AhR modulators and uses thereof

Abstract

The invention discloses an AhR regulator. The AhR regulator has the following structural formula (I) # imgabs0 #, wherein A is CR3 or N; ar is a 3-14-membered carbocyclic ring or a 3-14-membered heterocyclic ring, preferably a 3-10-membered alkane ring, a 3-10-membered heteroalkane ring, a 6-10-membered aromatic ring, a 3-10-membered heteroaromatic ring, a 10-14-membered fused aromatic ring or a 9-14-membered fused aromatic ring; n is a natural number from 0 to 5; q is deuterium, halogen, C is equivalent to N, hydroxyl, C1-C8 alkyl which is not substituted or substituted by 1-6 R ', C2-C8 alkenyl, C2-C8 alkynyl, C3-C10 cycloalkyl, C3-C10 cycloalkenyl, C1-C8 alkoxy, (C1-C8 alkyl) 1-2 amino, C2-C8 alkenyloxy, C2-C8 alkynyloxy, C1-C8 alkoxycarbonyl, C1-C8 carboxyl, S (O) 1-2R, N (R) 2, 6-10-membered aryl or 3-10-membered heteroaryl. The AhR regulator provided by the invention has good activity in drugs for treating cancers and obesity or in immune regulation, hematopoiesis, cell cycle or intestinal barrier.

Description

Technical Field

[0001] The present invention belongs to the technical field of compound synthesis, and in particular relates to an AhR regulator and application thereof. Background Art

[0002] Aryl hydrocarbon receptor (AHR) is a member of the bHLH-PAS (bHLH-PER-ARNT-SIM) subfamily of the basic helix-loop-helix (bHLH) superfamily and is the only receptor in the bHLH-PAS family that can be activated by ligands [Murray et al., Nat. Rev. Cancer, 2014, 14(12), 801-814; Bersten et al., Nat. Rev. Cancer, 2013, 13(12), 827-841].

[0003] AHR regulators are very extensive, including traditionally recognized aromatic hydrocarbon heterogeneities, such as 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). In the past two decades, many endogenous regulators have been gradually discovered, such as 2-(1'H-indole-3'-carbonyl)-thiazole-4-carboxylic acid methyl ester (ITE) and tryptophan metabolite kynurenic acid. AHR regulators can also come from food intake, such as indole-3-carbinol (I3C) from the cruciferous family, and indoleacetic acid (IAA), a metabolite of intestinal flora. After being activated by a regulator, AHR enters the nucleus from the cytoplasm and forms a heterodimer with the aryl hydrocarbon receptor nuclear translocator (ARNT), and binds to the xenobiotic response element (XRE) in DNA to start the transcription of XRE-regulated genes such as cytochrome CYP450 enzyme (CYP1A1). AHR can also regulate non-XRE-dependent gene expression and metabolism through protein interactions (such as with estrogen receptor, etc.).

[0004] The AHR pathway regulates many key innate and adaptive immune responses. Studies have found that some AHR regulators promote the differentiation of Th17 cells (T-helper cells) and the secretion of IL-17. Other AHR regulators can induce the horizontal differentiation of Th17 cells into Treg cells and enhance the inhibitory activity of Treg [Quintana et al., Nature, 2008, 453(7191), 65-71; Mezrich et al., J. Immunol., 2010, 185(6), 3190-3198;]. Studies have shown that AHR activation can inhibit the innate inflammatory response regulated by macrophages (for example, reducing the expression of IL-1b, IL-6, IL-12 and TNFalpha induced by lipopolysaccharide (LPS)) and inhibit dendritic cells (reducing the activation of dendritic cells and promoting the expression of IL-10) [Kimura et al., J. Exp. Med., 2009, 206(9), 2027-2035; Wang et al., Clin. Exp. Immunol., 2014, 177(2), 521-530; Wei et al., Lab. Invest., 2014, 94(5), 528-535; Nguyen et al., Proc. Natl. Acad. Sci. USA, 2010, 107(46), 19961-19966].

[0005] As an important key factor in immune regulation, AHR is considered to be a key target for immune-related diseases. Studies have shown that AHR has a regulatory effect in models such as septic shock, herpes simplex virus eye infection, and Toxoplasma gondii infection. AHR regulators also have a protective effect in immune diseases such as multiple sclerosis, inflammatory bowel disease, rheumatoid arthritis, and psoriasis. AHR-related immune regulatory activity also plays an important role in many inflammation-related metabolic diseases, such as atherosclerosis, type I diabetes, chronic kidney disease, bone metabolism-related diseases, and pulmonary hypertension. Therefore, the development of new regulators targeting AHR is of great significance in providing new drugs for the treatment of immune-related diseases. Summary of the invention

[0006] One aspect of the present invention is to provide an AhR modulator.

[0007] The AhR modulator of the present invention has the following general structural formula (I):

[0008]

[0009] A is CR 3 or N;

[0010] Ar is a 3- to 14-membered carbocyclic ring or a 3- to 14-membered heterocyclic ring, preferably a 3- to 10-membered alkane ring, a 3- to 10-membered heteroalkane ring, a 6- to 10-membered aromatic ring, a 3- to 10-membered heteroaromatic ring, a 10- to 14-membered condensed aromatic ring, or a 9- to 14-membered heterocondensed aromatic ring;

[0011] n is a natural number from 0 to 5;

[0012] Q is deuterium, halogen, C≡N, hydroxyl, C1-C8 alkyl, C2-C8 alkenyl, C2-C8 alkynyl, C3-C10 cycloalkyl, C3-C10 cycloalkenyl, C1-C8 alkoxy, (C1-C8 alkyl) which is unsubstituted or substituted by 1-6 R' 1-2 Amine, C2-C8 alkenyloxy, C2-C8 alkynyloxy, C1-C8 alkoxycarbonyl, C1-C8 carboxyl, S(O) 1～2 R、N(R) 2 , 6- to 10-membered aryl or 3- to 10-membered heteroaryl;

[0013] Each R 1 independently hydrogen, deuterium, halogen, unsubstituted or substituted with 1 to 6 R's, preferably 1 to 3 R's, more preferably 1 to 2 R's, C1-C2 alkyl, C2 alkenyl, C2 alkynyl, C3 cycloalkyl or C1 alkoxy;

[0014] R 2 is independently unsubstituted or substituted by 1 to 6 R' C1-C8 alkyl, C2-C8 alkenyl, C2-C8 alkynyl, C3-C10 cycloalkyl, C3-C10 cycloalkenyl or C1-C8 alkoxy;

[0015] R 3 are independently halogen, OH, C1-C8 alkoxy, C2-C8 alkenyloxy, C3-C10 cycloalkoxy, C3-C10 cycloalkenyloxy, (C1-C8 alkoxy)C1-C8 alkoxy, (C2-C8 alkenyl)C1-C8 alkoxy or (C2-C8 alkynyl)C1-C8 alkoxy;

[0016] Or, R 2 and R 3 Together they form a 3- to 10-membered carbocyclic ring or a 3- to 10-membered heterocyclic ring which is unsubstituted or substituted with 1 to 6 Rs;

[0017] R' is deuterium, halogen, R, OR, S(O) 1～2 R、N(R) 2 , C(O)OR, RC(O)O, unsubstituted or substituted with 1 to 6 R, 6-10 membered aryl or 3-10 membered heteroaryl; each R is independently hydrogen, deuterium, halogen, C1-C8 alkyl, C2-C8 alkenyl, C2-C8 alkynyl, C3-C10 cycloalkyl or C3-C10 cycloalkenyl;

[0018] Wherein, the heteroalkyl ring, the heteroaromatic ring, the heterocondensed aromatic ring, the heteroaryl group or the heterocyclic ring refers to a heteroalkyl ring, heteroaromatic ring, heterocondensed aromatic ring, heteroaryl group or heterocyclic ring containing 1 to 5 heteroatoms selected from C, O and S;

[0019] The carbocyclic ring and the heterocyclic ring refer to a carbocyclic ring and a heterocyclic ring which independently contain no or 1 to 5 double bonds, respectively; or the carbocyclic ring and the heterocyclic ring refer to an aromatic ring and a heteroaromatic ring, respectively.

[0020] When A is CR 3 , the general formula (Ⅰ) has the following general formula (Ⅱ)

[0021]

[0022] In a preferred embodiment of the present invention,

[0023] R 3 are independently halogen, OH, C1-C6 alkoxy, C2-C6 alkenyloxy, C3-C8 cycloalkoxy, C3-C8 cycloalkenyloxy, (C1-C6 alkoxy)C1-C6 alkoxy, (C2-C6 alkenyl)C1-C6 alkoxy or (C2-C6 alkynyl)C1-C6 alkoxy;

[0024] Preferably,

[0025] R 3 are independently halogen, OH, C1-C4 alkoxy, C2-C4 alkenyloxy, C3-C6 cycloalkoxy, C3-C6 cycloalkenyloxy, (C1-C4 alkoxy)C1-C4 alkoxy, (C2-C4 alkenyl)C1-C4 alkoxy or (C2-C4 alkynyl)C1-C4 alkoxy;

[0026] More preferably,

[0027] R 3 Independently, they are halogen, OH, C1-C3 alkoxy, C2-C3 alkenyloxy, C5-C6 cycloalkoxy, C5-C6 cycloalkenyloxy, (C1-C3 alkoxy)C1-C3 alkoxy, (C2-C3 alkenyl)C1-C3 alkoxy or (C2-C3 alkynyl)C1-C3 alkoxy.

[0028] In a preferred embodiment of the present invention,

[0029] Ar is a 3- to 10-membered carbocyclic ring or a 3- to 10-membered heterocyclic ring, preferably a 3- to 8-membered alkane ring, a 3- to 8-membered heteroalkane ring, a 6- to 8-membered aromatic ring, a 3- to 8-membered heteroaromatic ring, a 10- to 12-membered condensed aromatic ring, or a 9- to 12-membered heterocondensed aromatic ring;

[0030] Preferably, Ar is a 3- to 8-membered carbocyclic ring or a 3- to 8-membered heterocyclic ring, preferably a 3- to 6-membered alkyl ring, a 3- to 6-membered heteroalkyl ring, a 6-membered aromatic ring, a 5- to 6-membered heteroaromatic ring, a 10-membered condensed aromatic ring or a 9- to 10-membered heterocondensed aromatic ring.

[0031] In a preferred embodiment of the present invention,

[0032] n is a natural number of 0-3, preferably a natural number of 0-2, and more preferably a natural number of 0-1.

[0033] In a preferred embodiment of the present invention,

[0034] Q is deuterium, halogen, C≡N, hydroxyl, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C8 cycloalkyl, C3-C8 cycloalkenyl, C1-C6 alkoxy, (C1-C8 alkyl) which is unsubstituted or substituted by 1-3 R' 1-2 Amine, C2-C6 alkenyloxy, C2-C6 alkynyloxy, C1-C6 alkoxycarbonyl, C1-C6 carboxyl, S(O) 1～2 R、N(R) 2 , 6- to 8-membered aryl or 3- to 8-membered heteroaryl;

[0035] Preferably, Q is deuterium, halogen, C≡N, hydroxyl, C1-C4 alkyl, C2-C4 alkenyl, C2-C4 alkynyl, C3-C6 cycloalkyl, C3-C6 cycloalkenyl, C1-C4 alkoxy, (C1-C4 alkyl) which is unsubstituted or substituted by 1 to 2 R' 1-2 Amine, C2-C4 alkenyloxy, C2-C4 alkynyloxy, C1-C4 alkoxycarbonyl, C1-C4 carboxyl, S(O) 1～2 R、N(R) 2 , 6-membered aryl or 5- to 6-membered heteroaryl;

[0036] More preferably, Q is deuterium, halogen, C≡N, hydroxyl, C1-C3 alkyl, C2-C3 alkenyl, C2-C3 alkynyl, C5-C6 cycloalkyl, C5-C6 cycloalkenyl, C1-C3 alkoxy, (C1-C3 alkyl) which is unsubstituted or substituted with one R'. 1-2 Amine, C2-C3 alkenyloxy, C2-C3 alkynyloxy, C1-C3 alkoxycarbonyl, C1-C3 carboxyl, S(O) 1～2 R、N(R) 2 , 6-membered aryl or 5- to 6-membered heteroaryl.

