Proteolysis targeting chimera with 6,6-dimethylbicyclo[3.1.1]-2-methylamine as hydrophobic group, preparation method, pharmaceutical composition and application thereof

Abstract

The application belongs to the technical field of chemical drugs, and particularly relates to a protein degrading agent with 6,6-dimethylbicyclo[3.1.1]-2-methylamine as a hydrophobic group, a preparation method, a pharmaceutical composition and application thereof. The protein degrading agent with 6,6-dimethylbicyclo[3.1.1]-2-methylamine as a hydrophobic group has a structural formula as shown in formula I, and Linker is any chemically feasible connecting structure. The protein degrading agent provided by the application can effectively induce the degradation of AR and AR-V7 in a cancer cell line. Compared with the AR protein degrading agent based on adamantane as a hydrophobic tag reported by the previous person, the protein degrading agent has a significantly improved protein degradation effect and in-vivo drug efficacy, can exhibit excellent in-vivo anti-prostate cancer effect in a low drug dosage, has high safety, can be applied to the preparation of an AR degrading agent, and is suitable for the development of a cancer drug such as a prostate cancer drug. Formula I.

Description

TECHNICAL FIELD

[0001] The present application belongs to the technical field of chemical drugs, and particularly relates to a protein degrader with 6,6-dimethylbicyclo[3.1.1]-2-methylamine as a hydrophobic group, a preparation method, a pharmaceutical composition and an application thereof. BACKGROUND

[0002] Hydrophobic Tag (Hyt) bifunctional molecules are composed of three parts: target protein ligand, linker, and hydrophobic group. By connecting a large and hydrophobic group to a small molecule that can bind to the target, such a double-headed molecule, after binding to the target, will be mistakenly considered by the intracellular protein repair mechanism as a misfolded part of the target protein, and then it can be folded by chaperone proteins and degraded by proteasomes. The hydrophobic group in the Hyt molecule is usually small in molecular weight, and thus may have higher solubility and drugability. The research and development of degraders based on hydrophobic tags is still in the exploratory stage. On the one hand, there are fewer reported hydrophobic tag fragments, and there is still a lot of room for optimization in terms of degradation activity and physicochemical properties. On the other hand, the exact degradation mechanism has not been clearly defined. Therefore, exploring more hydrophobic fragments with high activity and excellent physicochemical properties and clarifying their corresponding degradation mechanisms are crucial for the development of hydrophobic tag fragments to clinical applications.

[0003] Androgen receptor (AR) is a clinically proven target for the treatment of human prostate cancer. Androgen receptor antagonists are effective for the treatment of metastatic castration-resistant prostate cancer (mCRPC), can significantly improve the survival of prostate cancer patients, and have good tolerance. However, current clinical androgen receptor antagonists (enzalutamide, etc.) are usually resistant to 18 months of patient medication. In most patients resistant to enzalutamide, the androgen receptor signaling pathway is still active, and traditional AR inhibitors and antagonists cannot affect the expression levels of AR protein and AR-V7 and other truncated protein. Therefore, targeting androgen receptor protein degradation may be a very promising treatment strategy, which may be more effective than androgen receptor antagonists. A team previously developed protein degraders SARD279 and SARD033 that can degrade AR based on the hydrophobic tag technology.

[0004]

[0005] Structure of SARD279, SARD033, and RU59063

[0006] These two degraders are obtained by connecting AR ligand RU59063 with adamantane as a hydrophobic tag through polyethylene glycol of different lengths (Formula 1), wherein SARD279 can degrade 50% of AR protein in LNCaP cells at a concentration of 1 μM (DC50 However, neither the conventional PROTAC degrader nor the hydrophobic tag degrader has a significant effect on the AR-V7 protein level. Moreover, the conventional AR degrader has a relatively low degradation effect, poor liver microsomal and plasma metabolic stability, poor solubility and is difficult to be orally taken, and is difficult to be used for further treatment and development in clinic. Therefore, it is necessary to develop a new AR protein degrader with better degradation effect to meet the clinical needs. SUMMARY

[0007] The present application aims to solve the problems of the prior art, and provides a protein degrader with 6,6-dimethylbicyclo[3.1.1]-2-methylamine as a hydrophobic group, a preparation method, a pharmaceutical composition and an application thereof. Specifically, the following technical solutions are adopted:

[0008] According to a first aspect of the present application, a protein degrader with 6,6-dimethylbicyclo[3.1.1]-2-methylamine as a hydrophobic group is provided, and the structural formula is shown as formula I:

[0009] Formula I; wherein Linker is any chemically feasible connecting structure.

[0010] The present application provides a protein degrader with 6,6-dimethylbicyclo[3.1.1]-2-methylamine as a hydrophobic group. In the bifunctional molecule, 6,6-dimethylbicyclo[3.1.1]-2-methylamine is used as a hydrophobic group, and enzalutamide parent nucleus is used as a target protein ligand. The present application can screen protein degraders capable of effectively degrading AR by replacing different Linkers. On this basis, the AR protein degrader prepared by the present application can efficiently induce the degradation of AR and AR-V7 in human prostate cancer cell lines (22Rv1 cell lines) in a dose-dependent manner, and exhibits extremely excellent in vivo anti-prostate cancer effect.

[0011] Preferably, the Linker is a saturated aliphatic chain or an unsaturated aliphatic chain. By selecting different types and lengths of ideal linkers, the purpose of not affecting the binding of the two proteins in space and maintaining their binding can be achieved.

[0012] Preferably, the protein degrader with 6,6-dimethylbicyclo[3.1.1]-2-methylamine as a hydrophobic group has a molecular structure of any one of formula II, formula III or formula IV:

[0013] Formula II;

[0014] Formula III;

[0015] Formula IV;

[0016] wherein, in formula II, m is any positive integer from 1 to 8; in formula III, n is any positive integer from 1 to 10; and in formula IV, o is any positive integer from 1 to 8.

[0017] More preferably, the molecular structure of the protein degrader with 6,6-dimethylbicyclo[3.1.1]-2-methylamine as the hydrophobic group is shown in formula V:

[0018] Formula V.

[0019] The experimental result data show that the compound shown in formula V above can efficiently induce the degradation of AR and AR-V7 in human prostate cancer cell line 22Rv1 in a dose-dependent manner, and exhibits the most excellent anti-prostate cancer effect in vivo, with a tumor inhibition rate of 94.2%.

