognitive changes, and prostate cancer. The occurrence of PADAM is related to the androgen environment and can be corrected by controlling the androgen environment.

As used herein, Osteoporosis (OP) is a systemic skeletal disorder characterized by low bone density and degeneration of bone tissue, which increases bone fragility and susceptibility to fracture. Femoral fractures are the most serious outcome of osteoporosis, with 5-20% of patients dying within one year and more than 50% of survivors losing their ability to live. Older people are at greatest risk for osteoporosis, and their incidence increases significantly with ageing. In addition, women are more likely to develop osteoporosis than men. The physiological concentrations of androgen and estrogen play an important role in maintaining bone homeostasis throughout the life cycle. Thus, when androgen or estrogen is lost, the result is an increase in the rate of bone remodeling, a tendency to resorb the balance of resorption and formation, and promote total loss of bone mass. Appropriate supplementation of androgen and androgen-like substances can increase bone density, improve osteoporosis, and reduce fractures.

As used herein, "prostatic hyperplasia" refers to a type of benign adenomatous hyperplasia of the prostate surrounding the urethra that causes varying degrees of bladder outflow obstructive disease or symptoms, also known as "benign prostatic hypertrophy." Prostatic hyperplasia is one of the most common diseases in urology and has become a "stealth killer" that threatens men's health. Clinical statistics show that men have a prevalence of benign prostatic hyperplasia of about 50% between the ages of 40 and 79, and up to 80% after age 80. As the pace of life continues to accelerate, the number of patients with benign prostatic hyperplasia is increasing, and it is younger. Benign prostatic hyperplasia can also cause a variety of potential complications, such as acute urinary retention, urinary tract infection, fleshy urine, bladder diverticulum, stones, hydronephrosis and renal failure. Studies have shown that dihydrotestosterone is the most important cause of benign prostatic hyperplasia in patients, and anti-androgen therapy can help to improve its symptoms.

As used herein, "prostate cancer" refers to a type of malignant tumor common to the male reproductive system, which is predominantly adenocarcinoma. Prostate cancer is a serious male senile disease with very high morbidity and mortality rates in Europe and America. High, accounting for the first place in male malignancies (Landis SH, Murray T, 1998, CA Cancer J. Clin. 48: 6-29). The incidence of prostate cancer in China is lower than in Europe and the United States, but in recent years, with the elderly The acceleration of the degree of regulation, the changes in the traditional diet structure, and the improvement of the diagnostic level of such diseases, the incidence rate has increased significantly. In addition to early prostate cancer can be surgically removed, anti-androgen therapy is the clinical choice.

As used herein, "women's hirsutism" refers to a hirsutous symptom in women caused by an increase in androgen secretion, that is, a long, thick, black hair, or hair, that grows in areas that should not have long hair. It is male-type, with thick and dark eyebrows and pubic hair developing to the abdomen and even the umbilicus.

As used herein, "severe androgen-dependent alopecia" refers to a type of severe seborrheic alopecia, also known as male pattern alopecia.

As used herein, "acne" refers to the chronic inflammation of a type of edema sebaceous gland, which occurs in the face, chest and back, and is characterized by acne, papules, pustules, nodules, carbuncle, etc., also known as youth acne.

As used herein, "Alzheimer's disease" is a group of primary degenerative brain degeneration diseases whose etiology is unknown. The pathological changes are mainly diffuse atrophy of the cortex, widening of the sulcus, enlargement of the ventricles, reduction of neurons, and visible Lesions of senile plaques, neurofibrillary tangles, granular vacuoles, and choline acetylase and acetylcholine were significantly reduced. Alzheimer's disease is more common in the elderly, latent onset, slow and irreversible progress (two years or longer), mainly based on intelligent damage. The onset of the disease before the age of 65 (pre-sensual), more family history of the same disease, the lesions develop faster, the front page and top page lesions are more significant, often aphasia and loss of use. In a group of placebo-controlled trials, cognitive impairment in patients with Alzheimer's disease improved after androgen replacement therapy, suggesting the clinical value of androgen receptors in the disease ( Cherrier MM, Matsumoto AM, Amory JK et al. 2005, Neurology, 64: 2063-2068).