[0037] In a preferred embodiment of the present invention,

[0038] R 2is independently unsubstituted or substituted by 1-3 R' C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C8 cycloalkyl, C3-C8 cycloalkenyl or C1-C6 alkoxy;

[0039] Preferably,

[0040] R 2 is independently unsubstituted or substituted by 1-2 R' C1-C4 alkyl, C2-C4 alkenyl, C2-C4 alkynyl, C3-C6 cycloalkyl, C3-C6 cycloalkenyl or C1-C4 alkoxy;

[0041] More preferably,

[0042] R 2 It is independently unsubstituted or substituted by 1-2 R's and is C1-C3 alkyl, C2-C3 alkenyl, C2-C3 alkynyl, C5-C6 cycloalkyl, C5-C6 cycloalkenyl or C1-C3 alkoxy.

[0043] In a preferred embodiment of the present invention,

[0044] R 2 and R 3 Together they form a 3- to 8-membered carbocyclic ring or a 3- to 8-membered heterocyclic ring which is unsubstituted or substituted with 1 to 3 Rs;

[0045] Preferably, R 2 and R 3 Together they form a 3- to 6-membered carbocyclic ring or a 3- to 6-membered heterocyclic ring which is unsubstituted or substituted with 1 to 2 Rs;

[0046] More preferably, R 2 and R 3 Together they form a 5- to 6-membered carbocyclic ring or a 5- to 6-membered heterocyclic ring which is unsubstituted or substituted with one R.

[0047] In a preferred embodiment of the present invention,

[0048] R' is deuterium, halogen, R, OR, S(O) 1～2 R、N(R) 2 , C(O)OR, RC(O)O, unsubstituted or substituted with 1 to 3 R, 6- to 8-membered aryl or 3- to 8-membered heteroaryl; each R is independently hydrogen, deuterium, halogen, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C8 cycloalkyl or C3-C8 cycloalkenyl;

[0049] Preferably, R' is deuterium, halogen, R, OR, S(O) 1～2 R、N(R) 2, C(O)OR, RC(O)O, unsubstituted or substituted with 1 to 2 R 6-membered aryl or 3 to 6-membered heteroaryl; each R is independently hydrogen, deuterium, halogen, C1-C4 alkyl, C2-C4 alkenyl, C2-C4 alkynyl, C3-C6 cycloalkyl or C3-C6 cycloalkenyl;

[0050] More preferably, R' is deuterium, halogen, R, OR, S(O) 1～2 R、N(R) 2 , C(O)OR, RC(O)O, unsubstituted or substituted with one R, 6-membered aryl or 3-6-membered heteroaryl; each R is independently hydrogen, deuterium, halogen, C1-C3 alkyl, C2-C3 alkenyl, C2-C3 alkynyl, C5-C6 cycloalkyl or C5-C6 cycloalkenyl.

[0051] In a preferred embodiment of the present invention,

[0052] The heteroaromatic ring, the hetero-condensed aromatic ring, the heteroaryl group or the heterocyclic ring refers to a heteroaromatic ring, hetero-condensed aromatic ring, heteroaryl group or heterocyclic ring containing 1 to 3 heteroatoms selected from C, O and S;

[0053] The carbocyclic ring and the heterocyclic ring refer to a carbocyclic ring and a heterocyclic ring that independently contain no or 1 to 3 double bonds, or the carbocyclic ring and the heterocyclic ring refer to an aromatic ring and a heteroaromatic ring, respectively;

[0054] Preferably,

[0055] The heteroaromatic ring, the hetero-condensed aromatic ring, the heteroaryl group or the heterocyclic ring refers to a heteroaromatic ring, hetero-condensed aromatic ring, heteroaryl group or heterocyclic ring containing 1 to 3 heteroatoms selected from C, O and S;

[0056] The carbocyclic ring and the heterocyclic ring refer to a carbocyclic ring and a heterocyclic ring which each independently contains no or 1 to 2 double bonds, or the carbocyclic ring and the heterocyclic ring refer to an aromatic ring and a heteroaromatic ring, respectively.

[0057] In a preferred embodiment of the present invention, the AhR modulator is

[0058]

[0059]

[0060]

[0061] Another aspect of the present invention is to provide the use of the AhR modulator of the general formula (I) of the present invention in the treatment of cancer, obesity or in immunoregulation, hematopoiesis, cell cycle or intestinal barrier.

[0062] Another aspect of the present invention is to provide the use of the AhR modulator of the general formula (I) of the present invention in the preparation of drugs for treating cancer, obesity, or in the preparation of drugs for immunomodulation, hematopoiesis, cell cycle or intestinal barrier.

[0063] The synthesis process of the AhR modulator of the general formula (I) of the present invention is as follows:

[0064] Synthesis process A of general formula (Ia):

[0065]

[0066] Synthesis process B of general formula (Ia):

[0067] DETAILED DESCRIPTION

[0068] Example 1

[0069]

[0070] Step 1: Synthesis of Intermediate 1-A

[0071] Dissolve 4-bromo-3,5-dihydroxybenzoic acid (45 g, 193 mmol) in methanol (500 mL), slowly add concentrated sulfuric acid (18.5 mL, 348 mmol) while stirring. The mixture was stirred and refluxed at 70 ° C for 16 hours. LCMS monitored the end of the reaction. The reaction solution was concentrated under reduced pressure, the residue was diluted with water (250 mL), and the aqueous phase was extracted with ethyl acetate (250 mL × 4). The combined organic phases were dried over sodium sulfate and concentrated under reduced pressure to obtain the product 4-bromo-3,5-dihydroxybenzoic acid methyl ester (1-A) (45 g, 182 mmol, yield 94.3%), an off-white solid.

[0072] LCMS(ESI):[M+H] + =247,249;

[0073] 1 H NMR (400 MHz, DMSO-d 6 ): δ10.45(s,2H),7.01(s,2H),3.80(s,3H).

[0074] Step 2: Synthesis of Intermediate 1-B

[0075] Dissolve methyl 4-bromo-3,5-dihydroxybenzoate (1-A) (25g, 101mmol) in dichloromethane (250mL), add N,N-diisopropylethylamine (44mL, 253mmol), and stir at 0℃ for 5min. Then slowly add methoxybromomethane (31.6g, 253mmol) at 0℃. The resulting mixture is stirred at 25℃ for 3 hours. LCMS monitors the end of the reaction, slowly pour the reaction solution into water (250mL) to quench, and extract the aqueous phase with ethyl acetate (500mL×3). Combine the organic phases, dry with sodium sulfate and concentrate under reduced pressure to obtain a crude product. The crude product is purified by column chromatography to obtain the product methyl 4-bromo-3,5-di[(methoxymethyl)oxy]benzoate (1-B) (28g, 83.5mmol, yield 82.6%), an off-white solid.

[0076] LCMS(ESI):[M+H] + =335,337;

[0077] 1 H NMR (400 MHz, DMSO-d 6 ): δ7.48(s,2H),5.30(s,4H),3.91(s,3H),3.53(s,6H).

[0078] Step 3: Synthesis of intermediate 1-C

[0079] Dissolve methyl 4-bromo-3,5-di[(methoxymethyl)oxy]benzoate (1-B) (12 g, 35.8 mmol) in toluene (160 mL), add 2-propenylboronic acid pinacol ester (9.02 g, 53.7 mmol), [1,1'-bis(diphenylphosphino)ferrocene]dichloropalladium (1.21 g, 1.6 mmol) and saturated sodium carbonate aqueous solution (40 mL), and reflux at 100 ° C for 20 hours under nitrogen protection. LCMS monitoring of the reaction was completed, and the reaction solution was diluted with ethyl acetate (100 mL) and filtered. The filtrate was extracted with ethyl acetate (300 mL×3), the organic phases were combined, dried over sodium sulfate, and concentrated to obtain a crude product. The crude product was purified by column chromatography to give the product methyl 3,5-di[(methoxymethyl)oxy]-4-(prop-1-en-2-yl)benzoate (1-C) (6 g, 20.2 mmol, yield 56.4%) as a colorless oil.

[0080] LCMS(ESI):[M+H] + =297.2;

[0081] 1 H NMR (400 MHz, DMSO-d 6): δ7.46(s,2H),5.36-5.31(m,1H),5.21(s,4H),4.92-4.85(m,1H),3.90(s,3H),3.47(s,6H),2.04(s,3H).

[0082] Step 4: Synthesis of intermediate 1-D

[0083] Dissolve 3,5-bis[(methoxymethyl)oxy]-4-(2-propenyl)benzoic acid methyl ester (1-C) (5g, 16.8mmol) in methanol (250mL), add 10% palladium / carbon (1g), and stir at 25°C in a hydrogen (1atm) atmosphere for 16 hours. LCMS monitors the end of the reaction, filter and concentrate the filtrate under reduced pressure to obtain a crude product. The crude product is purified by column chromatography to obtain the product 3,5-bis[(methoxymethyl)oxy]-4-isopropylbenzoic acid methyl ester (1-D) (4.60g, 15.4mmol, yield 91.4%), a colorless oil.

[0084] LCMS(ESI):[M+H] + =299.2;

[0085] 1 H NMR (400 MHz, DMSO-d 6 ): δ7.41 (s, 2H), 5.23 (s, 4H), 3.88 (s, 3H), 3.68 (p, J = 7.2Hz, 1H), 3.50 (s, 6H), 1.32 (d, J = 7.2Hz, 6H).

[0086] Step 5: Synthesis of Intermediate 1-E

[0087] A solution of methyl 3,5-di[(methoxymethyl)oxy]-4-isopropylbenzoate (1-D) (3g, 10mmol) in tetrahydrofuran (20mL) was slowly added dropwise to a suspension of lithium aluminum hydride (1.14g, 30.2mmol) in tetrahydrofuran (20mL) under ice bath, and the mixture was stirred at 0°C for 2 hours under nitrogen protection. The reaction was completed after LCMS monitoring, and sodium sulfate decahydrate (2.50g) was added to quench the reaction. Filter, and the filtrate was concentrated under reduced pressure to obtain a crude product. The crude product was purified by column chromatography to obtain the product 3,5-di[(methoxy)oxy]-4-isopropylbenzyl alcohol (1-E) (2.60g, 9.6mmol, yield 95.7%), an off-white solid.

[0088] LCMS(ESI):[M+H] + =271;

[0089] 1 H NMR (400 MHz, DMSO-d 6): δ6.80-6.74(m,2H),5.20-5.14(m,4H),4.60(s,2H),3.70-3.60(m,1H),3.48(s,6H),1.31(d,J=7.2Hz,6H).

[0090] Step 6: Synthesis of Intermediate 1-F

[0091] 3,5-bis[(methoxy)oxy]-4-isopropylbenzyl alcohol (1-E) (2.6 g, 9.6 mmol) was dissolved in dichloromethane (25 mL), cooled to 0 ° C, and Dess-Martin reagent (4.9 g, 11.5 mmol) was added under nitrogen protection. The temperature was naturally raised from 0 ° C to room temperature, and the reaction was stirred for 2 hours. LCMS monitored the completion of the reaction, and the reaction was quenched with saturated sodium thiosulfate aqueous solution (25 mL) and saturated sodium bicarbonate aqueous solution (25 mL). The reaction solution was extracted with ethyl acetate (100 mL × 3), the organic phases were combined and dried over sodium sulfate, and concentrated under reduced pressure to obtain a crude product. The crude product was purified by column chromatography to obtain the product 3,5-bis[(methoxymethyl)oxy]-4-isopropylbenzaldehyde (1-F) (2.2 g, 8.2 mmol, yield 85.2%), an off-white solid.