[0020] According to the second aspect of the present application, a preparation method of the above-mentioned protein degrader with 6,6-dimethylbicyclo[3.1.1]-2-methylamine as the hydrophobic group is also provided, and the preparation route is route one, route two or route three:

[0021] Route one:

[0022] ;

[0023] Route two:

[0024] ;

[0025] Route three:

[0026] .

[0027] The above route is simple, the raw materials are cheap and easy to obtain, and the overall yield of the reaction is high. In route one, enzalutamide nucleus A is used to substitute with different lengths of dibromopolyethylene glycol (B1-B4) to obtain intermediate (C1-C4) under the action of K2CO3, DMF, and then react with compound D under the action of TEA to obtain final product E1-E4; in route two, 6,6-dimethylbicyclo[3.1.1]-2-methylamine (D) is used to react with different lengths of dibromoalkane (F1-F3) under the action of TEA to obtain intermediate G1-G3, and then substitute with enzalutamide nucleus A under the action of K2CO3, DMF to obtain final product H1-H3; in route three, 6,6-dimethylbicyclo[3.1.1]-2-methylamine (D) is used to amide condense with different lengths of alkyne acid (I1-I5) under the action of HATU to obtain intermediate J1-J5, and then enzalutamide nucleus A and 2-azidoethyl-4-methylbenzenesulfonate (compound K) are substituted under the action of K2CO3, DMF to obtain intermediate L, and then Click reaction with J1-J5 to obtain final product M1-M5.

[0028] According to a third aspect of the present application, the above protein degrading agent with 6,6-dimethylbicyclo[3.1.1]-2-methylamine as a hydrophobic group or a pharmaceutically acceptable salt thereof can also be applied in the preparation of androgen receptor protein (AR) degrading agent.

[0029] Preferably, the above androgen receptor protein (AR) degrading agent can be used for the preparation of a drug for treating and / or preventing cancer related to AR abnormalities or resistance to traditional AR inhibitors. The related cancer includes prostate cancer, breast cancer, ovarian cancer, endometrial cancer. It is preferably used for the treatment of prostate cancer.

[0030] According to a fourth aspect of the present application, a pharmaceutical composition is also provided, which comprises the above protein degrading agent with 6,6-dimethylbicyclo[3.1.1]-2-methylamine as a hydrophobic group or a pharmaceutically acceptable salt thereof as the main active ingredient. The above pharmaceutical composition comprises one or more pharmaceutically acceptable excipients or carriers. The above excipients include at least one of gum arabic, sugar syrup, lanolin, and starch. The excipients are stable in nature, have no compatibility with the main drug, do not produce side effects, do not affect the therapeutic effect, are not easy to deform, dry, mold, and insect-bored at room temperature, are harmless to the human body, have no physiological effects, do not produce chemical or physical effects with the main drug, and do not affect the content determination of the main drug. The solvent includes water, glycerol or ethanol.

[0031] The protein degrading agent with 6,6-dimethylbicyclo[3.1.1]-2-methylamine as a hydrophobic group is obtained, the preparation process is simple and easy to operate, various protein degrading agents are obtained through different paths, and the protein degrading agent can be applied to preparation of an AR degrading agent, the obtained protein degrading agent has higher protein degrading activity effect compared with a positive control SARD279, and the drug efficacy can be effectively improved. In addition, the protein degrading agent can also constitute a drug composition, has a certain inhibitory effect on the proliferation of various tumor cells, and is suitable for the development of a cancer drug for treating prostate cancer and the like. BRIEF DESCRIPTION OF DRAWINGS

[0032] Figure 1 The figure shows the screening diagram of the concentration-dependent degradation of AR and AR-V7 protein of compounds E2, M3, M4, M5 and M6 in the 22Rv1 cell line;

[0033] Figure 2 The figure shows the concentration-dependent degradation of AR and AR-V7 protein of compound M4 at (A) five concentration gradients of 0, 1, 10, 100 and 1000 nM; (B) six concentration gradients of 0, 12.5, 25, 50, 100 and 200 nM; and (C) seven time gradients of 0, 2, 4, 8, 16, 24 and 48 h. DETAILED DESCRIPTION

[0034] The concept, specific structure and generated technical effects of the present application will be described clearly and completely in combination with the embodiments and the drawings below, so as to fully understand the purposes, schemes and effects of the present application. It should be noted that the embodiments in the present application and the features in the embodiments can be combined with each other without conflict.

[0035] Embodiment 1

[0036] A protein degrading agent with 6,6-dimethylbicyclo[3.1.1]-2-methylamine as a hydrophobic group

[0037] (1) Preparation of compound E1

[0038] The structure of compound E1 is as follows:

[0039] Compound E1; the preparation process is as follows:

[0040] Step 1: Preparation of compound C1

[0041] Compound A (405 mg, 1.00 mmol) and compound B1 (2,2'-dibromo diethyl ether, 232 mg, 1.00 mmol) were dissolved in N,N-dimethylformamide (3 mL), potassium carbonate (277 mg, 2.00 mmol) was added, and the reaction was allowed to proceed at room temperature overnight. After the reaction was completed, the reaction was diluted with water (12 mL) and extracted with ethyl acetate (3 x 10 mL), and then the organic phase was combined and washed with a saturated NaCl solution (12 mL). The organic phase was dried over anhydrous sodium sulfate, concentrated, and the obtained crude product was purified by silica gel column chromatography (petroleum ether: ethyl acetate = 6: 1) to obtain compound C1.

[0042] Compound A, compound B1, and compound C1 used in the above preparation process have the following structures:

[0043] Compound A, Compound B1,

[0044] Compound C1;

[0045] Compound C1 was detected, and the detection results are as follows: HRMS (ESI) calculated for C 23 H 22 BrF3N3O3S + [M+H] + : 556.0512, found. 556.0513.

[0046] Step 2: Preparation of compound E1

[0047] Compound C1 (278 mg, 0.50 mmol) and compound D (76.2 mg, 0.500 mmol) were dissolved in tetrahydrofuran (5 mL), triethylamine (139 μL, 1.00 mmol) was added, and the reaction was allowed to proceed at room temperature for 1 h. After the reaction was completed, the tetrahydrofuran was evaporated, diluted with water (8 mL), and extracted with ethyl acetate (3 x 8 mL), and then the organic phase was combined and dried over anhydrous sodium sulfate. The obtained crude product was purified by silica gel column chromatography (dichloromethane:methanol = 22: 1) to obtain compound E1.

