Novel compound for autophagy-mediated targeted degradation of androgen receptor and medical use thereof

Abstract

The present invention relates to a novel compound for autophagy-mediated targeted degradation of an androgen receptor and medical use thereof. The novel compound according to the present invention is an autophagosome-tethering compound (ATTEC) targeting an androgen receptor, which has not been previously reported, and exhibits excellent binding affinity for the androgen receptor and the autophagy marker LC3, and thus can activate autophagy and ultimately target and degrade the androgen receptor, thereby effectively treating androgen receptor-associated diseases.

Description

Novel compound for autophagy-mediated androgen receptor target degradation and its medical uses

[0001] The present invention relates to a novel compound for androgen receptor target degradation and its medical use, and more specifically, to a novel compound for autophagy-mediated androgen receptor target degradation and its medical use for androgen receptor-mediated diseases.

[0002] Prostate cancer is a malignant tumor that develops in the prostate gland. Age, race, and family history are reported to be factors that increase the risk of prostate cancer. In addition to these genetic predispositions, hormones, dietary habits, and chemicals such as herbicides are also known to play a significant role in its development. In Western countries, prostate cancer is the most common cancer among men, showing a high incidence rate, and in Korea as well, the frequency of prostate cancer has recently been increasing rapidly.

[0003] Drugs used to treat prostate cancer include hormones and cytotoxic anticancer agents. The primary treatment for prostate cancer is hormone therapy, and the representative hormone therapy drug currently available is Xtandi (active ingredient: Enzalutamide). It works by inhibiting the binding of active androgens to androgen receptors (ARs), and ranked 18th among global blockbuster drugs, recording sales of over $4.3 billion as of 2020. However, Xtandi exhibits limitations such as side effects and resistance.

[0004] Androgen receptors (ARs) play a crucial role in the prostate, primarily responding to androgens to provide gene products necessary for differentiation and growth. However, under abnormal conditions, they can also lead to the development of prostate cancer. As androgen receptor signaling is a unique characteristic of prostate cancer and can serve as a major therapeutic target for metastatic prostate cancer (mPC), androgen receptor target degradation strategies are being attempted to improve this.

[0005] The objective of the present invention is to provide a novel compound capable of targeting and degrading androgen receptors, which improves upon the disadvantages of conventional drug or protein degradation technologies.

[0006] Another objective of the present invention is to provide a therapeutic agent capable of treating related androgen receptor-related diseases using the novel compound described above.

[0007] To achieve the above objective, the present invention has a structure of an androgen receptor (AR) binding moiety-linker-autophagy labeling factor binding ligand, wherein the androgen receptor binding moiety is a compound represented by the following chemical formula A (compound A) or a pharmaceutically acceptable salt thereof;

[0008] <Chemical Formula A>

[0009]

[0010] The above autophagy labeling factor binding ligand is a compound represented by the following chemical formula B (Compound B) or a pharmaceutically acceptable salt thereof that binds to microtubule-associated protein light chain 3 (LC3);

[0011] <Chemical Formula B>

[0012]

[0013] The present invention provides a compound for targeting androgen receptor degradation, characterized in that the linker is a (C1-C20)alkylene connecting the nitrogen atom of the piperazine ring of compound A and the hydroxyl group of compound B, and one or more compounds A and B are connected to the linker.

[0014] The present invention provides a pharmaceutical composition for the prevention or treatment of androgen receptor-related diseases or cancer, comprising the above-mentioned compound for target degradation of androgen receptors as an active ingredient.

[0015] In addition, the present invention provides a food composition for preventing or improving androgen receptor-related diseases or cancer, comprising the above-mentioned compound for targeting androgen receptor degradation as an active ingredient.

[0016] The novel compound according to the present invention is an autophagosome-tethering compound (ATTEC) that targets androgen receptors and has not been previously reported. It has excellent binding affinity to androgen receptors and to LC3, an autophagy marker, so it can activate autophagy and ultimately target and degrade androgen receptors, thereby effectively treating androgen receptor-related diseases.

[0017] Figure 1 shows the androgen receptor (AR) binding affinity of compound A.

[0018] Figure 2 shows the LC3 binding strength of compounds B and C.

[0019] FIG. 3 shows compounds 1 and 2 synthesized according to one embodiment of the present invention.

[0020] Figure 4 evaluates the androgen receptor degradation efficacy of synthesized compounds 1 and 2.

[0021] The present invention will be described in detail below.

