Targeted SIRT3 agonist and application thereof in anti-tumor therapeutic drugs

Abstract

The invention relates to a targeted SIRT3 agonist and application of the targeted SIRT3 agonist in anti-tumor treatment drugs, and belongs to the technical field of tumor treatment drug discovery. The structure of an isomer of the compound or the pharmaceutically acceptable salt thereof is shown in a formula I, wherein Ar and R are described in the claims and the specification. The compound or the pharmaceutically acceptable salt, the isomer and the pharmaceutical composition of the compound have SIRT3 agonist activity. In addition, the compound has an obvious anti-tumor effect, especially has an obvious inhibition effect on breast cancer, and can be used for preparing anti-tumor drugs. # imgabs0 #

Description

Technical Field

[0001] The present invention relates to a targeted SIRT3 agonist and application thereof in anti-tumor therapeutic drugs, belonging to the technical field of tumor therapeutic drug discovery. Background Art

[0002] Cancer is one of the leading causes of death worldwide in recent years, and its threat to humanity has become increasingly prominent. There are approximately 20 million new cancer cases and 9.7 million deaths worldwide. About one in five people will develop cancer in their lifetime, which has become a major public health issue worldwide. Currently, cancer treatments include surgical resection, radiotherapy, chemotherapy, targeted drug therapy, and immunotherapy. Drug therapy has become an effective and common treatment for malignant tumors. There are many types of anti-tumor drugs used clinically, including those that directly act on DNA, interfere with DNA synthesis, inhibit cell mitosis, interfere with tumor signal transduction, and new anti-tumor drugs. For example, taxanes, anthracyclines, platinum compounds, and other compounds.

[0003] In the field of drug treatment for malignant tumors, how to discover new therapeutic targets, design new targeted therapeutic small molecule drugs in a targeted manner, and deeply explore their mechanism of action is the key to solving clinical problems and has important research significance and value. SIRT3 is an important oncogene that is abnormally low in malignant tumors. Designing targeted small molecule agonists to mediate tumor cell metabolic reprogramming is an important strategy for treating breast cancer, especially triple-negative breast cancer. Summary of the invention

[0004] The technical problem solved by the present invention is to provide a compound as a SIRT3 agonist.

[0005] The present invention provides a compound having a structural formula as shown in Formula I or a pharmaceutically acceptable salt or isomer thereof:

[0006]

[0007] Wherein, Ar is a 6-10 membered aryl or heteroaryl group, and the heteroaryl group contains 1-3 heteroatoms of N, O or S;

[0008] Ar is substituted by one or more of the following substituents: H, cyano, C1-C6 alkoxy, C1-C6 alkyl, halogen, C1-C6 ester, halogenated C1-C6 alkyl, phenyl, benzyloxy, methylsulfone;

[0009] R is substituted or unsubstituted phenyl, naphthyl, thienyl, furyl;

[0010] The substituent is C1-C6 alkoxy, C1-C6 alkyl, halogen, halogenated C1-C6 alkyl;

[0011] The present invention preferably comprises a compound having the following structure or a pharmaceutically acceptable salt or isomer thereof:

[0012] wherein Ar is naphthyl, 2,3-methylenedioxy-phenyl, pyridyl, or phenyl;

[0013] Ar is substituted by one or more of the following substituents: H, cyano, C1-C6 alkoxy, C1-C6 alkyl, halogen, halogenated C1-C4 alkyl, C1-C6 ester, phenyl, benzyloxy, methylsulfone;

[0014] R is substituted or unsubstituted phenyl, naphthyl, thienyl, furyl;

[0015] The substituent is C1-C4 alkoxy, C1-C4 alkyl, halogenated C1-C4 alkyl, halogen;

[0016] The present invention preferably comprises a compound having the following structure or a pharmaceutically acceptable salt or isomer thereof:

[0017] Wherein, Ar is naphthyl, 2,3-methylenedioxy-phenyl, 4,5-dichloropyridyl,

[0018] X1 is hydrogen, phenyl, C1-C6 alkyl, C1-C4 alkoxy, F, Cl, Br, I, benzyloxy, CF3, methylsulfonyl, cyano, methoxycarbonyl;

[0019] R is Naphthyl, thienyl, furanyl;

[0020] X2 is hydrogen, C1-C4 alkoxy, C1-C4 alkyl, halogenated C1-C4 alkyl, or halogen.

[0021] The present invention preferably comprises a compound having the following structure or a pharmaceutically acceptable salt or isomer thereof:

[0022] Wherein Ar is naphthyl, 2,3-methylenedioxy-phenyl, 4,5-dichloropyridyl or

[0023] X1 is monosubstituted or polysubstituted at positions 2-5;

[0024] R is

[0025] X2 is monosubstituted or polysubstituted at positions 3 to 5.

[0026] The present invention preferably comprises a compound having the following structure or a pharmaceutically acceptable salt or isomer thereof:

[0027] Wherein, Ar is 4,5-dichloropyridyl;

[0028] R is naphthyl, thienyl, furanyl,

[0029] X2 is C1-C4 alkoxy, C1-C4 alkyl, halogenated C1-C4 alkyl, halogen;

[0030] The present invention preferably comprises a compound having the following structure or a pharmaceutically acceptable salt or isomer thereof:

[0031] Among them, Ar is X1 is a methylsulfone group;

[0032] R is naphthyl, thienyl, furyl, X2 is hydrogen, C1-C4 alkoxy, C1-C4 alkyl, CF3, Br, Cl.

[0033] The present invention preferably comprises a compound having the following structure or a pharmaceutically acceptable salt or isomer thereof:

[0034]

[0035]

[0036]

[0037] The pharmaceutically acceptable salt of the compound described in the present invention may be nitrate, hydrochloride, sulfate, phosphate or citrate of the compound.

[0038] The present invention also provides use of the above compound or its pharmaceutically acceptable salt or isomer in the preparation of SIRT3 agonists.

[0039] The present invention also provides the use of the above compound or its pharmaceutically acceptable salt or isomer in the preparation of anti-tumor drugs.

[0040] Furthermore, the compounds of the present invention or their pharmaceutically acceptable salts and isomers can be used as SIRT3 agonists for the treatment of tumors and related diseases.

[0041] The tumor is breast cancer, and the breast cancer is further triple-negative breast cancer.

[0042] The present invention also provides a pharmaceutical composition, which is a preparation containing an effective dose of the above-mentioned compound or its pharmaceutically acceptable salt, isomer. The compounds of the present invention can be made into the following forms by methods known in the art: tablets, capsules, aqueous or oily solutions, suspensions, emulsions, creams, ointments, gels, nasal sprays, suppositories, finely dispersed powders or aerosols or sprays for inhalation, sterile aqueous or oily solutions or suspensions or sterile emulsions for extra-gastric administration (including intravenous, intramuscular or infusion). Sterile water or water-propylene glycol solutions can be used as solvents to prepare liquid preparations, and active ingredients can also be formulated in polyethylene glycol aqueous solutions. Aqueous solutions for oral administration can be prepared by dissolving the active ingredient in water and adding suitable colorants, flavoring agents, stabilizers and thickeners as needed. Aqueous suspensions for oral use may be prepared by dispersing the finely divided active component in water together with viscous material such as natural synthetic gums, resins, methylcellulose, carboxymethylcellulose and other suspending agents known in the pharmaceutical art.

[0043] The pharmaceutical composition may be in unit dosage form. In these forms, the composition is divided into unit doses containing appropriate amounts of the active ingredient. The unit dosage form may be a packaged preparation comprising separate amounts of the preparation, such as boxed tablets, capsules, and powders in vials or ampoules. The unit dosage form may also be a capsule or tablet or it may be any of these packaged forms in appropriate quantities.

[0044] The active ingredient of the pharmaceutical composition of the present invention may be the compound of the present invention alone, or may be combined with other anti-tumor compounds as the active ingredient.

[0045] In the process of treating tumors, the compound of the present invention or its pharmaceutically acceptable salt, isomer or pharmaceutical composition can be used in combination with other anti-tumor drugs for treatment.

[0046] In treating tumors, such combined therapy can be achieved by administering the various therapeutic components simultaneously, sequentially or separately. Such combination products employ the compounds of the present invention within an effective dosage range and the other pharmaceutically active agents within an approved dosage range. BRIEF DESCRIPTION OF THE DRAWINGS

[0047] Figure 1 The proliferation of BT-549 and MDA-MB-231 cells after treatment with compound 42 for 6, 12, 24, 36 and 48 hours was detected by MTT assay.

[0048] Figure 2 The proliferation of BT-549 and MDA-MB-231 cells treated with compound 42 for 24 and 48 hours was examined for 3D tumor spheroid assays.

[0049] Figure 3DNA replication in BT-549 and MDA-MB-231 cells treated with 0 μM and 2 μM compound 42 was examined for EdU assay.

[0050] Figure 4 Cell migration of BT-549 cells treated with 0 μM, 1 μM, 2 μM and 4 μM compound 42 for 24 hours and 48 hours was examined for the scratch assay.

[0051] Figure 5 The cell migration of MDA-MB-231 cells treated with 0 μM, 1 μM, 2 μM and 4 μM compound 42 for 24 hours and 48 hours was examined for the scratch assay.

[0052] Figure 6 Flow cytometry was used to detect the glucose uptake and lactose production of BT-549 and MDA-MB-231 cells treated with 0 μM, 1 μM, 2 μM and 4 μM of compound 42.

[0053] Figure 7 Flow cytometry was used to detect mitochondrial membrane potential and ATP production in BT-549 and MDA-MB-231 cells treated with 0 μM, 1 μM, 2 μM and 4 μM compound 42.

[0054] Figure 8 Flow cytometry was used to detect apoptosis in BT-549 and MDA-MB-231 cells treated with 0 μM, 1 μM, 2 μM and 4 μM of compound 42.

[0055] Fig. 9 The in vivo antitumor activity of compound 42 against the BT-549 xenograft nude mouse model.

[0056] Fig.10 These are the daily body weights and organ ratios of animals in the BT-549 xenograft nude mouse model. DETAILED DESCRIPTION

[0057] The specific implementation modes of the present invention are further described below in conjunction with embodiments, but the present invention is not limited to the scope of the embodiments.

[0058] Example 1 Synthesis of Compounds 1 to 50

[0059] Compounds 1 to 50 are synthesized using the following reaction formula:

[0060]

[0061] Synthesis route and conditions: (I) glacial acetic acid, 80°C, heated to reflux for 8h, (II) DIEA, dissolved in dichloromethane, reacted at room temperature for 10h; wherein Ar and R are as described above.

[0062] Preparation of Intermediate 1-23a

[0063] Differently substituted aromatic aldehydes or heteroaromatic aldehydes (8.92mmol), ethyl propiolate (17.84mmol) and ammonium acetate (8.92mmol) were dissolved in glacial acetic acid (5mL) and stirred under reflux at 80℃ for 8h. TLC was used to monitor the progress of the reaction. The reaction was stopped, the reaction bottle was brought to room temperature, 50mL of water was added to quench the reaction, and an appropriate amount of sodium hydroxide solid was slowly added while stirring, and the pH of the reaction solution was adjusted to 7-8, and then stirred at room temperature for 10min. The mixture was extracted with ethyl acetate (3×30mL), the organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated under reduced pressure to obtain a brown crude product, which was purified by silica gel column chromatography under PE:EA=3:1 elution system to obtain intermediate 1-23a as a yellow solid with a yield of 30-40%.

[0064] Intermediate 1a, 1 H-NMR (400MHz, CDCl3), δ (ppm): 7.55 (d, 2H, J = 8.33Hz), 7.46 (d, 2H, J = 8.33Hz), 7.37 ( d,2H,J=5.31Hz),6.53(s,1H),4.97(s,1H),4.03-4.13(m,4H),1.18(t,6H,J=7.14Hz), 13 C-NMR (100MHz, CDCl3), δ (ppm): 166.6 (2C), 152.0, 134.0 (2C), 131.9 (2C), 129.2 (2C), 119.2, 110.1, 107.7 (2C), 60.3 (2C), 38.2, 14.2 (2C).