[0048] Compound D used in the above preparation process has the following structure:

[0049] Compound D;

[0050] Compound E1 was detected, and the detection results are as follows: 1H NMR (400 MHz, Chloroform- d ) δ8.13 (d, J = 2.0 Hz, 1H), 8.03 (d, J = 7.5 Hz, 1H), 7.69 (dd, J = 7.5, 2.1 Hz, 1H),7.13 (s, 1H), 7.12 (q, J = 4.0 Hz, 1H), 6.89 – 6.81 (m, 2H), 4.13 (s, 2H), 3.67(d, J = 0.7 Hz, 2H), 3.63 – 3.49 (m, 2H), 2.90 – 2.74 (m, 3H), 2.53 (d, J = 0.6Hz, 2H), 1.75 (d, J = 13.0 Hz, 1H), 1.62 (d, J = 13.0 Hz, 1H), 1.58 – 1.46 (m,10H), 1.31 (d, J = 1.3 Hz, 1H), 1.30 – 1.22 (m, 2H), 0.97 (s, 3H), 0.92 (s, 3H). 13 C NMR (100 MHz, Chloroform- d ) δ 172.88, 171.30, 158.47, 136.87, 135.57,133.83, 133.64, 125.66, 125.33, 124.79, 124.48, 119.43, 117.44, HRMS (ESI) calculated for C 33 H 40 F3N4O3S + [M+H] + 629.2768 was found. 629.2766.

[0051] (2) Preparation of compound E2

[0052] The structure of compound E2 is as follows:

[0053] Compound E2; its preparation process is as follows:

[0054] The specific preparation method is: according to the synthesis steps of compound E1, only 2,2'-dibromo diethyl ether is changed to 1,2-bis (2-bromoethoxy) ethane to obtain compound C2 compared with compound E1, and then compound C2 and compound D are reacted to obtain compound E2.

[0055] The structure of compound C2 in the above preparation process is as follows:

[0056] Compound C2;

[0057] Compound E2 is detected, and the detection results are as follows: 1 H NMR (400 MHz, Chloroform- d ) δ8.13 (d, J = 2.0 Hz, 1H), 8.03 (d, J = 7.5 Hz, 1H), 7.69 (dd, J = 7.5, 2.1 Hz, 1H),7.13 (s, 1H), 7.12 (q, J = 4.0 Hz, 1H), 6.89 – 6.81 (m, 2H), 4.13 (s, 2H), 3.69– 3.49 (m, 9H), 2.89 – 2.74 (m, 3H), 2.53 (d, J = 0.6 Hz, 2H), 1.75 (d, J = 13.0Hz, 1H), 1.62 (d, J = 13.0 Hz, 1H), 1.58 – 1.46 (m, 9H), 1.34 – 1.22 (m, 3H),0.97 (s, 3H), 0.92 (s, 3H). 13 C NMR (100 MHz, Chloroform- d) δ 172.88, 171.30,158.47, 136.87, 135.57, 133.83, 133.64, 125.66, 125.33, 124.79, 124.48,119.43, 117.44, 106.85, 71.81, 71.45, 71.29, 70.72, 68.98, 66.95, 54.49,50.80, 48.08, 45.22, 41.41, 39.07, 33.95, 30.50, 27.59, 23.78, 23.75. HRMS(ESI): m / z calcd for C 35 H 44 F3N4O4S + [M+H] + : 673.3030; found 673.3033.

[0058] (3) Preparation of compound E3

[0059] The structure of compound E3 is as follows:

[0060] Compound E3; its preparation process is as follows:

[0061] The specific preparation method is: according to the synthesis steps of compound E1, compound E3 is obtained, compared with compound E1, only 2,2'-dibromo diethyl ether is changed to 1,11-dibromo-3,6,9-trioxaundecane, to obtain compound C3, then compound C3 and compound D are reacted to obtain compound E3.

[0062] The structure of compound C3 in the above preparation process is as follows:

[0063] Compound C3;

[0064] Compound E3 is detected, and the detection results are as follows: 1 H NMR (400 MHz, Chloroform- d ) δ8.13 (d, J = 2.0 Hz, 1H), 8.03 (d, J = 7.5 Hz, 1H), 7.69 (dd, J = 7.5, 2.1 Hz, 1H),7.12 (q, J= 4.0 Hz, 1H), 7.11 (s, 1H), 6.89 – 6.81 (m, 2H), 4.13 (s, 2H), 3.69– 3.49 (m, 13H), 2.89 – 2.74 (m, 3H), 2.53 (d, J = 0.6 Hz, 2H), 1.75 (d, J = 13.0Hz, 1H), 1.62 (d, J = 13.0 Hz, 1H), 1.58 – 1.46 (m, 9H), 1.34 – 1.22 (m, 3H),0.97 (s, 3H), 0.92 (s, 3H). 13 C NMR (100 MHz, Chloroform- d ) δ 172.88, 171.30,158.47, 136.87, 135.57, 133.83, 133.64, 125.66, 125.33, 124.79, 124.48,119.43, 117.44, 106.85, 71.81, 71.54, 71.44, 71.37, 71.29, 70.72, 68.98,66.95, 54.49, 50.80, 48.08, 45.22, 41.41, 39.07, 33.95, 30.50, 27.59, 23.78,23.75. HRMS (ESI): m / z calcd for C 37 H 48 F3N4O5S + [M+H] + : 717.3292; found 717.3289.

[0065] (4) Preparation of compound E4

[0066] The structure of compound E4 is as follows:

[0067] Compound E4; its preparation process is as follows:

[0068] The specific preparation method is: according to the synthesis steps of compound E1, compound E4 is obtained, compared with compound E1, only 2,2'-dibromo diethyl ether is changed to 1,14-dibromo-3,6,9,12-tetraoxatetradecane to obtain compound C4, and then compound C4 and compound D are reacted to obtain compound E4.