[0022]

[0023] The inventors developed an autophagy-mediated androgen receptor target degradation compound (AR-ATTEC) as a novel modality therapeutic agent for prostate cancer, and completed the present invention by confirming that the compound exhibits excellent degradation activity against androgen receptors and can act as a novel therapeutic agent for various cancer diseases related to androgen receptors.

[0024]

[0025] The present invention provides a novel compound for autophagy-mediated androgen receptor target degradation.

[0026] More specifically, the above compound may have the structure of an androgen receptor (AR) binding moiety-linker-autophagy labeling factor binding ligand.

[0027] The above androgen receptor (AR) binding moiety may be a compound represented by the following chemical formula A (Compound A) or a pharmaceutically acceptable salt thereof:

[0028] <Chemical Formula A>

[0029]

[0030] In this specification, "moiety" means part of a molecular functional group and may be used interchangeably with "substituent," "radical," "group," or "fragment."

[0031] In this specification, "pharmaceuticalally acceptable salt" means a salt having a safety and efficacy profile suitable for administration to humans, having no toxicity to cells or humans exposed to the compound.

[0032] The above salt may be used in the form of either a basic salt or an acidic salt that is pharmaceutically or food-grade acceptable. The basic salt may be used in the form of either an organic basic salt or an inorganic basic salt, and may be selected from the group consisting of sodium salt, potassium salt, calcium salt, lithium salt, magnesium salt, cesium salt, aluminum salt, ammonium salt, triethylaminium salt, and pyridinium salt. For the acidic salt, an acid addition salt formed by a free acid is useful. Inorganic and organic acids may be used as free acids. Inorganic acids may include hydrochloric acid, bromic acid, sulfuric acid, sulfite, phosphoric acid, diphosphate, nitric acid, etc., and organic acids may include citric acid, acetic acid, maleic acid, malic acid, fumaric acid, gluconic acid, methanesulfonic acid, benzenesulfonic acid, camphorsulfonic acid, oxalic acid, malonic acid, glutaric acid, acetic acid, glyconic acid, succinic acid, tartaric acid, 4-toluenesulfonic acid, galacturonic acid, emvonic acid, glutamic acid, citric acid, aspartic acid, stearic acid, etc., but are not limited thereto and may include all salts formed using various inorganic and organic acids commonly used in the industry.

[0033] In addition, the above-mentioned compound may include not only the above-mentioned salt, but also all salts, hydrates, solvates, derivatives, etc. that can be prepared according to conventional methods. The addition salt can be prepared by conventional methods, and can be prepared by dissolving it in a water-miscible organic solvent, such as acetone, methanol, ethanol, or acetonitrile, adding an excess amount of organic base or an aqueous solution of an inorganic base, and then precipitating or crystallizing it. Alternatively, the addition salt can be obtained by evaporating the solvent or the excess base from the mixture and then drying it, or by suction filtration of the precipitated salt.

[0034]

[0035] The above autophagy marker is a protein involved in the autophagy process and may include microtubule-associated protein light chain 3 (LC3).

[0036] In this specification, "autophagy" refers to a metabolic process in which eukaryotic cells digest their own organelles and old proteins, and signifies a protein degradation process essential for maintaining cellular homeostasis and genetic stability by breaking down aging or dysfunctional organelles and damaged or improperly constructed proteins.

[0037] The above autophagy labeling factor binding ligand is an LC3 binding ligand and may be a compound represented by the following chemical formula B that binds to LC3 (Compound B) or a pharmaceutically acceptable salt thereof:

[0038] <Chemical Formula B>

[0039]

[0040]

[0041] The above linker may be a carbon ring connecting the androgen receptor binding moiety and the autophagy labeling factor binding ligand, and preferably may be a (C1-C20)alkylene connecting the nitrogen atom of the piperazine ring of compound A and the hydroxyl group of compound B, but is not limited thereto.

[0042] One or more of the above androgen receptor binding moiety and autophagy marker binding ligand may be connected to the above linker.

[0043]

[0044] Preferably, the androgen receptor target degradation compound may comprise a compound represented by the following chemical formula 1 or 2 or a pharmaceutically acceptable salt thereof:

[0045] <Chemical Formula 1>

[0046]

[0047] <Chemical Formula 2>

[0048]

[0049]

