A nitric oxide donor type Omidenepag derivative and its preparation method and application

Abstract

The present invention discloses a nitric oxide donor omidate (Omidenepag) derivative with structural formula I, its preparation method, and application. These derivatives, representing different classes of NO donors, can release a certain amount of NO in aqueous humor in vitro while retaining strong EP2 receptor agonist activity. These two synergistically reduce intraocular pressure in New Zealand white rabbits without causing conjunctival irritation. The present invention relates to such NO donor Omidenepag derivatives, pharmaceutical compositions containing the target compounds, and their medical uses. These derivatives have promising applications, particularly in the prevention and treatment of ocular hypertension and glaucoma-related diseases. #imgabs0#

Description

Technical Field

[0001] The present invention relates to a compound, in particular to a nitric oxide donor type Omidenepag derivative and a preparation method and application thereof. Background Art

[0002] Glaucoma is a neurodegenerative disease. As the disease progresses, patients with glaucoma will experience apoptosis of retinal ganglion cells (RGCs), thinning of the retinal nerve fiber layer, and enlarged optic disc pits, ultimately leading to visual impairment and irreversible blindness (Clin. Exp. Ophthalmol., 2012, 40(4):341-349). Pathologically elevated intraocular pressure (IOP) is the main risk factor for the disease. Currently, the treatment for glaucoma mainly uses eye pressure-lowering drugs or laser or surgical blood pressure reduction to control and delay the progression of the disease (JAMA, J. Am. Med. Assoc., 2014, 311(18):1901-1911). According to statistics, there were 76.5 million glaucoma patients worldwide in 2020. With the aging population, this number is expected to increase to 110 million by 2040 (Prog. Retinal Eye Res., 2021, 83:100916). In 2020, there were 21 million glaucoma patients in my country, and 5.67 million people were blinded by the disease (Arch. Ophthalmol., 2002, 120(6):701-713). Glaucoma patients are mainly middle-aged and elderly people, but studies have shown that glaucoma is becoming younger (Drugs Aging, 2005, 22(1):1-21). Therefore, the increasing number of patients and the trend of younger patients are enough to attract people's attention to this disease.

[0003] The prostaglandin E2 receptor subtype EP2 is a G protein-coupled receptor that has been shown to be widely expressed in ocular tissues. It can exit the eye through the conventional outflow pathway, namely, the anterior ciliary veins that drain into the sclera through the trabecular meshwork (TM) and Schlemm's canal (SC), and the uveoscleral pathway, namely, entering the ciliary body and suprachoroidal space through the ciliary muscle bundle space and exiting the eye through scleral collagen or neurovascular spaces, thereby lowering IOP without causing serious side effects.

[0004] Omidenepag isopropyl (OMDI) is a selective prostaglandin E2 receptor 2 (EP2) agonist with a non-prostaglandin structure, developed by Ube Industries and Santen Pharmaceutical as an ophthalmic solution for the treatment of glaucoma and ocular hypertension. OMDI is an isopropyl ester derivative that hydrolyzes into its active metabolite, Omidenepag (OMD), during corneal penetration. It increases aqueous humor outflow through the uveoscleral and trabecular outflow pathways, thereby lowering IOP. In September 2018, OMDI eye drops 0.002% were approved in Japan for the treatment of glaucoma and ocular hypertension. On September 26, 2022, Santen and UBE jointly announced that the U.S. Food and Drug Administration (FDA) approved Omlonti 0.002% eye drops for lowering IOP in patients with primary open-angle glaucoma or ocular hypertension.

[0005] Molecular formula of free base: C 26 H 28 N6O4S, molecular weight: 520.61. Studies have shown that in in vitro studies using human recombinant prostaglandin receptors, OMD was shown to be able to selectively bind to the EP2 receptor (Ki = 3.6nM) and exhibit high agonist activity on the receptor (EC 50 =1.1 nM) (J Med Chem. 2018, 61(15): 6869-6891). OMDI is converted to the active form of OMD during corneal penetration, and OMD increases the intracellular level of cAMP, which is a second messenger (AORN J. 2019, 109(2): 256-261).

[0006] If OMDI's ability to lower patients' IOP can be further improved, it can be better used to prevent and treat high intraocular pressure and glaucoma-related diseases. Summary of the Invention

[0007] Objectives of the invention: The first objective of the present invention is to provide a nitric oxide donor-type Omidenepag derivative that improves the ability of OMDI to lower the patient's IOP, thereby improving the intraocular pressure-lowering activity and neuroprotective effect; the second objective of the present invention is to provide a method for preparing the nitric oxide donor-type Omidenepag derivative; the third objective of the present invention is the application of the nitric oxide donor-type Omidenepag derivative.

[0008] The nitric oxide donor Omidenepag derivative of the present invention has the structural formula:

[0009]

[0010] Wherein, R1 is selected from Or -H; R2 is selected from methylene, ethylene, propylene, butylene, pentylene, or isosorbide R3 is selected from or -ONO2.

[0011] Preferably, R1 is H.

[0012] Preferably, R1 is H, R2 is selected from methylene, ethylene, propylene, butylene or pentylene, and R3 is selected from

[0013] Preferably, the compound I is:

[0014] LO7: 4-(2-(((6-((N-(4-(1H-pyrazol-1-yl)benzyl)pyridine-3-sulfonamido)methyl)pyridin-2-yl)glycylamino)oxy)ethoxy)-3-(phenylsulfonyl)-1,2,5-oxadiazole 2-oxide, with the following structural formula:

[0015]

[0016] LO8: 4-(3-(((6-((N-(4-(1H-pyrazol-1-yl)benzyl)pyridine-3-sulfonamido)methyl)pyridin-2-yl)glycylamino)oxy)propoxy)-3-(phenylsulfonyl)-1,2,5-oxadiazole 2-oxide, with the following structural formula:

[0017]

[0018] LO9: 4-(4-(((6-(N-(4-(1H-pyrazol-1-yl)benzyl)pyridine-3-sulfonamido)methyl)pyridin-2-yl)glycylamino)oxy)butoxy)-3-(phenylsulfonyl)-1,2,5-oxadiazole 2-oxide, with the following structural formula:

[0019]

[0020] LO10: 4-(3-(((6-(N-(4-(1H-pyrazol-1-yl)benzyl)pyridine-3-sulfonamido)methyl)pyridin-2-yl)glycylamino)oxy)propoxy)-3-methyl-1,2,5-oxadiazole 2-oxide, with the following structural formula:

[0021]

[0022] LO11: 4-(4-(((6-(N-(4-(1H-pyrazol-1-yl)benzyl)pyridine-3-sulfonamido)methyl)pyridin-2-yl)glycylamino)oxy)butoxy)-3-methyl-1,2,5-oxadiazole 2-oxide, with the following structural formula:

[0023]

[0024] LO12: 4-((5-(((6-(N-(4-(1H-pyrazol-1-yl)benzyl)pyridine-3-sulfonamido)methyl)pyridin-2-yl)glycylamino)oxy)pentyl)oxy)-3-methyl-1,2,5-oxadiazole 2-oxide, with the following structural formula:

[0025]

[0026] LO13: 4-(3-(((6-(N-(4-(1H-pyrazol-1-yl)benzyl)pyridine-3-sulfonamido)methyl)pyridin-2-yl)glycylamino)oxy)propyloxy)-3-phenyl-1,2,5-oxadiazole 2-oxide, with the following structural formula:

[0027]

[0028] LO14: 2-(Nitro)ethyl (6-((N4-(1H-pyrazol-1-yl)phenyl)pyridine-3-sulfonylamino)methyl)pyridin-2-yl)glycine, with the following structural formula:

[0029]

[0030] LO15: 3-(Nitro)propyl(6-((N4-(1H-pyrazol-1-yl)phenyl)pyridine-3-sulfonylamino)methyl)pyridin-2-yl)glycine, with the following structural formula:

[0031]

[0032] LO16: (3-Furan-3-yl-3-aryl-6R-pyridine-3-sulfonamide-6As)-6-(nitrooxy)hexahydrofuran[3je2-]furan-3-yl(6-((N4-(1H-pyrazol-1-yl)phenyl)pyridine-3-sulfonamido)pyridin-2-yl)pyridin-2-yl)glycine, structural formula is as follows:

[0033]

[0034] LO17: (E)-10-(6-((N-(4-(1H-pyrazol-1-yl)benzyl)pyridine-3-sulfonamido)methyl)pyridin-2-yl)-13-(9H-fluoren-9-yl)-2-methyl-8,11-dioxo-5,7,12-trioxa-2,3,4,10-tetraazatrideca-3-ene-3-oxide, structural formula is as follows:

[0035]

[0036] LO18: (E)-4-(6-((N-(4-(1H-pyrazol-1-yl)benzyl)pyridine-3-sulfonamido)methyl)pyridin-2-yl)-12-ethyl-1-(9H-fluoren-9-yl)-3,6-dioxo-2,7,9-trioxa-4,10,11,12-tetraazatetradec-10-ene-11-oxide, structural formula is as follows:

[0037]

[0038] LO19: (E)-2-(((N-(6-(N-(4-(1H-pyrazol-1-yl)benzyl)pyridine-3-sulfonamido)methyl)pyridin-2-yl)-N-(((9H-fluoren-9-yl)methoxy)carbonyl)glycylamino)oxy)formyl)-1-morpholinodiazene-1-oxide, structural formula is as follows:

[0039]

[0040] LO20: (E)-2-(((N-(6-(N-(4-(1H-pyrazol-1-yl)benzyl)pyridine-3-sulfonamido)methyl)pyridin-2-yl)-N-(((9H-fluoren-9-yl)methoxy)carbonyl)glycylamino)oxy)formyl)-1-(4-hydroxypiperidin-1-yl)diazene-1-oxide, structural formula is as follows:

[0041]

[0042] More preferably, the structural formula of the compound I is:

[0043]

[0044] The preparation method of the nitric oxide donor type Omidenepag derivative comprises the following steps:

[0045] (1) Compound III Omidenepag and 9-fluorenylmethyl chloroformate undergo amide condensation reaction to obtain intermediate IV;

[0046] (2) Intermediate IV undergoes ester condensation with compound R3-R2-OH or nucleophilic substitution with compound R3-R2-Cl to obtain compound II;

[0047] (3) Compound II is deprotected from the 9-fluorenylmethoxycarbonyl protecting group to obtain the target compound I;

[0048] The synthetic route is as follows:

[0049]

[0050] Wherein, R1 is selected from Or -H; R2 is selected from methylene, ethylene, propylene, butylene, pentylene, or isosorbide R3 is selected from Or -ONO2; when R1 is When , step (3) is not performed.

[0051] Preferably, in step (1), the reaction temperature is -20 to 25°C, and the reaction solvent is one or more of anhydrous dichloromethane, chloroform, ethyl acetate, redistilled acetone, anhydrous tetrahydrofuran, anhydrous N,N-dimethylformamide, dimethyl sulfoxide or dioxane. The reaction in step (1) is carried out under alkaline conditions, and the base is selected from potassium carbonate, sodium carbonate, sodium bicarbonate, sodium hydroxide, potassium hydroxide, pyridine, 4-methylaminopyridine, triethylamine, N,N-diisopropylmethylamine, N-methylmorpholine, piperidine

[0052] Preferably, in step (2), the reaction temperature is -20 to 25° C., and the solvent is selected from one or more of anhydrous acetonitrile, anhydrous dichloromethane, chloroform, ethyl acetate, redistilled acetone, anhydrous tetrahydrofuran, anhydrous N,N-dimethylformamide, dimethyl sulfoxide or dioxane.

[0053] Preferably, in step (3), the reaction temperature is -20 to 25° C., and the solvent is selected from one or more of anhydrous piperidine, anhydrous acetonitrile, anhydrous dichloromethane, chloroform, ethyl acetate, redistilled acetone, anhydrous tetrahydrofuran, anhydrous N,N-dimethylformamide, dimethyl sulfoxide or dioxane.

[0054] The synthesis steps of compound III are as follows:

[0055]

[0056] The invention relates to an application of the nitric oxide donor type Omidenepag derivative in the preparation of a drug for preventing and / or treating diseases related to increased intraocular pressure.

[0057] The diseases associated with increased intraocular pressure include ocular hypertension, glaucoma and diabetic eye complications.

[0058] Invention Mechanism: The nitric oxide donor-type omidate derivative of the present invention can produce the active substance omidate (OMD) and release NO. OMD can selectively bind to EP2 receptors. NO has the activity to treat glaucoma. On the one hand, it can relax TM and promote the outflow of aqueous humor through the traditional pathway, and it can also inhibit Na + ,K +OMD inhibits the production of ATPase, reducing aqueous humor production and ultimately lowering IOP. On the other hand, NO can regulate ocular vascular tone and maintain ocular blood flow in the eye. Combining OMD with an NO donor has the dual effects of lowering IOP and protecting retinal ganglion cells, further enhancing the effectiveness of OMD in preventing and treating ocular hypertension and glaucoma-related diseases.

[0059] Beneficial effects: Compared with the prior art, the present invention has the following advantages: the present invention designs and synthesizes NO-donating Omidenepag derivatives, which are proven to stimulate EP2 receptors and release appropriate amounts of NO, producing the dual effects of lowering IOP and protecting retinal ganglion cells, and is non-irritating to the conjunctiva of New Zealand white rabbits; pharmaceutical combinations containing the target compounds and their medical uses have good application prospects, especially in the prevention and treatment of high intraocular pressure and glaucoma-related diseases. BRIEF DESCRIPTION OF THE DRAWINGS

[0060] Figure 1 This is the NO release graph of the compound in the rabbit aqueous humor in vitro;

[0061] Figure 2 This is the HPLC detection chart of the compound metabolized to Omidenepag;

[0062] Figure 3 This is a graph showing the stability of the compound at room temperature;

[0063] Figure 4 is the agonist activity of the compound on EP2 receptor;

[0064] Figure 5 The graph shows the intraocular pressure-lowering activity of the compound in normal intraocular pressure rabbits. DETAILED DESCRIPTION

[0065] The present invention will be further described below with reference to specific embodiments.

[0066] Example 1

[0067] The synthesis of the neokotalanol derivative LO7 of the present invention comprises the following steps:

[0068]

[0069] (1) Synthesis of intermediate III

[0070]

[0071] The synthesis of intermediate LO1-2 was as follows: Compound LO1-1 (2 g, 9.62 mmol, 1.0 equiv) was dissolved in anhydrous DMF and precooled at -20°C for 15 minutes. Then, sodium hydride (461 mg, 9.62 mmol, 1.2 equiv) was added and stirred for 15 minutes. Tert-butyl bromoacetate (2.25 g, 9.62 mmol, 1.2 equiv) was added, and the mixture was brought to room temperature under nitrogen atmosphere and stirred for 2 hours. The reaction was monitored by TLC (PE:EA = 10:1). After complete reaction, the mixture was extracted with water and ethyl acetate, washed with saturated brine, dried over anhydrous sodium sulfate, concentrated, and purified by column chromatography (PE:EA = 10:1) to yield approximately 1.41 g of LO1-2 as a colorless oil.