[0069] The structure of compound C4 in the above preparation process is as follows:

[0070] Compound C4;

[0071] Compound E4 was detected, and the detection results are as follows: 1 H NMR (400 MHz, Chloroform- d ) δ 8.13 (d, J = 2.0 Hz, 1H), 8.03 (d, J = 7.4 Hz, 1H), 7.69 (dd, J = 7.5, 2.0 Hz, 1H),7.16 – 7.08 (m, 2H), 6.89 – 6.81 (m, 2H), 4.13 (s, 2H), 3.64 (d, J = 3.5 Hz,2H), 3.65 – 3.49 (m, 14H), 2.84 (s, 1H), 2.82 (d, J = 0.8 Hz, 1H), 2.80 – 2.74(m, 1H), 2.53 (s, 2H), 1.75 (d, J = 13.0 Hz, 1H), 1.62 (d, J = 13.0 Hz, 1H), 1.56(d, J = 1.5 Hz, 4H), 1.54 – 1.46 (m, 6H), 1.31 (d, J = 1.3 Hz, 1H), 1.30 – 1.22(m, 2H), 0.97 (s, 3H), 0.92 (s, 3H). 13 C NMR (100 MHz, Chloroform- d) δ 172.88, 171.30, 158.47, 136.87, 135.57, 133.83, 133.64, 125.66, 125.33, 124.79, 124.48, 119.43, 117.44, 106.85, 71.81, 71.53, 71.44, 71.37, 71.29, 70.72, 68.98, 66.95, 54.49, 50.80, 48.08, 45.22, 41.41, 39.07, 33.95, 31.32, 30.50, 27.66, 27.59, 23.78, 23.75. HRMS (ESI): m / z calcd for C 39 H 52 F3N4O6S + [M+H] + :761.3554; found 761.3552.

[0072] (5) Preparation of compound H1

[0073] The structure of compound H1 is as follows:

[0074] Compound H1; its preparation process is as follows:

[0075] Step 1: Preparation of compound G1

[0076] Compound D (152 mg, 1.00 mmol) and compound F1 (1,4-dibromobutane, 216 mg, 1.00 mmol) were dissolved in tetrahydrofuran (8 mL), triethylamine (278 μL, 2.00 mmol) was added, and the reaction was carried out at room temperature for 1 h. After the reaction was completed, the tetrahydrofuran was spun dry, diluted with water (12 mL), and extracted with ethyl acetate (3x10 mL), then the organic phases were combined and dried with anhydrous sodium sulfate, concentrated, and the obtained crude product was purified by silica gel column chromatography (dichloromethane:methanol = 20:1) to obtain compound G1.

[0077] The structures of compound D, compound F1, and compound G1 in the above preparation process are as follows:

[0078] Compound D, Compound F1, Compound G1;

[0079] Compound G1 was tested and the results of the test are as follows: HRMS (ESI) calculated for C14H27BrN+ [M+H]+: 288.1321, found. 288.1323.

[0080] Step 2: Preparation of compound H1

[0081] Compound A (203 mg, 0.500 mmol) and compound G1 (144 mg, 0.500 mmol) were dissolved in N,N-dimethylformamide (6 mL), potassium carbonate (139 mg, 1.00 mmol) was added, and the reaction was allowed to proceed at room temperature overnight. After completion of the reaction, the reaction was diluted with water (12 mL) and extracted with ethyl acetate (3 x 10 mL), and then the organic phase was combined and washed with a saturated NaCl solution (12 mL). The organic phase was dried over anhydrous sodium sulfate, concentrated, and the obtained crude was purified by silica gel column chromatography (dichloromethane:methanol = 15: 1) to obtain compound H1.

[0082] The structural formula of compound A in the above preparation process is as follows:

[0083] ;

[0084] Compound H1 was tested and the results of the test are as follows: 1 H NMR (400 MHz, Chloroform- d ) δ 8.13 (d, J = 2.0 Hz, 1H), 8.03 (d, J = 7.5 Hz, 1H), 7.69 (dd, J = 7.5, 2.1 Hz, 1H), 7.13 – 7.06 (m, 2H), 6.86 – 6.78 (m, 2H), 4.00 (s, 2H), 2.96 (s, 1H), 2.74(d, J = 0.5 Hz, 2H), 2.60 (s, 1H), 2.54 – 2.46 (m, 1H), 1.86 – 1.78 (m, 2H),1.75 (d, J = 13.0 Hz, 1H), 1.69 – 1.46 (m, 13H), 1.31 (d, J = 1.3 Hz, 1H), 1.30 –1.22 (m, 2H), 0.97 (s, 3H), 0.92 (s, 3H). 13C NMR (100 MHz, Chloroform- d ) δ 172.88, 171.30, 159.04, 136.87, 135.57, 133.83, 133.64, 125.66, 125.33, 124.79, 124.42, 119.43, 117.54, 106.85, 78.03, 66.95, 53.83, 53.82, 50.80, 50.43, 45.22, 41.41, 39.15, 33.95, 30.88, 30.53, 28.47, 27.59, 23.78. HRMS (ESI): m / z calcd for C 33 H 40 F3N4O2S + [M+H] + : 613.2819; found 613.2817.

[0085] (6) Preparation of compound H2

[0086] The structure of compound H2 is as follows:

[0087] Compound H2; its preparation process is as follows:

[0088] The specific preparation method is: according to the synthesis steps of compound H1, compound H2 is obtained, compared with compound H1, only 1,4-dibromobutane is changed to 1,5-dibromopentane to obtain compound compound G2, then compound G2 and compound D are reacted to obtain compound H2.

[0089] The structure of compound G2 in the above preparation process is as follows:

[0090] Compound G2;

[0091] Compound H2 is detected, and the detection results are as follows: 1 H NMR (400 MHz, Chloroform- d ) δ 8.13 (d, J = 2.0 Hz, 1H), 8.03 (d, J = 7.4 Hz, 1H), 7.69 (dd, J= 7.5, 2.1 Hz, 1H), 7.13 – 7.06 (m, 2H), 6.86 – 6.78 (m, 2H), 4.01 (s, 2H), 2.96 (s, 1H), 2.67(d, J = 0.6 Hz, 2H), 2.60 (s, 1H), 2.54 – 2.46 (m, 1H), 1.80 – 1.71 (m, 3H),1.67 (s, 1H), 1.61 (t, J = 12.9 Hz, 1H), 1.58 – 1.46 (m, 13H), 1.31 (d, J = 1.3Hz, 1H), 1.30 – 1.22 (m, 2H), 0.97 (s, 3H), 0.92 (s, 3H). 13 C NMR (100 MHz,Chloroform- d ) δ 172.88, 171.30, 159.05, 159.04, 136.87, 135.57, 133.83,133.64, 125.66, 125.33, 124.79, 124.42, 119.43, 117.54, 106.85, 78.29, 66.95,52.46, 50.80, 49.25, 45.22, 41.41, 39.15, 33.95, 30.57, 30.53, 30.44, 27.59,26.28, 23.78. HRMS (ESI): m / z calcd for C 34 H 42 F3N4O2S + [M+H] + : 627.2975; found627.2977.