[0050] The compound represented by the above chemical formula 1 can be named N-((1r,4r)-4-(3-chloro-4-cyanophenoxy)cyclohexyl)-4-(4-(10-((5-hydroxy-2-oxo-4-phenyl-2H-chromen-7-yl)oxy)decyl)piperazin-1-yl)benzamide [N-((1r,4r)-4-(3-chloro-4-cyanophenoxy)cyclohexyl)-4-(4-(10-((5-hydroxy-2-oxo-4-phenyl-2H-chromen-7-yl)oxy)decyl)piperazin-1-yl)benzamide], and the compound represented by the above chemical formula 2 is It can be named as 4,4'-(4,4'-(((2-oxo-4-phenyl-2H-chromene-5,7-diyl)bis(oxy))bis(decane-10,1-diyl))bis(piperazine-4,1-diyl))bis(N-((1r,4r)-4-(3-chloro-4-cyanophenoxy)cyclohexyl)benzamide) [4,4'-(4,4'-(((2-oxo-4-phenyl-2H-chromene-5,7-diyl)bis(oxy))bis(decane-10,1-diyl))bis(piperazine-4,1-diyl))bis(N-((1r,4r)-4-(3-chloro-4-cyanophenoxy)cyclohexyl)benzamide)].

[0051]

[0052] The androgen receptor target degradation compound according to the present invention can act as an autophagosome-tethering compound (ATTEC) that targets androgen receptors. ATTEC is one of the targeted protein degradation (TPD) technologies, and it is differentiated from the existing representative TPD technology, the proteolysis-targeting chimera (PROTAC), in that the autophagosome is larger than the proteasome in size for accommodating the protein to be degraded, and its degradation activity is not dependent on E3 ligase.

[0053] The androgen receptor target degradation compound according to the present invention has excellent binding affinity to androgen receptors and binding affinity to LC3, an autophagy marker, so it can activate autophagy and ultimately target and degrade androgen receptors, thereby enabling the treatment of androgen receptor-related diseases.

[0054]

[0055] The present invention provides a pharmaceutical composition for the prevention or treatment of cancer, comprising the above-mentioned androgen receptor target degradation compound as an active ingredient.

[0056] Preferably, the cancer disease may be an androgen receptor-related cancer disease and may be selected from, for example, prostate cancer, breast cancer, or triple-negative breast cancer, but is not limited thereto.

[0057] Alternatively, the present invention provides a pharmaceutical composition for the prevention or treatment of androgen receptor-related diseases, comprising the above-mentioned compound for target degradation of androgen receptors as an active ingredient.

[0058] In addition to the above-mentioned cancer, the above-mentioned androgen receptor-related diseases include, but are not limited to, asthma, multiple sclerosis, Kenney's disease, ciliosis, cleft palate, diabetes, heart disease, hypertension, inflammatory bowel disease, obesity, refractive errors, infertility, Angelman syndrome, Canavan disease, chronic digestive disease, Charcot-Marie-Tooth disease, cystic fibrosis, Duchenne muscular dystrophy, hemochromatosis, hemophilia, Klinefelter's syndrome, neurofibromatosis, phenylketonuria, polycystic kidney disease, sickle cell disease, Tay-Sachs disease, and Turner syndrome.

[0059]

[0060] In this specification, the term "pharmaceutical composition" means a composition administered for the purpose of preventing or treating a specific disease, and for the purposes of the present invention, means administered for the treatment of cancer or androgen receptor-related diseases, or diseases or complications caused by such diseases.

[0061] The pharmaceutical composition according to the present invention may be prepared according to conventional methods in the pharmaceutical field. The pharmaceutical composition may be combined with a suitable pharmaceutically acceptable carrier depending on the formulation, and may be prepared by further including excipients, diluents, dispersants, emulsifiers, buffers, stabilizers, binders, disintegrants, solvents, etc., as needed. The suitable carrier, etc., may be selected differently depending on the dosage form and formulation, as it does not impair the activity and properties of the compound or salt thereof according to the present invention.

[0062] Examples of carriers, excipients, diluents, etc. that may be included in the above pharmaceutical composition include lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, acacia gum, alginate, gelatin, calcium phosphate, calcium silicate, cellulose, methyl cellulose, microcrystalline cellulose, polyvinylpyrrolidone, water, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate, mineral oil, etc.

[0063] The above pharmaceutical composition can be applied in any dosage form, and more specifically, can be formulated and used in oral dosage forms, topical preparations, suppositories, and parenteral dosage forms of sterile injectable solutions according to conventional methods, but is not limited thereto.

[0064] Among the above oral formulations, solid formulations may be in the form of tablets, pills, powders, granules, capsules, etc., and may be prepared by mixing at least one excipient, such as starch, calcium carbonate, sucrose, lactose, sorbitol, mannitol, cellulose, gelatin, etc., and may also include lubricants such as magnesium stearate and talc in addition to simple excipients. Furthermore, in the case of capsule formulations, in addition to the substances mentioned above, liquid carriers such as fatty oils may be further included. Among the above oral formulations, liquid formulations may include suspensions, liquid formulations, emulsions, syrups, etc., and may include various excipients, such as humectants, sweeteners, flavorings, and preservatives, in addition to commonly used simple diluents such as water and liquid paraffin.