[0065] Intermediate 2a, 1 H-NMR (400MHz, CDCl3), δ (ppm): 7.28 (d, 2H, J = 5.35Hz), 7.25 (d, 2H, J = 8.65Hz), 6.86 (brs, 1H) ,6.78(d,2H,J=8.65Hz),4.83(s,1H),4.02-4.13(m,4H),3.74(s,3H),1.20(t,6H,J=7.20Hz), 13 C-NMR (100MHz, CDCl3), δ (ppm): 167.4 (2C), 158.1, 139.6, 133.7 (2C), 129.3 (2C), 113.3 (2C), 108.4 (2C), 60.0 (2C), 55.1, 36.7, 14.2 (2C).

[0066] Intermediate 3a,1 H-NMR (400MHz, CDCl3), δ (ppm): 7.32 (d, 2H, J = 5.30Hz), 7.23 (d, 2H, J = 8.00Hz), 7.05 (d, 2H, J = 8.00Hz),6.62(brs,1H),4.86(s,1H),4.03-4.13(m,4H),2.28(s,3H),1.19(t,6H,J=7.12Hz), 13 C-NMR (100MHz, CDCl3), δ (ppm): 167.0 (2C), 144.0, 133.3 (2C), 128.7 (3C), 128.2 (2C), 108.9 (2C), 60.0 (2C), 39.1, 21.1, 14.2 (2C).

[0067] Intermediate 4a, 1 H-NMR (400MHz, CDCl3), δ (ppm): 7.36 (d, 2H, J = 8.40Hz), 7.32 (d, 2H, J = 5.32Hz), 7.22 ( d,2H,J=8.40Hz),6.75(s,1H),4.87(s,1H),4.03-4.13(m,4H),1.19(t,6H,J=7.14Hz), 13 C-NMR (100MHz, CDCl3), δ (ppm): 167.0 (2C), 146.0, 133.9 (2C), 131.0 (2C), 130.1 (2C), 120.3, 108.1 (2C), 60.2 (2C), 37.3, 14.2 (2C).

[0068] Intermediate 5a, 1 H-NMR (400MHz, CDCl3), δ (ppm): 7.31 (d, 2H, J = 5.26Hz), 6.78-6.83 (m, 1H), 6.72 (t, 1H, J = 7.69Hz), 6. 62-6.83(m,1H),6.22(brs,1H),5.95(s,2H),5.02(s,1H),4.02-4.14(m,4H),1.19(t,6H,J=7.15Hz), 13 C-NMR (100MHz, CDCl3), δ (ppm): 167.0 (2C), 147.1, 145.1, 133.8 (2C), 128.6, 123.0, 121.0, 107.1, 106.6, 100.6, 60.0 (2C), 33.6, 14.2 (2C).

[0069] Intermediate 6a, 1H-NMR (400MHz, CDCl3), δ (ppm): 7.92 (d, 2H, J = 8.36Hz), 7.42 (d, 2H, J = 8.36Hz), 7.36 (d, 2H, J = 5.32Hz),6.44(brs,1H),4.97(s,1H),4.00-4.12(m,4H),3.88(s,3H),1.18(t,6H,J=7.19Hz), 13 C-NMR (100MHz, CDCl3), δ (ppm): 167.2, 166.8 (2C), 151.9, 133.8 (2C), 129.4 (2C), 128.5 (2C), 128.2, 108.2 (2C), 60.2 (2C), 52.0, 37.9, 14.2 (2C).

[0070] Intermediate 7a, 1 H-NMR (400MHz, CDCl3), δ (ppm): 7.56 (d, 2H, J = 8.27Hz), 7.32 (d, 2H, J = 5.26Hz), 7. 09(d,2H,J=8.27Hz),6.47(brs,1H),4.85(s,1H),4.03-4.13(m,4H),1.19(t,6H,J=

[0071] 7.17Hz), 13 C-NMR (100MHz, CDCl3), δ (ppm): 166.9 (2C), 146.6, 137.0 (2C), 133.7 (2C), 130.5 (2C), 108.2 (2C), 92.0, 60.2 (2C), 37.4, 14.2 (2C).

[0072] Intermediate 8a, 1 H-NMR (400MHz, CDCl3), δ (ppm): 8.26 (d, 1H, J = 2.16Hz), 7.72 (d, 1H, J = 2.16Hz), 7. 38(d,2H,J=5.33Hz),6.59(brs,1H),4.94(s,1H),4.07-4.15(m,4H),1.22(t,6H,J=

[0073] 7.08Hz), 13 C-NMR (100MHz, CDCl3), δ (ppm): 166.2 (2C), 147.2 (2C), 142.7, 138.8, 134.3 (3C), 107.1 (2C), 60.5 (2C), 35.0, 14.2 (2C).

[0074] Intermediate 9a,1 1H-NMR (400 MHz, CDCl3), δ (ppm): 7.70 - 7.80 (m, 4H), 7.50 - 7.57 (m, 1H), 7.71 - 7.75 (m, 2H), 7.37 - 7.43 (m, 2H), 7.36 (d, 2H, J = 5.33 Hz), 6.42 (brs, 1H), 5.07 (s, 1H), 4.00 - 4.09 (m, 4H), 1.17 (t, 6H, J = 7.10 Hz), 13 13C-NMR (100 MHz, CDCl3), δ (ppm): 167.0 (2C), 144.5, 133.4 (2C), 132.4, 127.9, 127.5 (2C), 127.2, 126.9, 126.4, 125.6, 125.3, 108.8 (2C), 60.1 (2C), 37.8, 14.2 (2C). Intermediate 10a, 1 1H-NMR (400 MHz, CDCl3), δ (ppm): 7.31 (d, 2H, J = 5.26 Hz), 6.98 (dd, 1H, J = 3.17 Hz), 6.78 - 6.83 (m, 1H), 6.74 (dd, 1H, J = 4.54 Hz), 6.28 (brs, 1H), 5.20 (s, 1H), 4.00 - 4.11 (m, 4H), 3.83 (s, 3H), 1.17 (t, 6H, J = 7.09 Hz), 13 13C-NMR (100 MHz, CDCl3), δ (ppm): 167.0 (2C), 153.3, 134.1 (2C), 117.6, 117.4, 113.7, 113.4, 111.7, 108.0 (2C), 59.9 (2C), 56.3, 32.5, 14.2 (2C).

[0075] Intermediate 11a, 1 1H-NMR (400 MHz, CDCl3), δ (ppm): 7.31 - 7.34 (m, 4H), 7.23 (d, 2H, J = 7.47 Hz), 7.14 (t, 1H, J = 6.92 Hz), 6.58 (brs, 1H), 4.89 (s, 1H), 4.02 - 4.13 (m, 4H), 1.19 (t, 6H, J = 7.16 Hz), 13 13C-NMR (100 MHz, CDCl3), δ (ppm): 167.2 (2C), 146.9, 133.7 (2C), 128.3 (2C), 128.0 (2C), 126.4, 108.5 (2C), 60.1 (2C), 37.6, 14.2 (2C).

[0076] Intermediate 12a, 1 H-NMR (400MHz, CDCl3), δ (ppm): 7.55 (d, 2H, J = 7.34Hz), 7.47 (d, 2H, J = 8.26Hz), 7.37-7.41 (m, 4H), 7.34 (d, 2 H,J=5.23Hz),7.30(d,1H,J=7.34Hz),6.55(brs,1H),4.94(s,1H),4.03-4.14(m,4H),1.20(t,6H,J=7.10Hz), 13 C-NMR (100MHz, CDCl3), δ (ppm): 167.1 (2C), 146.0, 141.2, 139.2, 133.7 (2C), 128. 7(2C),128.6(2C),127.0(3C),126.8(2C),108.5(2C),60.1(2C),37.3,14.2(2C).

[0077] Intermediate 13a, 1 H-NMR (400MHz, CDCl3), δ (ppm): 7.27-7.36 (m, 4H), 6.89-6.96 (t, 2H, J = 8.78 Hz),6.33(brs,1H),4.89(s,1H),4.01-4.15(m,4H),1.19(t,6H,J=6.96Hz), 13 C-NMR (100MHz, CDCl3), δ (ppm): 166.9 (2C), 133.4 (2C), 129.8 (2C), 114.8 (2C), 114.6 (2C), 108.7 (2C), 60.1 (2C), 37.0, 14.2 (2C).

[0078] Intermediate 14a, 1 H-NMR (400MHz, CDCl3), δ (ppm): 7.29 (d, 2H, J = 5.28Hz), 7.23 (d, 2H, J = 8.53Hz), 6.77 (d, 2H, J = 8.58Hz), 6.58 (brs,1H),4.83(s,1H),4.02-4.12(m,4H),3.94-4.00(m,2H),1.37(t,3H,J=7.01Hz),1.19(t,6H,J=7.13Hz), 13C-NMR (100MHz, CDCl3), δ (ppm): 167.3 (2C), 157.5, 139.4, 133.4 (2C), 129.3 (2C), 113.9 (2C), 108.7 (2C), 63.3, 60.0 (2C), 36.7, 14.9, 14.2 (2C).

[0079] Intermediate 15a, 1 H-NMR (400MHz, CDCl3), δ (ppm): 7.34-7.43 (m, 4H), 7.28-7.33 (m, 3H), 7.24 (s, 1H), 6.86 (d, 2H, J=8.72Hz),6.21(brs,1H),5.00(s,2H),4.85(s,1H),4.03-4.13(m,4H),1.19(t,6H,J=7.11Hz), 13 C-NMR (100 MHz, CDCl3), δ (ppm): 167.1 (2C), 157.4, 139.7, 137.2, 133.2 (2C), 129.3 (2C), 128.5 (2C), 127.9, 127.6 (2C), 114.2 (2C), 109.0 (2C), 70.0, 60.0 (2C), 36.7, 14.2 (2C). Intermediate 16a, 1 H-NMR (600MHz, CDCl3), δ (ppm): 7.39 (d, 1H, J = 2.02Hz), 7.35 (d, 2H, J = 5.13Hz), 7.31 (d, 1H, J = 8.21H z),7.20(dd,1H,J=2.00Hz),6.42(brs,1H),4.88(s,1H),4.04-4.15(m,4H),1.21(t,6H,J=7.15Hz), 13 C-NMR (100MHz, CDCl3), δ (ppm): 166.6 (2C), 147.1, 133.8 (2C), 132.0, 130.4, 130.3, 129.8, 127.9, 108.0 (2C), 60.3 (2C), 37.2, 14.2 (2C).

[0080] Intermediate 17a, 11H-NMR (400 MHz, CDCl3), δ (ppm): 7.38 (d, 1H, J = 7.92 Hz), 7.34 (d, 2H, J = 5.24 Hz), 6.95 (d, 1H, J = 1.85 Hz), 6.78 (dd, 1H, J = 1.85 Hz), 6.17 (brs, 1H), 4.87 (s, 1H), 4.06 - 4.12 (m, 6H), 1.44 (t, 3H, J = 6.98 Hz), 1.20 (t, 6H, J = 7.12 Hz), 13 13C-NMR (100 MHz, CDCl3), δ (ppm): 166.8 (2C), 154.8, 147.5, 133.4 (2C), 132.7 (2C), 121.6 (2C), 113.8, 108.5, 64.7, 60.2 (2C), 37.5, 14.7, 14.3 (2C).

[0081] Intermediate 18a, 1 1H-NMR (400 MHz, CDCl3), δ (ppm): 7.68 (d, 1H, J = 2.06 Hz), 7.33 (d, 2H, J = 5.33 Hz), 7.29 (dd, 1H, J = 2.06 Hz), 6.70 (d, 1H, J = 8.59 Hz), 6.25 (brs, 1H), 4.81 (s, 1H), 4.02 - 4.17 (m, 4H), 3.83 (s, 3H), 1.21 (t, 6H, J = 7.04 Hz), 13 13C-NMR (100 MHz, CDCl3), δ (ppm): 166.9 (2C), 156.6, 141.5, 139.2, 133.4 (3C), 129.5, 110.3, 108.6, 85.7, 60.1 (2C), 56.3, 36.5, 14.2 (2C).