[0072] Intermediate LO1-3 was synthesized by dissolving compound LO1-2 (1.41 g, 4.37 mmol, 1.0 equiv) in anhydrous acetonitrile, adding N-bromosuccinimide (818 mg, 4.59 mmol, 1.05 equiv), and stirring at 40°C for 3 hours under a nitrogen atmosphere. The reaction was monitored by TLC (PE:EA = 20:1). After the reaction was complete, the product was extracted with water and ethyl acetate, washed with saturated brine, dried over anhydrous sodium sulfate, concentrated, and purified by column chromatography (PE:EA = 50:1) to yield approximately 1.21 g of LO1-3 as a white solid powder.

[0073] Intermediate LO1 was synthesized by dissolving compound LO1-3 (1.21 g, 3.03 mmol, 1.0 equiv) in anhydrous 1,2-dichloroethane, adding N-bromosuccinimide (565 mg, 3.18 mmol, 1.05 equiv) and azobisisobutyronitrile (50 mg, 0.3 mmol, 0.1 equiv). The reaction was incubated at 90°C under nitrogen for 2 hours. The reaction was monitored by TLC (PE:EA = 20:1). After complete reaction, the product was extracted with water and ethyl acetate, washed with saturated brine, dried over anhydrous sodium sulfate, concentrated, and purified by column chromatography (PE:EA = 50:1) to yield approximately 1.03 g of LO1 as a white solid powder.

[0074] The synthesis of intermediate LO2-2 is as follows: Compound LO2-1 (5 g, 29.58 mmol, 1.0 equiv) was dissolved in anhydrous tetrahydrofuran. Lithium aluminum hydroxide (1 M solution in THF) (35.5 mmol, 1.2 equiv) was added to a dropping funnel, the atmosphere was replaced with nitrogen, and the mixture was pre-cooled to 0°C for 15 minutes before being added dropwise. The mixture was brought to room temperature and stirred for 3 hours. The reaction was monitored by TLC (PE:EA = 1:2). After the reaction was complete, 15% aqueous sodium hydroxide solution was added at 0°C. The mixture was extracted with water and dichloromethane, washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated to a white solid powder. The mixture was then slurried with dichloromethane / n-hexane to remove impurities, yielding approximately 2.36 g of LO2-2 as a white solid powder.

[0075] Intermediate LO2 was synthesized by dissolving compound LO2-2 (2.36 g, 13.64 mmol, 1.0 equiv) in anhydrous dichloromethane, adding triethylamine (2.76 g, 27.28 mmol, 2.0 equiv), and then adding 3-pyridinesulfonyl chloride (2.41 g, 13.64 mmol, 1.0 equiv) at 0°C. The mixture was stirred for 30 minutes, then brought to room temperature and reacted overnight under a nitrogen atmosphere. The reaction was monitored by TLC (PE:EA = 1:2). After complete reaction, the solvent was evaporated and purified by column chromatography (PE:EA = 1:2) to yield approximately 1.98 g of LO2 as a white solid powder.

[0076] Intermediate LO3 was synthesized by dissolving compound LO2 (1.98 g, 6.31 mmol, 1.0 equiv) in anhydrous DMF, adding LO1 (3.01 g, 6.31 mmol, 1.0 equiv) and anhydrous potassium carbonate (870.2 mg, 6.31 mmol, 1.0 equiv), and stirring at room temperature for 3.5 hours. The reaction was monitored by TLC (PE:EA = 3:1). After the starting materials reacted completely, a large amount of water was added to the reaction solution, which was then extracted with ethyl acetate, washed with saturated brine, dried over anhydrous sodium sulfate, concentrated, and purified by column chromatography (PE:EA = 5:1) to yield approximately 2.86 g of LO3 as a pale yellow solid powder.

[0077] Intermediate LO4 was synthesized by dissolving compound LO3 (2.86 g, 4.02 mmol, 1.0 equiv) in ethanol, adding palladium on carbon (286 mg, 0.42 mmol, 0.1 equiv) and triethylamine (812 mg, 8.04 mmol, 2.0 equiv). The atmosphere was then replaced with nitrogen and then hydrogen, and the reaction was allowed to proceed at room temperature for 4 hours. The reaction was monitored by TLC (PE:EA = 1:1). After complete reaction, the starting material was filtered through celite, concentrated, and purified by column chromatography (PE:EA = 1:1) to yield approximately 2 g of LO4 as a pale yellow foamy solid.

[0078] The synthesis steps of intermediate III are as follows: compound LO4 (1.98 g, 0.39 mmol, 1.0 equiv) is dissolved in anhydrous dichloromethane, hydrogen chloride / 1,4-dioxane solution (4 M) is added, and the reaction is carried out at room temperature for 14 hours. The reaction is monitored by TLC (PE:EA=1:1). After the raw materials are completely reacted, the solvent is dried and a large amount of dichloromethane is added. At this time, a white solid powder is precipitated. After ultrasonication, the insoluble matter is filtered out, and the filter cake is washed with dichloromethane several times and infrared drying is carried out at 60°C to obtain about 1.48 g of LO5.

[0079] (2) Synthesis of Intermediate IV

[0080] Compound III (200 mg, 0.39 mmol, 1.0 equiv) was dissolved in 10 mL of anhydrous dichloromethane. The reaction flask was placed at 0°C and N-methylmorpholine (129 μL, 1.95 mmol, 3.0 equiv) was slowly added dropwise. The mixture was stirred vigorously at this temperature for 10 minutes. Once the compound was completely dissolved, 9-fluorenylmethyl chloroformate (102 mg, 0.39 mmol, 3.0 equiv) was added in small portions. The solution turned from light yellow to dark green. Under nitrogen atmosphere, the mixture was stirred at room temperature for 2 hours. The reaction was monitored by TLC (DCM / MeOH = 25:1). After the starting material reacted completely, the dichloromethane was removed by rotary evaporation. The product was purified by column chromatography (DCM / MeOH = 25:1) and concentrated to afford Intermediate IV.

[0081] (3) Synthesis of Intermediate II

[0082] Intermediate IV (100 mg, 0.143 mmol, 1.0 equiv) was dissolved in 20 mL of anhydrous dichloromethane. The reaction flask was placed at 0°C, and 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (33 mg, 0.172 mmol, 1.2 equiv) and 1-hydroxybenzotriazole (24 mg, 0.172 mmol, 1.2 equiv) were added. After stirring at this temperature for 1 hour, 2-[[5-oxyl-4-(phenylsulfonyl)-1,2,5-oxadiazol-3-yl]oxy]ethanol (50 mg, 0.172 mmol, 1.2 equiv) was added. The mixture was stirred at room temperature overnight under nitrogen. The reaction was monitored by TLC (DCM / MeOH = 25:1). After complete reaction of the starting material, the dichloromethane was removed by rotary evaporation, and the mixture was purified by column chromatography (DCM / MeOH = 25:1) and concentrated to afford Intermediate II.

[0083] (4) Synthesis of LO7

[0084] Intermediate II (50 mg, 0.052 mmol, 1.0 equiv) was dissolved in 1 mL of dichloromethane, and 0.5 mL of piperidine was added. The mixture was allowed to react at room temperature under nitrogen for 2 hours. The reaction was monitored by TLC (DCM / MeOH = 25:1). After complete reaction, the dichloromethane was removed by rotary evaporation, and the product was purified by column chromatography (DCM / MeOH = 25:1). The product was concentrated to afford 35 mg (87%) of a pale yellow solid.

[0085] LO7 1 H NMR (400MHz, CDCl3) δ9.01 (s, 1H), 8.74 (d, J = 8.6Hz, 1H), 8.08 (m, 2H),

[0086] 7.97(d,J=12.6Hz,,1H),7.93(d,J=4.6Hz,1H),7.76(m,2H),7.62(m,4H),7.38(m,4H) ,6.49(m,2H),6.33(d,J=11.8Hz,1H),4.67(m,6H),4.36(s,2H),4.02(d,J=7.6Hz,2H). 13 CNMR (75MHz, CDCl3) δ170.99,158.60,152.73,148.08,141.22,139.82,137.81,135.77,134.71,133.71,129.92,129.7 3,128.63,126.79,123.41,119.23,112.90,107.79,107.68,68.75,61.61,51.23,51.11,43.34.MS(ESI)m / z:747[M+H] + .HRMS(ESI)m / z calcdfor C 33 H 30 N8O9S2,747.16554; found,747.16335[M+H] + .