[0065] The parenteral formulations described above may include sterile aqueous solutions, non-aqueous solvents, suspensions, emulsions, lyophilized formulations, and suppositories. As non-aqueous solvents and suspensions, propylene glycol, polyethylene glycol, vegetable oils such as olive oil, and injectable esters such as ethyl oleate may be used. As bases for suppositories, witepsol, macrogol, Tween 61, cacao oil, laurin oil, glycerogelatin, etc., may be used. However, they are not limited thereto, and any suitable formulation known in the art may be used.

[0066]

[0067] The pharmaceutical composition according to the present invention can be administered in a pharmaceutically effective amount.

[0068] In this specification, "pharmaceuticalally effective amount" means an amount sufficient to treat a disease with a reasonable benefit / risk ratio applicable to medical treatment and that does not cause adverse effects.

[0069] The effective dose level of the above pharmaceutical composition may be determined differently depending on factors including the purpose of use, the patient's age, gender, weight and health status, type and severity of the disease, drug activity, sensitivity to the drug, method of administration, time of administration, route of administration and elimination rate, duration of treatment, drugs used in combination or concurrently, and other factors well known in the medical field. For example, although not constant, it may generally be administered at a dose of 0.001 to 1000 mg / kg, preferably 0.01 to 100 mg / kg, once or several times daily. The above dosage does not limit the scope of the present invention in any way.

[0070] The above pharmaceutical composition may be administered to any animal that may develop cancer or androgen receptor-related diseases, and said animals may include, for example, humans and primates, as well as livestock such as cattle, pigs, horses, and dogs.

[0071] The above pharmaceutical composition may be administered via a suitable route of administration depending on the formulation form, and may be administered via various oral or parenteral routes as long as it can reach the target tissue. The method of administration may be administered by conventional methods, such as oral, rectal or intravenous, intramuscular, topical application, respiratory inhalation, intradural or intracerebroventricular injection, without needing to be particularly limited.

[0072] The above pharmaceutical composition may be used alone for the prevention or treatment of cancer or androgen receptor-related diseases, or may be used in combination with surgery or other drug treatments.

[0073]

[0074] In addition, the present invention provides a food composition for preventing or improving cancer, comprising the above-mentioned androgen receptor target degradation compound as an active ingredient.

[0075] Alternatively, the present invention provides a food composition for preventing or improving androgen receptor-related diseases, comprising the above-mentioned compound for degrading androgen receptor targets as an active ingredient.

[0076] The food composition according to the present invention may include all forms such as functional food, nutritional supplement, health food, and food additives.

[0077] The term "health functional food" above refers to a food manufactured and processed using raw materials or ingredients that have functional properties beneficial to the human body, and is a food with high medical or medical effects that is processed to efficiently exhibit biological regulatory functions in addition to providing nutrition; it may be used interchangeably with terms known in the industry, such as functional food.

[0078] The food composition according to the present invention may be prepared as a powder, granule, tablet, capsule, syrup, or beverage, etc., for the purpose of preventing or improving cancer or androgen receptor-related diseases. There are no restrictions on the form in which the food may take place, and it may include all foods in the conventional sense. For example, beverages and various drinks, fruits and their processed foods (canned fruit, jam, etc.), fish, meat and its processed foods (ham, bacon, etc.), breads and noodles, cookies and snacks, dairy products (butter, cheese, etc.), etc., are possible, and it may include all functional foods in the conventional sense. In addition, it may also include foods used as animal feed.

[0079] The above food composition may be prepared by further including food-grade acceptable food additives commonly used in the industry and appropriate other auxiliary ingredients. Unless otherwise stipulated, suitability as a food additive may be determined by the specifications and standards for the relevant item in accordance with the general provisions and general test methods of the Food Additives Codex approved by the Ministry of Food and Drug Safety. Items listed in the above 'Food Additives Codex' may include, for example, chemically synthesized products such as ketones, glycine, calcium citrate, nicotinic acid, and cinnamon acid; natural additives such as persimmon dye, licorice extract, crystalline cellulose, sorghum dye, and guar gum; and mixed preparations such as L-sodium glutamate preparations, alkaline noodle additives, preservative preparations, and tar dye preparations.