As used herein, "Parkinson's disease" is a common central nervous system degenerative disease in middle-aged and elderly people, mainly characterized by slow patient movement, tremors in the hands, feet or other parts of the body, loss of softness in the body, and muscle stiffness. Parkinson's disease is second only to tumors and cardiovascular and cerebrovascular diseases, and is called the "third killer" and "chronic cancer". Okun et al confirmed that androgen supplementation helps improve Symptoms of the disease ( Okun MS, Walter BL, Mcdonald WM, et al. 2002, Arch. Neurol. 59: 1750-1753).

As used herein, an "effective amount" of a compound for treating a particular condition refers to an amount sufficient to ameliorate or to some extent alleviate the symptoms associated with the disease. This dose can be administered in a single dose or in a therapeutic regimen. This dose cures the disease, but is typically administered to improve the condition. Repeated administration to improve symptoms may be desirable.

As used herein, "pharmaceutically acceptable salts, esters or other derivatives" include any salt, ester or derivative which is readily prepared by those skilled in the art by known methods. The compounds thus derived and produced can be administered to animals and humans without toxic effects. The compound is either pharmaceutically active or a prodrug.

As used herein, "treatment" means that the disease and symptoms are improved in any way, or other beneficial changes. Treatment also includes the use of the compounds of the invention in medicine.

As used herein, administration of a particular pharmaceutical composition to "improve" the symptoms of a particular disease means any reduction, whether permanent, temporary, prolonged, transient, can be attributed to or associated with the pharmaceutical composition. Relevant application.

As used herein, "substantially pure" means sufficient homogeny to detect impurities by standard analytical methods used by those skilled in the art for evaluating purity, such as thin layer chromatography (TLC), gel electrophoresis. And high performance liquid chromatography (HPLC). Or sufficiently pure also means that even further purification does not alter the physicochemical properties detectable by the substance, such as enzymatic activity and biological activity. Methods for producing substantially chemically pure compounds for purification are well known to those skilled in the art. However, substantially chemically pure compounds can be stereoisomers or mixtures of isomers. In this case, further purification may increase the specific activity of the compound.

As used herein, "prodrug" refers to a compound administered in vivo that can be metabolized or converted to a biologically, pharmaceutically or therapeutically active form. To produce a prodrug, the pharmaceutically active compound will be modified to reproduce the active compound by metabolic processes. Prodrugs can be designed to alter their metabolic stability, or transport properties of precursors, to mask their side effects or toxicity, improve The taste of the drug, or other characteristics. By virtue of knowledge of pharmacokinetics and metabolism of the drug in the body, once the pharmaceutically active compound is known, one skilled in the art can design a prodrug of the compound. [See Medicinal Chemistry A Biochemical Approach, Oxford University Press, New York, 1985, pages 388-392].

The terms "substantially" are the same or are homogenous or similar, and the understanding of the relevant art can vary in the context, and is generally at least 70%, preferably at least 80%, more preferably at least 90%, in accordance with the understanding of the relevant art. Optimally at least 95% identical.

As used herein, "composition" refers to any mixture. It can be a solution, suspension, liquid, powder, ointment, aqueous, non-aqueous or any combination thereof.

As used herein, "union" refers to any association between two or more.

The term "subject" as used herein includes humans and animals, for example, dogs, cats, cows, pigs, rodents, etc., preferably humans. An experienced practitioner should understand that the subject is suitable and willing to suffer from orrogen and/or androgen receptor dysfunction or associated with androgen partial deficiency syndrome, osteoporosis, muscle wasting, anorexia, Treatment and prevention of diseases and conditions such as female hirsutism, severe androgen-dependent alopecia, acne, dysentery, benign prostatic hyperplasia, prostate cancer, breast cancer, Alzheimer's disease and Parkinson's disease.

Any of the protective groups, abbreviations for amino acids and other compounds used herein, are consistent with their common, recognized abbreviations or the biochemical nomenclature issued by the IUPAC-IUB committee, unless otherwise stated.