[0087] Example 2

[0088] The synthesis of the neokotalanol derivative LO8 of the present invention comprises the following steps:

[0089]

[0090] (1) Synthesis of Intermediate IV is the same as in Example 1

[0091] (2) Synthesis of Intermediate II

[0092] Intermediate IV (100 mg, 0.143 mmol, 1.0 equiv) was dissolved in 20 mL of anhydrous dichloromethane. The reaction flask was placed at 0°C, and 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (33 mg, 0.172 mmol, 1.2 equiv) and 1-hydroxybenzotriazole (24 mg, 0.172 mmol, 1.2 equiv) were added. After stirring at this temperature for 1 hour, 3-[[5-oxido-4-(phenylsulfonyl)-1,2,5-oxadiazol-3-yl]oxy]-1-propanol (52 mg, 0.172 mmol, 1.2 equiv) was added. The mixture was stirred at room temperature overnight under nitrogen. The reaction was monitored by TLC (DCM / MeOH = 25:1). After complete reaction, the dichloromethane was removed by rotary evaporation, and the mixture was purified by column chromatography (DCM / MeOH = 25:1) and concentrated to afford Intermediate II.

[0093] (3) Synthesis of LO8

[0094] Intermediate II (50 mg, 0.052 mmol, 1.0 equiv) was dissolved in 1 mL of dichloromethane, and 0.5 mL of piperidine was added. The mixture was allowed to react at room temperature under nitrogen for 2 hours. The reaction was monitored by TLC (DCM / MeOH = 25:1). After the reaction was complete, the dichloromethane was removed by rotary evaporation, and the product was purified by column chromatography (DCM / MeOH = 25:1). The product was concentrated to yield 32 mg (85%) of a pale yellow solid.

[0095] LO8 1 H NMR (400MHz, CDCl3) δ9.01 (d, J=1.5Hz, 1H), 8.73 (dd, J=4.2, 1.2Hz, 1H),

[0096] 8.07(m,2H),7.96(m,2H),7.74(m,2H),7.64(m,4H),7.38(m,5H),6.48(m,2H),6.33(d,J=6.4H z,1H),4.64(s,2H),4.50(t,2H),4.39(t,2H),4.34(m,2H),3.97(d,J=6.1Hz,2H),2.25(m,2H). 13C NMR (75MHz, CDCl3) δ171.12,158.80,157.14,153.31,152.66,148.09,141.22,139.82,137.84,137.82,137.37,135.73,134.70,133.7 3,129.91,129.73,128.57,126.78,123.33,119.22,107.79,107.55,68.01,61.26,51.19,51.05,43.42,27.92.MS(ESI)m / z:761[M+H] + .HRMS(ESI)m / z calcd for C 34 H 32 N8O9S2,761.18119; found,761.17992[M+H] + .

[0097] Example 3

[0098] The synthesis of the neokotalanol derivative LO9 of the present invention comprises the following steps:

[0099]

[0100] (1) Synthesis of Intermediate IV is the same as in Example 1

[0101] (2) Synthesis of Intermediate II

[0102] Intermediate IV (100 mg, 0.143 mmol, 1.0 equiv) was dissolved in 20 mL of anhydrous dichloromethane. The reaction flask was placed at 0°C, and 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (33 mg, 0.172 mmol, 1.2 equiv) and 1-hydroxybenzotriazole (24 mg, 0.172 mmol, 1.2 equiv) were added. After stirring at this temperature for 1 hour, 4-[[5-oxido-4-(phenylsulfonyl)-1,2,5-oxadiazol-3-yl]oxy]-1-butanol (54 mg, 0.172 mmol, 1.2 equiv) was added. The mixture was stirred at room temperature overnight under nitrogen. The reaction was monitored by TLC (DCM / MeOH = 25:1). After complete reaction of the starting material, the dichloromethane was removed by rotary evaporation. The mixture was purified by column chromatography (DCM / MeOH = 25:1) and concentrated to afford Intermediate II.

[0103] (3) Synthesis of LO9

[0104] Intermediate II (50 mg, 0.052 mmol, 1.0 equiv) was dissolved in 1 mL of dichloromethane, and 0.5 mL of piperidine was added. The mixture was allowed to react at room temperature under nitrogen for 2 hours. The reaction was monitored by TLC (DCM / MeOH = 25:1). After complete reaction, the dichloromethane was removed by rotary evaporation, and the product was purified by column chromatography (DCM / MeOH = 25:1). The product was concentrated to yield 33 mg (81%) of a pale yellow solid.

[0105] LO9 1 H NMR(400MHz, CDCl3)δ9.01(d,J=1.6Hz,1H),8.74(d,J=4.2Hz,1H),

[0106] 8.08(m,2H),7.98(m,2H),7.78,(m,2H),7.67(m,4H),7.41(m,5H),6.49(m,2H),6.33(d,J=6.6Hz, 1H), 4.65 (s, 2H), 4.46 (t, 2H), 4.34 (s, 2H), 4.29 (t, 2H), 3.95 (d, J = 6.2Hz, 2H), 1.86-1.99 (m, 4H). 13 C NMR (75MHz, CDCl3) δ170.90,158.91,152.74,148.07,141.23,139.84,137.90,135.70,134.73,133.65,129.98,129.71128.56,126. 79,123.41,119.23,112.81,110.50,107.80,107.57,70.86,64.50,53.47,51.25,43.47,34.99,25.13,25.01.MS(ESI)m / z:775[M+H] + .HRMS(ESI)m / z calcd for C 35 H 34 N8O9S2,775.19684; found,775.19595[M+H] + .

[0107] Example 4

[0108] The synthesis of the neokotalanol derivative LO10 of the present invention comprises the following steps:

[0109]

[0110] (1) Synthesis of Intermediate IV is the same as in Example 1

[0111] (2) Synthesis of Intermediate II

[0112] Intermediate IV (100 mg, 0.143 mmol, 1.0 equiv) was dissolved in 20 mL of anhydrous dichloromethane. The reaction flask was placed at 0°C, and 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (33 mg, 0.172 mmol, 1.2 equiv) and 1-hydroxybenzotriazole (24 mg, 0.172 mmol, 1.2 equiv) were added. After stirring at this temperature for 1 hour, 4-(3-hydroxypropoxy)-3-methyl-1,2,5-oxadiazole 2-oxide (30 mg, 0.172 mmol, 1.2 equiv) was added. The mixture was stirred at room temperature overnight under a nitrogen atmosphere. The reaction was monitored by TLC (DCM / MeOH = 25:1). After complete reaction of the starting material, the dichloromethane was removed by rotary evaporation. The mixture was purified by column chromatography (DCM / MeOH = 25:1) and concentrated to afford Intermediate II.

[0113] (3) Synthesis of LO10

[0114] Intermediate II (50 mg, 0.052 mmol, 1.0 equiv) was dissolved in 1 mL of dichloromethane, and 0.5 mL of piperidine was added. The mixture was allowed to react at room temperature under nitrogen for 2 hours. The reaction was monitored by TLC (DCM / MeOH = 25:1). After complete reaction, the dichloromethane was removed by rotary evaporation, and the product was purified by column chromatography (DCM / MeOH = 25:1). The product was concentrated to afford 25 mg (61%) of a pale yellow solid.