[0080] The above other auxiliary ingredients may additionally contain, for example, flavoring agents, natural carbohydrates, sweeteners, vitamins, electrolytes, coloring agents, pectic acid, alginic acid, organic acids, protective colloidal thickeners, pH adjusters, stabilizers, preservatives, glycerin, alcohol, carbonating agents, etc. In particular, the above natural carbohydrates may include monosaccharides such as glucose and fructose, disaccharides such as maltose and sucrose, polysaccharides such as dextrin and cyclodextrin, and sugar alcohols such as xylitol, sorbitol, and erythritol, and as sweeteners, natural sweeteners such as taumatin and stevia extract, or synthetic sweeteners such as saccharin and aspartame may be used.

[0081] The effective dose of the compound or its salt contained in the above food composition can be appropriately adjusted according to the purpose of use, such as the prevention or improvement of cancer or androgen receptor-related diseases. The above composition has the advantage of being made from food and thus free from side effects that may occur with the long-term use of general medicines, and due to its excellent portability, it can be consumed as an adjuvant for the prevention or improvement of cancer or androgen receptor-related diseases.

[0082] Hereinafter, the present invention will be described in detail with reference to examples to aid in understanding. However, the following examples are merely illustrative of the content of the present invention and the scope of the present invention is not limited to the following examples. The examples of the present invention are provided to more completely explain the present invention to those with average knowledge in the art.

[0083]

[0084] <Preliminary Experimental Example 1> Evaluation of Androgen Receptor (AR) Binding Affinity (AR-Luciferase Reporter assay)

[0085] The AR binding affinity of compound A was verified using the androgen receptor-luciferase reporter assay.

[0086] [Compound A]

[0087]

[0088] First, AR-luciferase reporter MDA-MB-453 stable breast cancer cell line (Cat# SL-0008) was cultured in a white-bottomed 96-well plate (10 4 The plates were treated with 2 nM of the AR cofactor [Dihydrotestosterone (DHT)] and the drug (Compound A) at evaluation concentrations for 24 hours. The medium was changed with PBS (100 μL / well), and an equal amount of detection reagent (ONE-Glo Luciferase assay) was added. The plates were then shaken at room temperature for 5 minutes. Luciferase activity was measured using a Tecan i-control plate reader. The binding affinity to the DHT competitive androgen receptor (AR) was evaluated using the RLU (Relative Luciferase Unit) value.

[0089] As a result, as shown in Fig. 1, the IC of the above compound 50 The value was evaluated to be 0.687 μM, and based on this, the synthesis of an androgen receptor target compound was designed.

[0090]

[0091] <Preliminary Experimental Example 2> LC3 Purification and Evaluation of LC3 Binding Strength

[0092] The LC3B (1-125) gene was cloned into the pGEX41 vector, and a transformed plasmid was obtained using BL21 (DE3) competent cells. To overexpress the plasmid, the cells were incubated at 37°C, followed by the addition of 0.5 mM IPTG at an OD of 0.6 and incubation at 20°C for 20 hours. After a cell lysis pulse of 2 seconds, the cells were centrifuged at 18,000 rpm for 1 hour to obtain only the soluble protein. The protein was purified in the first stage using affinity (GST tag) buffer 20 mM Tris-HCl (pH 7.5), 150 mM NaCl, and 10 mM GSH to obtain the GST-tagged protein. For the second stage of purification, thrombin was added to cleave the GST tag, and after tag removal via dialysis, the protein was obtained using an affinity column. Finally, for the third stage of purification, pure protein was obtained using SEC (S75 column) buffer 20 mM Tris-HCl (pH 7.5) and 150 mM NaCl.

[0093] The binding strength of the following two compounds (B, C) was measured using ITC (Isothermal Titration Calorimetry). 0.05 mM of LC3B (1-125) protein and 0.5 mM of DP and GW compounds were dissolved in 50 mM HEPES pH 7.5 100 mM NaCl 2% DMSO, and the compounds were analyzed by titrating 200 μL at 25°C 25 times.

[0094] [Compound B]

[0095]

[0096] [Compound C]

[0097]

[0098] As a result, as shown in Fig. 2, the two compounds are each Kd It was evaluated to be 7.46 and 10.2 μM, and among these, compound B (K showing a stronger binding affinity) d The synthesis of an androgen receptor target compound was designed using 7.46 μM.

[0099]

[0100] <Example 1> Synthesis of a Novel Compound

[0101] Based on the above preliminary experimental examples 1 and 2, a novel androgen receptor target compound was synthesized according to the following reaction scheme 1.