Formula and dosage

According to the invention, the compounds of the invention, alone or in combination with other agents, carriers or excipients, are formulated for any suitable route of administration, such as intraluminal, subcutaneous, intravenous, intramuscular, intradermal injection. Oral or topical. The method can be administered by injection, in a single dose, in an ampoule, or in a multi-dose container with an additional buffer. The formulations may take such forms as suspensions, solutions or emulsions in oily or aqueous vehicles. The formulations may contain formulating agents such as suspending, stabilizing and/or dispersing agents. In addition, the active ingredient can be in powder form and suitable before use. The carrier, sterile non-pyrogenic water or other solvent constitutes a dosage form. The topical medicament of the present invention may be a foam, a gel, an ointment, an ointment, a transdermal patch or a paste.

Pharmaceutical compositions and methods for administration which can be used in the present invention include, but are not limited to, those disclosed in U.S. Patent Nos. 5,736,154, 6,197,801 B1, 5,741,511, 5,886,039, 5,941,868, 6,258,374, and 5,686,102.

The size of the dose to be treated or prevented will vary depending on the severity of the condition and the route of administration. The frequency of dosing and medication will vary with age, weight, health status, and individual patient response.

It should be pointed out that (the doctor should also know), according to toxicity and side effects, necessary measures must be taken to terminate, interrupt or reduce the therapeutic dose. Conversely, if the clinical response is not obvious (excluding toxicity and side effects), the doctor should adjust the treatment plan to increase the dose.

Any suitable route of administration can be employed. Dosage forms include tablets, troches, soy gums, dispersing agents, suspending agents, solutions, capsules, films and the like.

In a practical application, the compound of the present invention, alone or in combination with other preparations, may be in accordance with general pharmaceutical mixing techniques with pharmaceutical carriers or excipients such as β-cyclodextrin and 2-hydroxy-propyl-β-cyclodextrin. Finely mixed. Depending on the needs of the administration, a special carrier, a local or parenteral route, may be employed. For the preparation of parenteral dosage forms, such as compositions for intravenous injection or infusion, similar pharmaceutical vehicles can be employed, water, glycols, oils, buffers, sugars, preservatives, liposomes, etc., which are well known to those skilled in the art. . Examples of such parenteral compositions include, but are not limited to, 5% w/v dextrose, physiological saline or other solutions. The total dose of the compound of the present invention, alone or in combination with other preparations, can be administered in vials of intravenous injection, in a volume of from about 1 ml to about 2000 ml. The amount of diluent will vary depending on the total dose administered.

The invention also provides a kit for achieving a therapeutic regimen. The kit comprises an effective amount of a compound of the invention, in a pharmaceutically acceptable form, alone or in combination with other agents, in one or more containers. A preferred pharmaceutical form is in combination with sterile saline, dextrose solution, buffered solution, or other pharmaceutically acceptable sterile liquid. Alternatively, the composition can be lyophilized or dried; in this case, the kit Optionally, a pharmaceutically acceptable solution, preferably a sterile solution, is contained in a container to reconstitute the complex to form a solution for injection purposes. Typical pharmaceutically acceptable solutions are saline solutions and dextrose solutions.

In another embodiment, the kit of the present invention further comprises a needle or syringe and/or a packaged alcohol pad for injection of the composition, preferably in sterile form. Instructions for use by a doctor or patient may optionally be included.

The invention is further illustrated by the following specific examples, without limiting the invention.