[0082] Intermediate 19a, 1 1H-NMR (600 MHz, CDCl3), δ (ppm): 7.50 (d, 2H, J = 8.25 Hz), 7.46 (d, 2H, J = 8.25 Hz), 7.36 (d, 2H, J = 5.30 Hz), 6.53 (brs, 1H), 4.98 (s, 1H), 4.04 - 4.12 (m, 4H), 1.19 (t, 6H, J =

[0083] 7.15 Hz), 13C-NMR (100MHz, CDCl3), δ (ppm): 166. (2C), 150.5, 133.8 (3C), 128.7 (3C), 125.0 (2C), 108.2 (2C), 60.2 (2C), 37.7, 14.2 (2C).

[0084] Intermediate 20a, 1 H-NMR (500MHz, CDCl3), δ (ppm): 7.82 (d, 2H, J = 8.55Hz), 7.56 (d, 2H, J = 8.36Hz), 7.39 (d, 2H, J = 5.28Hz),6.60(brs,1H),5.03(s,1H),4.04-4.13(m,4H),3.03(s,3H),1.19(t,6H,J=7.10Hz), 13 C-NMR (100MHz, CDCl3), δ (ppm): 166.6 (2C), 152.9, 138.3, 134.1 (2C), 129.4 (2C), 127.2 (2C), 107.7 (2C), 60.3 (2C), 44.6, 38.0, 14.2 (2C).

[0085] Intermediate 21a, 1 H-NMR (400MHz, CDCl3), δ (ppm): 7.33 (d, 2H, J = 5.24Hz), 7.23 (d, 2H, J = 7.97Hz), 7. 04(d,2H,J=8.01Hz),6.12(brs,1H),4.87(s,1H),4.02-4.14(m,4H),2.53(t,2H,J=

[0086] 7.76Hz),1.53-1.60(m,4H),1.19(t,6H,J=7.20Hz),0.87(t,3H,J=7.15Hz), 13 C-NMR (100MHz, CDCl3), δ (ppm): 166.6 (2C), 150.5, 133.7 (2C), 128.7 (2C), 125.0, 124.9,124.4,108.2(2C),60.2(2C),37.7,35.0,31.5,30.1,22.7,14.2(2C),14.1.

[0087] Intermediate 22a, 1H-NMR (600MHz, DMSO-d6), δ (ppm): 7.33 (d, 2H, J = 5.45Hz), 7.07 (d, 2H, J = 8.10Hz), 7.03 (d, 2H, J = 8.10Hz), 4.67 ( s,1H),3.93-4.04(m,4H),1.48-1.58(m,2H),1.14-1.28(m,2H),1.12(t,6H,J=7.15Hz),0.87(t,3H,J=7.15Hz), 13 C-NMR (100MHz, DMSO-d6), δ (ppm): 166.9 (2C), 145.1, 135.3 (2C), 128.3 (2C) ,127.9(2C),106.4(2C),61.8,59.8(2C),37.4,36.9,24.5,14.6(2C),14.2.

[0088] Intermediate 23a, 1 H-NMR (600MHz, DMSO-d6), δ (ppm): 7.34 (d, 2H, J = 5.16Hz), 7.08 (d, 2H, J = 7.97Hz), 7.03 (d, 2H, J = 7.97Hz), 4.68 ( s,1H),3.94-4.04(m,4H),1.47-1.52(m,2H),1.14-1.39(m,4H),1.12(t,6H,J=7.14Hz),0.87(t,3H,J=7.30Hz), 13 C-NMR (100MHz, DMSO-d6), δ (ppm): 166.9 (2C), 145.0, 140.6, 135.3 (2C), 128.3 ( 2C),127.9(2C),106.4(2C),59.7(2C),36.9,34.9,33.6,22.3,14.6(2C),14.2.

[0089] General Synthesis Method of Compounds 1-50

[0090] In a 50 mL round-bottom flask, the intermediate 1-23a (0.17 mmol) was dissolved in anhydrous dichloromethane (7 mL), triethylamine (0.85 mmol) was added, and then a solution of different substituted acyl chlorides (0.26 mmol) dissolved in a small amount of anhydrous DCM was slowly added dropwise, and stirred at room temperature overnight. The reaction was monitored by TLC, and 25 mL of water was added to quench the reaction, and stirred at room temperature for 10 min. The mixture was extracted with ethyl acetate (3×15 mL), the organic phases were combined and washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated under reduced pressure to obtain a brown crude product. The final product 1-51 was purified by silica gel column chromatography in an elution system of PE:EA=2:1, as a white solid, with a yield of 35-40%.

[0091] The following are the NMR test results of compound 1-50.

[0092] Compound 1, 1 H-NMR (600MHz, DMSO-d6), δ (ppm): 7.96 (s, 2H), 7.77 (d, 2H, J = 8.27Hz), 7.56 (d, 2H, J = 8.27Hz), 7.06(s,2H),4.84(s,1H),3.99-4.08(m,4H),3.82(s,6H),3.77(s,3H),1.09(t,6H,J=7.27Hz), 13 C-NMR (100MHz, CDCl3), δ (ppm): 167.4, 165.3 (2C), 153.5 (2C), 153.4, 149.1, 132.5, 132.2 (2C), 132.0 (2C), 12 9.3(2C),128.4,125.9,114.1,106.9(2C),106.8,61.1(3C),56.5(2C),39.2,14.1(2C).HRMS(ESI+)m / z:calcd for C 28 H 28 N2O8([M+H] + ),521.1918; found,521.1900; HPLC t R =12.612min(96.63%purity).

[0093] Compound 2, 11H-NMR (600 MHz, DMSO-d6), δ (ppm): 7.91 (s, 2H), 7.22 (d, 2H, J = 8.74 Hz), 7.04 (s, 2H), 6.84 (d, 2H, J = 8.77 Hz), 4.71 (s, 1H), 4.00 - 4.08 (m, 4H), 3.82 (s, 6H), 3.77 (s, 3H), 3.71 (s, 3H), 1.11 (t, 6H, J = 7.09 Hz), 13 13C-NMR (150 MHz, DMSO-d6), δ (ppm): 167.6, 165.6 (2C), 158.6, 153.3 (3C), 141.2, 136.4, 131.2, 129.9 (2C), 127.0, 114.5, 114.0 (2C), 107.3 (3C), 60.8 (4C), 56.7 (2C), 37.9, 14.4 (2C). HRMS (ESI+) m / z: calcd for C 28 H 31 NO9 ([M+Na] + ), 548.1891; found, 548.1876;

[0094] HPLC t R = 14.741 min (100.00% purity).

[0095] Compound 3, 1 1H-NMR (400 MHz, CDCl3), δ (ppm): 8.11 (s, 2H), 7.23 (d, 2H, J = 8.08 Hz), 7.09 (d, 2H, J = 7.95 Hz), 6.90 (s, 2H), 4.91 (s, 1H), 4.07 - 4.017 (m, 4H), 3.94 (s, 3H), 3.89 (s, 6H), 2.30 (s, 3H), 1.20 (t, 6H, J = 7.10 Hz), 13 13C-NMR (100 MHz, CDCl3), δ (ppm): 167.6, 165.8 (2C), 153.4 (2C), 142.0, 141.0, 136.7, 131.3 (2C), 129.0 (2C), 128.3 (2C), 126.5, 115.6, 106.8 (3C), 61.1, 60.8 (2C), 56.4 (2C), 38.4, 21.1, 14.4 (2C). HRMS (ESI+) m / z: calcdfor C 28 H 31 NO8 ([M+H] +),510.2122; found,510.2108; HPLC t R =15.186min (97.91% purity).

[0096] Compound 4, 1 H-NMR (600MHz, CDCl3), δ (ppm): 8.13 (s, 2H), 7.41 (d, 2H, J = 8.21Hz), 7.22 (d, 2H, J = 8.21Hz), 6 .88(s,2H),4.92(s,1H),4.08-4.17(m,4H),3.94(s,3H),3.89(s,6H),1.20(t,6H,J=7.15Hz), 13 C-NMR (100MHz, CDCl3), δ (ppm): 167.5, 165.5 (2C), 153.4 (2C), 143.0, 142.1, 131.6 (2C), 131.4 (3C), 130. 2(3C),126.2,121.0,114.8,106.8(2C),61.1,60.9(2C),56.4(2C),38.5,14.1(2C).HRMS(ESI+)m / z:calcd for C 27 H 28 BrNO8([M+H] + ),574.1071; found,574.1060; HPLC t R =19.486min (95.20% purity).

[0097] Compound 5, 1 H-NMR (600MHz, DMSO-d6), δ (ppm): 7.92 (s, 2H), 7.01 (s, 2H), 6.77-6.81 (m, 3H), 5.99 (s ,2H),4.86(s,1H),4.01-4.09(m,4H),3.82(s,6H),3.77(s,3H),1.11(t,6H,J=7.11Hz), 13 C-NMR (150MHz, DMSO-d6), δ (ppm): 167.4, 165.5 (2C), 153.4 (3C), 147.5, 145.8, 141.3, 132.2, 126.9, 125.4, 12 3.4,121.7,112.3,107.8,107.2(3C),101.2,60.9(2C),60.8,56.7(2C),34.2,14.4(2C).HRMS(ESI+)m / z:calcd for C 28H 29 NO 10 ([M+Na] + ),562.1684; found,562.1674; HPLC t R =13.065min (98.23% purity).

[0098] Compound 6, 1 H-NMR (600MHz, DMSO-d6), δ (ppm): 7.96 (s, 2H), 7.89 (d, 2H, J = 8.33Hz), 7.49 (d, 2H, J = 8.34Hz), 7.06 ( s,2H),4.85(s,1H),3.98-4.08(m,4H),3.83(s,3H),3.82(s,6H),3.77(s,3H),1.09(t,6H,J=7.13Hz), 13 C-NMR (150MHz, DMSO-d6), δ (ppm): 167.6, 166.5, 165.4 (2C), 153.3 (3C), 149.6, 141.3, 132.0, 129.6 (2C), 129.4 ( 2C),128.8,126.9,113.5,107.4(3C),61.0(2C),60.8,56.7(2C),52.6,39.1,14.4(2C).HRMS(ESI+)m / z:calcdfor C 29 H 31 NO 10 ([M+H] + ),554.2021; found,554.2005; HPLC t R =13.440min (95.47% purity).

[0099] Compound 7, 1 H-NMR (400MHz, CDCl3), δ (ppm): 8.13 (s, 2H), 7.61 (d, 2H, J = 8.31Hz), 7.09 (d, 2H, J = 8.31Hz), 6 .87(s,2H),4.90(s,1H),4.06-4.17(m,4H),3.94(s,3H),3.89(s,6H),1.20(t,6H,J=7.11Hz), 13C-NMR (150MHz, DMSO-d6), δ (ppm): 167.6, 165.4 (2C), 153.3 (2C), 147.6, 144.2, 141.2, 138.4 (2C), 137.4 (2C), 131.3(2C),126.9,118.9,107.4(2C),93.6,61.0(2C),60.8,56.8(2C),38.6,14.4(2C).HRMS(ESI+)m / z:calcd for C 27 H 28 INO8([M+H] + ),622.0932; found,622.0916;

[0100] HPLC R =20.515min (98.14% purity).

[0101] Compound 8, 1 H-NMR (600MHz, DMSO-d6), δ (ppm): 8.42 (d, 1H, J = 2.13Hz), 8.16 (d, 1H, J = 2.13Hz), 7.97 (s, 2H), 7.09(s,2H),4.85(s,1H),4.00-4.11(m,4H),3.83(s,6H),3.78(s,3H),1.10(t,6H,J=7.11Hz), 13 C-NMR (150MHz, DMSO-d6), δ (ppm): 167.6, 165.2 (2C), 153.3 (3C), 148.6, 146.4, 141.5, 141.2, 139.8, 13 2.8,129.5,126.9,111.8,107.5(3C),61.1(2C),60.8,56.7(2C),36.2,14.3(2C).HRMS(ESI+)m / z:calcd for C 26 H 26 Cl2N2O8([M+H] + ),565.1139; found,565.1123; HPLC t R =15.196min (98.92% purity).