[0092] (7) Preparation of compound H3

[0093] The structure of compound H3 is as follows:

[0094] Compound H3; its preparation process is as follows:

[0095] The specific preparation method is: according to the synthesis steps of compound H1, compound H3 is obtained, compared with compound H1, only 1,4-dibromobutane is changed to 1,6-dibromohexane to obtain compound G3, then compound G3 and compound D are reacted to obtain compound H3.

[0096] The structure of compound G3 in the above preparation process is as follows:

[0097] Compound G3;

[0098] Compound H3 was detected, and the detection results are as follows: 1 H NMR (400 MHz, Chloroform- d ) δ 8.13 (d, J = 2.0 Hz, 1H), 8.03 (d, J = 7.4 Hz, 1H), 7.69 (dd, J = 7.5, 2.1 Hz, 1H),7.13 – 7.06 (m, 2H), 6.86 – 6.78 (m, 2H), 4.01 (s, 2H), 2.96 (s, 1H), 2.69 –2.62 (m, 2H), 2.60 (s, 1H), 2.54 – 2.46 (m, 1H), 1.78 (s, 2H), 1.75 (d, J =13.0 Hz, 1H), 1.66 (s, 1H), 1.62 (d, J = 13.0 Hz, 1H), 1.58 – 1.44 (m, 11H),1.45 (s, 2H), 1.36 (d, J = 0.6 Hz, 2H), 1.31 (d, J = 1.3 Hz, 1H), 1.30 – 1.22 (m,2H), 0.97 (s, 3H), 0.92 (s, 3H). 13 C NMR (100 MHz, Chloroform- d ) δ 172.88,171.30, 159.04, 136.87, 135.57, 133.83, 133.64, 125.66, 125.33, 124.79,124.42, 119.43, 117.54, 106.85, 78.29, 66.95, 52.46, 50.80, 49.16, 45.22,41.41, 39.15, 33.95, 31.01, 30.91, 30.53, 29.01, 27.59, 26.99, 23.78, 23.75.HRMS (ESI): m / z calcd for C 35 H44 F3N4O2S + [M+H] + : 641.3132; found 641.3136.

[0099] (8) Preparation of compound M1

[0100] The structure of compound M1 is as follows:

[0101] Compound M1; its preparation process is as follows:

[0102] Step 1: Preparation of compound J1

[0103] Compound D (307 mg, 2.00 mmol) was dissolved in N,N-dimethylformamide (6 mL) under ice bath, N,N,N',N'-tetramethyl-O-(7-azabenzotriazol-1-yl)urea hexafluorophosphate (912 mg, 2.40 mmol), diisopropylethylamine (388 mg, 3.00 mmol) and compound I1 (propynoic acid, 140 mg, 2.00 mmol) were added respectively, 5 minutes later, the ice bath was removed, and the bottle was stirred overnight at room temperature. The liquid in the bottle was diluted with ethyl acetate (20 mL), then washed with 1 N HCl solution (20 mL), saturated sodium bicarbonate solution (20 mL) and saturated brine (20 mL) respectively, the combined organic phase was dried over anhydrous sodium sulfate, concentrated and purified by silica gel flash column chromatography (petroleum ether: ethyl acetate = 3:1) to obtain compound J1.

[0104] The structures of compound D, compound I1 and compound J1 in the above preparation process are as follows:

[0105] Compound D, Compound I1, Compound J1;

[0106] Compound J1 was detected, and the detection results were as follows: HRMS (ESI) calculated for C13H20NO+[M+H]+: 206.3085, found. 206.3083.

[0107] Step 2: Preparation of compound L

[0108] Compound A (811 mg, 2.00 mmol) and compound K (483 mg, 2.00 mmol) were dissolved in N,N-dimethylformamide (6 mL), and potassium carbonate (553 mg, 4.00 mmol) was added. The mixture was reacted overnight at room temperature. After the reaction was completed, the mixture was diluted with water (30 mL) and extracted with ethyl acetate (3 × 20 mL). The organic phases were then combined and washed with saturated NaCl solution (30 mL). The organic phases were dried with anhydrous sodium sulfate and concentrated. The crude product was purified by silica gel column chromatography (dichloromethane:methanol = 25:1) to give compound L.

[0109] The structures of compounds A, K, and L in the above preparation process are shown below:

[0110] Compound A Compound K, Compound L;

[0111] The detection results for compound L are as follows: HRMS (ESI) calculated for C 21 H 18 F3N6O2S + [M+H] + : 475.1159, found. 475.1161.

[0112] Step 3: Preparation of compound M1

[0113] L (237 mg, 0.500 mmol) and compound J1 (103 mg, 0.500 mmol) were dissolved in... t In a BuOH / H2O (67 mL, 1:1) mixed solvent, CuSO4•5H2O (62.5 mg, 0.250 mmol) and L-ascorbic acid sodium (89.1 mg, 0.450 mmol) were added. The mixture was reacted overnight at room temperature. After the reaction was completed, CuSO4•5H2O was removed by filtration. The filtrate was concentrated under vacuum and separated by silica gel rapid column chromatography (dichloromethane:methanol = 18:1) to obtain compound M1.