[0092] LCMS(ESI):[M+H] + =269.0;

[0093] 1 H NMR (400 MHz, DMSO-d 6 ): δ9.87(s,1H),7.27(s,2H),5.25(s,4H),3.75-3.66(m,1H),3.50(s,6H),1.33(d,J=7.2Hz,6H).

[0094] Step 7: Synthesis of Intermediate 1-G

[0095] 3,5-bis[(methoxymethyl)oxy]-4-isopropylbenzaldehyde (1-F) (800 mg, 3 mmol) was dissolved in anhydrous methanol (10 mL), and (1-diazo-2-oxopropyl)-phosphonic acid dimethyl ester (0.9 mL, 6 mmol) and potassium carbonate (828 mg, 6 mmol) were added, and the reaction was stirred at 25 ° C for 16 hours. LCMS monitoring of the reaction was completed, and the reaction was quenched with saturated brine (10 mL) and filtered. The filtrate was extracted with ethyl acetate (20 mL × 3), and the organic phases were combined and dried over sodium sulfate, and concentrated under reduced pressure to obtain a crude product. The crude product was purified by column chromatography to obtain the product 1,3-bis[(methoxymethyl)oxy]-5-ethynyl-2-isopropylbenzene (1-G) (720 mg, 2.7 mmol, yield 91.4%), a light yellow solid.

[0096] LCMS(ESI):[M+H] + =265;

[0097] 1 H NMR (400 MHz, DMSO-d 6 ): δ6.91(s,2H),5.16(s,4H),3.62(dt,J=14.0,6.8Hz,1H),3.48(s,6H),2.99(s,1H),1.30(d,J=7.2Hz,6H).

[0098] Step 8: Synthesis of intermediate 1-H

[0099] 1,3-bis[(methoxymethyl)oxy]-5-ethynyl-2-isopropylbenzene (1-G) (260 mg, 0.98 mmol) was dissolved in anhydrous toluene (10 mL), 2'-iodo-benzyl alcohol (345 mg, 1.48 mmol), cuprous iodide (19 mg, 0.10 mmol, 0.1 eq), triphenylphosphine (38 mg, 0.15 mmol), bistriphenylphosphine palladium dichloride (12 mg, 0.02 mmol) and triethylamine (2.05 mL, 14.8 mmol) were added, and the mixture was heated at 80°C for 18 hours under nitrogen protection. The reaction was completed by LCMS monitoring, the reaction solution was diluted with ethyl acetate (50 mL) and filtered, the filtrate was washed with saturated brine (50 mL×3), the organic phases were combined, dried over anhydrous sodium sulfate, and concentrated under reduced pressure to obtain a crude product. The crude product was purified by column chromatography to give 2-({3,5-di[(methoxymethyl)oxy]-4-isopropylphenyl}ethynyl)benzyl alcohol (1-H) (280 mg, 0.76 mmol, yield 76.8%) as a light yellow solid.

[0100] LCMS(ESI):[M+H] + =371;

[0101] 1 H NMR (400 MHz, DMSO-d 6 ): δ7.86-7.77(m,2H),7.67(qd,J=6.8,2.4Hz,2H),7.62-7.56(m,2H),5.52(s,4H ), 5.22 (s, 2H), 3.97 (dt, J = 14.4, 7.2Hz, 1H), 3.82 (s, 6H), 1.64 (d, J = 7.2Hz, 6H).

[0102] Step 9: Synthesis of Compound 1

[0103] 2-({3,5-di[(methoxymethyl)oxy]-4-isopropylphenyl}ethynyl)benzyl alcohol (1-H) (110 mg, 0.30 mmol) was dissolved in anhydrous tetrahydrofuran (2.5 mL), cooled to 0°C in an ice bath, and 4M hydrogen chloride / dioxane solution (2.5 mL) was slowly added dropwise. The temperature was naturally raised to room temperature under nitrogen protection and stirred for 2.5 hours. TLC monitored the completion of the reaction, and the reaction solution was placed in an ice-water bath, and saturated sodium bicarbonate solution (5 mL) was slowly added dropwise to neutralize. The aqueous phase was separated and extracted with ethyl acetate (20 mL × 3), the combined organic phases were dried over sodium sulfate, and concentrated under reduced pressure to obtain a crude product. The crude product was purified by prep-HPLC (0.1% formic acid addition) to obtain the product 2-({3,5-dihydroxy-4-isopropylphenyl}ethynyl)benzyl alcohol (compound 1) (3 mg, 0.01 mmol, yield 3.6%), a white solid.

[0104] LCMS(ESI):[M+H] + =283;

[0105] 1 H NMR (400 MHz, DMSO-d 6 ): δ9.16(s,2H),7.24(td,J=6.8,2.4Hz,1H),7.15(q,J=7.6Hz,3H),6.64(s,2H) ,6.32(s,1H),5.14(s,2H),3.44(dd,J=14.0,7.2Hz,1H),1.23(d,J=7.2Hz,6H).

[0106] Example 2

[0107]

[0108] Step 1: Synthesis of intermediate 2-A

[0109] 1,3-bis[(methoxymethyl)oxy]-5-ethynyl-2-isopropylbenzene (1-G) (20 mg, 0.08 mmol) was dissolved in anhydrous toluene (2 mL), 2'-iodo-benzoic acid methyl ester (17 mg, 0.07 mmol), cuprous iodide (0.6 mg, 0.003 mmol), triphenylphosphine (2.48 mg, 0.01 mmol), bistriphenylphosphine palladium dichloride (0.78 mg, 0.001 mmol) and triethylamine (0.088 mL, 0.631 mmol) were added, and the mixture was heated at 80°C for 18 hours under nitrogen protection. The reaction was completed after LCMS monitoring, and the reaction solution was diluted with ethyl acetate (5 mL) and filtered. The filtrate was washed with saturated brine (5 mL×3), the organic phases were combined, dried over sodium sulfate, and concentrated under reduced pressure to obtain a crude product. The crude product was purified by column chromatography to give the product methyl 2-({3,5-di[(methoxymethyl)oxy]-4-isopropylphenyl}ethynyl)benzoate (2-A) (23 mg, 0.06 mmol, yield 91.6%) as a light yellow solid.

[0110] LCMS(ESI):[M+H] + =399,[M+Na] + =421;

[0111] 1 H NMR (400 MHz, DMSO-d 6 ): δ7.96(dd,J=8.0,0.8Hz,1H),7.62(dd,J=7.6,0.8Hz,1H),7.48(td,J=7.6,1.2Hz,1H),7.37(td,J=7.6,1.2H z,1H),6.99(s,2H),5.20(s,4H),3.97(s,3H),3.65(dt,J=14.0,7.2Hz,1H),3.50(s,6H),1.32(d,J=7.2Hz,6H).

[0112] Step 2: Synthesis of Compound 2

[0113] Dissolve methyl 2-({3,5-di[(methoxymethyl)oxy]-4-isopropylphenyl}ethynyl)benzoate (2-A) (90 mg, 0.23 mmol) in anhydrous 1,4-dioxane (2.5 mL), cool to 0°C in an ice bath, slowly drop 4M hydrogen chloride / dioxane solution (2.5 mL), and stir at 0°C for 5 hours under nitrogen protection. After the reaction is completed by TLC monitoring, slowly drop saturated sodium bicarbonate solution (5 mL) to neutralize. Separate the aqueous phase and extract with ethyl acetate (20 mL × 3), combine the organic phases and dry with sodium sulfate, and concentrate under reduced pressure to obtain a crude product. The crude product was purified by prep-HPLC (0.1% formic acid addition) to obtain the product methyl 2-[(3,5-dihydroxy-4-isopropylphenyl)ethynyl]benzoate (Compound 2) (18 mg, 0.06 mmol, yield 25.7%).

[0114] LCMS(ESI):[M+H] + =311,[2M+Na] + =643;

[0115] 1 H NMR (400 MHz, DMSO-d 6 ): δ9.33(s,2H),7.89(d,J=7.6Hz,1H),7.62(d,J=7.6Hz,2H),7.50(d,J=6.8 Hz, 1H), 6.46 (s, 2H), 3.89 (s, 3H), 3.50-3.43 (m, 1H), 1.23 (d, J = 6.8Hz, 6H).

[0116] According to the synthesis method of Example 2, the compounds in the following table were prepared

[0117]

[0118]

[0119]

[0120]

[0121]

[0122] Example 3

[0123]

[0124] Step 1: Synthesis of intermediate 3-A

[0125] Potassium carbonate (5.59 g, 40.4 mmol) and 4-bromobut-1-ene (2.73 g, 20.2 mmol) were added to a solution of methyl 4-bromo-3,5-dihydroxybenzoate (5.0 g, 20.2 mmol) in N,N-dimethylformamide (50 mL). The reaction was stirred overnight at 90 ° C. LCMS detected the formation of the product. The reaction solution was concentrated in vacuo. Ethyl acetate and water were added to the residue and the liquid was extracted. The organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, filtered and concentrated in vacuo. The residue was purified by column chromatography to obtain the product methyl 4-bromo-3-(but-3-enyloxy)-5-hydroxybenzoate (3-A) (2.1 g, 6.97 mmol, yield 34.5%), a colorless oil.

[0126] LCMS(ESI):[M+H] + =301.1;

[0127] Step 2: Synthesis of intermediate 3-B

[0128] 4-Bromo-3-(but-3-enyloxy)-5-hydroxybenzoic acid methyl ester (3-A) (2.1 g, 6.9 mmol) was dissolved in dichloromethane (20 mL), and N,N-diisopropylethylamine (3.45 mL, 20.9 mmol, 3.0 eq) was added. At 0°C, methoxybromomethane (1.74 g, 13.9 mmol) was added dropwise. The reaction was stirred at 0°C for 1 hour. LCMS detected that the reaction was complete. Dichloromethane and water were added to the reaction solution, and the liquid was extracted. The organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, filtered and concentrated in vacuo. The residue was purified by column chromatography to obtain the product 4-bromo-3-(but-3-enyloxy)-5-[(methoxymethyl)oxy]benzoic acid methyl ester (3-B) (1.60 g, 4.64 mmol, yield 66.5%), a colorless oil.

[0129] 1 H NMR (400 MHz, CDCl 3 ): δ7.43(d,J=1.6Hz,1H),7.26(s,1H),6.00-5.90(m,1H),5.30(s,2H),5.20-5.1 0(m,2H),4.13(t,J=6.8Hz,2H),3.91(s,3H),3.52(s,3H),2.61(q,J=6.8Hz,2H).

[0130] Step 3: Synthesis of intermediate 3-C

[0131] Azobisisobutyronitrile (0.05 g, 0.319 mmol) and tri-n-butyltin hydrogen (1.86 g, 6.37 mmol) were added to a solution of 4-bromo-3-(but-3-enyloxy)-5-[(methoxymethyl)oxy]benzoic acid methyl ester (3-B) (1.10 g, 3.18 mmol) in toluene (20 mL), and the reaction was stirred at 100 ° C overnight. LCMS detected that the reaction was complete. The reaction solution was cooled and concentrated in vacuo. The reactants were diluted with ethyl acetate and water, and the extracts were separated. The organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, filtered and concentrated in vacuo. The residue was purified by column chromatography to give the product 5-(methoxymethyl)oxy-4-methylbenzopyran-7-carboxylic acid methyl ester (3-C) (460 mg, 1.72 mmol, yield 54.2%), a colorless oil.