[0115] LO10 1 H NMR(400MHz, CDCl3)δ8.98(dd,J=4.2,1.4Hz,1H),8.71(m,1H),7.94(m,2H),7.74(m,1H),7.66(m,2H),7.37(m,6H),6.47(m ,2H),6.31(dd,J=6.4,1.6Hz,1H),4.63(d,J=9.6Hz,2H),4.54(t,2H),4.42(t,2H),4.33(m,2H),3.78(s,3H),2.20(m,2H). 13C NMR (75MHz, CDCl3) δ171.10,153.29,152.63,148.08,141.23,139.84,137.86,137.41,134.72,133.71,129.92,129.89,126.78 ,123.34,119.27,119.23112.88,107.81,107.55,66.48,61.12,52.27,51.06,43.41,30.14,27.99,6.78.MS(ESI)m / z:635[M+H] + .HRMS(ESI)m / zcalcd for C 29 H 30 N8O7S,635.20364; found,635.20167[M+H] + .

[0116] Example 5

[0117] The synthesis of the neokotalanol derivative LO11 of the present invention comprises the following steps:

[0118]

[0119] (1) Synthesis of Intermediate IV is the same as in Example 1

[0120] (2) Synthesis of Intermediate II

[0121] Intermediate IV (100 mg, 0.143 mmol, 1.0 equiv) was dissolved in 20 mL of anhydrous dichloromethane. The reaction flask was placed at 0°C, and 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (33 mg, 0.172 mmol, 1.2 equiv) and 1-hydroxybenzotriazole (24 mg, 0.172 mmol, 1.2 equiv) were added. After stirring at this temperature for 1 hour, 4-(4-hydroxybutoxy)-3-methyl-1,2,5-oxadiazole 2-oxide (33 mg, 0.172 mmol, 1.2 equiv) was added. The mixture was stirred at room temperature overnight under a nitrogen atmosphere. The reaction was monitored by TLC (DCM / MeOH = 25:1). After complete reaction of the starting material, the dichloromethane was removed by rotary evaporation. The mixture was purified by column chromatography (DCM / MeOH = 25:1) and concentrated to afford Intermediate II.

[0122] (3) Synthesis of LO11

[0123] Intermediate II (50 mg, 0.052 mmol, 1.0 equiv) was dissolved in 1 mL of dichloromethane, and 0.5 mL of piperidine was added. The mixture was allowed to react at room temperature under nitrogen for 2 hours. The reaction was monitored by TLC (DCM / MeOH = 25:1). After complete reaction, the dichloromethane was removed by rotary evaporation, and the product was purified by column chromatography (DCM / MeOH = 25:1). The product was concentrated to yield 27 mg (73%) of a white solid.

[0124] LO11 1 H NMR (400MHz, CDCl3) δ9.01 (m, 1H), 8.74 (d, J = 4.5Hz, 1H), 7.98 (m, 2H),

[0125] 7.74(m,1H),7.64(m,2H),7.38(m,4H),6.49(m,2H),6.33(m,1H),4.64(m,2H),4.39( m,2H),4.24(m,2H),3.94(m,2H),3.77(s,2H),2.10(s,3H)1.87(m,2H),1.65(m,2H). 13 C NMR (75MHz, CDCl3) δ171.11,163.37,157.13,153.26,152.65,148.08,141.23,141.21,139.84,137.84,137.37,134.69,133.82,133.73,130 .02,129.92,126.77,123.33,123.29,119.25,112.81,107.81,69.57, 64.52,53.48,52.25,43.42,28.74,25.25,6.79.MS(ESI)m / z:649[M+H] + .HRMS(ESI)m / z calcd for C 30 H 32 N8O7S,649.21929; found,649.21661[M+H] + .

[0126] Example 6

[0127] The synthesis of the neokotalanol derivative LO12 of the present invention comprises the following steps:

[0128]

[0129] (1) Synthesis of Intermediate IV is the same as in Example 1

[0130] (2) Synthesis of Intermediate II

[0131] Intermediate IV (100 mg, 0.143 mmol, 1.0 equiv) was dissolved in 20 mL of anhydrous dichloromethane. The reaction flask was placed at 0°C, and 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (33 mg, 0.172 mmol, 1.2 equiv) and 1-hydroxybenzotriazole (24 mg, 0.172 mmol, 1.2 equiv) were added. After stirring at this temperature for 1 hour, 4-(4-hydroxybutoxy)-3-methyl-1,2,5-oxadiazole 2-oxide (35 mg, 0.172 mmol, 1.2 equiv) was added. The mixture was stirred at room temperature overnight under nitrogen. The reaction was monitored by TLC (DCM / MeOH = 25:1). After complete reaction, the dichloromethane was removed by rotary evaporation, and the mixture was purified by column chromatography (DCM / MeOH = 25:1) and concentrated to afford Intermediate II.

[0132] (3) Synthesis of LO12

[0133] Intermediate II (50 mg, 0.052 mmol, 1.0 equiv) was dissolved in 1 mL of dichloromethane, and 0.5 mL of piperidine was added. The mixture was allowed to react at room temperature under nitrogen for 2 hours. The reaction was monitored by TLC (DCM / MeOH = 25:1). After complete reaction, the dichloromethane was removed by rotary evaporation, and the product was purified by column chromatography (DCM / MeOH = 25:1). The product was concentrated to yield 21 mg (68%) of a gray solid.

[0134] LO12 1 H NMR(400MHz, CDCl3)δ9.01(d,J=1.3Hz,1H),8.74(d,J=4.2Hz,1H),

[0135] 7.98(m,2H),7.73(d,J=1.2Hz,,1H),7.64(d,J=7.8Hz,2H),7.38(m,4H),6.49(m,2H),6.33(d,J=6.9Hz,1H) ,4.65(s,2H),4.39(m,4H),4.20(t,2H),3.92(d,J=4.6Hz,2H),3.77(s,2H),2.10(s,3H),1.45-1.89(m,6H). 13C NMR (75MHz, CDCl3) δ163.44,152.70,148.09,141.24,139.86,134.71,129.98,126.77,123.36,119.23, 112.81,107.81,107.52,69.92,64.97,62.56,53.47,43.46,29.71,28.45,6.82.MS(ESI)m / z:663[M+H] + .HRMS(ESI)m / z calcd for C 31 H 34 N8O7S,663.23494; found,663.23184[M+H] + .

[0136] Example 7

[0137] The synthesis of the neokotalanol derivative LO13 of the present invention comprises the following steps:

[0138]

[0139] (1) Synthesis of Intermediate IV is the same as in Example 1

[0140] (2) Synthesis of Intermediate II

[0141] Intermediate IV (100 mg, 0.143 mmol, 1.0 equiv) was dissolved in 20 mL of anhydrous dichloromethane. The reaction flask was placed at 0°C, and 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (33 mg, 0.172 mmol, 1.2 equiv) and 1-hydroxybenzotriazole (24 mg, 0.172 mmol, 1.2 equiv) were added. After stirring at this temperature for 1 hour, 4-(3-hydroxypropoxy)-3-phenyl-1,2,5-oxadiazole 2-oxide (41 mg, 0.172 mmol, 1.2 equiv) was added. The mixture was stirred at room temperature overnight under a nitrogen atmosphere. The reaction was monitored by TLC (DCM / MeOH = 25:1). After complete reaction, the dichloromethane was removed by rotary evaporation, and the mixture was purified by column chromatography (DCM / MeOH = 25:1) and concentrated to afford Intermediate II.

[0142] (3) Synthesis of LO13

[0143] Intermediate II (50 mg, 0.052 mmol, 1.0 equiv) was dissolved in 1 mL of dichloromethane, and 0.5 mL of piperidine was added. The mixture was allowed to react at room temperature under nitrogen for 2 hours. The reaction was monitored by TLC (DCM / MeOH = 25:1). After complete reaction, the dichloromethane was removed by rotary evaporation, and the product was purified by column chromatography (DCM / MeOH = 25:1). The product was concentrated to afford 28 mg (77%) of a pale yellow solid.