Example 1

Synthesis of 3-phenyl-3-(4-nitroanilino)-1-(4-chlorophenyl)-1-propanone (MWW6003)

Add 10.60 g (0.1 mol) of benzaldehyde, 13.81 g (0.1 mol) of 4-nitroaniline and 150 ml of absolute ethanol to the reaction flask. After stirring at room temperature for 10 minutes, add 14.46 g of 4-chloroacetophenone (0.1 mol). And a catalytic amount of concentrated hydrochloric acid, and then the reaction was stirred at room temperature for 20 hours. After completion of the reaction, the reaction solution was cooled overnight, and the precipitated solid was filtered, and washed with anhydrous ethanol. The obtained solid was suspended in 150 ml of 95% ethanol, stirred at room temperature for 2 hours, neutralized with saturated NaHC0 ₃ to basicity, stirring was continued for 1 hour, suction filtration, and the filter cake was washed with a small amount of anhydrous ethanol, and the crude product was mixed with ethanol/water. (1: 1) Recrystallization, obtaining 29.09 g of needle crystals, yield 76.4%. ifiNMRi CDC1 ₃ ): 8.00 ( 2H, d, J = 9.0 Hz, Ar-H ), 7.81 ( 2H, d, J = 8.1 Hz, Ar-H ), 7.42 ( 2H, d, J = 8.1 Hz, Ar- H), 7.23-7.36 ( 5H, m, Ar-H ), 6.52 ( 2H, d, J = 8.7 Hz, Ar-H ), 5.08 ( 1H, t, J = 6.3 Hz, CH ), 3.49 ( 2H, d, J = 6.0 Hz, CH ₂ ); MS (FAB): 382 (M+H).

Example 2 Synthesis of 3-phenyl-3-(4-nitroanilino)-1-(4-bromophenyl)-1-propanone (MWW6015)

Add 10.60 g (0.1 mol) of benzaldehyde, 13.81 g (0.1 mol) of 4-nitroaniline, 150 ml of absolute ethanol to the reaction flask, stir at room temperature for 10 minutes, then add 4-91 acetophenone 19.91 g (0.1 mol And a catalytic amount of concentrated hydrochloric acid, and then the reaction was stirred at room temperature for 20 hours. After completion of the reaction, the reaction solution was cooled overnight, and the precipitated solid was filtered, and washed with anhydrous ethanol. The obtained solid was suspended in 150 ml of 95% ethanol, stirred at room temperature for 2 hours, neutralized with saturated NaHC0 ₃ to basicity, stirring was continued for 1 hour, suction filtration, and the filter cake was washed with a small amount of anhydrous ethanol, and the crude product was mixed with ethanol/water. The solvent (1:1) was recrystallized to obtain 30.66 g of needle crystals, yield 72.1%. ifiNMRi CDC1 ₃ ): 8.00 ( 2H, d, J = 9.3 Hz, Ar-H ), 7.73 ( 2H, d, J = 8.4 Hz, Ar-H ), 7.58 ( 2H, d, J = 8.4 Hz, Ar- H), 7.23-7.39 ( 5H, m, Ar-H ), 6.52 ( 2H, d, J = 9.3 Hz, Ar-H ), 5.09 ( 1H, t, J = 6.0 Hz, CH ), 3.49 ( 2H, d, J = 7.2 Hz, CH ₂ ); MS (FAB): 426 (M+H). Example 3

Synthesis of 3-phenyl-3-(4-nitroanilino)-l-(4-nitrophenyl)-1-propanone (MWW6009)

Benzaldehyde (1.016 ml, 10 mmol), 4-nitroaniline (1.181 g, 10 mmol), toluene

50 ml was added to the reaction flask, a water separator was installed, and water was refluxed for 32 hours. After completion of the reaction, the reaction solution was concentrated in an appropriate amount and allowed to stand overnight. The precipitated solid was filtered and washed with a small amount of toluene to give a yellow solid which was directly transferred to the next reaction. 15 ml of the obtained solid, absolute ethanol was added to the reaction flask, and the reaction flask was placed in a water bath and heated to reflux. 4-Nitroacetophenone ( 1.652 g, 10 mmol) was dissolved in 15 ml of absolute ethanol, and 2~3 drops of concentrated hydrochloric acid were added dropwise, and then added dropwise to the refluxing reaction flask, and reflux was continued for 3 hours. The temperature is 50~60. C. After completion of the reaction, the reaction solution was cooled overnight, and the precipitated solid was filtered, and washed with anhydrous ethanol. The obtained solid was suspended in 10 ml of 95% ethanol, stirred at room temperature for 30 minutes, neutralized with a 10% aqueous NaHCO ₃ solution until basic, stirring was continued for 30 minutes, suction filtration, and the filter cake was washed with a small amount of anhydrous ethanol. The water mixed solvent (1:1) was recrystallized to obtain 0.567 g of needle crystals, yield 36.6%.