[0132] LCMS(ESI):[M+H] + =267.1;

[0133] Step 4: Synthesis of intermediate 3-D

[0134] 5-(Methoxymethyl)oxy-4-methylbenzopyran-7-carboxylic acid methyl ester (3-C) (460 mg, 1.73 mmol) was dissolved in tetrahydrofuran (10 mL), and lithium aluminum hydride (131 mg, 3.45 mmol) was added in batches at 0°C. The reaction was stirred at 0°C for 2 hours. LCMS detected that the reaction was complete. Sodium sulfate decahydrate was added to the reaction solution to quench the reaction. Filter and concentrate the filtrate in vacuo to obtain the product 5-[(methoxymethyl)oxy]-4-methyl-3,4-dihydro-2H-chromen-7-ylmethanol (3-D) (350 mg, 1.47 mmol, yield 85.0%), a colorless oil.

[0135] LCMS(ESI):[M+H] + =239.2;

[0136] Step 5: Synthesis of intermediate 3-E

[0137] {5-[(Methoxymethyl)oxy]-4-methyl-3,4-dihydro-2H-chromen-7-yl}methanol (3-D) (350 mg, 1.47 mmol) was dissolved in dichloromethane (10 mL), Dess-Martin reagent (1.24 g, 2.93 mmol) was added at 0°C, and the reaction was stirred at 0°C for 2 hours. LCMS detected that the reaction was complete. Saturated sodium thiosulfate solution was added to the reaction solution to quench. Dichloromethane and water were added to the reaction solution, and the liquid was extracted. The organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, filtered and concentrated in vacuo. The residue was purified by column chromatography to obtain the product 5-[(methoxymethyl)oxy]-4-methyl-3,4-dihydro-2H-chromen-7-yl acetaldehyde (3-E) (210 mg, 0.889 mmol, yield 60.5%), a colorless oil.

[0138] LCMS(ESI):[M+H] + =237.2;

[0139] Step 6: Synthesis of intermediate 3-F

[0140] 5-[(Methoxymethyl)oxy]-4-methyl-3,4-dihydro-2H-chromene-7-carboxyacetaldehyde (3-E) (210 mg, 0.89 mmol) was dissolved in anhydrous methanol (4 mL), and dimethyl (1-diazo-2-oxopropyl)-phosphonate (0.27 mL, 1.78 mmol) and K 2 CO 3 (246 mg, 1.78 mmol), stirred at 25 ° C for 16 hours. LCMS monitoring reaction was completed, quenched with saturated brine (4 mL) and filtered, the filtrate was extracted with ethyl acetate (10 mL × 3), the organic phases were combined and dried over sodium sulfate, and concentrated under reduced pressure to obtain a crude product. The crude product was purified by column chromatography to obtain the product 5-[(methoxymethyl)oxy]-4-methyl-3,4-dihydro-2H-chromen-7-yl acetylene (3-F) (186 mg, 0.8 mmol, yield 90.1%), a light yellow oil.

[0141] LCMS(ESI):[M+H] + =233.1;

[0142] Step 7: Synthesis of intermediate 3-G

[0143] 5-[(Methoxymethyl)oxy]-4-methyl-3,4-dihydro-2H-chromen-7-yl acetylene (3-F) (186 mg, 0.8 mmol) was dissolved in anhydrous toluene (10 mL), and iodobenzene (345 mg, 1.2 mmol), cuprous iodide (15 mg, 0.08 mmol), triphenylphosphine (31 mg, 0.12 mmol), bistriphenylphosphine palladium dichloride (10 mg, 0.016 mmol) and triethylamine (1.65 mL, 11.8 mmol) were added, and the mixture was heated at 80°C for 18 hours under nitrogen protection. The reaction was completed after LCMS monitoring, and the reaction solution was diluted with ethyl acetate (50 mL) and filtered. The filtrate was washed with saturated brine (50 mL×3), and the organic phases were combined and dried over anhydrous sodium sulfate, and concentrated under reduced pressure to obtain a crude product. The crude product was purified by column chromatography to give 5-[(methoxymethyl)oxy]-4-methyl-7-[(2-phenyl)ethynyl]-3,4-dihydro-2H-chromene (3-G) (185 mg, 0.6 mmol, yield 74.8%) as a light yellow solid.

[0144] LCMS(ESI):[M+H] + =309.1;

[0145] Step 8: Synthesis of Compound 3

[0146] 5-[(Methoxymethyl)oxy]-4-methyl-3,4-dihydro-2H-chromen-7-yl(2-phenyl)acetylene (3-G) (80 mg, 0.25 mmol) was dissolved in 1,4-dioxane (2 mL) and 4 M hydrogen chloride / dioxane (2 mL) was slowly added at 0°C. The reaction was stirred at 0°C for 1 hour and then at room temperature for 2 hours. LCMS showed that the reaction was complete. The reaction solution was cooled and concentrated in vacuo. The reaction was diluted with ethyl acetate and water and the extracts were separated. The organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, filtered and concentrated in vacuo. The residue was purified by prep-HPLC (0.1% formic acid addition) to give the product 4-methyl-7-(2-phenylethynyl)-3,4-dihydro-2H-chromen-5-ol as an off-white solid (12 mg, yield 17.6%).

[0147] LCMS(ESI):[M+H] + =265.2;

[0148] 1 HNMR: (400MHz, DMSO-d 6): δ9.45(s,1H),7.57(d,J=7.6Hz,2H),7.35(t,J=7.6Hz,2H),7.24(t,J=7.2Hz,1H),7.08(s,1H),6.95(s,1H),4.17(d,J= 10.8Hz, 1H), 4.02 (t, J = 10.8Hz, 1H), 3.02-2.91 (m, 1H), 1.97-1.85 (m, 1H), 1.63 (d, J = 14.0Hz, 1H), 1.21 (d, J = 6.8Hz, 3H).

[0149] Example 4

[0150]

[0151] Step 1: Synthesis of intermediate 4-A

[0152] 4-Bromo-3,5-dihydroxybenzoic acid methyl ester (5g, 20.2mmol) was dissolved in N,N-dimethylformamide (50mL), potassium carbonate (5.6g, 40.5mmol) was added and stirred at room temperature for 10 minutes, and then allyl bromide (2.28g, 18.8mmol) was slowly added dropwise while stirring. The mixture was heated at 85°C for 3 hours, and the reaction was completed by LCMS monitoring. The reaction solution was diluted with ethyl acetate (100mL) and filtered, and the filtrate was washed with saturated brine (100mL×5). The organic phase was collected and dried over sodium sulfate, concentrated under reduced pressure, and the crude product was purified by column chromatography to obtain the product 3-(allyloxy)-4-bromo-5-hydroxybenzoic acid methyl ester (4-A) (1.9g, 6.6mmol, yield 32.7%), an off-white solid.

[0153] LCMS(ESI):[M+H] + =287.0,289.0;

[0154] 1 H NMR (400 MHz, CDCl 3 ): δ7.35(d,J=1.6Hz,1H),7.14(d,J=1.6Hz,1H),6.14-6.00(m,1H),5.75(s,1H),5.50(dd ,J=17.2,1.2Hz,1H),5.33(d,J=10.4Hz,1H),4.66(d,J=5.2Hz,2H),3.92(d,J=6.4Hz,3H).

[0155] Step 2: Synthesis of intermediate 4-B

[0156] 3-(Allyloxy)-4-bromo-5-hydroxybenzoic acid methyl ester (4-A) (1.91g, 6.7mmol) was dissolved in dichloromethane (20mL), N,N-diisopropylethylamine (2.2mL, 13.3mmol) was added, and stirred at 0°C for 5min. Then methoxymethyl bromide (0.82mL, 10mmol) was slowly added dropwise at 0°C. The resulting mixture was stirred at room temperature for 3 hours. LCMS monitored the completion of the reaction, the reaction solution was slowly poured into water (50mL) to quench, and the aqueous phase was extracted with ethyl acetate (100mL×3). The organic phases were combined, dried over sodium sulfate and concentrated under reduced pressure to obtain a crude product. The crude product was purified by column chromatography to obtain the product 3-allyloxy-5-[(methoxymethyl)oxy]-4-bromobenzoic acid methyl ester (4-B) (2g, 6mmol, yield 90.1%), an off-white solid.

[0157] LCMS(ESI):[M+H] + =331,333;

[0158] 1 H NMR (400 MHz, CDCl 3 ): δ7.44(d,J=1.6Hz,1H),7.27(d,J=1.6Hz,1H),6.14-6.00(m,1H),5.52(dd,J=17.2,1.6Hz,1H), 5.33(dd,J=10.8,1.6Hz,1H),5.30(s,2H),4.67(dt,J=4.8,1.6Hz,2H),3.91(s,3H),3.53(s,3H).

[0159] Step 3: Synthesis of intermediate 4-C

[0160] Dissolve 3-allyloxy-5-[(methoxymethyl)oxy]-4-bromobenzoic acid methyl ester (4-B) (1g, 3mmol) in toluene (10mL), add tributyltin hydride (1.6mL, 6mmol) and azobisisobutyronitrile (0.02g, 0.15mmol), and react at 80°C for 2 hours under nitrogen protection. LCMS monitoring of the reaction was completed, the reaction solution was diluted with ethyl acetate (10mL), washed with water (20mL×3), the organic phase was separated and dried over sodium sulfate, and concentrated under reduced pressure to obtain a crude product. The crude product was purified by column chromatography to obtain the product 4-[(methoxymethyl)oxy]-3-methyl-2,3-dihydrobenzofuran-6-carboxylic acid methyl ester (4-C) (680mg, 2.70mmol, yield 90%), a colorless oil.

[0161] 1 H NMR (400 MHz, CDCl 3): δ7.29(s,1H),7.13(s,1H),5.25(d,J=5.6Hz,2H),4.71 -4.64(m,1H),4.23-4.14(m,1H),3.88(s,3H),3.71-3.57(m,1H),3.49(s,3H),1.36(d,J=6.8Hz,3H).

[0162] Step 4: Synthesis of intermediate 4-D

[0163] Under ice bath, a solution of 4-[(methoxymethyl)oxy]-3-methyl-2,3-dihydrobenzofuran-6-carboxylic acid methyl ester (4-C) (753 mg, 3.0 mmol) in tetrahydrofuran (1 mL) was slowly added dropwise to a suspension of lithium aluminum hydride (340 mg, 9 mmol) in tetrahydrofuran (4 mL), and the mixture was stirred at 0 ° C for 2 hours under nitrogen protection. LCMS monitored the end of the reaction and sodium sulfate decahydrate (2.8 g) was added to quench the reaction. Filter, and the filtrate was concentrated under reduced pressure to obtain a crude product. The crude product was purified by column chromatography to obtain the product 4-[(methoxymethyl)oxy]-3-methyl-6-hydroxymethyl-2,3-dihydrobenzofuran (4-D) (600 mg, 2.67 mmol, yield 90.0%), a light brown solid.

[0164] LCMS(ESI):[M+H] + =225;

[0165] Step 5: Synthesis of intermediate 4-E

[0166] 4-[(Methoxymethyl)oxy]-3-methyl-6-hydroxymethyl-2,3-dihydrobenzofuran (4-D) (650mg, 2.9mmol) was dissolved in dichloromethane (8mL), cooled to 0°C, and Dess-Martin reagent (1.6g, 3.8mmol) was added under nitrogen protection, and the temperature was naturally raised to room temperature and stirred for 2 hours. LCMS monitored the completion of the reaction, and the reaction was quenched with saturated sodium thiosulfate aqueous solution (10mL) and saturated sodium bicarbonate aqueous solution (10mL). The reaction solution was extracted with ethyl acetate (30mLx3), the combined organic phases were dried over sodium sulfate, and concentrated under reduced pressure to obtain a crude product. The crude product was purified by column chromatography to obtain the product 4-[(methoxymethyl)oxy]-3-methyl-2,3-dihydrobenzofuran-6-carboxaldehyde (4-E) (400mg, 1.8mmol, yield 62.1%), a light yellow solid.