[0144] LO13 1 H NMR (400MHz, CDCl3) δ8.99(d,J=2.2Hz,1H),8.72(m,1H),8.08(m,2H),7.93(m,2H),7.74(d,J=4.6Hz,1H),7.63(m,2H),7.48(m,3 H),7.34(m,5H),6.48(m,2H),6.33(d,J=11.8Hz,1H),4.61(s,2H),4.55(t,2H),4.38(t,2H),3.93(d,J=8.3Hz,2H),2.28(m,2H). 13 C NMR (75MHz, CDCl3) δ170.78,162.10,152.70,148.07,141.22,139.82,134.70,133.62,130.57,129.91,128.93,126.7 7,126.09,123.37,122.30,119.21,112.88,107.79,107.53,67.24,61.35,53.53,51.19,28.05.MS(ESI)m / z:697[M+H] + .HRMS(ESI)m / zcalcd for C 34 H 32 N8O7S,697.21929; found,697.21712[M+H] + .

[0145] Example 8

[0146] The synthesis of the neokotalanol derivative LO14 of the present invention comprises the following steps:

[0147]

[0148] (1) Synthesis of Intermediate IV is the same as in Example 1

[0149] (2) Synthesis of Intermediate II

[0150] Intermediate IV (100 mg, 0.143 mmol, 1.0 equiv) was dissolved in 20 mL of anhydrous dichloromethane. The reaction flask was placed at 0°C, and 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (33 mg, 0.172 mmol, 1.2 equiv) and 1-hydroxybenzotriazole (24 mg, 0.172 mmol, 1.2 equiv) were added. After stirring at this temperature for 1 hour, 2-hydroxyethyl nitrate (19 mg, 0.172 mmol, 1.2 equiv) was added. Under nitrogen, the mixture was stirred at room temperature overnight. The reaction was monitored by TLC (DCM / MeOH = 25:1). After complete reaction, the dichloromethane was removed by rotary evaporation, and the mixture was purified by column chromatography (DCM / MeOH = 25:1) and concentrated to afford Intermediate II.

[0151] (3) Synthesis of LO14

[0152] Intermediate II (50 mg, 0.052 mmol, 1.0 equiv) was dissolved in 1 mL of dichloromethane, and 0.5 mL of piperidine was added. The mixture was allowed to react at room temperature under nitrogen for 2 hours. The reaction was monitored by TLC (DCM / MeOH = 25:1). After complete reaction, the dichloromethane was removed by rotary evaporation, and the product was purified by column chromatography (DCM / MeOH = 25:1). The product was concentrated to afford 28 mg (77%) of a pale yellow solid.

[0153] LO14 1 H NMR (300MHz, CDCl3) δ9.00(s,1H),8.73(d,J=4.6Hz,1H),7.93(m,2H),7.74(d,J=1.5Hz,1H),7.62(d,H=8.9Hz,2H),7.33(m,4H ),6.46(m,2H),6.32(d,J=6.4Hz,1H),4.63(m,2H),4.48(t,J=2.8Hz,2H),4.33(s,2H),4.27(t,2H),3.94(m,2H),2.04(m,2H). 13 C NMR (75MHz, CDCl3) δ170.97,157.06,153.26,152.66,148.08,141.23,139.83,137.89,137.34,134.69,133.72,129.9 0,126.79,123.34,119.23,112.86,107.80,107.59,70.34,61.23,51.11,53.49,43.38,26.32.MS(ESI)m / z:568[M+H] + .HRMS(ESI)m / zcalcd for C 26H 27 N7O7S,568.16144; found,568.15933[M+H] + .

[0154] Example 9

[0155] The synthesis of the neokotalanol derivative LO15 of the present invention comprises the following steps:

[0156]

[0157] (1) Synthesis of Intermediate IV is the same as in Example 1

[0158] (2) Synthesis of Intermediate II

[0159] Intermediate IV (100 mg, 0.143 mmol, 1.0 equiv) was dissolved in 20 mL of anhydrous dichloromethane. The reaction flask was placed at 0°C, and 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (33 mg, 0.172 mmol, 1.2 equiv) and 1-hydroxybenzotriazole (24 mg, 0.172 mmol, 1.2 equiv) were added. After stirring at this temperature for 1 hour, 3-hydroxypropyl nitrate (21 mg, 0.172 mmol, 1.2 equiv) was added. Under nitrogen, the mixture was stirred at room temperature overnight. The reaction was monitored by TLC (DCM / MeOH = 25:1). After complete reaction, the dichloromethane was removed by rotary evaporation, and the mixture was purified by column chromatography (DCM / MeOH = 25:1) and concentrated to afford Intermediate II.

[0160] (3) Synthesis of LO15

[0161] Intermediate II (50 mg, 0.052 mmol, 1.0 equiv) was dissolved in 1 mL of dichloromethane, and 0.5 mL of piperidine was added. The mixture was allowed to react at room temperature under nitrogen for 2 hours. The reaction was monitored by TLC (DCM / MeOH = 25:1). After complete reaction, the dichloromethane was removed by rotary evaporation, and the product was purified by column chromatography (DCM / MeOH = 25:1). The product was concentrated to yield 23 mg (77%) of a pale yellow solid.

[0162] LO15 1H NMR (300MHz, CDCl3) δ9.00(s,1H),8.73(d,J=4.6Hz,1H),7.93(m,2H),7.74(d,J=1.5Hz,1H),7.62(d,H=8.9Hz,2H),7.33(m,4H ),6.46(m,2H),6.32(d,J=6.4Hz,1H),4.63(m,2H),4.48(t,J=2.8Hz,2H),4.33(s,2H),4.27(t,2H),3.94(m,2H),2.04(m,2H). 13 C NMR (75MHz, CDCl3) δ170.97,157.06,153.26,152.66,148.08,141.23,139.83,137.89,137.34,134.69,133.72,129.9 0,126.79,123.34,119.23,112.86,107.80,107.59,70.34,61.23,51.11,53.49,43.38,26.32.MS(ESI)m / z:582[M+H] + .HRMS(ESI)m / zcalcd for C 26 H 27 N7O7S,582.17709; found,582.17450[M+H] + .

[0163] Example 10

[0164] The synthesis of the neokotalanol derivative LO16 of the present invention comprises the following steps:

[0165]

[0166] (1) Synthesis of Intermediate IV is the same as in Example 1

[0167] (2) Synthesis of Intermediate II

[0168] Intermediate IV (100 mg, 0.143 mmol, 1.0 equiv) was dissolved in 20 mL of anhydrous dichloromethane. The reaction flask was placed at 0°C, and 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (33 mg, 0.172 mmol, 1.2 equiv) and 1-hydroxybenzotriazole (24 mg, 0.172 mmol, 1.2 equiv) were added. After stirring at this temperature for 1 hour, 3-hydroxypropyl nitrate (21 mg, 0.172 mmol, 1.2 equiv) was added. Under nitrogen, the mixture was stirred at room temperature overnight. The reaction was monitored by TLC (DCM / MeOH = 25:1). After complete reaction, the dichloromethane was removed by rotary evaporation, and the mixture was purified by column chromatography (DCM / MeOH = 25:1) and concentrated to afford Intermediate II.

[0169] (3) Synthesis of LO16

[0170] Intermediate II (50 mg, 0.052 mmol, 1.0 equiv) was dissolved in 1 mL of dichloromethane, and 0.5 mL of piperidine was added. The mixture was allowed to react at room temperature under nitrogen for 2 hours. The reaction was monitored by TLC (DCM / MeOH = 25:1). After complete reaction, the dichloromethane was removed by rotary evaporation, and the product was purified by column chromatography (DCM / MeOH = 25:1). The product was concentrated to afford 18 mg (52%) of a pale yellow solid.