ifiNMl CDC1 ₃ ): 8.31 ( 2H, d, J=8.4 Hz, Ar-H ), 8.02 ( 2H, d, J=8.4 Hz, Ar-H ), 8.02-7.98 ( 4H, m, Ar-H ), 7.37-7.27 ( 5H, m, Ar-H ), 6.54 ( 2H, d, J=9.3 Hz, Ar-H ), 5.13 ( 1H, t, J=5.7 Hz, CH ) , 3.58 ( 2H, d, J = 9.6 Hz, CH ₂ ); MS (ESI): 392.20 (M+H).

Example 4

Synthesis of 3-phenyl-3-(4-nitroanilino)-l-(4-fluorophenyl)-1-propanone (MWW6040)

Add benzaldehyde (0.254 ml, 2.5 mmol), 4-nitroaniline (0.276 g, 2.0 mmol), and 7 ml of absolute ethanol to the reaction tube. After stirring at room temperature for 10 minutes, add 4-fluoroacetophenone (0.268). g, 2.0 mmol) and 0.12 ml of concentrated hydrochloric acid were placed in a microwave reactor and stirred for 20 minutes at a reaction temperature of 60. C. After completion of the reaction, the reaction solution was cooled overnight, and the precipitated solid was filtered, and washed with anhydrous ethanol. The resulting solid was suspended in 10 ml 95% ethanol was stirred at room temperature for 30 minutes, 10% NaHC0 ₃ aqueous solution to alkaline and stirring was continued for 30 minutes, filtered off with suction, the filter cake washed with a small amount of ethanol, the crude product from ethanol / The water mixed solvent (1:1) was recrystallized to obtain 0.567 g of needle crystals, yield 65.7%.

iHNMRi CDCI3 ): 8.13 ( 2H, d, J = 9.3 Hz, Ar-H ), 7.90 ( 2H, d, J = 8.1 Hz, Ar-H ), 7.42 ( 2H, d, J = 8.4 Hz, Ar-H ) , 7.35-7.24 ( 5H, m, Ar-H ) , 6.58 ( 2H, d, J = 8.7 Hz, Ar-H ) , 4.99 ( 1H, t , J=6.3 Hz, CH ) , 3.51 ( 2H, d, J=6.0 Hz, CH ₂ ); MS ( ESI ): 365.32 ( M+H

Example 5

Biological activity test

Material equipment

1.1 Plasmids and cell lines: Androgen receptor expression plasmid and luciferase reporter plasmid were constructed by National New Drug Screening Center; human cervical cancer epithelial cell line HeLa, monkey kidney epithelial cell CV-1, human breast cancer cell line MDA-MB -453 and human prostate cancer cell line LNCaP were purchased from ATCC, USA.

1.2 Reagents: Fetal bovine serum (FBS, GIBCO/BRL, USA); Activated carbon and dextran treatment of fetal bovine serum (CD-FBS, Hyclone, USA); DMEM and RPMI1640 medium (GIBCO/BRL, USA) IMEM medium (Bioresource, USA), Luciferase Assay Kit (Promega Corporation, USA); Fugene 6 (Roche Ltd., USA);

[ ³ H] Diarsonone (Dehydrotestosterone, DHT, Amersham, UK); Scintillation fluid (SuperMixTM, PerkinElmer, USA); Androgen receptor protein is the expression product of this receptor gene in insect cells.