[0114] The results of the detection of compound M1 are as follows: 1 H NMR (400 MHz, Chloroform- d ) δ8.25 (s, 1H), 7.98 – 7.94 (m, 2H), 7.83 (dd, J = 8.3, 2.1 Hz, 1H), 7.21 (d, J= 8.8 Hz, 2H), 7.17 (d, J = 6.1 Hz, 1H), 7.01 (d, J = 8.9 Hz, 2H), 4.84 (t, J = 4.8Hz, 2H), 4.42 (t, J = 4.9 Hz, 2H), 3.45 (dt, J = 7.9, 6.3 Hz, 2H), 2.38 – 2.29(m, 2H), 2.03 – 1.87 (m, 5H), 1.70 (d, J = 14.2 Hz, 1H), 1.56 (s, 6H), 1.25 (s,1H), 1.19 (s, 3H), 1.07 (s, 3H). 13 C NMR (100 MHz, Chloroform- d ) δ 180.2,175.0, 159.9, 158.3, 143.5, 137.1, 135.2, 133.8(q, J (CF) = 33.1 Hz), 132.2,130.9, 128.5(q, J (CF) = 221.1 Hz), 127.2(q, J (CF) = 5.1 Hz), 126.3, 123.2, 115.6,114.8, 110.2, 66.3, 66.3, 49.9, 44.7, 43.7, 41.4, 41.3, 33.2, 27.9, 25.9,23.6, 23.2, 19.8. 19 F NMR (376 MHz, Chloroform- d ) δ -61.9. HRMS (ESI): m / zcalcd for C 34 H 37 F3N7O3S + [M+H] + : 680.2625; found 680.2629.

[0115] (9) Preparation of compound M2

[0116] The structure of compound M2 is as follows:

[0117] Compound M2; its preparation process is as follows:

[0118] The specific preparation method is: according to the synthesis steps of compound M1, only propynoic acid is changed to 1-butynoic acid to obtain compound J2 compared with compound M1, and then compound J2 and compound L are reacted to obtain compound M2.

[0119] The structure of compound J2 in the above preparation process is as follows:

[0120] Compound J2;

[0121] Compound M2 is detected, and the detection result is as follows: 1 H NMR (400 MHz, Chloroform- d ) δ8.13 (d, J = 2.0 Hz, 1H), 8.03 (d, J = 7.5 Hz, 1H), 7.69 (dd, J = 7.5, 2.1 Hz, 1H),7.57 (s, 1H), 7.15 – 7.06 (m, 3H), 6.89 – 6.81 (m, 2H), 4.48 – 4.40 (m, 2H),4.38 (s, 2H), 3.79 – 3.68 (m, 2H), 3.12 (d, J = 12.5 Hz, 1H), 3.07 – 2.99 (m,1H), 1.75 (d, J = 13.0 Hz, 1H), 1.62 (d, J = 13.0 Hz, 1H), 1.58 – 1.49 (m, 8H),1.46 (d, J = 2.5 Hz, 2H), 1.33 – 1.22 (m, 3H), 0.97 (s, 3H), 0.92 (s, 3H). 13 CNMR (100 MHz, Chloroform- d) δ 172.88, 171.54, 171.30, 158.13, 136.87, 135.57,134.14, 133.83, 133.64, 126.63, 125.66, 125.33, 124.79, 124.48, 119.43,117.70, 106.85, 71.91, 66.95, 57.23, 49.17, 47.93, 45.22, 41.42, 41.02,37.84, 34.60, 29.64, 27.59, 23.78, 23.75. HRMS (ESI): m / z calcd for C 35 H 39 F3N7O3S + [M+H] + : 694.2782; found 694.2783.

[0122] (10) Preparation of compound M3

[0123] The structure of compound M3 is as follows:

[0124] Compound M3; its preparation process is as follows:

[0125] The specific preparation method is: according to the synthesis steps of compound M1, compound M3 is obtained, compared with compound M1, only propynic acid is changed to 1-pentynoic acid to obtain compound compound J3, then compound J3 and compound L are reacted to obtain compound M3.

[0126] The structure of compound J3 in the above preparation process is as follows:

[0127] Compound J3;

[0128] Compound M3 is detected, and the detection result is as follows: 1 H NMR (400 MHz, Chloroform- d ) δ 8.03 – 7.92 (m, 2H), 7.83 (dd, J = 8.2, 1.9 Hz, 1H), 7.56 (s, 1H), 7.20 (d, J =8.9 Hz, 2H), 6.99 (d, J = 8.9 Hz, 2H), 4.74 (t, J = 5.0 Hz, 2H), 4.39 (t, J= 5.0Hz, 2H), 3.27 – 3.11(m, 2H), 3.04 (t, J = 7.1 Hz, 2H), 2.59 (t, J = 7.1 Hz, 2H),1.97 – 1.75 (m, 6H), 1.53 (s, 6H), 1.15 (s, 3H), 0.99 (s, 3H), 0.84 (q, J =10.0, 9.0 Hz, 4H). HRMS (ESI): m / z calcd for C 36 H 41 F3N7O3S + [M+H] + : 708.2938;found 708.2939.

[0129] (11) Preparation of compound M4

[0130] The structure of compound M4 is as follows:

[0131] Compound M4; its preparation process is as follows:

[0132] The specific preparation method is: according to the synthesis steps of compound M1, compound M4 is obtained, compared with compound M1, only propynic acid is changed to 1-hexynoic acid to obtain compound J4, then compound J4 and compound L are reacted to obtain compound M4.

[0133] The structure of compound J4 in the above preparation process is as follows:

[0134] Compound J4;

[0135] Compound M4 was detected, and the detection results were as follows: 1 H NMR (400 MHz, Chloroform- d ) δ7.96 (d, J = 9.6 Hz, 2H), 7.83 (d, J = 8.3 Hz, 1H), 7.53 (s, 1H), 7.21 (d, J = 8.0Hz, 2H), 7.01 (d, J = 8.2 Hz, 2H), 5.93 (s, 1H), 4.76 (s, 2H), 4.40 (s, 2H),3.25 (tt, J= 13.6, 6.5 Hz, 2H), 2.76 (t, J = 7.0 Hz, 2H), 2.34 (d, J = 7.7 Hz,1H), 2.23 (d, J = 6.4 Hz, 2H), 2.19 – 2.12 (m, 1H), 2.03 – 1.97 (m, 2H), 1.89(d, J = 6.0 Hz, 4H), 1.25 (s, 6H), 1.17 (s, 3H), 1.02 (s, 3H), 0.87 (d, J = 4.3Hz, 2H). 13 C NMR (100 MHz, Chloroform- d ) δ 180.20, 175.00, 172.56, 158.60,137.16, 135.33, 133.71 (q, J F-C = 303.8 Hz),133.36 (q, J F-C = 30.5 Hz), 130.92,130.66, 128.39, 128.13, 127.17(q, J F-C = 2.0 Hz), 122.26, 115.65, 114.80,110.20, 66.73, 66.32, 53.43, 49.53, 45.21, 43.89, 41.40, 35.66, 33.19, 29.69,27.97, 26.00, 24.58, 23.64, 19.85, 14.10. HRMS (ESI): m / z calcd for C 37 H 43 F3N7O3S + [M+H] + : 722.3095; found 722.3097.