[0167] 1 H NMR (400 MHz, CDCl 3): δ9.86(s,1H),7.14(s,1H),6.96(s,1H),5.26(d,J=5.2Hz,2H),4.70(t,J=8.8Hz,1H ), 4.23 (dd, J = 8.8, 5.4Hz, 1H), 3.70-3.60 (m, 1H), 3.50 (s, 3H), 1.37 (d, J = 6.8Hz, 3H).

[0168] Step 6: Synthesis of intermediate 4-F

[0169] 4-[(Methoxymethyl)oxy]-3-methyl-2,3-dihydrobenzofuran-6-carbaldehyde (4-E) (400 mg, 1.8 mmol) was dissolved in anhydrous methanol (10 mL), and (1-diazo-2-oxopropyl)-phosphonic acid dimethyl ester (0.54 mL, 3.6 mmol) and K 2 CO 3 (497 mg, 3.6 mmol), stirred at 25 ° C for 16 hours. LCMS monitoring reaction was completed, quenched with saturated brine (10 mL) and filtered, the filtrate was extracted with ethyl acetate (20 mL × 3), the organic phases were combined and dried over sodium sulfate, and concentrated under reduced pressure to obtain a crude product. The crude product was purified by column chromatography to obtain the product 6-ethynyl-4-[(methoxymethyl)oxy]-3-methyl-2,3-dihydrobenzofuran (4-F) (344 mg, 1.58 mmol, yield 87.8%), a light yellow solid.

[0170] LCMS(ESI):[M+H] + =219.1;

[0171] Step 7: Synthesis of intermediate 4-G

[0172] 6-Ethynyl-4-[(methoxymethyl)oxy]-3-methyl-2,3-dihydrobenzofuran (4-F) (174 mg, 0.8 mmol) was dissolved in anhydrous toluene (10 mL), and iodobenzene (345 mg, 1.2 mmol), cuprous iodide (15 mg, 0.08 mmol), triphenylphosphine (31 mg, 0.12 mmol), bistriphenylphosphine palladium dichloride (10 mg, 0.016 mmol) and triethylamine (1.65 mL, 11.8 mmol) were added, and the mixture was heated at 80°C for 18 hours under nitrogen protection. The reaction was completed after LCMS monitoring, and the reaction solution was diluted with ethyl acetate (50 mL) and filtered. The filtrate was washed with saturated brine (50 mL×3), and the organic phases were combined and dried over anhydrous sodium sulfate, and concentrated under reduced pressure to obtain a crude product. The crude product was purified by column chromatography to give 4-[(methoxymethyl)oxy]-3-methyl-6-[(2-phenyl)ethynyl]-2,3-dihydrobenzofuran (4-G) (170 mg, 0.58 mmol, yield 72.5%) as an off-white solid.

[0173] LCMS(ESI):[M+H] + =295.1;

[0174] Step 8: Synthesis of compound 4

[0175] 4-[(Methoxymethyl)oxy]-3-methyl-6-[(2-phenyl)ethynyl]-2,3-dihydrobenzofuran (4-G) (85 mg, 0.29 mmol) was dissolved in 1,4-dioxane (2 mL) and 4M hydrogen chloride / dioxane (2 mL) was slowly added at 0°C. The reaction was stirred at 0°C for 1 hour and then at room temperature for 2 hours. LCMS detected that the reaction was complete. The reaction solution was cooled and concentrated in vacuo. The reactants were diluted with ethyl acetate (80 mL) and water (10 mL) and the extracts were separated. The organic phase was washed with saturated brine (10 mL×3), dried over anhydrous sodium sulfate, filtered and concentrated in vacuo. The residue was purified by prep-HPLC (0.1% formic acid addition) to give the product 3-methyl-6-[(2-phenyl)ethynyl]-2,3-dihydrobenzofuran-4-ol as a white solid (19 mg, yield 26.2%).

[0176] LCMS(ESI):[M+H] + =251.1;

[0177] 1 H NMR (400 MHz, DMSO-d 6)δ9.47(s,1H),7.57(d,J=7.2Hz,2H),7.35(t,J=7.6Hz,2H),7.25(t,J=7.2Hz,1H),7.12(s,1H),6.95 (s,1H),4.57(t,J=8.8Hz,1H),4.07(dd,J=8.8,5.2Hz,1H),3.56-3.42(m,1H),1.23(d,J=6.8Hz,3H).

[0178] Example 5

[0179]

[0180] Step 1: Synthesis of Compound 5-A

[0181] Anhydrous aluminum chloride (7.6 g, 57 mmol) was suspended in dichloromethane (50 mL), cooled to 0 ° C, and acetyl chloride (3.72 ml, 52 mmol) was added dropwise. After the addition was completed, the reaction system was stirred at 0 ° C until it became transparent, and 1-bromo-3,5-dimethoxybenzene (10.3 g, 47.6 mmol) was dissolved in dichloromethane (30 mL) and added thereto. The reaction system was stirred at 0 ° C for 2 hours. TLC monitored the completion of the reaction. The reaction system was quenched with saturated ammonium chloride (40 mL) and extracted with dichloromethane (50 mL × 3). The organic phase was washed with saturated brine (10 mL × 3), dried over anhydrous sodium sulfate, filtered and concentrated in vacuo. The crude product was purified by silica gel column to obtain the product 1-(4-bromo-2,6-dimethoxyphenyl)ethanone (5-A), a colorless oil (4.86 g, yield 39.6%).

[0182] 1 H NMR (400 MHz, CDCl 3 ): δ6.70(s,2H),3.46(s,6H),2.45(s,3H).

[0183] Step 2: Synthesis of compound 5-B

[0184] 1-(4-bromo-2,6-dimethoxyphenyl)ethanone (5-A) (4.86 g, 18.8 mmol) was dissolved in anhydrous tetrahydrofuran (40 mL), and 3 M methylmagnesium chloride Grignard reagent (18.8 mL, 56.6 mmol) was slowly added dropwise at 0°C. The reaction system was stirred at 0°C for 30 minutes and then stirred at room temperature overnight. The reaction was complete by LCMS. The reaction system was cooled to 0°C, quenched with saturated ammonium chloride (40 mL), and diluted with ethyl acetate (150 mL). The organic phase was washed with saturated brine (15 mL × 3), dried over anhydrous sodium sulfate, filtered and concentrated in vacuo. The crude product was purified by silica gel column to obtain the product 2-(4-bromo-2,6-dimethoxyphenyl)propyl-2-ol (5-B), a colorless oil (4.74 g, yield 92.1%).

[0185] LCMS(ESI):[M+H] + =274.0;

[0186] Step 3: Synthesis of Compound 5-C

[0187] 2-(4-bromo-2,6-dimethoxyphenyl)propyl-2-ol (5-B) (4.74 g, 17.3 mmol) was dissolved in dichloromethane (70 mL), trifluoroacetic acid (3.94 mL, 51.9 mmol) was slowly added at -30 ° C, and then triethylsilane (8.27 mL, 51.9 mmol) was slowly added dropwise. After the addition was completed, the reaction was continued to stir at -30 ° C for 30 minutes, and then stirred at room temperature overnight. LCMS detected that the reaction was complete. Water (25 mL) was added to dilute the reaction, and the aqueous phase was extracted with dichloromethane (30 mL × 3). The organic phases were combined and washed with saturated brine (10 mL × 3), dried over anhydrous sodium sulfate, filtered and concentrated in vacuo. The crude product was purified by silica gel column to obtain the product 5-bromo-2-isopropyl-1,3-dimethoxybenzene (5-C), a colorless oil (3.72 g, yield 83.2%).

[0188] 1 H NMR (400 MHz, CDCl 3 ): δ6.76 (s, 2H), 3.76 (s, 6H), 3.50-3.42 (m, 1H), 1.22 (d, J = 7.2Hz, 6H).

[0189] Step 4: Synthesis of compound 5-D

[0190] 5-Bromo-2-isopropyl-1,3-dimethoxybenzene (5-C) (3.72 g, 14.4 mmol) was dissolved in dichloromethane (14 mL). After slowly adding 1 M boron tribromide / dichloromethane solution (14.4 mL, 14.4 mmol) at -10 °C, the reaction was heated to room temperature and stirred overnight. LC-MS detected that the reaction was complete. Water (10 mL) was added to quench the reaction, and the aqueous phase was extracted with dichloromethane (40 mL × 3). The combined organic phases were washed with saturated brine (10 mL × 3), dried over anhydrous sodium sulfate, filtered and concentrated in vacuo. The crude product was purified by silica gel column to obtain the product 5-bromo-2-isopropyl-1,3-dihydroxybenzene (5-D) as a white solid (1.78 g, yield 53.7%).

[0191] LCMS(ESI):[M+H] + =231,233;

[0192] 1 H NMR (400 MHz, DMSO-d 6 )δ9.45(s,2H),6.43(s,2H),3.48-3.39(m,1H),1.21(d,J=6.8Hz,6H).

[0193] Step 5: Synthesis of Compound 5

[0194] 5-Bromo-2-isopropyl-1,3-dihydroxybenzene (5-D) (230 mg, 1 mmol) was dissolved in toluene (8 mL), and cyclohexyl acetylene (162 mg, 1.5 mmol), cuprous iodide (19 mg, 0.1 mmol), triethylamine (0.56 mL, 4 mmol), and bistriphenylphosphine palladium dichloride (70 mg, 0.1 mmol) were added, and the mixture was heated to 100 ° C for 16 hours. LCMS detected that the reaction was complete. After the reaction system was concentrated under reduced pressure, dichloromethane (60 mL) and water (10 mL) were added to dilute it. The organic phase was washed with saturated brine (10 mL × 3), dried over anhydrous sodium sulfate, and concentrated under reduced pressure to obtain a crude product. The crude product was purified by silica gel column to obtain the product 5-(cyclohexylethynyl)-2-isopropyl-1,3-benzenediol (Compound 5) (124 mg, yield 48.1%).

[0195] LCMS(ESI):[M+H] + =259.2;

[0196] 1 H NMR (400 MHz, DMSO-d 6)δ9.10(s,2H),6.25(s,2H),3.44–3.35(m,1H),2.60–2.53(m,1H),1.82–1.73(m,2 H),1.71–1.60(m,2H),1.52–1.38(m,3H),1.37–1.28(m,3H),1.20(d,J=7.2Hz,6H).

[0197] Example 6

[0198] Step 1: Synthesis of compound 6

[0199] 5-Bromo-2-isopropyl-1,3-dihydroxybenzene (5-D) (230 mg, 1 mmol) was dissolved in toluene (8 mL), and cyclopropylacetylene (132 mg, 2 mmol), cuprous iodide (19 mg, 0.1 mmol), triethylamine (0.56 mL, 4 mmol), and bistriphenylphosphine palladium dichloride (70 mg, 0.1 mmol) were added, and the mixture was heated to 100 ° C for 16 hours. The reaction was completed by LCMS. After the reaction system was concentrated under reduced pressure, dichloromethane (60 mL) and water (10 mL) were added to dilute the mixture. The organic phase was washed with saturated brine (10 mL × 3), dried over anhydrous sodium sulfate, and concentrated under reduced pressure to obtain a crude product. The crude product was purified by silica gel column to obtain the product 5-(cyclopropylethynyl)-2-isopropyl-1,3-benzenediol (compound 6) (86 mg, yield 39.8%).

[0200] LCMS(ESI):[M+H] + =217.2;

[0201] 1 H NMR (400 MHz, DMSO-d 6 )δ9.12(s,2H),6.23(s,2H),3.44–3.34(m,1H),1.52–1.42(m,1H),1.20(d,J=7.2Hz,6H),0.88–0.80(m,2H),0.70–0.62(m,2H).