[0102] Compound 9, 11H-NMR (500 MHz, CDCl3), δ (ppm): 8.18 (s, 2H), 7.76 - 7.80 (m, 4H), 7.50 - 7.52 (m, 1H), 7.43 - 7.46 (m, 2H), 6.94 (s, 2H), 5.13 (s, 1H), 4.05 - 4.13 (m, 4H), 3.94 (s, 3H), 3.88 (s, 6H), 1.18 (t, 6H, J = 7.42 Hz), 13 13C-NMR (125 MHz, CDCl3), δ (ppm): 167.6, 165.7 (2C), 153.4 (2C), 142.0, 141.4, 133.4, 132.6, 131.4, 127.9, 127.8, 127.6, 127.3, 126.8, 126.4, 126.1, 125.8, 115.4 (2C), 106.9 (3C), 61.1, 60.9 (2C), 56.4 (2C), 39.0, 14.1 (2C). HRMS (ESI+) m / z: calcd for C 31 H 31 NO8([M + Na] + ), 568.1942; found, 568.1926; HPLC t R = 19.345 min (96.57% purity).

[0103] Compound 10, 1 1H-NMR (400 MHz, CDCl3), δ (ppm): 8.11 (s, 2H), 7.01 - 7.07 (m, 1H), 6.96 (s, 2H), 6.86 - 6.93 (m, 1H), Hz), 6.78 - 6.82 (m, 1H), 5.10 (s, 1H), 4.06 - 4.18 (m, 4H), 3.94 (s, 3H), 3.89 (s, 6H), 3.80 (s, 3H), 1.20 (t, 6H, J = 7.11 Hz), 13 13C-NMR (125 MHz, CDCl3), δ (ppm): 167.4, 165.8 (2C), 153.4 (2C), 142.0, 132.5 (2C), 126.5, 118.4, 118.2, 114.5, 114.3, 113.5, 112.5 (2C), 107.0 (3C), 61.1, 60.7 (2C), 56.5, 56.4 (2C), 35.8, 14.1 (2C). HRMS (ESI+) m / z: calcd for C 28 H 30 FNO9([M + H] +), 544.1977; found, 544.1959; HPLC t R = 15.501 min (99.38% purity).

[0104] Compound 11, 1 1H-NMR (600 MHz, DMSO-d6), δ (ppm): 7.97 (s, 2H), 7.64 (d, 2H, J = 7.62 Hz), 7.59 (d, 2H, J = 8.19 Hz), 7.45 (t, 2H, J = 7.62 Hz), 7.41 (d, 2H, J = 8.19 Hz), 7.35 (t, 1H, J = 7.62 Hz), 7.07 (s, 2H), 4.82 (s, 1H), 4.01 - 4.12 (m, 4H), 3.83 (s, 6H), 3.77 (s, 3H), 1.12 (t, 6H, J = 7.09 Hz), 13 13C-NMR (150 MHz, DMSO-d6), δ (ppm): 167.1, 165.0 (2C), 152.3 (2C), 142.9, 140.6, 139.7, 138.7, 131.1, 128.9 (2C), 128.8 (4C), 126.5 (5C), 126.4, 113.5, 106.8 (2C), 60.3 (2C), 60.2, 56.2 (2C), 37.9, 13.8 (2C). HRMS (ESI+) m / z: calcd for C 33 H 33 NO8 ([M+Na] + ), 594.2098; found, 594.2088; HPLC t R = 17.496 min (97.50% purity).

[0105] Compound 12, 1 1H-NMR (400 MHz, CDCl3), δ (ppm): 8.12 (s, 2H), 7.28 - 7.32 (m, 2H), 6.97 (t, 2H, J = 7.61 Hz), 6.88 (s, 2H), 4.94 (s, 1H), 4.07 - 4.18 (m, 4H), 3.94 (s, 3H), 3.89 (s, 6H), 1.20 (t, 6H, J = 7.11 Hz), 13C-NMR (150MHz, CDCl3), δ (ppm): 167.5, 165.6 (2C), 153.4 (2C), 142.0, 139.7, 131.4 (2C), 130.0, 129.9, 12 6.2,115.2(2C),115.1(2C),106.8(3C),61.1,60.9(2C),56.4(2C),38.2,14.1(2C).HRMS(ESI+)m / z:calcd for C 27 H 28 FNO8([M+H] + ),514.1872; found,514.1859; HPLC t R =15.920min(97.56%purity).

[0106] Compound 13, 1 H-NMR (600MHz, DMSO-d6), δ (ppm): 7.90 (s, 2H), 7.20 (d, 2H, J = 8.72Hz), 7.04 (s, 2H), 6.82 (d, 2H, J = 8.72Hz), 4.70 (s, 1H),4.00-4.08(m,4H),3.95-3.99(m,2H),3.82(s,6H),3.77(s,3H),1.29(t,3H,J=7.05Hz),1.11(t,6H,J=7.08Hz), 13 C-NMR (100MHz, CDCl3), δ (ppm): 167.6, 165.8 (2C), 158.0, 153.4 (2C), 141.9, 136.0, 131.1 (2C), 129.4 (3C), 126 .5,115.7,114.2(2C),106.8(2C),63.4,61.1,60.8(2C),56.4(2C),38.0,14.9,14.2(2C).HRMS(ESI+)m / z:calcd for C 29 H 33 NO9([M+Na] + ),562.2048; found,562.2035; HPLC t R =16.967min (98.28% purity).

[0107] Compound 14, 11H-NMR (600 MHz, CDCl3), δ (ppm): 8.10 (s, 2H), 7.30 - 7.43 (m, 5H), 7.24 (s, 1H), 7.04 (s, 2H), 6.89 (d, 4H, J = 8.15 Hz), 5.02 (s, 2H), 4.90 (s, 1H), 4.06 - 4.20 (m, 4H), 3.94 (s, 3H), 3.88 (s, 6H), 1.20 (t, 6H, J = 7.15 Hz), 13 13C-NMR (150 MHz, DMSO-d6), δ (ppm): 167.0, 165.0 (2C), 157.2, 152.7 (2C), 140.6, 137.0, 136.1, 130.6, 129.4 (2C), 128.3 (3C), 127.7, 127.6 (3C), 126.5, 114.3 (2C), 113.9, 106.7 (2C), 69.0, 60.3 (2C), 60.2, 56.2 (2C), 37.3, 13.8 (2C). HRMS (ESI+) m / z: calcd for C 34 H 35 NO9 ([M + Na] + ), 624.2204; found, 624.2195; HPLC t R = 17.183 min (97.65% purity).

[0108] Compound 15, 1 1H-NMR (500 MHz, CDCl3), δ (ppm): 8.14 (s, 2H), 7.34 - 7.38 (m, 3H), 7.21 - 7.23 (m, 1H), 6.88 (s, 2H), 4.92 (s, 1H), 4.10 - 4.18 (m, 4H), 3.95 (s, 3H), 3.90 (s, 6H), 1.22 (t, 6H, J = 7.02 Hz), 13 13C-NMR (100 MHz, CDCl3), δ (ppm): 167.5, 165.4 (2C), 153.5 (2C), 144.2, 132.4, 131.9 (2C), 131.1, 130.3, 130.2, 128.1, 126.1, 114.3 (2C), 106.8 (3C), 61.1, 61.0 (2C), 56.4 (2C), 38.3, 14.1 (2C). HRMS (ESI+) m / z: calcd for C 27 H 27 Cl2NO8 ([M + H] +), 564.1186; found, 564.1169; HPLC t R =

[0109] 21.063 min (99.08% purity).

[0110] Compound 16, 1 1H-NMR (600 MHz, DMSO-d6), δ (ppm): 7.93 (s, 2H), 7.48 (d, 1H, J = 8.10 Hz), 7.03 (s, 2H), 6.97 (d, 1H, J = 2.16 Hz), 6.84 (m, 1H, J = 2.15 Hz), 4.76 (s, 1H), 4.03 - 4.11 (m, 6H), 3.82 (s, 6H), 3.78 (s, 3H), 1.34 (t, 3H, J = 6.91 Hz), 1.11 (t, 6H, J = 7.24 Hz), 13 13C-NMR (150 MHz, DMSO-d6), δ (ppm): 167.0, 165.0 (2C), 154.1, 152.7 (3C), 144.9, 140.7, 13.9 (2C), 132.6, 131.2, 126.3, 121.8, 114.0, 113.0, 109.5, 106.8 (3C), 64.3, 60.4 (2C), 60.2, 56.1 (2C), 38.2, 14.4. HRMS (ESI+) m / z: calcd for C 29 H 32 BrNO9 ([M + H] + ), 618.1333; found, 618.1319; HPLC t R = 17.176 min (95.00% purity).

[0111] Compound 17, 1 1H-NMR (500 MHz, CDCl3), δ (ppm): 8.10 (s, 2H), 7.66 (d, 1H, J = 2.09 Hz), 7.33 (dd, 1H, J = 2.14 Hz), 6.90 (s, 2H), 6.74 (d, 1H, J = 8.43 Hz), 4.85 (s, 1H), 4.09 - 4.18 (m, 4H), 3.94 (s, 3H), 3.91 (s, 6H), 3.85 (s, 3H), 1.22 (t, 6H, J = 7.05 Hz), 13C-NMR (125MHz, CDCl3), δ (ppm): 167.6, 165.6 (2C), 157.1, 153.4 (2C), 142.0, 139.1, 138.3, 131.5 (2C), 129.9 ( 2C),126.3,115.0,110.5,106.8(3C),61.1,60.9(2C),56.5(2C),56.3,37.6,14.2(2C).HRMS(ESI+)m / z:calcd for C 28 H 30 INO9([M+Na] + ),674.0857; found,674.0837; HPLCt R =16.208min (96.57% purity).

[0112] Compound 18, 1 H-NMR (400MHz, CDCl3), δ (ppm): 8.17 (s, 2H), 7.55 (d, 2H, J = 8.17Hz), 7.46 (d, 2H, J = 8.17Hz), 6 .89(s,2H),5.03(s,1H),4.08-4.18(m,4H),3.95(s,3H),3.89(s,6H),1.20(t,6H,J=7.11Hz), 13 C-NMR (125MHz, CDCl3), δ (ppm): 167.5, 165.4 (2C), 153.5 (2C), 147.7, 142.2, 131.8 (2C), 128.8 (3C), 1 26.1,125.3(2C),114.6,106.9(4C),61.1,61.0(2C),56.5(2C),38.9,14.1(2C).HRMS(ESI+)m / z:calcd for C 28 H 28 F3NO8([M+H] + ),564.1840; found,564.1833; HPLC t R =18.793min (96.44% purity).

[0113] Compound 19, 11H-NMR (400 MHz, CDCl3), δ (ppm): 8.18 (s, 2H), 7.87 (d, 2H, J = 8.30 Hz), 7.55 (d, 2H, J = 8.30 Hz), 6.88 (s, 2H), 5.06 (s, 1H), 4.06 - 4.19 (m, 4H), 3.95 (s, 3H), 3.90 (s, 6H), 3.05 (s, 3H), 1.20 (t, 6H, J = 7.15 Hz), 13 13C-NMR (150 MHz, CDCl3), δ (ppm): 167.4, 165.3 (2C), 153.5 (2C), 150.0, 142.2, 139.2, 132.0 (2C), 129.5 (3C), 127.5 (2C), 125.9, 114.1, 106.9 (2C), 61.1 (3C), 56.5 (2C), 44.5, 39.0, 14.1 (2C). HRMS (ESI+) m / z: calcd for C 28 H 31 NO 10 S ([M + H] + ), 574.1741; found, 574.1732; HPLC t R = 10.389 min (97.96% purity).