1.3 Instruments: Envision 2101 Multilabel Reader (PerkinElmer, USA); Carbon Dioxide Incubator (Forma, USA); Wallac MicroBeta® TriLux 1450 (PerkinElmer, USA); VERS A Microplate Reader (Molecular Devices, USA) „

2. Experimental methods and results

2.1 receptor binding activity test

The gradient solution of DHT and the compound of the present invention shown in Figure 1 was prepared in DMSO, and the DHT and compound concentrations were 0, 0.128, 0.64, 3.2, 16, 80, 400, 2000 依次, respectively, and 5 μΐ each gradient DHT or The test compound solution is placed in each well of the tube. The androgen receptor protein is added to a protease inhibitor such as 1 μμμ aprotinin and Leupeptin. In a buffer [25 mM Na ₃ P0 ₄ , 10% glycerol (Glycerol), 10 mM NaMo0 ₄ , 10 mM KF, pH 7.5], add [ ³ H]DHT at a final concentration of 5 nM, mix well, Immediately add 195 μM per well to the tube and incubate overnight at 4 °C. After the incubation, add 50 μM hydroxyapatite ( ΗΑ ) solution (25%, 25 mM Na ₃ P0 ₄ , pH=7.4) to each well of the tube, mix by shaking, incubate for 10 minutes, and oscillate every three minutes. once. Centrifuge at 2500 rpm for 3 minutes, aspirate the supernatant and retain the pellet. Add 200 μM of the above buffer to each well, avoid vibration precipitation as much as possible, and centrifuge again for 3 minutes. The supernatant was aspirated, the pellet was retained, and the centrifugation was repeated once. The supernatant was aspirated, the pellet was retained, 300 μM of scintillation fluid was added to each well, and the mixture was shaken and read with a Wallac MicroBeta® TriLux 1450. Two of the compounds, MWW6003 and M WW6015, have an affinity for the androgen receptor comparable to the positive drug DHT, with IC ₅₀ values less than 10 nM (see Figure 1, Table 1).

Table 1 Test of binding activity of compounds to androgen receptor

Compound receptor binding (IC ₅₍₎ , nM )

DHT 7.75

MWW6003 2.23

MWW6009 21.60

MWW6015 3.48

MWW6040 14.40

2.2 cytotoxicity experiment

HeLa cells were cultured in RPMI1640 medium containing 10% FBS and 2 mM L-glutamine, and were grown to 90% confluence, and then trypsinized and added to the 96-well plate at 4000/well, 37 Incubate overnight at °C. The test compound was diluted to a certain concentration and added to the cells. The concentrations of the compounds MWW6003 and MWW6015 were 0, 2.56, 12.8, 64.0, 320, 1600, 8000, 40,000 依次, and the MTT solution (5 mg/mL) was added after 68 hours of culture. , 20μΙ7 well, measured 560nm light absorption value, reference The wavelength is 690 nM. The experimental results show that the compounds MWW6003 and MWW6015 have only low cytotoxicity to HeLa cells, IC _{5 of} MWW6003. With a value of 2.8 μΜ, the MWW6015 has an IC ₅₀ value between 10 and 40 μΜ (see Figure 2).

2.3 Report gene expression detection

CV-1 cells were cultured in DMEM medium containing 10% FBS and 2 mM L-glutamine. The day before transfection, the cells were replaced with DMEM medium containing 10% CD-FBS, and transfected with Fugene 6 reagent. The AR expression plasmid, the 4 reporter gene vector and Fugene 6 were uniformly added dropwise to the cells at a ratio of 1:10:30, and cultured at 37 ° C and 5% CO ₂ for 6 hours. After the cells were digested, the cells were inserted into a 96-well culture plate at 8000/100 μM/well, and cultured in DMEM medium containing 10% CD-FBS at 37 ° C for 16 hours. Add the compound to be tested. In the agonist mode, the DHT concentrations were 0, 0.01, 0.1, 1, 10, 100, and 1000 依次, and the concentrations of the 4 conjugates MWW6003 and MWW6015 were 0, 2.6, 13, 64, 320, 1600, 8000 ΉΜ, The concentrations of the compounds MWW6009 and MWW6040 were 0, 3.2, 16, 80, 400, 2000, 10000 依次. In the antagonistic mode, the concentrations of Bicalutamide (Casodex) were 0, 0.01, 0.1, 1, and 10. 100, 1000, 10000 ΉΜ, the concentration of compound MWW6009 was 0, 3.2, 16, 80, 400, 2000, 10000 依次, and 2 ΉΜ DHT was added to the cells as an agonist. After incubation for 24 hours, the luciferase assay kit was used to detect the enzyme activity to evaluate the pharmacological activity of the compound on the androgen receptor. The agonistic activities of the compounds MWW6003 and MWW6015 were stronger, with EC ₅ o of 70.8 ΉΜ and 122.1 分别, respectively, and the potency was 122.7% and 107.0%, respectively, which was equivalent to DHT, while the agonistic activity of compound MWW6009 was relatively weak. EC ₅ o was 1206 nM, performance is 26.2%. MWW6009 also showed a certain antagonistic activity with an IC ₅₀ of 80.43 ΉΜ, while the IC ₅₀ of Casodex was 100.1 同样 under the same conditions (see Figure 3).