[0202] Example 7

[0203]

[0204] Step 1: Synthesis of compound 7

[0205] 2-[(3,5-dihydroxy-4-isopropylphenyl)ethynyl]-2-carbaldehyde (70 mg, 0.25 mmol) and ethoxycarbonylmethylenetriphenylphosphine (130 mg, 0.375 mmol) were dissolved in tetrahydrofuran (5 mL) and stirred at 60 ° C for 3 hours. LCMS detected that the reaction was complete. Saturated ammonium chloride solution (10 mL) and ethyl acetate (80 mL) were added to the reaction system, and the organic phase was washed with saturated brine (10 mL × 3), dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The residue was purified by silica gel column to obtain the product (E)-3-(2-((3,5-dihydroxy-4-isopropylphenyl)ethynylethynyl)phenyl) ethyl acrylate (Compound 7) (60 mg, yield 68.6%).

[0206] LCMS(ESI):[M+H] + =351.2;

[0207] 1 H NMR (400 MHz, DMSO-d 6 )δ9.39(s,2H),8.11-8.07(d,J=16.0Hz,1H),7.96-7.94(m,1H),7.60-7.58(m,1H),7.49-7.38(m,1H) ,6.79-6.75(d,J=16.0Hz,1H),6.48(s,1H),4.24-4.19(m,2H),3.49-3.34(m,1H),1.29-1.23(m,9H).

[0208] Example 8

[0209]

[0210] Step 1: Synthesis of Compound 8-A

[0211] 5-Bromo-1,3-difluoro-2-iodobenzene (15 g, 47.2 mmol) was dissolved in anhydrous methanol (150 mL), and 30% sodium methoxide (15 g, 83.3 mmol) was added at room temperature, and then heated to 60 ° C and stirred for 15 hours. The reaction system was concentrated under reduced pressure, and the residue was diluted with water (150 mL) and filtered. The filter cake was dissolved in dichloromethane (300 mL), and the organic phase was washed three times with saturated sodium chloride solution (80 mL), dried over anhydrous sodium sulfate and concentrated under reduced pressure. The residue was purified by silica gel column to obtain the product 5-bromo-1-fluoro-2-iodo-3-methoxybenzene (8-A) (11.4 g, 34.5 mmol, yield 73.1%), a white solid.

[0212] 1H NMR (400MHz, CDCl3): δ6.90 (dd, J=7.2, 1.6Hz, 1H), 6.75 (m, 1H), 3.90 (s, 3H).

[0213] Step 2: Synthesis of compound 8-B

[0214] 5-Bromo-1-fluoro-2-iodo-3-methoxybenzene (8-A) (11.4 g, 34.5 mmol) was added to dioxane (200 mL) and water (20 mL), and then diboric acid pinacol ester (13.2 g, 51.8 mmol), potassium carbonate (9.52 g, 69 mmol) and [1,1'-bis(diphenylphosphino)ferrocene] palladium dichloride (2.5 g, 3.45 mmol) were added. The reaction system was stirred at 100 ° C for 16 hours under nitrogen. The reactants were poured into saturated ammonium chloride solution (80 mL) and extracted twice with ethyl acetate (250 mL). The organic phases were combined and washed three times with saturated sodium chloride solution (80 mL), dried over anhydrous sodium sulfate and concentrated under reduced pressure. The residue was purified by silica gel column to give the product 5-bromo-1-fluoro-3-methoxy-2-isopropenylbenzene (8-B) (6.86 g, 28.1 mmol, yield 81.4%) as a colorless oil.

[0215] 1 H NMR (400MHz, CDCl3): δ6.89 (dd, J=8.4, 1.6Hz, 1H), 6.81 (m, 1H), 5.35 (m, 1H), 4.96 (s, 1H), 3.82 (s, 3H), 2.01 (s, 3H).

[0216] Step 3: Synthesis of Compound 8-C

[0217] 5-Bromo-1-fluoro-3-methoxy-2-(1-propene-2-yl)benzene (8-B) (6.86 g, 28.1 mmol) was dissolved in ethyl acetate (10 mL) and platinum dioxide (1 g) was added. The reaction system was stirred at room temperature for 12 hours under a hydrogen atmosphere. LCMS detected that the reaction was complete. The reactants were filtered, the filtrate was concentrated under reduced pressure, and the residue was purified by silica gel column to obtain the product 5-bromo-1-fluoro-3-methoxy-2-isopropylbenzene (8-C) (6.68 g, yield 96.6%).

[0218] 1 H NMR (400 MHz, CDCl 3 )δ6.84-6.81(m,1H),6.77-6.75(m,1H),3.81(s,3H),3.45-3.38(m,1H),1.27-1.25(m,6H).

[0219] Step 4: Synthesis of compound 8-D

[0220] 5-Bromo-1-fluoro-3-methoxy-2-isopropylbenzene (8-C) (6.68 g, 27.2 mmol) was dissolved in dichloromethane (30 mL). After 1 M boron tribromide / dichloromethane solution (27.2 mL, 27.2 mmol) was slowly added at 0 ° C, the reaction was heated to room temperature and stirred overnight. LC-MS detected that the reaction was complete. Water (30 mL) was added to quench the reaction, and the aqueous phase was extracted with dichloromethane (40 mL × 3). The combined organic phases were washed with saturated brine (10 mL × 3), dried over anhydrous sodium sulfate, filtered and concentrated in vacuo. The crude product was purified by silica gel column to obtain the product 5-bromo-1-fluoro-3-hydroxy-2-isopropylbenzene (8-D) as a white solid (3.86 g, yield 61%).

[0221] Step 5: Synthesis of compound 8-E

[0222] 5-Bromo-1-fluoro-3-hydroxy-2-isopropylbenzene (8-D) (3.86 g, 16.6 mmol) was dissolved in dichloromethane (20 mL), N,N-diisopropylethylamine (5.8 mL, 33.2 mmol) was added, and stirred at 0 ° C for 5 min. Then methoxybromomethane (2.02 mL, 24.9 mmol) was slowly added dropwise at 0 ° C. The resulting mixture was stirred at room temperature for 3 hours. LCMS monitored the completion of the reaction, the reaction solution was slowly poured into water (30 mL) to quench, and the aqueous phase was extracted with ethyl acetate (80 mL × 3). The organic phases were combined, dried over anhydrous sodium sulfate and concentrated under reduced pressure to obtain a crude product. The crude product was purified by column chromatography to obtain the product 5-bromo-1-fluoro-2-isopropyl-3-(methoxymethyleneoxy)benzene (8-E) (4.15 g, yield 90.4%), a colorless oil.

[0223] LCMS(ESI):[M+H] + =277,279;

[0224] Step 6: Synthesis of Compound 8-F

[0225] 5-Bromo-1-fluoro-2-isopropyl-3-(methoxymethyleneoxy)benzene (8-E) (4.15 g, 15 mmol) was dissolved in toluene (80 mL), trimethylsilyl acetylene (8.5 mL, 60 mmol), cuprous iodide (285 mg, 1.5 mmol), triethylamine (8.4 mL, 60 mmol), bistriphenylphosphine palladium dichloride (842 mg, 1.2 mmol) were added, and the mixture was heated to 100 °C for 16 hours. The reaction was completed by LCMS. After the reaction system was concentrated under reduced pressure, dichloromethane (150 mL) and water (15 mL) were added to dilute the mixture. The organic phase was washed with saturated brine (30 mL × 3), dried over anhydrous sodium sulfate, and concentrated under reduced pressure to obtain a crude product. The crude product was purified by silica gel column to give the product ((3-fluoro-4-isopropyl-5-(methoxymethyleneoxy)phenyl)ethynyl)trimethylsilane (8-F) (3.44 g, yield 78%).

[0226] LCMS(ESI):[M+H] + =295.2;

[0227] Step 7: Synthesis of Compound 8-G

[0228] ((3-Fluoro-4-isopropyl-5-(methoxymethyleneoxy)phenyl)ethynyl)trimethylsilane (8-F) (3.44 g, 11.7 mmol) was dissolved in methanol (20 mL) and tetrahydrofuran (20 mL), potassium carbonate (4.84 g, 35.1 mmol) was added, and the mixture was stirred at room temperature for 4 hours. LCMS detected that the reaction was complete. After the reaction system was concentrated under reduced pressure, dichloromethane (100 mL) and water (20 mL) were added to dilute it. The organic phase was washed with saturated brine (30 mL×3), dried over anhydrous sodium sulfate, and concentrated under reduced pressure to obtain a crude product. The crude product was purified by silica gel column to obtain the product 5-ethynyl-1-fluoro-2-isopropyl-3-(methoxymethyleneoxy)benzene (8-G) (2.47 g, yield 94.9%).

[0229] LCMS(ESI):[M+H] + =223.2;

[0230] Step 8: Synthesis of Compound 8-H

[0231] 5-Ethynyl-1-fluoro-2-isopropyl-3-(methoxymethyleneoxy)benzene (8-G) (222 mg, 1 mmol) was dissolved in toluene (8 mL), and bromobenzene (0.12 mL, 1.1 mmol), cuprous iodide (19 mg, 0.1 mmol), triethylamine (0.56 mL, 4 mmol), and bistriphenylphosphine palladium dichloride (70 mg, 0.1 mmol) were added, and the mixture was heated to 100 ° C for 16 hours. LCMS detected that the reaction was complete. After the reaction system was concentrated under reduced pressure, dichloromethane (60 mL) and water (10 mL) were added to dilute the mixture. The organic phase was washed with saturated brine (10 mL × 3), dried over anhydrous sodium sulfate, and concentrated under reduced pressure to obtain a crude product. The crude product was purified by silica gel column to obtain the product 1-fluoro-2-isopropyl-3-(methoxymethyleneoxy)-5-(phenylethynyl)benzene (8-H) (156 mg, yield 52.3%).

[0232] LCMS(ESI):[M+H] + =298.2;

[0233] Step 9: Synthesis of Compound 8

[0234] 1-Fluoro-2-isopropyl-3-(methoxymethyleneoxy)-5-(phenylethynyl)benzene (8-H) (156 mg, 0.52 mmol) was dissolved in 1,4-dioxane (5 mL), cooled to 0°C in an ice bath, and 4M hydrogen chloride / dioxane solution (5 mL) was slowly added dropwise. The mixture was stirred at 0°C for 5 hours under nitrogen protection. The reaction was complete by LCMS. Saturated sodium bicarbonate solution (5 mL) was slowly added dropwise to neutralize. The aqueous phase was separated and extracted with ethyl acetate (30 mL × 3), the combined organic phases were dried over sodium sulfate, and concentrated under reduced pressure to obtain a crude product. The crude product was purified by prep-HPLC (0.1% formic acid addition) to obtain the product 3-fluoro-2-isopropyl-5-(phenylethynyl)phenol (compound 8) (47 mg, yield 35.6%).

[0235] LCMS(ESI):[M+H] + =255.2;

[0236] 1 H NMR (400 MHz, DMSO-d 6 )δ9.68(s,1H),7.65-7.60(m,2H),7.42-7.36(m,3H),7.30(s,1H),6.73(m,2H),2.96-2.90(m,1H),1.27(d,J=7.2Hz,6H).

[0237] Example 9

[0238]

[0239] Step 1: Synthesis of Compound 9-A

[0240] 5-Bromo-2-isopropyl-1,3-dimethoxybenzene (5-C) (1g, 3.86mmol) was dissolved in dichloromethane (10mL), cooled to 0℃ in an ice bath, and titanium tetrachloride (0.55mL, 5.02mmol) and methoxydichloromethane (0.454mL, 5.02mmol) were slowly added dropwise. After stirring at 0℃ for 30 minutes, the mixture was stirred at room temperature for 3 hours. LCMS detected that the reaction was complete. The reaction system was diluted with water (5mL) and dichloromethane (80mL), and the organic phase was washed with saturated brine (10mL×3), dried over anhydrous sodium sulfate, and concentrated under reduced pressure to obtain a crude product. The crude product was purified by silica gel column to obtain the product 6-bromo-3-isopropyl-2,4-dimethoxybenzaldehyde (9-A) (881mg, yield 79.5%), a colorless oil.