[0114] Compound 20, 1 1H-NMR (600 MHz, DMSO-d6), δ (ppm): 7.91 (s, 2H), 7.19 (d, 2H, J = 8.15 Hz), 7.09 (d, 2H, J = 8.15 Hz), 7.02 (s, 2H), 4.72 (s, 1H), 3.98 - 4.07 (m, 4H), 3.81 (s, 6H), 3.75 (s, 3H), 1.49 - 1.54 (m, 2H), 1.14 - 1.35 (m, 6H), 1.08 (t, 6H, J = 7.15 Hz), 0.83 (t, 3H, J = 7.03 Hz), 13 13C-NMR (125 MHz, CDCl3), δ (ppm): 167.6, 165.8 (2C), 153.4 (2C), 141.7, 141.0, 131.3 (2C), 128.3 (3C), 128.2 (2C), 126.5, 115.6, 106.8 (3C), 61.1, 60.8 (2C), 56.4 (2C), 38.4, 35.6, 31.6, 31.0, 22.5, 14.1 (2C), 14.0. HRMS (ESI+) m / z: calcd for C 32 H 39NO8([M + Na] + ), 588.2568; found, 588.2550; HPLC t R = 20.996 min (95.46% purity).

[0115] Compound 21, 1 1H-NMR (400 MHz, CDCl3), δ (ppm): 8.11 (s, 2H), 7.24 (d, 2H, J = 8.15 Hz), 7.09 (d, 2H, J = 8.15 Hz), 6.90 (s, 2H), 4.92 (s, 1H), 4.07 - 4.18 (m, 4H), 3.94 (s, 3H), 3.89 (s, 6H), 2.54 (t, 2H, J = 7.25 Hz), 1.58 - 1.64 (m, 2H), 1.20 (t, 6H, J = 7.08 Hz), 0.87 (t, 3H, J = 7.27 Hz), 13 13C-NMR (125 MHz, CDCl3), δ (ppm): 167.6, 165.8 (2C), 153.4, 141.9, 141.4, 141.1, 131.3, 128.4 (3C), 128.2 (3C), 126.5, 115.6, 106.8 (3C), 61.1, 60.8 (2C), 56.4 (2C), 38.4, 37.7, 24.4, 14.1 (2C), 13.9. HRMS (ESI+) m / z: calcd for C 30 H 35 NO8([M + H] + ), 538.2435; found, 538.2433; HPLC t R = 18.097 min (97.56% purity).

[0116] Compound 22, 1 1H-NMR (600 MHz, CDCl3), δ (ppm): 8.11 (s, 2H), 7.23 (d, 2H, J = 8.09 Hz), 7.09 (d, 2H, J = 8.09 Hz), 6.89 (s, 2H), 4.92 (s, 1H), 4.07 - 4.19 (m, 4H), 3.94 (s, 3H), 3.89 (s, 6H), 2.52 - 2.59 (m, 2H), 1.23 - 1.45 (m, 4H), 1.20 (t, 6H, J = 7.17 Hz), 0.91 (t, 3H, J = 7.36 Hz), 13C-NMR (150MHz, CDCl3), δ (ppm): 167.6, 165.8 (2C), 153.4 (2C), 142.0, 141.7, 141.0, 131.3 (2C), 128.4 (2C), 128.2 (2C), 126.5,115.6(2C),106.8(2C),61.1,60.8(2C),56.4(2C),38.4,35.3,33.5,22.4,14.1(2C),14.0.HRMS(ESI+)m / z:calcd for C 31 H 37 NO8([M+Na] + ),574.2411; found,574.2400;

[0117] HPLC R =19.629min (95.19% purity).

[0118] Compound 23, 1 H-NMR (600MHz, CDCl3), δ (ppm): 8.15 (s, 2H), 7.88 (d, 2H, J = 8.42Hz), 7.63-7.67 (m, 3H), 7.55- 7.57(m,4H),4.89(s,1H),5.06(s,1H),4.07-4.16(m,4H),3.05(s,3H),1.19(t,6H,J=7.11Hz), 13 C-NMR (150MHz, CDCl3), δ (ppm): 167.9, 165.3 (2C), 149.9, 139.1, 133.0, 131.9, 131.3, 129.5 (3C),129.1(4C),127.5(3C),114.3,61.1(2C),44.5,39.0,14.1(2C).HRMS(ESI+)m / z:calcd for C 25 H 25 NO7S([M+H] + ),484.1424; found,484.1383; HPLC t R =11.029min (96.00% purity).

[0119] Compound 24, 11H-NMR (400 MHz, CDCl3), δ (ppm): 8.00 (s, 2H), 7.87 (d, 2H, J = 8.37 Hz), 7.52 (d, 2H, J = 8.37 Hz), 7.45 - 7.49 (m, 1H), 7.33 - 7.36 (m, 3H), 5.03 (s, 1H), 4.04 - 4.14 (m, 4H), 3.04 (s, 3H), 2.39 (s, 3H), 1.18 (t, 6H, J = 7.14 Hz), 13 13C-NMR (150 MHz, CDCl3), δ (ppm): 168.2, 165.2 (2C), 149.7, 139.1, 131.4, 131.3, 130.9, 129.6 (4C), 127.4 (5C), 126.4, 114.7, 61.1 (2C), 44.5, 39.1, 19.5, 14.1 (2C). HRMS (ESI+) m / z: calcd for C 26 H 27 NO7S ([M + H] + ), 498.1580; found, 498.1540; HPLC t R = 13.703 min (95.14% purity).

[0120] Compound 25, 1 1H-NMR (600 MHz, CDCl3), δ (ppm): 8.15 (s, 2H), 7.88 (d, 2H, J = 8.26 Hz), 7.56 (d, 4H, J = 7.52 Hz), 7.35 (d, 2H, J = 8.07 Hz), 5.06 (s, 1H), 4.08 - 4.16 (m, 4H), 3.05 (s, 3H), 2.47 (s, 3H), 1.20 (t, 6H, J = 7.12 Hz), 13 13C-NMR (150 MHz, CDCl3), δ (ppm): 167.9, 165.4 (2C), 150.1, 144.0, 139.1, 132.2 (2C), 129.7 (2C), 129.5 (2C), 129.3 (2C), 127.5 (2C), 114.0, 61.0 (2C), 44.5, 39.0, 21.7, 14.1 (2C). HRMS (ESI+) m / z: calcd for C 26 H 27 NO7S ([M + H] + ), 498.1580; found, 498.1534; HPLC t R = 13.242 min (98.95% purity).

[0121] Compound 26, 1 H-NMR (400MHz, CDCl3), δ (ppm): 8.15 (s, 2H), 7.88 (d, 2H, J = 8.11Hz), 7.55 (d, 2H, J = 8.19Hz), 7.45 (t, 1H, J = 8 .05Hz),7.15-7.19(m,3H),5.05(s,1H),4.07-4.16(m,4H),3.88(s,3H),3.05(s,3H),1.20(t,6H,J=7.18Hz), 13 C-NMR (125MHz, CDCl3), δ (ppm): 167.7, 165.3 (2C), 160.1, 149.9, 139.1, 132.6, 131.9, 130.1, 129.5 (3C ),127.5(3C),120.9,118.8,114.4,114.3,61.1(2C),55.6,44.5,39.0,14.1(2C).HRMS(ESI+)m / z:calcd for C 26 H 27 NO8S([M+H] + ),514.1529; found,514.1490; HPLC t R =11.500min(97.39%purity).

[0122] Compound 27, 1 H-NMR (400MHz, CDCl3), δ (ppm): 8.16 (s, 2H), 7.88 (d, 2H, J = 8.31Hz), 7.66 (d, 2H, J = 8.78Hz), 7.56 (d, 2H, J = 8.3 1Hz),7.03(d,2H,J=8.78Hz),5.06(s,1H),4.07-4.17(m,4H),3.91(s,3H),3.05(s,3H),1.20(t,6H,J=7.15Hz), 13 C-NMR (100MHz,CDCl3),δ(ppm):

[0123] 167.5,165.5(2C),163.5,150.2,139.1,132.5(2C),131.7(2C),129.5(2C),127.5(3C), 123.1,114.4(2C),113.7,61.0(2C),55.6,44.5,39.0,14.1(2C).HRMS(ESI+)m / z:calcd for C26 H 27 NO8S([M+H] + ), 514.1529; found, 514.1486; HPLC t R = 11.626 min (98.25% purity).

[0124] Compound 28, 1 H-NMR (500 MHz, CDCl3), δ (ppm): 8.17 (s, 2H), 7.88 (d, 2H, J = 8.38 Hz), 7.57 (d, 2H, J = 8.38 Hz), 7.27 (s, 1H), 7.21 - 7.25 (m, 1H), 6.97 (d, 2H, J = 8.36 Hz), 5.07 (s, 1H), 4.08 - 4.17 (m, 4H), 3.98 (s, 3H), 3.95 (s, 3H), 3.05 (s, 3H), 1.20 (t, 6H, J = 7.15 Hz), 13 C-NMR (125 MHz, CDCl3), δ (ppm): 167.5, 165.4 (2C), 153.3, 150.2, 149.6, 139.1, 132.5 (2C), 129.5 (2C), 127.7, 127.5 (2C), 123.2 (2C), 113.8, 112.5, 110.5, 61.0 (2C), 56.2 (2C), 44.5, 39.0, 14.2 (2C). HRMS (ESI+) m / z: calcd for C 27 H 29 NO9S([M+H] + ), 544.1635; found, 544.1588; HPLC t R = 9.810 min (88.72% purity).

[0125] Compound 29, 1 H-NMR (400 MHz, CDCl3), δ (ppm): 8.09 (s, 2H), 7.89 (d, 2H, J = 8.35 Hz), 7.84 (d, 2H, J = 8.35 Hz), 7.76 (d, 2H, J = 8.18 Hz), 7.55 (d, 2H, J = 8.35 Hz), 5.06 (s, 1H), 4.07 - 4.17 (m, 4H), 3.05 (s, 3H), 1.19 (t, 6H, J = 7.10 Hz), 1313C-NMR(125MHz, CDCl3), δ(ppm): 166.5, 165.1(2C), 149.5, 139.3, 131.1, 129.6(4C), 129.4(4C), 127.6(4C), 126.2, 115.2, 61.2(2C), 44.5, 39.1, 14.1(2C). HRMS(ESI+) m / z: calcd for C 26 H 24 F3NO7S ([M+H] + ), 552.1298; found, 552.1250; HPLC t R =

[0126] 14.396 min (95.41% purity).

[0127] Compound 30, 1 1H-NMR(400MHz, CDCl3), δ(ppm): 8.12(s, 2H), 7.88(d, 2H, J = 8.35Hz), 7.68 - 7.72(m, 2H), 7.55(d, 2H, J = 8.35Hz), 7.24(d, 2H, J = 7.44Hz), 5.06(s, 1H), 4.07 - 4.17(m, 4H), 3.05(s, 3H), 1.20(t, 6H, J = 7.10Hz), 13 13C-NMR(100MHz, CDCl3), δ(ppm): 166.8, 165.2(2C), 149.8, 139.2, 131.9(2C), 131.7(3C), 129.5(3C), 127.5(3C), 116.6, 116.4, 114.5, 61.1(2C), 44.5, 39.0, 14.1(2C). HRMS(ESI+) m / z: calcd for C 25 H 24 FNO7S ([M+H] + ), 502.1330; found, 502.1285; HPLC t R =11.600 min (96.56% purity).

[0128] Compound 31, 11H-NMR (400 MHz, CDCl3), δ (ppm): 8.10 (s, 2H), 7.88 (d, 2H, J = 8.33 Hz), 7.61 (d, 2H, J = 8.52 Hz), 7.53 - 7.56 (m, 4H), 5.06 (s, 1H), 4.07 - 4.17 (m, 4H), 3.05 (s, 3H), 1.20 (t, 6H, J = 7.06 Hz), 13 13C-NMR (125 MHz, CDCl3), δ (ppm): 167.0, 166.2 (2C), 149.7, 139.2, 132.5 (3C), 131.5, 130.6 (3C), 130.2, 129.5 (2C), 128.0, 127.5 (2C), 114.7, 61.2 (2C), 44.5, 39.0, 14.1 (2C). HRMS (ESI+) m / z: calcd for C 25 H 24 ClNO7S ([M + H] + ), 518.1034; found, 518.0987; HPLC t R = 13.905 min (98.35% purity).