[0136] (12) Preparation of compound M5

[0137] The structure of compound M5 is as follows:

[0138] Compound M5; its preparation process is as follows:

[0139] The specific preparation method is: obtaining compound M5 according to the synthesis steps of compound M1, only replacing propynoic acid with 1-heptynoic acid compared with compound M1 to obtain compound J5, and then reacting compound J5 and compound L to obtain compound M5.

[0140] The structure of compound J5 in the above preparation process is as follows:

[0141] Compound J5;

[0142] Compound M5 is detected, and the detection result is as follows: 1 H NMR (400 MHz, Chloroform- d ) δ8.08 – 7.90 (m, 2H), 7.83 (dd, J = 8.2, 2.0 Hz, 1H), 7.51 (s, 1H), 7.21 (d, J =8.7 Hz, 2H), 7.08 – 6.95 (m, 2H), 5.60 (s, 1H), 4.75 (t, J = 5.0 Hz, 2H), 4.40(t, J = 4.9 Hz, 2H), 3.33 – 3.16 (m, 2H), 2.74 (t, J = 6.5 Hz, 2H), 2.35 (ddd, J =11.6, 5.2, 2.4 Hz, 1H), 2.20 (q, J = 6.1, 4.4 Hz, 2H), 1.88 (dq, J = 14.8, 8.7,6.2 Hz, 4H), 1.60 (d, J = 9.8 Hz, 1H), 1.53 (s, 6H), 1.26 (d, J = 11.6 Hz, 2H),1.17 (s, 3H), 1.02 (s, 3H), 0.91 – 0.81 (m, 4H), 0.81 – 0.70 (m, 1H). HRMS(ESI): m / z calcd for C 38 H 45 F3N7O3S + [M+H] + : 736.3251; found 736.3253.

[0143] (13) Preparation of compound M6

[0144] The structure of compound M6 is as follows:

[0145] Compound M6; its preparation method is as follows:

[0146] The specific preparation method is: according to the synthesis steps of compound M1, only propynoic acid is replaced by 1-octynoic acid to obtain compound J6 compared with compound M1, and then compound J6 and compound L are reacted to obtain compound M6.

[0147] The structure of compound J6 in the above preparation process is as follows:

[0148] Compound J6;

[0149] Compound M6 is detected, and the detection result is as follows: 1 H NMR (400 MHz, Chloroform- d ) δ7.98 – 7.94 (m, 2H), 7.83 (dd, J = 8.2, 2.1 Hz, 1H), 7.48 (s, 1H), 7.21 (d, J =8.9 Hz, 2H), 7.00 (d, J = 8.9 Hz, 2H), 5.55 (t, J = 5.8 Hz, 1H), 4.75 (t, J = 5.0Hz, 2H), 4.39 (t, J = 4.9 Hz, 2H), 3.23 (ddd, J = 13.5, 9.6, 5.5 Hz, 2H), 2.71(t, J = 7.6 Hz, 2H), 2.33 (td, J = 6.2, 3.1 Hz, 1H), 2.15 (t, J = 7.5 Hz, 3H), 1.89(ddt, J = 6.8, 4.8, 2.7 Hz, 4H), 1.71 – 1.64 (m, 4H), 1.56 (s, 6H), 1.53 (s,1H), 1.49 – 1.30 (m, 4H), 1.17 (s, 3H), 1.01 (s, 3H). 13 C NMR (100 MHz,Chloroform-d ) δ 180.2, 175.0, 172.9, 158.6, 137.1, 135.3, 135.2, 133.4(q, J (CF) = 32.5 Hz), 132.2, 130.9, 130.7(q, J (CF) = 274.7 Hz), 128.3, 127.2(q, J (CF) = 4.6Hz), 121.9, 115.6, 114.8, 110.2, 66.8, 66.4, 49.5, 45.1, 43.8, 41.4, 41.3,38.7, 36.7, 33.2, 28.7, 27.9, 25.9, 25.4, 25.4, 23.6, 23.2, 19.8. 19 F NMR (376MHz, Chloroform- d ) δ -61.9. HRMS (ESI): m / z calcd for C 39 H 47 F3N7O3S + [M+H] + :750.3408; found 750.3412.

[0150] Example 2

[0151] This example evaluated the ability of the above protein degraders with 6,6-dimethylbicyclo[3.1.1]-2-methylamine as the hydrophobic group (the protein degraders prepared in Example 1, E1-E4, H1-H3, M1-M6) to degrade AR and AR-V7 in human prostate cancer cell line 22Rv1 using Western blotting method, with the literature reported AR protein degrader SARD279 as the positive control, and the concentration of the compounds used was 1 μM. The degradation efficiency results are shown in Table 1.

[0152] Table 1 Degradation efficiency of compounds on AR and AR-V7 in human prostate cancer cell line 22Rv1

[0153]

[0154] As can be seen from the table, each compound has a certain ability to degrade AR and AR-V7, among which compounds E2, M3, M4, M5, M6 show comparable or relatively higher efficiency in degrading AR and AR-V7 than the positive drug SARD279. E2, M3, M4, M5, M6 are selected for further evaluation of degradation effect, among which compounds E2, M6 with moderate degradation effect at a concentration of 10 μM are set at three concentration gradients of 0, 0.1, 1 μM, and compounds M3, M4, M5 with excellent degradation effect at a concentration of 10 μM are set at three concentration gradients of 0, 10, 50 nM, to determine whether these preferred compounds can degrade AR and AR-V7 in a concentration-dependent manner. The results are shown in Figure 1 . The results show that compounds E2, M3, M4, M5, M6 can degrade AR and AR-V7 in a concentration-dependent manner, and the compound M4 has the highest degradation efficiency. Therefore, compound M4 is selected as the preferred compound for subsequent evaluation.

[0155] Compound M4 is set at more detailed concentration gradients for screening, group one is set at five concentration gradients of 0, 1, 10, 100, 1000 nM, and the degradation effect is determined at 24 hours. The results are shown in A of Figure 2 , and group two is set at six concentration gradients of 0, 12.5, 25, 50, 100, 200 nM, and the degradation effect is determined at 24 hours. The results are shown in B of Figure 2 . The results show that compound M4 can degrade AR and AR-V7 in a concentration-dependent manner at high and low concentrations.