[0102] [Reaction Equation 1]

[0103]

[0104] a) TFA, DCM; b) DIPEA, HATU, DMF; c) DIPEA, DMF; d) MsCl, TEA, DMF; e) K2CO3, KI, DMF

[0105]

[0106] 1. Preparation of Compounds 1 and 2

[0107] 1) Step 1

[0108]

[0109] 1 g (2.85 mmol) of tert-butyl((1r,4r)-4-(3-chloro-4-cyanophenoxy)cyclohexyl)carbamate was dissolved in 15 mL of dichloromethyl, and trifluoroacetic acid (8.0 eq) was slowly added dropwise to the reaction mixture. The mixture was then stirred at room temperature for 3 hours. After the reaction was complete, the pH of the reaction mixture was adjusted to 7.0 using a saturated sodium bicarbonate solution, and the mixture was extracted with dichloromethyl. The resulting dichloromethyl layer was washed with water. Then, the organic layer was dehydrated with anhydrous sodium sulfate and filtered, and the solvent was removed from the filtrate under reduced pressure. The resulting mixture was separated and purified by MPLC to obtain 431 mg of 4-(((1r,4r)-4-aminocyclohexyl)oxy)-2-chlorobenzonitrile with a yield of 60.3%.

[0110]

[0111] 2) Step 2

[0112]

[0113] 70 mg (0.279 mmol) of 4-(((1r,4r)-4-aminocyclohexyl)oxy)-2-chlorobenzonitrile was dissolved in 1 mL of dimethylformamide, and N,N-diisopropylethylamine (3.0 eq), 4-(4-(tert-butoxycarbonyl)piperazine-1-yl)benzoic acid (1.5 eq), and 1-[bis(dimethylamino)methylene]-1H-1,2,3-triazolo[4,5-b]pyridinium 3-oxide hexafluorophosphate (1.5 eq) were added to the reaction mixture. The mixture was stirred at 70°C for 5 hours. After the reaction was complete, 1 M hydrochloric acid was added and the mixture was extracted with dichloromethyl, and the resulting dichloromethyl layer was washed with water. Then, anhydrous sodium sulfate was added to the organic layer to dehydrate it, and the mixture was filtered. The filtrate was evaporated under reduced pressure and separated and purified by MPLC to obtain 99 mg of tert-butyl 4-(4-(((1r,4r)-4-(3-chloro-4-cyanophenoxy)cyclohexyl)carbamoyl)phenyl)piperazine-1-carboxylate with a yield of 65.8%.

[0114]

[0115] 3) Step 3

[0116]

[0117] 41.6 mg (0.077 mmol) of tert-butyl 4-(4-(((1r,4r)-4-(3-chloro-4-cyanophenoxy)cyclohexyl)carbamoyl)phenyl)piperazine-1-carboxylate prepared in Step 2 above was dissolved in 1 mL of dichloromethyl, and then trifluoroacetic acid (8.0 eq) was slowly added dropwise to the reaction mixture. The mixture was then stirred at room temperature for 3 hours. After the reaction was complete, the pH of the reaction mixture was adjusted to 7.0 using a saturated sodium bicarbonate solution, and the mixture was extracted with dichloromethyl. The resulting dichloromethyl layer was washed with water. Then, the organic layer was dehydrated with anhydrous sodium sulfate and filtered, and the solvent of the filtrate was removed under reduced pressure. The mixture obtained at this time was separated and purified by MPLC to obtain 31.7 mg of N-((1r,4r)-4-(3-chloro-4-cyanofenoxy)cyclohexyl)-4-(piperazine-1-yl)benzamide with a yield of 92%.

[0118]

[0119] 4) Step 4

[0120]

[0121] 10 mg (0.023 mmol) of N-((1r,4r)-4-(3-chloro-4-cyanofenoxy)cyclohexyl)-4-(piperazine-1-yl)benzamide prepared in Step 3 above was dissolved in 1 mL of dimethylformamide, and N,N-diisopropylethylamine (3.0 eq) was added. Then, 10-bromodecan-1-ol (1.3 eq) was added to the reaction mixture and stirred at 60°C for 6 hours. After the reaction was completed, the solvent was removed from the reaction mixture under reduced pressure. This was separated and purified by MPLC to obtain 9 mg of N-((1r,4r)-4-(3-chloro-4-cyanofenoxy)cyclohexyl)-4-(4-(10-hydroxydecyl)piperazine-1-yl)benzamide with a yield of 65%.