[0241] LCMS(ESI):[M+H] + =287,289;

[0242] 1 H NMR (400 MHz, DMSO-d 6 )δ10.12(s,1H),7.16(s,1H),3.90(s,3H),3.73(s,3H),3.42(m,1H),1.25(d,J=6.8Hz,6H).

[0243] Step 2: Synthesis of compound 9-B

[0244] 6-Bromo-3-isopropyl-2,4-dimethoxybenzaldehyde (9-A) (640 mg, 2.23 mmol) was dissolved in dichloromethane (15 mL), and 1M boron tribromide / dichloromethane solution (8.9 mL, 8.9 mmol) was slowly added dropwise at -30 ° C, stirred at -30 ° C for 30 minutes, and stirred at room temperature for 15 hours. LCMS detected that the reaction was complete. The reaction system was cooled to 0 ° C, quenched with water (5 mL), diluted with dichloromethane (80 mL), and the organic phase was washed with saturated brine (10 mL × 3), dried over anhydrous sodium sulfate, and concentrated under reduced pressure to obtain a crude product. The crude product was purified by silica gel column to obtain the product 6-bromo-2,4-dihydroxy-3-isopropylbenzaldehyde (9-B) (460 mg, yield 79.7%), a colorless oil.

[0245] LCMS(ESI):[M+H] + =259.2,261.2;

[0246] Step 3: Synthesis of compound 9

[0247] 6-Bromo-2,4-dihydroxy-3-isopropylbenzaldehyde (9-B) (460 mg, 1.78 mmol) was dissolved in N,N-dimethylformamide (10 mL), and cuprous iodide (33.8 mg, 0.178 mmol), tetrakis(triphenylphosphine)palladium (103 mg, 0.09 mmol), triethylamine (5 mL, 36 mmol), phenylacetylene (544 mg, 5.33 mmol) were added, and the mixture was heated to 75 °C and stirred for 15 hours. LCMS detected that the reaction was complete. The reaction system was cooled and filtered, and the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column to obtain the product 2,4-dihydroxy-6-phenylethynyl-3-isopropylbenzaldehyde (Compound 9) (270 mg, yield 54.3%), a yellow solid.

[0248] LCMS(ESI):[M+H] + =259.2,261.2;

[0249] 1 H NMR (400 MHz, DMSO-d 6 )δ12.57(s,1H),11.05(br,1H),10.21(s,1H),7.66-7.63(m,2H),7.46-7.44(m,3H),6.69(s,1H),3.50-3.43(m,1H),1.27(d,J=7.2Hz,6H).

[0250] Effect Example 1 Human Aryl Hydrocarbon Receptor (AHR) Regulator Luciferase Experiment

[0251] Cytochrome P450 family member A1 (CYP1A1) is one of the iconic genes regulated by the AHR signaling pathway. The upstream promoter of the CYP1A1 gene contains a specific sequence called dioxin responsive element (DRE), which can bind to the activated AhR to initiate CYP1A1 expression. In order to detect the activation of AhR, the gene sequence from -1200bp to 0bp upstream of CYP1A1 was placed in front of the Nano-luc luciferase reporter gene, and the reporter gene plasmid was constructed according to the conventional method in the prior art. According to the conventional method, the reporter gene plasmid was introduced into human liver cancer cells HepG2 by liposome transfection, and the stable expression cells were selected by puromycin. The screened cells were diluted to a concentration of 5 cells / mL and inoculated into a 96-well plate at 100 μL / well, and monoclones were obtained after growth. A monoclonal clone with normal morphology, normal growth and the highest induction signal was selected for the human AHR regulator luciferase experiment.

[0252] The HepG2 monoclonal cell line stably expressing the reporter gene was counted on the first day and the cells were 1-8×104 The cells were seeded at a density of 100 μL / well in a white opaque 96-well plate using complete culture medium without tryptophan and phenol red (basal culture medium MEM without tryptophan and phenol red, 10% fetal bovine serum, 1× non-essential amino acids, 1× sodium pyruvate and 1× GlutaMax) and cultured in an incubator maintained at 37°C and 5% carbon dioxide for 24 hours.

[0253] The test compounds were prepared into 10 mM stock solutions with dimethyl sulfoxide (DMSO) and stored at 4°C in the dark. Before treatment, the compound stock solution and the positive control compounds (such as dioxins, ITE, kynurenine, etc.) and the negative control (DMSO) were diluted with complete culture medium without tryptophan and phenol red for an initial dilution (determined according to preliminary experiments) and then 3-20 times serial dilutions were made for 11 concentration points to obtain a 2× dilution. When treating cells, 50 μL of culture medium was first aspirated from the 96-well plate inoculated with cells, and then 50 μL of the serially diluted 2× dilution was added. Two replicate experiments were set for each compound, and one well of each replicate experiment was set to a concentration of 0 (i.e., only culture medium) to determine the base number. After treatment, the reporter cells were returned to the incubator for incubation for 4-24 hours.

[0254] After treatment, add 100 μL / well of Nano-Glo luciferase assay reagent to the 96-well plate and measure the RLU (relative fluorescence intensity) of each well. The baseline Ave RLU is calculated by taking the average of the RLU with a concentration of 0 in each 96-well plate. Vehicle The RLU of the test compound at different concentrations in the experimental group TestCmpd and baseline Ave RLU Vehicle The ratio of , was used to calculate the activity of AhR under the action of different concentrations of the test compound according to formula (1) to determine the activation multiple.

[0255]

[0256] The activation factor and the corresponding compound concentration were fitted using Graphpad Prism 9 [Agonist] vs. response-variable slope (four parameters) to calculate the EC of the compound to activate AhR 50 The EC of each compound 50 The values ​​are shown in Table 2, where A represents EC 50 ≤100nM, B means 100nM<EC 50 ≤2.0μM, C means 2.0μM<EC 50 ≤100μM.

[0257] Effect Example 2 Mouse Aryl Hydrocarbon Receptor (AhR) Modulator Luciferase Experiment

[0258] In order to detect the activation of AhR, 6 xenobiotic responsive elements (XRE, see Buckley, SMK et al., Sci. Rep. 2015; 5: 11842.) were placed in front of the Nano-luc luciferase reporter gene, and the reporter gene plasmid was constructed according to the conventional method in the prior art. According to the conventional method, the reporter gene plasmid was introduced into mouse liver cancer cells Hepa1-6 by liposome transfection, and the stable expression cells were selected by puromycin for mouse AhR regulator luciferase experiment.

[0259] On the first day, the Hepa1-6 cell line stably expressing the reporter gene was counted and the number of cells was 1-8×10 4 The cells were seeded into a white opaque 96-well plate at a density of 100 μL / well using phenol red-free DMEM medium (phenol red-free DMEM, 10% fetal bovine serum), and cultured in an incubator maintained at 37° C. and 5% carbon dioxide for 24 hours.

[0260] The test compounds were prepared into 10 mM stock solutions with dimethyl sulfoxide (DMSO) and stored at 4°C in the dark. Before treatment, the compound stock solution and the positive control compounds (such as dioxins, ITE, kynurenine, etc.) and the negative control (DMSO) were diluted with complete culture medium without tryptophan and phenol red for an initial dilution (determined according to preliminary experiments) and then 3-20 times serial dilutions were made to obtain 11 concentration points to obtain a 10× dilution. When treating cells, 10 μL of culture medium was first aspirated from the 96-well plate inoculated with cells, and then 10 μL of the serially diluted 10× dilution was added. Two replicate experiments were set for each compound, and one well of each replicate experiment was set to a concentration of 0 (i.e., only culture medium) to determine the base number. After treatment, the reporter cells were returned to the incubator for incubation for 4-24 hours.

[0261] After treatment, add 100 μL / well of Nano-Glo luciferase assay reagent to the 96-well plate and measure the RLU (relative fluorescence intensity) of each well. The baseline Ave RLU is calculated by taking the average of the RLU with a concentration of 0 in each 96-well plate. Vehicle The RLU of the test compound at different concentrations in the experimental group TestCmpd and baseline Ave RLU Vehicle The ratio of , was used to calculate the activity of AhR under the action of different concentrations of the test compound according to formula (1) to determine the activation multiple.

[0262]

[0263] The activation fold and the corresponding compound concentration were fitted using Graphpad Prism 9 [Agonist] vs. response-variable slope (four parameters) to calculate the EC of the compound to activate AHR 50 The EC of each compound 50 The values ​​are shown in Table 2, where A represents EC 50 ≤100nM, B means 100nM<EC 50 ≤2.0μM, C means 2.0μM<EC 50 ≤100μM.

[0264] Effect Example 3 Rat-derived Aryl Hydrocarbon Receptor (AHR) Modulator Luciferase Experiment

[0265] In order to detect the activation of AhR, 6 Nano-luc luciferase reporter genes associated with xenobiotic responsive elements (XRE, see Buckley, SMK et al., Sci. Rep. 2015; 5: 11842.) were synthesized and cloned into the lentiviral transfer plasmid pGWLV11-new through two restriction sites, KpnI and Xbal. According to the conventional method in the prior art, the transfer plasmid and the packaging plasmid were co-transfected into 293T cells and purified to obtain a lentivirus containing the reporter gene. According to the conventional method, the reporter gene was introduced into the rat hepatoma cell H-4-II-E genome by lentiviral infection, and the cells were stably expressed by puromycin to screen the reporter gene cells for rat AhR regulator luciferase experiments.

[0266] The H-4-II-E cell line stably expressing the reporter gene was counted on the first day and seeded into a white opaque 96-well plate at a density of 1-8×104 / well using 100 μL of complete culture medium without tryptophan and phenol red (basal culture medium MEM without tryptophan and phenol red, 10% fetal bovine serum, 1× non-essential amino acids, 1× sodium pyruvate and 1× GlutaMax), and cultured in an incubator maintained at 37°C and 5% carbon dioxide for 24 hours.

[0267] The test compounds were prepared into 10 mM stock solutions with dimethyl sulfoxide (DMSO) and stored at 4°C in the dark. Before treatment, the compound stock solution and the positive control compounds (such as dioxins, ITE, kynurenine, etc.) and the negative control (DMSO) were diluted with complete culture medium without tryptophan and phenol red for an initial dilution (determined according to preliminary experiments) and then 3-20 times serial dilutions were made for 11 concentration points to obtain a 2× dilution. When treating cells, 50 μL of culture medium was first aspirated from the 96-well plate inoculated with cells, and then 50 μL of the serially diluted 2× dilution was added. Two replicate experiments were set for each compound, and one well of each replicate experiment was set to a concentration of 0 (i.e., only culture medium) to determine the base number. After treatment, the reporter cells were returned to the incubator for incubation for 4-24 hours.

[0268] After the treatment, 100 μL / well of Nano-Glo luciferase detection reagent was added to the 96-well plate, and the RLU (relative fluorescence intensity) of each well was measured. The RLU of each 96-well plate with a concentration of 0 was averaged to calculate the baseline Ave RLUVehicle. The activity of AhR under the action of different concentrations of the test compound was calculated according to formula (1) based on the ratio of RLUTestCmpd of the test compound of different concentrations in the experimental group to the baseline Ave RLUVehicle, and the activation multiple was determined.

[0269]

[0270] The activation fold and the corresponding compound concentration were fitted using Graphpad Prism 9 [Agonist] vs. response-variable slope (four parameters) to calculate the EC50 of the compound activating AHR. The EC50 values ​​of each compound are shown in Table 2, where A represents EC 50 ≤10nM, B means 10nM<EC 50 ≤2.0μM, C means 2.0μM<EC50≤100μM.

[0271] Table 2 EC values ​​of each compound 50 value

[0272]

[0273]

[0274] As can be seen from Table 2, the compounds of the present invention activate the expression of the cytochrome P450 family member A1 gene.