[0129] Compound 32, 1 1H-NMR (400 MHz, CDCl3), δ (ppm): 8.10 (s, 2H), 7.88 (d, 2H, J = 8.20 Hz), 7.71 (d, 2H, J = 8.52 Hz), 7.53 - 7.56 (m, 4H), 5.06 (s, 1H), 4.07 - 4.17 (m, 4H), 3.05 (s, 3H), 1.20 (t, 6H, J = 7.12 Hz), 13 13C-NMR (125 MHz, CDCl3), δ (ppm): 167.0, 165.2 (2C), 149.7, 139.3, 132.5 (3C), 131.5, 130.6 (3C), 129.5 (3C), 127.5 (3C), 114.7, 61.2 (2C), 44.5, 39.0, 14.1 (2C). HRMS (ESI+) m / z: calcd for C 25 H 24 BrNO7S ([M + H] + ), 562.0529; found, 562.0481; HPLC t R = 14.658 min (97.92% purity). Compound 33, 11H-NMR (600 MHz, CDCl3), δ (ppm): 8.07 - 8.08 (m, 2H), 7.96 - 7.98 (m, 1H), 7.91 (d, 3H, J = 8.20 Hz), 7.55 - 7.65 (m, 7H), 5.05 (s, 1H), 4.02 - 4.10 (m, 4H), 3.05 (s, 3H), 1.14 (t, 6H, J = 7.18 Hz), 13 13C-NMR (150 MHz, CDCl3), δ (ppm): 167.7, 165.2 (2C), 149.7, 139.1, 133.7, 132.2, 129.6 (4C), 127.5 (3C), 129.0, 128.2 (2C), 127.2 (2C), 126.6, 124.8, 124.3, 114.7, 61.0 (2C), 44.5, 39.1, 14.0 (2C). HRMS (ESI+) m / z: calcd for C 29 H 27 NO7S ([M + H] + ), 534.1580; found, 534.1527; HPLC t R = 14.407 min (97.56% purity).

[0130] Compound 34, 1 1H-NMR (500 MHz, CDCl3), δ (ppm): 8.21 (s, 2H), 7.95 - 8.01 (m, 3H), 7.90 - 7.92 (m, 3H), 7.66 - 7.69 (m, 2H), 7.59 - 7.64 (m, 3H), 5.09 (s, 1H), 4.06 - 4.15 (m, 4H), 3.06 (s, 3H), 1.18 (t, 6H, J = 7.17 Hz), 13 13C-NMR (125 MHz, CDCl3), δ (ppm): 168.0, 165.3 (2C), 150.0, 139.2, 132.4, 132.0 (2C), 130.6 (2C), 129.6 (3C), 129.1 (2C), 128.9, 128.0, 127.6, 127.5 (2C), 124.5 (2C), 114.3, 61.1 (2C), 44.5, 39.1, 14.1 (2C). HRMS (ESI+) m / z: calcd for C 29 H 27 NO7S ([M + H] + ), 534.1580;

[0131] found, 534.1535; HPLC tR = 15.321 min (96.17% purity).

[0132] Compound 35, 1 1H-NMR (600 MHz, CDCl3), δ (ppm): 8.44 (s, 2H), 7.85 (d, 2H, J = 8.44 Hz), 7.72 - 7.73 (m, 1H), 7.55 (d, 1H, J = 8.39 Hz), 7.44 - 7.46 (dd, 2H), 6.68 - 6.69 (m, 1H), 5.06 (s, 1H), 4.11 - 4.19 (m, 4H), 3.03 (s, 3H), 1.23 (t, 6H, J = 7.09 Hz), 13 13C-NMR (150 MHz, CDCl3), δ (ppm): 165.4 (2C), 155.9, 149.7, 147.2, 145.2, 139.1, 131.6, 129.6, 127.5 (3C), 122.2, 114.7 (2C), 112.8 (2C), 61.1 (2C), 44.5, 38.9, 14.1 (2C). HRMS (ESI+) m / z: calcd for C 23 H 23 NO8S ([M + H] + ), 474.1216; found, 474.1170; HPLC t R = 8.937 min (95.75% purity).

[0133] Compound 36, 1 1H-NMR (500 MHz, CDCl3), δ (ppm): 8.32 (s, 2H), 7.87 (d, 2H, J = 8.27 Hz), 7.78 (d, 1H, J = 4.96 Hz), 7.66 (d, 2H, J = 3.89 Hz), 7.55 (d, 2H, J = 8.36 Hz), 7.19 - 7.25 (m, 1H), 5.08 (s, 1H), 4.10 - 4.19 (m, 4H), 3.04 (s, 3H), 1.22 (t, 6H, J = 7.06 Hz), 13 13C-NMR (125 MHz, CDCl3), δ (ppm): 165.3 (2C), 161.0, 149.8, 139.2, 134.2, 133.5, 132.0 (2C), 129.5 (2C), 128.1 (2C), 127.6 (2C), 114.5 (2C), 61.1 (2C), 44.5, 39.0, 14.1 (2C). HRMS (ESI+) m / z: calcd for C 23 H23 NO7S2([M+H] + ), 490.0988; found, 490.0947; HPLC t R = 10.925 min (96.64% purity).

[0134] Compound 37, 1 1H-NMR (500 MHz, CDCl3), δ (ppm): 8.29 (d, 1H, J = 1.87 Hz), 8.14 (s, 2H), 7.73 (d, 1H, J = 1.87 Hz), 7.62 - 7.68 (m, 3H), 7.54 - 7.59 (m, 2H), 4.96 (s, 1H), 4.09 - 4.18 (m, 4H), 1.22 (t, 6H, J = 7.03 Hz), 13 13C-NMR (125 MHz, CDCl3), δ (ppm): 167.8, 165.1 (2C), 147.4 (2C), 139.9, 138.8, 133.0 (2C), 132.2 (2C), 131.2, 130.3, 129.1 (4C), 113.3, 61.3 (2C), 36.0, 14.1 (2C). HRMS (ESI+) m / z: calcd for C 23 H 20 Cl2N2O5([M+H] + ), 475.0821; found, 475.0777; HPLC t R = 19.403 min (96.76% purity).

[0135] Compound 38, 1 1H-NMR (500 MHz, CDCl3), δ (ppm): 8.26 (d, 1H, J = 1.87 Hz), 8.01 (s, 2H), 7.69 - 7.70 (m, 1H), 7.45 - 7.49 (m, 1H), 7.31 - 7.37 (m, 3H), 4.93 (s, 1H), 4.07 - 4.16 (m, 4H), 2.38 (s, 3H), 1.20 (t, 6H, J = 7.06 Hz), 13 13C-NMR (125 MHz, CDCl3), δ (ppm): 168.1, 165.0 (2C), 147.4 (2C), 138.8 (2C), 139.8, 131.5 (2C), 131.3 (2C), 130.2, 127.4 (2C), 126.5 (2C), 113.8, 61.2 (2C), 36.1, 19.5, 14.1 (2C). HRMS (ESI+) m / z: calcd for C24 H 22 Cl2N2O5([M+H] + ), 489.0978; found, 489.0921; HPLC t R = 20.906 min (95.12% purity).

[0136] Compound 39, 1 1H-NMR (500 MHz, CDCl3), δ (ppm): 8.29 (d, 1H, J = 2.15 Hz), 8.15 (s, 2H), 7.73 (d, 1H, J = 2.15 Hz), 7.55 (d, 2H, J = 8.13 Hz), 7.35 (d, 2H, J = 8.13 Hz), 4.95 (s, 1H), 4.10 - 4.18 (m, 4H), 2.46 (s, 3H), 1.22 (t, 6H, J = 7.08 Hz), 13 13C-NMR (125 MHz, CDCl3), δ (ppm): 167.9, 165.1 (2C), 147.4, 144.1, 140.0, 138.8, 132.5 (2C), 129.8 (3C), 129.3 (3C), 128.2, 113.0 (2C), 61.2, 36.0, 21.7, 14.1 (2C). HRMS (ESI+) m / z: calcd for C 24 H 22 Cl2N2O5([M+H] + ), 489.0978; found, 489.0929; HPLC t R = 21.877 min (98.24% purity).

[0137] Compound 40, 1 1H-NMR (500 MHz, CDCl3), δ (ppm): 8.28 (d, 1H, J = 2.17 Hz), 8.15 (s, 2H), 7.72 (d, 1H, J = 2.17 Hz), 7.41 - 7.48 (m, 1H), 7.11 - 7.20 (m, 3H), 4.95 (s, 1H), 4.10 - 4.18 (m, 4H), 3.88 (s, 3H), 1.22 (t, 6H, J = 7.13 Hz), 13C-NMR (125 MHz, CDCl3), δ (ppm): 167.7, 165.0 (2C), 160.1, 147.4 (2C), 139.9, 138.8 (2C), 132.3 (2C), 130.1, 120.9, 119.0 (2C), 114.3 (2C), 113.3, 61.2 (2C), 55.6, 36.0, 14.1 (2C). HRMS (ESI+) m / z: calcd for C 24 H 22 Cl2N2O6 ([M+H] + ), 505.0927; found, 505.0870; HPLC t R = 18.930 min (98.53% purity).

[0138] Compound 41, 1 H-NMR (500 MHz, CDCl3), δ (ppm): 8.29 (d, 1H, J = 2.10 Hz), 8.15 (s, 2H), 7.74 (d, 1H, J = 2.09 Hz), 7.65 (d, 2H, J = 8.79 Hz), 7.03 (d, 2H, J = 8.79 Hz), 4.96 (s, 1H), 4.10 - 4.19 (m, 4H), 3.91 (s, 3H), 1.22 (t, 6H, J = 7.09 Hz), 13 C-NMR (125 MHz, CDCl3), δ (ppm): 167.4, 165.2 (2C), 163.6, 147.3, 140.2, 138.8, 132.8 (2C), 131.7 (3C), 130.2, 122.9, 114.5 (3C), 112.8, 61.2 (2C), 55.7, 36.0, 14.1 (2C). HRMS (ESI+) m / z: calcd for C 24 H 22 Cl2N2O6 ([M+H] + ), 505.0927; found, 505.0879; HPLC t R = 18.940 min (99.69% purity).

[0139] Compound 42, 11H-NMR (600 MHz, CDCl3), δ (ppm): 8.30 (d, 1H, J = 2.06 Hz), 8.17 (s, 2H), 7.74 (d, 1H, J = 2.06 Hz), 7.24 - 7.26 (m, 1H), 6.97 (d, 2H, J = 8.22 Hz), 4.97 (s, 1H), 4.11 - 4.19 (m, 4H), 3.98 (s, 3H), 3.95 (s, 3H), 1.23 (t, 6H, J = 7.09 Hz), 13 13C-NMR (150 MHz, CDCl3), δ (ppm): 167.4, 165.2 (2C), 153.3, 149.5, 147.5, 147.3 (2C), 140.2, 138.8, 132.8 (2C), 130.3, 123.3, 123.0, 112.8, 112.4, 110.5, 61.2 (2C), 56.2 (2C), 36.0, 14.2 (2C). HRMS (ESI+) m / z: calcd for C 25 H 24 Cl2N2O7 ([M + H] + ), 535.1033; found, 535.0975; HPLC t R = 17.440 min (96.17% purity). Compound 43, 1 1H-NMR (500 MHz, CDCl3), δ (ppm): 8.28 (d, 1H, J = 2.11 Hz), 8.09 (s, 2H), 7.84 (d, 2H, J = 8.22 Hz), 7.77 (d, 2H, J = 8.22 Hz), 7.72 (d, 2H, J = 2.11 Hz), 4.96 (s, 1H), 4.11 - 4.18 (m, 4H), 1.22 (t, 6H, J = 7.16 Hz), 13 13C-NMR (125 MHz, CDCl3), δ (ppm): 166.5, 164.8 (2C), 147.7, 147.4 (2C), 139.5 (2C), 138.8 (2C), 134.7, 131.5, 130.3, 129.4 (3C), 126.3, 126.2, 114.2, 61.4 (2C), 36.1, 14.1 (2C). HRMS (ESI+) m / z: calcd for C 24 H 19 Cl2F3N2O5 ([M + H] + ), 543.0695; found, 543.0642; HPLC t R = 21.626 min (98.11% purity).