MDA-MB-453 cells were cultured in IMEM medium containing 10% FBS and 2 mM L-glutamine. The day before transfection, the cells were replaced with IMEM medium containing 5% CD-FBS, and Fugene 6 reagent was used for transfection. Mix the reporter gene carrier and Fugene 6 in a 1:3 ratio and add them dropwise. The cells were cultured for 6 hours at 37 ° C and 5% CO ₂ . After the cells were digested, the cells were inserted into a 96-well culture plate at 20,000 cells/100 μM/well, and cultured in IMEM medium containing 5% CD-FBS at 37 ° C for 2 hours. Add the compound to be tested. When the agonist mode is detected, the concentrations of DHT, MWW6003 and MWW6015 are 0, 0.01, 0.1, 1, 10, 100, 1000, 10000 依次, and the concentrations of MWW6009 are 0, 3.2, 16, 80, 400, 2000, 10000 ΉΜ. When the antagonistic mode is detected, the concentrations of Casodex and MWW6003 are 0, 0.01, 0.1, 1, 10, 100, 1000, 10000 依次, and the concentrations of MWW6015 are 0, 3.2, 16, 80, 400, 2000, 10000 依次, At the same time, 5 ΉΜ DHT was used as an agonist in the cells. After incubation for 24 hours, the luciferase assay kit was used to detect the enzyme activity to evaluate the pharmacological activity of the compound on the androgen receptor. Compounds MWW6003, MWW6009 and MWW6015 showed partial AR agonistic activity in agonistic mode with potency of 19.5%, 9.2% and 20.4%, respectively, and EC ₅₀ of 239.3 ΉΜ, 561.7 ΉΜ and 102.1 分别, respectively, under the same conditions. ₅ . It was 0.6 ΉΜ; in the antagonistic mode, both compounds MWW6003 and MWW6015 showed certain AR antagonistic activities with IC ₅ o of 812.6 nM and 84.8 分别, respectively, and the inhibition rates were 99.1% and 23.9%, respectively, while Casodex under the same conditions. The IC ₅₀ is 400.4 nM (see Figure 4).

2.4 Prostate cancer cell proliferation assay

LNCaP cells were cultured in RPMI 1640 medium containing 10% FBS and 2 mM L-glutamine. One day before the experiment, the cells were replaced with RPMI1640 medium containing 5% CD-FBS, and were incubated until 90%. After trypsinization, the cells were added to a 96-well plate at 4000/90 μΙ7 well, and cultured at 37 ° C overnight. The test compound was diluted to a certain concentration and added to the cells at 10 μΙ7 well. The concentration of DHT, MWW6003 and MWW6015 was 0, 0.01, 0.1, 1, 10, 100, 1000, 10000 依次 in the agonist mode; antagonistic pattern detection At the same time, the concentrations of Casodex and the compounds MWW6003 and MWW6015 were 0, 0.01, 0.1, 1, 10, 100, 1000, 10000 依次, and 5 ηΜ DHT was added as an agonist to the cells. The culture was carried out for 6 days at 37 ° C, and the dressing was changed once on the third day. Before the end of the culture, MTT solution (5 mg/mL) was added, 20 μΙ 7 wells, and the absorbance at 560 nm was measured, and the reference wavelength was 690 nm. The experimental data is shown in Figure 5. Compounds MWW6003 and MWW6015 are both agonistic and antagonistic. Show a certain activity trend (see Figure 5).