Claims

1. An AhR modulator, characterized in that It has the following structural formula (Ⅰ) A is CR3 or N; Ar is a 3- to 14-membered carbocyclic ring or a 3- to 14-membered heterocyclic ring, preferably a 3- to 10-membered alkane ring, a 3- to 10-membered heteroalkane ring, a 6- to 10-membered aromatic ring, a 3- to 10-membered heteroaromatic ring, a 10- to 14-membered condensed aromatic ring, or a 9- to 14-membered heterocondensed aromatic ring; n is a natural number from 0 to 5; Q is deuterium, halogen, C≡N, hydroxyl, C1-C8 alkyl, C2-C8 alkenyl, C2-C8 alkynyl, C3-C10 cycloalkyl, C3-C10 cycloalkenyl, C1-C8 alkoxy, (C1-C8 alkyl) which is unsubstituted or substituted by 1-6 R' 1-2 Amine, C2-C8 alkenyloxy, C2-C8 alkynyloxy, C1-C8 alkoxycarbonyl, C1-C8 carboxyl, S(O) 1～2 R, N(R)2, 6-10 membered aryl or 3-10 membered heteroaryl; Each R1 is independently hydrogen, deuterium, halogen, carbonyl, unsubstituted or substituted with 1 to 6 R's, preferably 1 to 3 R's, more preferably 1 to 2 R's, C1-C2 alkyl, C2 alkenyl, C2 alkynyl, C3 cycloalkyl or C1 alkoxy; R2 is independently unsubstituted or substituted by 1-6 R' C1-C8 alkyl, C2-C8 alkenyl, C2-C8 alkynyl, C3-C10 cycloalkyl, C3-C10 cycloalkenyl or C1-C8 alkoxy; R3 is independently halogen, OH, B(OH)2, C1-C8 alkoxy, C2-C8 alkenyloxy, C3-C10 cycloalkoxy, C3-C10 cycloalkenyloxy, (C1-C8 alkoxy)C1-C8 alkoxy, (C2-C8 alkenyl)C1-C8 alkoxy or (C2-C8 alkynyl)C1-C8 alkoxy; Alternatively, R2 and R3 together form a 3-10 membered carbocyclic ring or a 3-10 membered heterocyclic ring which is unsubstituted or substituted with 1-6 R's; or R2 and R3 together form a C1-C5 carbon chain which is unsubstituted or substituted with 1-6 R's and has BO inserted therein; R' is deuterium, halogen, R, OR, S(O) 1～2 R, N(R)2, C(O)OR, RC(O)O, unsubstituted or substituted with 1 to 6 R, 6-10 membered aryl or 3-10 membered heteroaryl; each R is independently hydrogen, deuterium, halogen, C1-C8 alkyl, C2-C8 alkenyl, C2-C8 alkynyl, C3-C10 cycloalkyl or C3-C10 cycloalkenyl; Wherein, the heteroalkyl ring, the heteroaromatic ring, the heterocondensed aromatic ring, the heteroaryl group or the heterocyclic ring refers to a heteroalkyl ring, heteroaromatic ring, heterocondensed aromatic ring, heteroaryl group or heterocyclic ring containing 1 to 5 heteroatoms selected from C, O and S; The carbocyclic ring and the heterocyclic ring refer to a carbocyclic ring and a heterocyclic ring which independently contain no or 1 to 5 double bonds, respectively; or the carbocyclic ring and the heterocyclic ring refer to an aromatic ring and a heteroaromatic ring, respectively.

2. The AhR modulator according to claim 1, characterized in that A is CR3, and the general formula (I) has the following general formula (II) 3. The AhR modulator according to claim 1 or 2, characterized in that R3 is independently halogen, OH, B(OH)2, C1-C6 alkoxy, C2-C6 alkenyloxy, C3-C8 cycloalkoxy, C3-C8 cycloalkenyloxy, (C1-C6 alkoxy)C1-C6 alkoxy, (C2-C6 alkenyl)C1-C6 alkoxy or (C2-C6 alkynyl)C1-C6 alkoxy; Preferably, R3 is independently halogen, OH, B(OH)2, C1-C4 alkoxy, C2-C4 alkenyloxy, C3-C6 cycloalkoxy, C3-C6 cycloalkenyloxy, (C1-C4 alkoxy)C1-C4 alkoxy, (C2-C4 alkenyl)C1-C4 alkoxy or (C2-C4 alkynyl)C1-C4 alkoxy; More preferably, R3 is independently halogen, OH, B(OH)2, C1-C3 alkoxy, C2-C3 alkenyloxy, C5-C6 cycloalkoxy, C5-C6 cycloalkenyloxy, (C1-C3 alkoxy)C1-C3 alkoxy, (C2-C3 alkenyl)C1-C3 alkoxy or (C2-C3 alkynyl)C1-C3 alkoxy.

4. The AhR modulator according to claim 1 or 2, characterized in that Ar is a 3- to 10-membered carbocyclic ring or a 3- to 10-membered heterocyclic ring, preferably a 3- to 8-membered alkane ring, a 3- to 8-membered heteroalkane ring, a 6- to 8-membered aromatic ring, a 3- to 8-membered heteroaromatic ring, a 10- to 12-membered condensed aromatic ring, or a 9- to 12-membered heterocondensed aromatic ring; Preferably, Ar is a 3- to 8-membered carbocyclic ring or a 3- to 8-membered heterocyclic ring, preferably a 3- to 6-membered alkyl ring, a 3- to 6-membered heteroalkyl ring, a 6-membered aromatic ring, a 5- to 6-membered heteroaromatic ring, a 10-membered condensed aromatic ring or a 9- to 10-membered heterocondensed aromatic ring.

5. The AhR modulator according to claim 1 or 2, characterized in that n is a natural number of 0-3, preferably a natural number of 0-2, and more preferably a natural number of 0-1.

6. The AhR modulator according to claim 1 or 2, characterized in that Q is deuterium, halogen, C≡N, hydroxyl, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C8 cycloalkyl, C3-C8 cycloalkenyl, C1-C6 alkoxy, (C1-C8 alkyl) which is unsubstituted or substituted by 1-3 R' 1-2 Amine, C2-C6 alkenyloxy, C2-C6 alkynyloxy, C1-C6 alkoxycarbonyl, C1-C6 carboxyl, S(O) 1～2 R, N(R)2, 6- to 8-membered aryl or 3- to 8-membered heteroaryl; Preferably, Q is deuterium, halogen, C≡N, hydroxyl, C1-C4 alkyl, C2-C4 alkenyl, C2-C4 alkynyl, C3-C6 cycloalkyl, C3-C6 cycloalkenyl, C1-C4 alkoxy, (C1-C4 alkyl) which is unsubstituted or substituted by 1 to 2 R' 1-2 Amine, C2-C4 alkenyloxy, C2-C4 alkynyloxy, C1-C4 alkoxycarbonyl, C1-C4 carboxyl, S(O) 1～2 R, N(R)2, 6-membered aryl or 5-6-membered heteroaryl; More preferably, Q is deuterium, halogen, C≡N, hydroxyl, C1-C3 alkyl, C2-C3 alkenyl, C2-C3 alkynyl, C5-C6 cycloalkyl, C5-C6 cycloalkenyl, C1-C3 alkoxy, (C1-C3 alkyl) which is unsubstituted or substituted with one R'. 1-2 Amine, C2-C3 alkenyloxy, C2-C3 alkynyloxy, C1-C3 alkoxycarbonyl, C1-C3 carboxyl, S(O) 1～2 R, N(R)2, 6-membered aryl or 5-6-membered heteroaryl.

7. The AhR modulator according to claim 1 or 2, characterized in that R2 is independently unsubstituted or substituted by 1-3 R' C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C8 cycloalkyl, C3-C8 cycloalkenyl or C1-C6 alkoxy; Preferably, R2 is independently unsubstituted or substituted by 1-2 R' C1-C4 alkyl, C2-C4 alkenyl, C2-C4 alkynyl, C3-C6 cycloalkyl, C3-C6 cycloalkenyl or C1-C4 alkoxy; More preferably, R2 is independently C1-C3 alkyl, C2-C3 alkenyl, C2-C3 alkynyl, C5-C6 cycloalkyl, C5-C6 cycloalkenyl or C1-C3 alkoxy which is unsubstituted or substituted with 1-2 R's.

8. The AhR modulator according to claim 1 or 2, characterized in that R2 and R3 together form a 3-8 membered carbocyclic ring or a 3-8 membered heterocyclic ring which is unsubstituted or substituted by 1-3 R's; or R2 and R3 together form a C1-C3 carbon chain which is unsubstituted or substituted by 1-3 R's and has BO inserted therein; Preferably, R2 and R3 together form a 3-6 membered carbocyclic ring or a 3-6 membered heterocyclic ring which is unsubstituted or substituted by 1-2 R's; or R2 and R3 together form a C1-C2 carbon chain which is unsubstituted or substituted by 1-2 R's and has BO inserted therein; More preferably, R2 and R3 together form a 5-6 membered carbocyclic ring or a 5-6 membered heterocyclic ring which is unsubstituted or substituted with one R'; or R2 and R3 together form a carbon chain of C1 which is unsubstituted or substituted with one R' and has BO inserted therein.

9. The AhR modulator according to claim 1 or 2, characterized in that R' is deuterium, halogen, R, OR, S(O) 1～2 R, N(R)2, C(O)OR, RC(O)O, unsubstituted or substituted with 1 to 3 R, 6- to 8-membered aryl or 3- to 8-membered heteroaryl; each R is independently hydrogen, deuterium, halogen, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C8 cycloalkyl or C3-C8 cycloalkenyl; Preferably, R' is deuterium, halogen, R, OR, S(O) 1～2 R, N(R)2, C(O)OR, RC(O)O, unsubstituted or substituted with 1 to 2 R, 6-membered aryl or 3 to 6-membered heteroaryl; each R is independently hydrogen, deuterium, halogen, C1-C4 alkyl, C2-C4 alkenyl, C2-C4 alkynyl, C3-C6 cycloalkyl or C3-C6 cycloalkenyl; More preferably, R' is deuterium, halogen, R, OR, S(O) 1～2 R, N(R)2, C(O)OR, RC(O)O, unsubstituted or substituted with one R, 6-membered aryl or 3-6-membered heteroaryl; each R is independently hydrogen, deuterium, halogen, C1-C3 alkyl, C2-C3 alkenyl, C2-C3 alkynyl, C5-C6 cycloalkyl or C5-C6 cycloalkenyl.

10. The AhR modulator according to claim 1 or 2, characterized in that The heteroaromatic ring, the hetero-condensed aromatic ring, the heteroaryl group or the heterocyclic ring refers to a heteroaromatic ring, hetero-condensed aromatic ring, heteroaryl group or heterocyclic ring containing 1 to 3 heteroatoms selected from C, O and S; The carbocyclic ring and the heterocyclic ring refer to a carbocyclic ring and a heterocyclic ring that independently contain no or 1 to 3 double bonds, or the carbocyclic ring and the heterocyclic ring refer to an aromatic ring and a heteroaromatic ring, respectively; Preferably, The heteroaromatic ring, the hetero-condensed aromatic ring, the heteroaryl group or the heterocyclic ring refers to a heteroaromatic ring, hetero-condensed aromatic ring, heteroaryl group or heterocyclic ring containing 1 to 3 heteroatoms selected from C, O and S; The carbocyclic ring and the heterocyclic ring refer to a carbocyclic ring and a heterocyclic ring which each independently contains no or 1 to 2 double bonds, or the carbocyclic ring and the heterocyclic ring refer to an aromatic ring and a heteroaromatic ring, respectively.

11. The AhR modulator according to claim 2, characterized in that The AhR modulator is 12. Use of the AhR modulator of claim 1 in the preparation of drugs for treating cancer or obesity or in the preparation of drugs for immunomodulation, hematopoiesis, cell cycle or intestinal barrier.

13. Use of the AhR modulator of claim 1 in the treatment of cancer, obesity, or in immunoregulation, hematopoiesis, cell cycle or intestinal barrier.

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