[0156] Subsequently, compound M4 is subjected to practical gradient screening, and at a concentration of 500 nM, seven time gradients of 0, 2, 4, 8, 16, 24, 48 h are set to determine the degradation effect. The results are shown in C of Figure 2 . The results show that compound M4 can degrade AR and AR-V7 in a time-dependent manner.

[0157] In summary, compared with the positive drug SARD279, compound M4 shows activity far superior to its degradation effect, and the degradation activity is increased by about 20 times.

[0158] Example 3

[0159] The anti-tumor cell proliferation experiment of the above-mentioned protein degrading agent with 6,6-dimethylbicyclo[3.1.1]-2-methylamine as a hydrophobic group (compound synthesized in examples 1-13) was evaluated, and the experiment was carried out in human prostate cancer cell 22Rv1 cell line, AR protein degrading agent SARD279 reported in the literature was used as a positive control, and the number of cells was counted after incubation with 100 nM of the drug for 6 days, and the results are shown in Table 2. It was found that the number of cells decreased to different degrees, and the number of cells of compound M4 decreased most obviously, which was consistent with the degradation efficiency result in example 2.

[0160] In order to more intuitively compare the activity difference of the compounds, the remaining proportion of the number of cells was divided into four categories: 80%<proportion<100% (*), 50%<proportion<80% (**), 50%>proportion (***). The specific test results are shown in Table 2 below:

[0161] Table 2 AR and AR-V7 degradation efficiency of compounds in human prostate cancer cell line 22Rv1

[0162] Compound No. Cell number remaining ratio E4 ** E5 ** E6 * E7 * H1 * H2 * H3 * M1 ** M2 * M3 *** M4 *** M5 *** M6 ** SARD279 *

[0163] Example 4

[0164] In this example, the hERG cardiotoxicity of compound M4 was evaluated.

[0165] HEK293 cells stably transfected and expressing human cardiac HERG ion channels were used. The cultured cells were placed under an inverted microscope, and the recording electrode was brought into contact with the cell surface by a micromanipulator. Subsequently, membrane capacitance compensation and series resistance compensation were performed to smooth the current line for subsequent testing. A magnetic valve-controlled 8-channel perfusion dosing system was used. The prepared compound M4 stock solution was diluted to 30 μM, 10 μM, 3 μM, 1 μM, and 0.3 μM according to the proportion, and was added to the perfusion dosing system. The flow rate was controlled by gravity to continuously perfuse the cells. The dosing electrode was connected and adjusted to the left upper position of the cells. After recording the current of the cells, the control group (without compound M4) was perfused. After the current was stable, compound M4 was used for perfusion, and its effect was observed. Through Clampfit software analysis, the hERG cardiotoxicity IC 50 >25 μM, showing high safety.

[0166] Example 5

[0167] In this example, the in vivo efficacy of compound M4 was evaluated. 22Rv1 cells were used to establish a xenotransplantation model, and SARD279 reported in the literature was used as a positive control.

[0168] Specific method is: 22Rv1 is injected into nu / nu immune deficiency mice. When the average diameter of the tumor reaches 3 mm, it is randomly divided into a control group (5), a low-dose compound M4 intravenous administration group (1 mpk, 5), a high-dose compound M4 intravenous administration group (3 mpk, 5) and a SARD279 intravenous administration group (10 mpk, 5). After 18 days, the body weight of the mice in the administration group and the body weight of the mice in the control group do not fluctuate significantly and the state of the mice is not significantly abnormal, indicating that the compound M4 has good safety, the tumor weight and the tumor volume of the mice in the administration group are significantly lower than those in the control group, and the tumor inhibition rate ((1-tumor weight of the administration group / tumor weight of the control group)*100%) is shown in Table 3. The results show that, compared with SARD279, the compound M4 has stronger activity and has a better effect than the high-dose positive drug in the lower administration dose group.

[0169] Table 3 TGI of 22Rv1 cell xenograft model

[0170] Group TGI M4 low dose group (1 mpk) 84.1% M4 high dose group (3 mpk) 94.2% SARD279 group (10 mpk) 62.1%

[0171] Although the description of the present application has been quite detailed and particularly described with respect to several described embodiments, it is not intended to be limited to any of these details or embodiments or any particular embodiment, but should be considered to be a broad interpretation of the possibilities provided by the appended claims in view of the prior art, so as to effectively encompass the intended scope of the present application. Furthermore, the present application has been described above with respect to embodiments that the inventors can foresee, the purpose of which is to provide a useful description, and non-essential modifications to the present application that have not yet been foreseen can still represent equivalent modifications of the present application.

Claims

1. A proteolysis targeting chimera with 6,6-dimethylbicyclo[3.1.1]-2-methylamine as a hydrophobic group, characterized in that, The protein degrader with 6,6-dimethylbicyclo[3.1.1]-2-methylamine as the hydrophobic group is as shown in any one of the following compounds E1-compound E2, compound M1-compound M6: Compound E1, compound E2, Compound M1, Compound M2, Compound M3, Compound M4, Compound M5, Compound M6.

2. A method of preparing a proteolysis inhibitor according to claim 1 with 6,6-dimethylbicyclo[3.1.1]-2-methylamine as the hydrophobic group, characterized in that, The preparation route is route one or route two: Route one: ； Route two: 。 3. Use of the protein degrader with 6,6-dimethylbicyclo[3.1.1]-2-methylamine as the hydrophobic group or a pharmaceutically acceptable salt thereof according to claim 1 in the preparation of an androgen receptor protein (AR) degrader.

4. Use according to claim 3, characterized in that, The androgen receptor protein (AR) degrader can be used in the preparation of a drug for treating and / or preventing a cancer related to AR abnormalities.

5. Use according to claim 4, characterized in that, The related cancer is prostate cancer.

6. A pharmaceutical composition, characterized by, The protein degrader with 6,6-dimethylbicyclo[3.1.1]-2-methylamine as the hydrophobic group or a pharmaceutically acceptable salt thereof according to claim 1. The pharmaceutical composition comprises one or more pharmaceutically acceptable excipients or carriers.

7. The pharmaceutical composition of claim 6, wherein, The pharmaceutical composition comprises one or more pharmaceutically acceptable excipients or carriers.

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