[0122]

[0123] 5) Step 5

[0124]

[0125] 38 mg (0.064 mmol) of N-((1r,4r)-4-(3-chloro-4-cyanofenoxy)cyclohexyl)-4-(4-(10-hydroxydecyl)piperazine-1-yl)benzamide was dissolved in 1 mL of dimethylformamide, and TEA (2.0 eq) was added. Then, after cooling to 0°C, Mesyl-Cl (1.1 eq) was added to the reaction mixture. The above mixture was stirred at room temperature for 1 hour. After the reaction was complete, the solvent was removed from the reaction mixture under reduced pressure, and 36 mg of 10-(4-(4-(((1r,4r)-4-(3-chloro-4-cyanofenoxy)cyclohexyl)carbamoyl)phenyl)piperazine-1-yl)decyl methanesulfonate was obtained. The yield was 83%.

[0126]

[0127] 6) Step 6

[0128]

[0129] 30 mg (0.045 mmol) of 10-(4-(4-(((1r,4r)-4-(3-chloro-4-cyanofenoxy)cyclohexyl)carbamoyl)phenyl)piperazine-1-yl)decyl methanesulfonate prepared in Example 5 above was dissolved in 1 mL of dimethylformamide, and potassium carbonate (5.0 eq) and potassium iodide (0.1 eq) were added. Then, the mixture was stirred at 60°C for 18 hours. After the reaction was complete, water was added and the mixture was filtered. After drying the filter cake under vacuum, it was purified by MPLC and PrepLC to obtain 4 mg, yield 12% N-((1r,4r)-4-(3-chloro-4-cyanofenoxy)cyclohexyl)-4-(4-(10-((5-hydroxy-2-oxo-4-phenyl-2H-cromen-7-yl)oxy)decyl)piperazine-1-yl)benzamide (Compound 1; DN302554-1) and 5 mg, yield 8% 4,4'-((((2-oxo-4-phenyl-2H-cromen-5,7-diyl)bis(oxy))bis(decane-10,1-diyl))bis(piperazine-4,1-diyl))bis(N-((1r,4r)-4-(3-chloro-4-cyanofenoxy)cyclohexyl)benzamide)(compound 2; DN302555-1) was obtained (Fig. 3).

[0130]

[0131] Compound 1: 1H NMR (400 MHz, CDCl3)δ12.52 (s, 1H), 7.72 (d,J= 8.7 Hz, 1H), 7.56 (d,J= 8.7 Hz, 1H), 7.35 - 7.34 (m, 3H), 7.25 - 7.23 (m, 2H), 6.99 (sd,J= 2.3 Hz, 1H), 6.91 (d,J= 8.7 Hz, 2H), 6.85 (dd,J= 8.8, 2.3 Hz, 1H), 6.48 (sd,J= 2.0 Hz, 1H), 6.30 (sd,J= 1.9 Hz, 1H), 6.07 (d,J= 7.8 Hz, 1H), 5.92 (s, 1H), 4.33 - 4.27 (m, 1H), 4.09 - 4.01 (m, 1H), 3.82 - 3.71 (m, 4H), 3.63 (t,J= 6.3 Hz, 2H), 3.43 - 3.38 (m, 2H), 3.04 - 2.96 (m, 4H), 2.24 - 2.17 (m, 4H), 1.73 - 1.63 (m, 8H), 1.50 - 1.40 (m, 3H), 1.21 - 1.18 (m, 2H), 1.15 - 1.05 (m, 5H), 0.92 - 0.85 (m, 2H). LCMS: 831 [M+H + ].

[0132] 화합물2: 1H NMR (400 MHz, CDCl3)δ12.95 (s, 2H), 7.70 (d,J= 8.5 Hz, 4H), 7.56 (d,J= 8.7 Hz, 2H), 7.36 - 7.35 (m, 3H), 7.26 - 7.24 (m, 2H), 6.99 (sd,J= 2.3 Hz, 2H), 6.89 (d,J= 8.5 Hz, 4H), 6.85 (dd,J= 8.8, 2.3 Hz, 2H), 6.48 (sd,J= 2.2 Hz, 1H), 6.21 (sd,J= 2.1 Hz, 1H), 6.05 (d,J=7.2 Hz, 2H), 5.96 (s, 1H), 4.33 - 4.26 (m, 2H), 4.08 - 3.99 (m, 4H), 3.78 - 3.76 (m, 8H), 3.64 (t,J= 6.2 Hz, 2H), 3.42 - 3.36 (m, 4H), 3.07 - 3.03 (m, 4H), 2.97 - 2.92 (m, 4H), 2.22 - 2.16 (m, 10H), 1.83 - 1.77 (m, 6H), 1.72 - 1.62 (m, 4H), 1.47 - 1.32 (m, 18H), 1.24 - 1.20 (m, 2H), 1.16 - 1.12 (m, 2H), 1.10 - 1.02 (m, 4H), 0.86 - 0.79 (m, 2H). LCMS: 1409 [M+H + ].