[0140] Compound 44, 1 H-NMR (600MHz, CDCl3), δ (ppm): 8.30 (d, 1H, J = 2.20Hz), 8.12 (s, 2H), 7.74 (d, 1H, J = 2.20Hz), 7.67-7.72(m,2H),7.24-7.26(m,2H),4.97(s,1H),4.10-4.20(m,4H),1.22(t,6H,J=7.13Hz), 13 C-NMR (100MHz, CDCl3), δ (ppm): 166.7, 165.0 (2C), 147.6, 147.3 (2C), 139.8, 138.8 (2C), 132.1 (2C) ),131.9,131.8,130.2,116.6,116.4,113.5(2C),61.3(2C),36.0,14.1(2C).HRMS(ESI+)m / z:calcd for C 23 H 19 Cl2FN2O5([M+H] + ),493.0727; found,493.0677; HPLC t R =18.929min (96.07% purity).

[0141] Compound 45, 1 H-NMR (500MHz, CDCl3), δ (ppm): 8.28 (s, 1H), 8.10 (s, 2H), 7.72 (s, 1H), 7.60 (d, 2H, J = 8 .42Hz),7.54(d,2H,J=8.42Hz),4.96(s,1H),4.10-4.19(m,4H),1.22(t,6H,J=7.12Hz), 13 C-NMR (125MHz, CDCl3), δ (ppm): 166.8, 164.9 (2C), 147.6, 147.3, 139.7, 139.6, 138.8, 13 1.9(2C),130.5(3C),129.5(4C),113.7,61.3(2C),36.0,14.1(2C).HRMS(ESI+)m / z:calcd for C 23 H 19 Cl3N2O5([M+H] + ),509.0432; found,509.0376; HPLC t R =21.342min (98.26% purity). Compound 46,1 1H-NMR (600 MHz, CDCl3), δ (ppm): 8.28 (d, 1H, J = 2.17 Hz), 8.10 (s, 2H), 7.72 (d, 1H, J = 2.17 Hz), 7.71 (d, 2H, J = 8.56 Hz), 7.52 (d, 2H, J = 8.56 Hz), 4.96 (s, 1H), 4.11 - 4.19 (m, 4H), 1.22 (t, 6H, J = 7.15 Hz), 13 13C-NMR (150 MHz, CDCl3), δ (ppm): 166.9, 164.9 (2C), 147.6, 147.3, 139.7, 138.8, 132.5 (3C), 131.9, 130.6 (3C), 130.3, 129.9, 128.1, 113.7, 61.4 (2C), 36.0, 14.1 (2C). HRMS (ESI+) m / z: calcd for C 23 H 19 BrCl2N2O5 ([M + H] + ), 552.9926; found, 552.9901; HPLC t R = 22.202 min (98.28% purity).

[0142] Compound 47, 1 1H-NMR (500 MHz, CDCl3), δ (ppm): 8.29 (d, 1H, J = 2.11 Hz), 8.04 - 8.12 (m, 2H), 7.98 (d, 1H, J = 8.31 Hz), 7.85 (s, 2H), 7.72 (d, 1H, J = 2.11 Hz), 7.57 - 7.70 (m, 4H), 4.94 (s, 1H), 4.05 - 4.11 (m, 4H), 1.17 (t, 6H, J = 7.05 Hz), 13 13C-NMR (125 MHz, CDCl3), δ (ppm): 167.7, 164.9 (2C), 147.5, 147.4 (2C), 139.7 (2C), 138.8 (2C), 133.6, 132.3, 131.5, 130.2, 129.0, 128.3, 127.2 (3C), 126.7, 124.9, 124.1, 113.8, 61.2 (2C), 36.1, 14.0 (2C). HRMS (ESI+) m / z: calcd for C 27 H 22 Cl2N2O5 ([M + H] + ), 525.0978; found, 525.0920; HPLC tR = 22.831 min (99.04% purity). Compound 48, 1 1H-NMR (500 MHz, CDCl3), δ (ppm): 8.29 (d, 1H, J = 2.11 Hz), 8.07 - 8.09 (m, 1H), 7.98 (d, 1H, J = 8.31 Hz), 7.85 (s, 2H), 7.72 (d, 1H, J = 2.11 Hz), 7.54 - 7.74 (m, 4H), 4.94 (s, 1H), 4.09 - 4.17 (m, 4H), 1.17 (t, 6H, J = 7.05 Hz), 13 13C-NMR (125 MHz, CDCl3), δ (ppm): 167.9, 165.1 (2C), 147.6, 147.4, 140.0, 138.9 (2C), 135.2, 132.4 (2C), 130.7, 130.3, 129.2, 129.1, 128.9, 128.2, 128.0, 127.6, 124.5 (2C), 113.3, 61.2 (2C), 36.1, 14.1 (2C). HRMS (ESI+) m / z: calcd for C 27 27 22 27Cl2N2O5 ([M + H] + ), 525.097; found, 525.0927; HPLC t R = 23.130 min (96.28% purity). Compound 49, 1 1H-NMR (500 MHz, CDCl3), δ (ppm): 8.44 (s, 2H), 8.29 (d, 1H, J = 2.17 Hz), 7.73 (s, 1H), 7.70 (d, 1H, J = 2.17 Hz), 7.46 (d, 1H, J = 3.52 Hz), 6.68 - 6.69 (dd, 1H), 4.96 (s, 1H), 4.13 - 4.21 (m, 4H), 1.25 (t, 6H, J = 7.11 Hz), 13 13C-NMR (125 MHz, CDCl3), δ (ppm): 165.1 (2C), 155.8, 147.5, 147.3 (2C), 145.1, 139.7, 138.7 (2C), 132.0 (2C), 122.4 (2C), 113.7, 112.9, 61.3 (2C), 35.9, 14.1 (2C). HRMS (ESI+) m / z: calcd for C 21 27 18 27Cl2N2O6 ([M + H] +),465.0614; found,465.0566; HPLC t R =16.576min (98.27% purity).

[0143] Compound 50, 1 H-NMR (500MHz, CDCl3), δ (ppm): 8.31 (s, 2H), 8.29 (d, 1H, J = 2.10Hz), 7.79 (d, 1H, J = 4.90Hz), 7.73 (d, 1H, J = 2. 10Hz),7.65(d,1H,J=3.76Hz),7.23(t,1H,J=4.11Hz),4.98(s,1H),4.12-4.21(m,4H),1.24(t,6H,J=7.21Hz), 13 C-NMR (125MHz, CDCl3), δ (ppm): 165.1 (2C), 160.9, 147.6, 147.3, 139.8, 138.7, 134.4, 133.6 ,132.4(2C),130.3,128.2(2C),113.5(2C),61.3(2C),36.0,14.1(2C).HRMS(ESI+)m / z:calcd forC 21 H 18 Cl2N2O5S([M+H] + ),481.0385; found,481.0331; HPLC t R =17.981min (99.15% purity).

[0144] Test Example 1 SIRT3 Enzyme Activation Activity of Compounds 1 to 50

[0145] The purpose of this experiment is to detect the in vitro SIRT3 agonist activity of the compounds of the present invention (Table 1).

[0146] All enzymatic reactions were performed at 37°C for 30 minutes. The 50ul reaction mixture contained 25mM Tris, pH 8.0, 1mMMgCl2, 0.1mg / ml BSA, 1 37mM NaCl, 2.7mM KCl, SIRT3 and enzyme substrate. The compounds were diluted with 10% DMSO and 5ul of diluent was added to the 50ul reaction, making the final concentration of DMSO in all reactions 1%. The analysis was performed by quantifying the amount of fluorescent product of the enzyme reaction in solution. Fluorescence analysis was then performed using a SpectraMax M5 microtiter plate reader at excitation wavelengths of 350-360nm and emission wavelengths of 450-460nm. IC 50Values ​​were calculated using nonlinear regression and normalized dose-response fitting using Prism GraphPad software.

[0147] Table 1 agonistic activity of compounds 1-50 on SIRT3

[0148]

[0149]

[0150] Test Example 2 Compound 42 inhibits proliferation of MDA-MB-231 and BT-549 cells

[0151] BT-549 and MDA-MB-231 cells were digested with trypsin, placed in DMEM culture medium and centrifuged, the supernatant was discarded, culture medium was added and gently blown to disperse evenly, inoculated in a 6-well plate, and cultured in a 5% CO2, 37°C incubator. The cell growth was observed every day, and when the cells entered the logarithmic growth phase, the DMSO solution of compound 42 (experimental group) and DMSO (blank control group) were added and cultured. When obvious clones appeared, the culture medium was aspirated, washed twice with PBS, and 4% polymethanol was added to fix the cells. After 15 minutes, the upper methanol was gently aspirated, and then crystal violet was added to stain the cells. The staining solution was washed off for 30 minutes, and the number of cell clones was observed by taking pictures. The clone formation rate was calculated by counting the number of clones greater than 10 cells.

[0152] Next, we selected compound 42 with the best agonist activity for antiproliferative activity study. First, we tested the antiproliferative ability of compound 42 on triple-negative breast cancer cells. The experimental results showed that compound 42 had an antiproliferative effect on MDA-MB-231 cells (6hIC 50 =60.76±1.784μM; 12h IC 50 =11.16±1.047μM; 24h IC 50 =1.61±0.207μM; 36hIC 50 =0.94±0.270μM; 48h IC 50 =0.76±0.121 μM) and BT-549 cells (6h IC 50 =4.272±0.631μM; 12h IC 50 =1.81±0.259μM; 24h IC 50 =1.95±0.289μM; 36h IC 50 =1.47±0.168μM; 48hIC 50 =0.93±0.030μM) has good anti-proliferative activity. The experimental results show that BT-549 cells are more sensitive to compound 42 ( Figure 1 ).

[0153] After the prepared 2% low-soluble agarose solution was dissolved in a 60°C water bath, 50 μL / well was quickly added to the 96-well plate and allowed to cool and solidify at room temperature. Logarithmically growing BT-549 and MDA-MB-231 cells were digested with trypsin and the cell concentration was adjusted to 1×10 4 / mL, take 200μL of cell suspension and add it to a 96-well plate pre-embedded with 2% low-soluble agarose. Incubate in a 37℃ incubator containing 5% CO2. The next day, use a 100μL pipette to gently aspirate 100μL of old culture medium along the wall of the well, and then add 100μL of fresh 1640 culture medium containing FBS. After 3 days of culture, tumor cells were observed to aggregate into spheres with regular edges. After the 3D tumor sphere model was successfully established, 100μL of old culture medium was aspirated and discarded. After adding compound 42, the tumor sphere morphology at 0h, 24h, and 48h was observed and recorded under a microscope.

[0154] The 3D tumor sphere experiment can more accurately simulate the growth process of tumors in vivo. Therefore, we tested the effect of compound 42 on the formation of 3D tumor spheres of BT-549 and MDA-MB-231 cells. The experimental results showed that compound 42 inhibited the formation of 3D tumor spheres in a dose-dependent manner ( Figure 2 ). Therefore, it was further confirmed that compound 42 could significantly inhibit the proliferation of triple-negative breast cancer.

[0155] BT-549 and MDA-MB-231 cells in the logarithmic growth phase were inoculated in a 96-well plate and cultured until the cells adhered to the wall. 2 μM compound 42 was added for cell treatment, and EdU culture medium was prepared and added to the wells for incubation at 37°C for 2 hours. The culture medium was discarded, and the cells in the wells were washed with PBS on a shaker. Cell fixative was added and incubated at room temperature for 30 minutes, the fixative was discarded, glycine was added, and the cells were incubated on a shaker for 5 minutes. The glycine solution was discarded, and the cells in the wells were washed with PBS on a shaker, PBS was discarded, and the permeabilization agent was added and incubated on a shaker for 10 minutes, and PBS was washed; after adding Apollo staining reaction solution, it was incubated in the dark, at room temperature, and on a shaker for 30 minutes, the staining reaction solution was discarded, and the permeabilization agent was added and washed on a shaker for 30 minutes, and the permeabilization agent was discarded; DAPI reaction solution was prepared, and after incubation in the dark, at room temperature, and on a shaker for 30 minutes, the staining reaction solution was discarded and PBS was added for washing; observation was made under a fluorescence microscope, and 3 fields of view were randomly selected from each well for photography.