[0171] LO16 1 H NMR (400MHz, CDCl3) δ9.01(m,1H),8.74(m,1H),7.98(m,2H),7.93(d,J=4.6Hz,1H),7.74(m,1H),7.67(m,2H),7.33(m,3H ),6.49(m,2H),6.33(d,J=12.2Hz,1H),5.29(m,1H),4.61(s,2H),4.48(d,J=7.8Hz,1H),4.33(s,2H),3.86-4.04(m,8H). 13 C NMR (75MHz, CDCl3) δ174.46,152.75,148.15,141.24,139.94,134.71,133.59,129.87,126.80,123.32,119 .25,113.01,107.81,86.43,81.62,81.20,78.05,73.38,69.34,53.47,51.10,43.43.MS(ESI)m / z:652[M+H] + .HRMS(ESI)m / z calcd for C 29 H 29N7O9S,652.18257; found,652.18130[M+H] +

[0172] Example 11

[0173] The synthesis of the neokotalanol derivative LO17 of the present invention comprises the following steps:

[0174]

[0175] (1) Synthesis of Intermediate IV is the same as in Example 1

[0176] (2) Synthesis of LO17

[0177] Intermediate IV (100 mg, 0.143 mmol, 1.0 equiv) was dissolved in 20 mL of anhydrous acetonitrile, and potassium carbonate (99 mg, 0.715 mmol, 5.0 equiv) was added. The mixture was stirred at room temperature for 1 hour. (E)-1-(Chloromethoxy)-3,3-dimethyltriazene-1-ene 2-oxide (27 mg, 0.172 mmol, 1.2 equiv) was added, and the mixture was allowed to react overnight at room temperature under a nitrogen atmosphere. The reaction was monitored by TLC (DCM / MeOH = 25:1). After complete reaction of the starting material, the dichloromethane was removed by rotary evaporation, and the product was purified by column chromatography (DCM / MeOH = 25:1). The product was concentrated to yield 22 mg (56%) of a pale yellow solid.

[0178] LO17 1 H NMR (400MHz, CDCl3) δ9.00(s,1H),8.71(s,1H),7.92(m,2H),7.81(m,2H),7.73(d,J=2.2Hz,1H),7.59(m,4H),7.44(m,3H),7.35(m,3H),7. 27(m,4H),6.88(m,1H),6.49(m,1H),5.76(s,2H),4.66(d,J=4.2Hz,1H),4.51(d,J=3.6Hz,2H),4.38(s,2H),4.29(m,2H),2.904.38(s,6H). 13C NMR (75MHz, CDCl3) δ168.19,154.20,153.35,152.15,147.86,143.53,141.39,141.23,139.82,138.23,134.61,133.34,129.84,12 7.89,127.25,126.81,124.75,120.09,119.20,118.68,107.80,87.62,68.12,51.69,51.54,46.96,42.11.MS(ESI)m / z:840[M+Na] + .HRMS(ESI)m / z calcd for C 41 H 39 N9O8S,840.25400; found,840.25303[M+Na] + .

[0179] Example 12

[0180] The synthesis of the neokotalanol derivative LO18 of the present invention comprises the following steps:

[0181]

[0182] (1) Synthesis of Intermediate IV is the same as in Example 1

[0183] (2) Synthesis of LO18

[0184] Intermediate IV (100 mg, 0.143 mmol, 1.0 equiv) was dissolved in 20 mL of anhydrous acetonitrile, and potassium carbonate (99 mg, 0.715 mmol, 5.0 equiv) was added. The mixture was stirred at room temperature for 1 hour. (E)-1-(Chloromethoxy)-3,3-diethyltriazene-1-ene 2-oxide (32 mg, 0.172 mmol, 1.2 equiv) was added, and the mixture was allowed to react overnight at room temperature under a nitrogen atmosphere. The reaction was monitored by TLC (DCM / MeOH = 25:1). After complete reaction of the starting material, the dichloromethane was removed by rotary evaporation, and the product was purified by column chromatography (DCM / MeOH = 25:1) and concentrated to yield 27 mg (54%) of a white solid.

[0185] LO18 1H NMR (400MHz, CDCl3) δ9.00(s,1H),8.72(m,1H),8.08(m,2H),7.93(m,2H),7.79(d,J=6.6Hz,2H),7.72(d,J=1.8Hz,1H),7.57(t,4H),7.24-7.46( m,7H),6.93(m,1H),6.48(t,1H),5.77(s,2H),4.68(d,J=7.8Hz,2H),4.4 7(d,J=12.8Hz,4H),4.38(s,2H),4.29(t,1H),3.11(q,4H),0.99(t,6H). 13 C NMR (75MHz, CDCl3) δ168.10,154.15,153.37,152.94,152.12,147.89,14 3.52,141.38,141.20,139.81,138.21,134.57,133.30,129.84,127.88,1 27.25,126.80,124.72,123.60,120.08,119.19,118.70,116.65,107.78, 87.80,68.10,51.84,51.61,47.67,46.96,31.52.MS(ESI)m / z:868[M+Na] + .HRMS(ESI)m / zcalcd for C 43 H 43 N9O8S,868.28530; found,868.28380[M+Na] + .

[0186] Example 13

[0187] The synthesis of the neokotalanol derivative LO19 of the present invention comprises the following steps:

[0188]

[0189] (1) Synthesis of Intermediate IV is the same as in Example 1

[0190] (2) Synthesis of LO19

[0191] Intermediate IV (100 mg, 0.143 mmol, 1.0 equiv) was dissolved in 20 mL of anhydrous acetonitrile, and potassium carbonate (99 mg, 0.715 mmol, 5.0 equiv) was added. The mixture was stirred at room temperature for 1 hour. (Z)-2-(Chloromethoxy)-1-morpholinodiazene-1-oxide (34 mg, 0.172 mmol, 1.2 equiv) was added, and the mixture was reacted at room temperature overnight under a nitrogen atmosphere. The reaction was monitored by TLC (DCM / MeOH = 25:1). After complete reaction of the starting material, the dichloromethane was removed by rotary evaporation, and the product was purified by column chromatography (DCM / MeOH = 25:1) and concentrated to afford 26 mg (51%) of a gray solid.

[0192] LO19 1 H NMR (400MHz, CDCl3) δ8.98 (s, 1H), 8.71 (d, J = 1.7Hz, 1H), 7.93 (m, 2H), 7.79 (d ,J=7.6Hz,2H),7.72(d,J=2.2Hz,1H),7.57(t,4H),7.44(m,3H),7.24-7.36(m, 6H),6.93(m,1H),6.48(m,1H),5.75(s,2H),4.68(d,J=7.9Hz,2H),4.52(d,J= 12.5Hz,4H),4.38(s,2H),4.29(t,1H),3.80(m,2H),3.54(m,4H),3.16(m,2H). 13 C NMR (75MHz, CDCl3) δ168.13,154.21,153.37,152.13,148.31,143.51,141.40,141.27,139.81,138.26,134.63,133.45,129.86,127.9 1,127.27,126.94,124.74,120.10,119.39,118.66,107.84,87.66,68.12,65.29,51.65,51.56,47.69,46.97.MS(ESI)m / z:882[M+Na] + .HRMS(ESI)m / zcalcd for C 43 H 41 N9O9S,882.26456; found,882.26379[M+Na] + .