3. Experimental conclusion

(1) Compounds MWW6003 and MWW6015 showed good binding activity in the androgen receptor competitive binding activity test, and their IC ₅ o values were less than 10 nM, which was comparable to DHT; compounds MWW6009 and MWW6040 also showed certain AR binding activity. .

(2) Compounds MWW6003 and MWW6015 showed low cytotoxicity in HeLa cytotoxicity experiments.

(3) Compounds MWW6003 and MWW6015 showed better agonistic activity in CV-1 cells co-transfected with AR expression vector and luciferase reporter gene expression vector, and their potency was comparable to DHT; MWW6009 also showed weak AR activation And antagonistic activity. Compounds MWW6003 and MWW6015 showed good androgen receptor agonistic/antagonistic activity in the MDA-MB-453 cell reporter assay; a certain agonistic/antagonistic activity trend was also demonstrated in the LNCaP proliferation assay. These results suggest that such compounds are useful as androgen receptor modulators and have certain utility in the prevention/treatment of androgen and androgen receptor related diseases.

Claims

Claim

A compound having the structure of the following formula I or a pharmaceutically acceptable salt thereof

I

Where R is F, Cl, Br or NO; the details are as follows:

3-phenyl-3-(4-nitroanilino)-1--3-phenyl-3-(4-nitroanilino)-1-(4-chlorophenyl)-1-propanone (4- Bromophenyl)-1-propanone

3-phenyl-3-(4-nitroanilino)-1- 3-phenyl-3-(4-nitroanilino)-1-(4-fluorophenyl)acetone (4-nitro Phenyl) small acetone.

The compound according to claim 1 or a pharmaceutically acceptable salt thereof, wherein the pharmaceutically acceptable salt is a compound of the formula I and hydrochloric acid, hydrobromic acid, nitric acid, sulfuric acid, a salt of phosphoric acid, formic acid, acetic acid, propionic acid, benzoic acid, maleic acid, fumaric acid, succinic acid, tartaric acid, citric acid, alkylsulfonic acid, arylsulfonic acid.

3. A compound according to any one of claims 1 to 2, or a pharmaceutically acceptable salt thereof, as a non-steroidal androgen receptor modulator for the prevention and/or treatment of androgen and androgen receptor dysfunction Use in symptoms or disease medications.

4. A compound according to any one of claims 1 to 2, or a pharmaceutically acceptable salt thereof, as a non-steroidal androgen receptor agonist for the prevention and/or treatment of androgen and androgen receptor dysfunction Use in symptoms or disease medications.

5. A compound according to any one of claims 1 to 2, or a pharmaceutically acceptable salt thereof, as a non-steroidal androgen receptor antagonist for the prevention and/or treatment of androgen and androgen receptor dysfunction Use in symptoms or disease medications.

The use according to any one of claims 3 to 5, wherein the symptoms or diseases caused by the androgen and androgen receptor dysfunction include androgen partial deficiency syndrome, osteoporosis, muscle consumption, Female sexual dysfunction, anorexia, female hirsutism, severe androgen-dependent alopecia, acne, dysentery, benign prostatic hyperplasia, prostate cancer, breast cancer, Alzheimer's disease, and Parkinson's syndrome.

7. A pharmaceutical composition having a prophylactic or/or therapeutic effect on a condition or a disease caused by androgen and androgen receptor dysfunction, comprising one or more of a therapeutically effective amount as an androgen receptor modulator A compound according to any one of claims 1 to 2, or a pharmaceutically acceptable salt thereof.

8. The pharmaceutical composition according to claim 7, wherein the pharmaceutical composition further comprises one or more pharmaceutically acceptable carriers or excipients.

The pharmaceutical composition according to claim 7 or 8, wherein the compound or a pharmaceutically acceptable salt thereof is contained as an active ingredient in an amount of from 50% to 99.5% by weight.