[0156] Then, we used EdU assay to detect the effect of compound 42 on DNA replication activity of BT-549 and MDA-MB-231 cells. Figure 5As shown in Figure 2, compound 42 can significantly reduce the green fluorescence intensity of EdU in BT-549 and MDA-MB-231 cells, indicating that compound 42 significantly inhibited the proliferation activity of TNBC cells ( Figure 3 ). In short, the experimental data prove that compound 42 can effectively inhibit the proliferation of TNBC cells.

[0157] Experimental Example 3 Compound 42 inhibits the migration of BT-549 and MDA-MB-231 cells

[0158] BT-549 and MDA-MB-231 cells were digested with trypsin, placed in DMEM culture medium and centrifuged, the supernatant was discarded, the culture medium was added and gently blown to make it evenly dispersed, and inoculated in a 6-well plate, divided into a blank control group and an experimental group (0μM, 1μM, 2μM, 4μM 42), and cultured in a 5% CO2, 37°C incubator. Observe the cell condition, when the cells enter the logarithmic phase, use a pipette to draw a cross in the well, aspirate the culture medium, wash with PBS 3 times, observe under a microscope and take pictures, and record the width of the scratch at 0h. Add serum-free culture medium, observe and take pictures of the cell scratch healing at 24h and 48h.

[0159] TNBC is a malignant tumor with the highest incidence in women. Cancer cells proliferate rapidly and tend to grow to other parts, leading to recurrence and metastasis. Therefore, the next step was to test whether compound 42 could inhibit the migration of BT-549 and MDA-MB-231 cells. First, BT-549 and MDA-MB-231 cells were treated with compound 42 at different concentrations (0μM, 1μM, 2μM, 4μM) for 24h and 48h, and then cell scratches were detected to verify whether compound 42 would inhibit the migration of tumor cells. The results of the cell scratch experiment showed that compound 42 inhibited the migration of BT-549 and MDA-MB-231 cells in a dose-dependent manner ( Figure 4 , 5 ).

[0160] Experimental Example 4 Compound 42 inhibits glycolysis in BT-549 and MDA-MB-231 cells by activating SIRT3

[0161] Cellular glucose uptake was determined by flow cytometric analysis of 2-NBDG uptake. BT-549 and MDA-MB-231 cells were seeded in 96-well plates with black transparent bottoms and cultured for 24 h. After the original culture medium was discarded, growth medium containing 1% FBS was added for starvation treatment for 10 h. The starvation culture medium was discarded and culture medium containing 0 μM, 1 μM, 2 μM, and 4 μM compound 42 was added. 10 min before the end of the intervention, the intervention culture medium was discarded, 2-NBDG was added to the sugar-free culture medium, and 100 μL was added to each well of the culture plate. The culture plate was shaken to mix thoroughly, and then incubated for another 30 min at 37°C in a humidified atmosphere containing 5% CO2 and 95% air. At the end of the culture, the samples were analyzed on the EasyCyte Plus flow cytometer system (Millipore, Chicago, IL, USA).

[0162] To measure lactate production, BT-549 and MDA-MB-231 cells were seeded in 24-well plates for 24 h in triplicate and then refreshed with 1 mM glucose medium overnight. The medium was harvested and lactate concentration was measured by lactate test strips and a lactate dosing meter (Nova Biomedical). Lactate production was normalized by cellular protein mass.

[0163] In order to verify whether compound 42 inhibits the glycolysis effect of TNBC cells by activating SIRT3, we detected the glucose uptake capacity and lactate production of BT-549 and MDA-MB-231 cells after treatment with different concentrations of compound 42 for 24 hours (0μM, 1μM, 2μM, 4μM). Glucose uptake capacity and lactate production are key characteristics of glycolysis effect. The experimental results showed that compound 42 inhibited the glucose uptake capacity of BT-549 and MDA-MB-231 cells in a dose-dependent manner ( Figure 6 A, C) and lactic acid production ( Figure 6 B, D). In short, compound 42 can inhibit the glycolysis of TNBC cells by activating SIRT3.

[0164] Experimental Example 5 Compound 42 induces mitochondrial metabolic abnormalities and apoptosis in BT-549 and MDA-MB-231 cells by activating SIRT3

[0165] Prepare the JC-1 working solution according to the instructions of the kit. Take BT-549 and MDA-MB-231 cells in the logarithmic growth phase and inoculate them in a 6-well plate, and culture them in a 5% CO2, 37°C incubator. When the cells are attached and 80% fused, replace the culture medium containing (0μM, 1μM, 2μM, 4μM) compound 42. There are 3 replicates in each group, and they are placed in the cell culture incubator again and incubated at a constant temperature for 48 hours. Wash the cells 3 times with PBS, digest with trypsin, terminate the digestion with complete culture medium, and collect the cells in a 1.5mL centrifuge tube. Centrifuge at 4℃1000rpm / min for 5min, remove the supernatant, add 500μL of JC-1 solution to resuspend, and incubate in a cell culture incubator for 15min. Centrifuge again, remove the supernatant containing JC-1, and wash twice with 1×IncubationBuffer. Finally, add 500 μL of 1× Incubation Buffer, gently suspend the cells, and detect on a flow cytometer.

[0166] BT-549 and MDA-MB-231 cells in the logarithmic growth phase were inoculated in a 6-well plate and cultured in a 5% CO2, 37°C incubator. When the cells adhered to the wall and grew, the culture medium containing different concentrations of compound 42 was replaced, which were: 0μM, 1μM, 2μM, and 4μM. Three replicates were added in each group and incubated at a constant temperature for 48 hours. The culture medium was removed, washed with PBS three times, digested with trypsin, and digested with complete culture medium. The cells were collected in a centrifuge tube, centrifuged and discarded the supernatant, and the extract was added according to the instructions and mixed thoroughly. The cells were ultrasonically disrupted for 1min, and centrifuged at 4°C 10000g for 10min. The supernatant was placed in a new EP tube, 500μL chloroform was added, mixed thoroughly, centrifuged again for 3min, and the supernatant was extracted in the EP tube and placed on ice for later use. The ATP standard was diluted according to the instructions and the working solution was prepared, and the ATP content was measured in an ELISA instrument.

[0167] Since tumor cell mitochondrial proteins are highly acetylated, we then tested the effect of compound 42 on mitochondria in BT-549 and MDA-MB-231 cells. First, BT-549 and MDA-MB-231 cells were treated with compound 42 at different concentrations (0 μM, 1 μM, 2 μM, 4 μM) for 24 hours, and then the cell membrane potential was detected by flow cytometry (JC-1). The results of the JC-1 experiment showed that compound 42 induced a decrease in the mitochondrial membrane potential of BT-549 and MDA-MB-231 cells in a dose-dependent manner ( Figure 7 A, C). At the same time, we also further detected the ATP content in the cells. The experimental results showed that compound 42 significantly reduced the ATP content in BT-549 and MDA-MB-231 cells ( Figure 7 B, D).

[0168] Since apoptosis is the most classic and widely studied mode of cell death, we then tested whether compound 42 could induce apoptosis in BT-549 and MDA-MB-231 cells. After 24 hours of exposure to compound 42 at different concentrations (0 μM, 1 μM, 2 μM, 4 μM) on BT-549 and MDA-MB-231 cells, flow cytometry was used to perform Annexin V-PI staining. The results of the Annexin V-PI staining experiment showed that compound 42 induced apoptosis in BT-549 and MDA-MB-231 cells in a dose-dependent manner ( Figure 8 ).

[0169] Experimental Example 6 Study on the in vivo anti-TNBC activity of compound 42

[0170] BALB / c mice (female, 3-4 weeks old, 19±1g) were provided by Sbefor (Beijing) Biotechnology Co., Ltd. (certificate number: SCXK[Jing]2019-0010). Based on the in vitro cytotoxicity assay and the results of the preliminary experiment, the following treatment plan was determined: Under standard experimental conditions, after the nude mice had adapted to the new environment and were fed normally for one week, 12 female BALB / c mice were subcutaneously inoculated with 5×106 BT-549 cells. When the tumor volume reached 0.2cm 3 At the time of treatment, we randomly divided the mice into two groups: control group (n=6) and treatment group (n=6, 20 mg / kg 42 intraperitoneal injection, daily). During the whole experiment, the tumor volume and body weight of the mice were recorded daily. After 7 days of treatment, the mice were killed by cervical dislocation, and the tumor tissues were taken and the tumor volume and weight were recorded, and then fixed in 4% paraformaldehyde for preservation. The heart, liver, spleen, lung and kidney were taken and the weight data were recorded.

[0171] The antitumor activity of compound 42 in nude mice with BT-549 cell xenografts was evaluated from the aspects of tumor growth rate, tumor volume and tumor weight. The experimental results showed that the tumor proliferation rate and volume indicated that compound 42 could effectively slow down the proliferation of BT-549 tumor cells ( Fig. 9 AD). Next, the effect of compound 42 on the proliferation marker Ki-67 in tumor tissue was evaluated by immunohistochemistry. The experimental results showed that the expression of the proliferation marker Ki-67 was reduced after treatment with compound 42, and there was a statistically significant difference ( Fig. 9 EF). The daily body weight and organ ratio of the animals showed that compound 42 had no significant toxic effect on BT-549 cell xenografted nude mice ( Fig.10 ).

Claims

1. The compound represented by formula I or its pharmaceutically acceptable salt or isomer: in, Ar is a 6-10 membered aryl or heteroaryl group, wherein the heteroaryl group contains 1-3 heteroatoms of N, O or S; Ar is substituted by one or more of the following substituents: H, cyano, C1-C6 alkoxy, C1-C6 alkyl, halogen, C1-C6 ester, halogenated C1-C6 alkyl, phenyl, benzyloxy, methylsulfone; R is substituted or unsubstituted phenyl, naphthyl, thienyl, furyl; The substituent is C1-C6 alkoxy, C1-C6 alkyl, halogen, or halogenated C1-C6 alkyl.

2. The compound of formula I according to claim 1 or its pharmaceutically acceptable salt or isomer: in, Ar is naphthyl, 2,3-methylenedioxy-phenyl, pyridyl, or phenyl; The Ar is substituted by one or more of the following substituents: H, cyano, C1-C6 alkoxy, C1-C6 alkyl, halogen, halogenated C1-C4 alkyl, C1-C6 ester, phenyl, benzyloxy, methylsulfone.

3. The compound represented by formula I of claim 1 or 2 or its pharmaceutically acceptable salt or isomer: in, Ar is naphthyl, 2,3-methylenedioxy-phenyl, 4,5-dichloropyridyl, X1 is hydrogen, phenyl, C1-C6 alkyl, C1-C4 alkoxy, F, Cl, Br, I, benzyloxy, CF3, methylsulfonyl, cyano, or methoxycarbonyl.

4. The compound of formula I according to any one of claims 1 to 3 or its pharmaceutically acceptable salt or isomer: in, R is Naphthyl, thienyl, furanyl; X2 is hydrogen, C1-C4 alkoxy, C1-C4 alkyl, halogenated C1-C4 alkyl, or halogen.

5. The following compounds or their pharmaceutically acceptable salts or isomers:

6. A method for preparing the compound of formula I or its pharmaceutically acceptable salt or isomer according to claim 1: in, Ar and R are as described in claim 1.

7. A pharmaceutical composition comprising a compound of formula I according to any one of claims 1 to 5 or a pharmaceutically acceptable salt, isomer thereof and a pharmaceutically acceptable carrier or excipient.

8. Use of the compound of formula I according to any one of claims 1 to 5 or its pharmaceutically acceptable salt, isomer or the pharmaceutical composition according to claim 7 in the preparation of a SIRT3 agonist.

9. Use of the compound of formula I according to any one of claims 1 to 5 or its pharmaceutically acceptable salt, isomer or the pharmaceutical composition according to claim 7 in the preparation of anti-tumor drugs.

10. The use according to claim 9, characterized in that: The tumor is breast cancer, preferably triple-negative breast cancer.

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