[0133]

[0134] <Experimental Example 1> Evaluation of Androgen Receptor Degradation Efficacy (AR Western Blot assay)

[0135] The MDA-MB-453 cell line was cultured in 6-well plates (5×10⁶). 5 / well). Cell lysates were obtained and quantified after 24 hours of treatment with drugs at different concentrations [compounds 1 and 2 (TFA salts), and as positive controls, enzalutamide, an AR-DHT binding inhibitor, and ARV-110, a proteasome-mediated AR degradation drug (AR-PROTAC)]. Protein samples were each mixed with 4× sample buffer and incubated at 95°C for 5 minutes. SDS-PAGE was performed at 100 V for 80 minutes, followed by membrane transfer at 100 V for 70 minutes. Membrane blocking was performed using 5% skim milk for 60 minutes, and the samples were incubated at 4°C for 16 hours after treatment with the AR antibody (Cat#. AB133273). After treatment with Anti-Rabbit IgG HRP-conjugated antibody (Cat#. HAF008) and incubation at room temperature for 60 minutes, the results were analyzed using a LAS4000 instrument.

[0136] After an initial evaluation of the two compounds, DN302554 and DN302555, at a single concentration of 10 μM, and additional evaluations at 0.1 and 1 μM, the on-target efficacy of the compounds could be confirmed by comparing the concentration-dependent degradation phenomenon of AR with ARV-110, as shown in Figure 4.

[0137]

[0138] Foregoing, specific parts of the present invention have been described in detail. It is evident to those skilled in the art that such specific descriptions are merely preferred embodiments and do not limit the scope of the invention. That is, the actual scope of the invention is defined by the appended claims and their equivalents.

Claims

1. It has the structure of an androgen receptor (AR) binding moiety-linker-autophagy marker binding ligand, and The above androgen receptor binding moiety is a compound represented by the following chemical formula A (compound A) or a pharmaceutically acceptable salt thereof; <Chemical Formula A> The above autophagy labeling factor binding ligand is a compound represented by the following chemical formula B (Compound B) or a pharmaceutically acceptable salt thereof that binds to microtubule-associated protein light chain 3 (LC3); <Chemical Formula B> A compound for targeting androgen receptor degradation, characterized in that the linker is a (C1-C20)alkylene connecting the nitrogen atom of the piperazine ring of compound A and the hydroxyl group of compound B, and one or more compounds A and B are connected to the linker.

2. In Paragraph 1, The above-mentioned androgen receptor target degradation compound is, A compound characterized by comprising a compound represented by the following chemical formula 1 or 2 or a pharmaceutically acceptable salt thereof: <Chemical Formula 1> <Chemical Formula 2> 3. In Paragraph 1 or 2, The above-mentioned androgen receptor target degradation compound is, A compound characterized by acting as an autophagosome-tethering compound (ATTEC) by targeting androgen receptors.

4. A pharmaceutical composition for the prevention or treatment of cancer, comprising as an active ingredient a compound for targeting androgen receptor degradation according to claim 1 or 2.

5. In Paragraph 4, The above cancer disease is, A pharmaceutical composition characterized by being an androgen receptor-related cancer.

6. A pharmaceutical composition for the prevention or treatment of androgen receptor-related diseases, comprising as an active ingredient a compound for targeting androgen receptor degradation according to claim 1 or 2.

7. In Paragraph 6, The above-mentioned androgen receptor-related disease is, A pharmaceutical composition characterized by being selected from asthma, multiple sclerosis, Kenney's disease, ciliosis, cleft palate, diabetes mellitus, heart disease, hypertension, inflammatory bowel disease, obesity, refractive error, infertility, Angelman syndrome, Canavan disease, chronic digestive disease, Charcot-Marie-Tooth disease, cystic fibrosis, Duchenne muscular dystrophy, hemochromatosis, hemophilia, Klinefelter's syndrome, neurofibromatosis, phenylketonuria, polycystic kidney disease, sickle cell disease, Tay-Sachs disease, or Turner syndrome.

8. A food composition for preventing or improving cancer, comprising as an active ingredient a compound for targeting androgen receptor degradation according to claim 1 or 2.

9. A food composition for preventing or improving androgen receptor-related diseases, comprising as an active ingredient a compound for targeting androgen receptor degradation according to claim 1 or 2.

Images