[0193] Example 14

[0194] The synthesis of the neokotalanol derivative LO20 of the present invention comprises the following steps:

[0195]

[0196] (1) Synthesis of Intermediate IV is the same as in Example 1

[0197] (2) Synthesis of LO20

[0198] Intermediate IV (100 mg, 0.143 mmol, 1.0 equiv) was dissolved in 20 mL of anhydrous acetonitrile, and potassium carbonate (99 mg, 0.715 mmol, 5.0 equiv) was added. The mixture was stirred at room temperature for 1 hour. (Z)-2-(Chloromethoxy)-1-(4-hydroxypiperidin-1-yl)diazene-1-oxide (36 mg, 0.172 mmol, 1.2 equiv) was added, and the mixture was reacted at room temperature overnight under a nitrogen atmosphere. The reaction was monitored by TLC (DCM / MeOH = 25:1). After complete reaction of the starting material, the dichloromethane was removed by rotary evaporation, and the product was purified by column chromatography (DCM / MeOH = 25:1) and concentrated to afford 33 mg (67%) of a pale yellow solid.

[0199] LO20 1 H NMR (400MHz, CDCl3) δ9.01(s,1H),8.71(s,1H),7.93(m,2H),7.80(d,J=7.9Hz,2H),7.72(d,J=2.1Hz,1H),7.57(m,4H),7.44(m,3H),7.24-7.36( m,6H),6.93(m,1H),6.48(m,1H),5.76(s,2H),4.68(d,J=7.4Hz,2H),4.5 2(d,J=12.9Hz,4H),4.38(s,2H),4.29(t,1H),3.68(m,4H),3.27(t,4H). 13 C NMR (75MHz, CDCl3) δ168.19,154.17,153.34,152.96,152.16,147.90,143.52,141.39,141.25,139.84,138.25,134.57,133.29,129.83,127. 92,127.27,126.77,124.75,120.10,119.19,118.66,107.83,87.61,68 .16,65.89,51.64,51.48,51.05,46.96,29.72.MS(ESI)m / z:896[M+Na] + .HRMS(ESI)m / z calcdfor C 44 H 43N9O9S,896.28021; found,896.27883[M+Na] + .

[0200] Performance Characterization

[0201] 1. NO release of NO-donating omidatepag derivatives in aqueous humor in vitro

[0202] Experimental method: Using the aqueous humor of New Zealand white rabbits (1-2 kg, half male and half female), a 500 μM NaNO2 solution was added to a 96-well plate (triplicate wells, 80 μL added to each empty well). Then, a gradient dilution (using blank solution, diluting twice each time) was performed to obtain NaNO2 solutions with concentrations of 500, 250, 125, 62.5, 31.25, 15.625, and 7.8125 μM, respectively. In addition, a blank control group (three wells for each concentration, 40 μL in each well) was required. Subsequently, 40 μL of Griess reagent was added, and the absorbance at 540 nm was measured using a microplate reader. A linear fit was performed between the absorbance and concentration to obtain a standard curve. The prepared aqueous humor solution (200 μM) of the test compound and the control group of aqueous humor solution without the test compound were placed in a shaker (37°C, 120 rpm) and incubated for 2 hours. 40 μL was taken with a pipette and added to a 96-well plate (triplicate wells). Then 40 μL of Griess reagent was added and placed in a microplate reader to measure the absorbance at 540 nm. The absorbance was substituted into the standard curve to calculate the concentration. Three independent experiments were performed for each test compound. The experimental results are shown in Figure 2. Figure 1 shown.

[0203] like Figure 1 As shown, all compounds released NO in vitro in rabbit aqueous humor. Different NO donors released varying amounts of NO, with azoiniums releasing the most, furazan N-oxides releasing a moderate amount, and nitrates releasing the least. Furthermore, the compounds released NO in a time-dependent manner over 8 hours, and different NO donor types released different amounts of NO at different rates. This indicates that the compounds can release NO in vitro in rabbit aqueous humor, and that different NO donor types can release varying amounts of NO at varying rates.

[0204] 2. Metabolism of NO-donating Omidenepag derivatives to the original drug after eye drop administration

[0205] Experimental method: New Zealand white rabbits (1-2 kg, half male and half female) were used. Before the experiment, the rabbits were adapted to feeding for 3 days (temperature 25°C, humidity 40%-70%), and their eyes were examined to ensure that the eyes were normal. 25 μL of 2 mM test compound was administered to the left eye. 2 hours after administration, 100 μL of aqueous humor from the left eye of the rabbit was drawn and temporarily stored in liquid nitrogen. 100 μL of mass spectrometry methanol was added, and the supernatant was collected after centrifugation and filtered through a 0.22 μm organic filter membrane. The compound concentration was detected by HPLC using the internal standard method. The experimental results are as follows. Figure 2 shown.

[0206] like Figure 2 As shown, no test compounds were detected after eye drop administration, indicating that all test compounds can release the original drug Omidenepag after penetrating the cornea after eye drop administration, among which compounds LO7, LO10, LO12, LO16, LO17 and LO19 can release more original drug after eye drop administration.

[0207] 3. Stability of NO-donating Omidenepag derivatives in solvents

[0208] Experimental method: Prepare 2mM pure water stock solution of the test compound, take another EP tube, add 3600μL solvent, and then add 400μL stock solution to obtain 4000μL of 200μM test compound solution. Place it in a shaker (25℃, 120rpm) and incubate for 7 days. At 0min, 15min, 30min, 1h, 2h, 4h, 8h, 1d, 2d, 3d, 4d, 5d, 6d and 7d after incubation, take 200μL with a pipette, add 200μL of mass spectrometry methanol, take the supernatant after centrifugation, filter with 0.22μM organic filter membrane, detect by HPLC, and calculate the concentration. The experimental results are as follows: Figure 3 shown.

[0209] like Figure 3 As shown, all compounds still had more than 80% remaining after 7 days, and compared with the positive drug OMDI, the compounds connected with different types of NO donors still retained higher stability, that is, the compounds had a certain stability in the solvent at room temperature.

[0210] 4. NO-donating Omidenepag derivatives have EP2 receptor agonist effects in vitro

[0211] Experimental Methods: Stably expressing EP2 receptor cells were incubated with various concentrations of test compounds, and the agonist activity of the compounds at the EP2 receptor was measured using a TR-FRET cAMP assay. HEK cells stably expressing EP2 were cultured in DMEM supplemented with 10% fetal bovine serum and 0.1 mg / mL Hygromycin B at 37°C and 5% CO2. Cells were passaged by removing the old medium and washing once with PBS, followed by addition of 1 mL of TrypLE TM Express solution, incubate at 37°C for about 2 minutes. When the cells detach from the bottom of the dish, add about 2 mL of complete culture medium preheated at 37°C. Gently blow the cell suspension with a pipette to separate the aggregated cells. Transfer the cell suspension to a sterile centrifuge tube and centrifuge at 1000 rpm for 5 minutes. To maintain the physiological activity of the cells, the experimental cell confluence is controlled at about 80%. Prepare 1×Stimulation Buffer according to the kit instructions for use. Use DMSO to gradiently dilute the positive compound and the test compound to 10 concentrations, then use 1×Stimulation Buffer to dilute the compound to 10×, shake and mix well for use. Stable transfected cells are cultured to 80% confluence, trypsin digestion is used to collect the cells, and after counting, 9 μL / well is inoculated into a 384-well plate in an exciting mode. Take 1 μL of the 10× compound diluted in step 2 and add it to the corresponding experimental wells, among which 1 μL of 10× first concentration positive compound is added to PC well and 1 μL of 10× DMSO buffer (1%) is added to VC well. After centrifugation, place it at 37℃ and incubate for 30 minutes. Dilute Eu-cAMP to the working concentration with detection buffer and take 5 μL / well and add it to the corresponding experimental wells. TM Dilute the anti-cAMP antibody to the working concentration in detection buffer and add 5 μL / well to the corresponding experimental wells; centrifuge and incubate at room temperature for 1 hour. After incubation, read the values ​​at 665 nm and 620 nm using a microplate reader under 330 nm excitation. The formula for % activation is:

[0212]

[0213] Where: represents the mean value of positive control; The experimental results are shown in Table 2. Figure 4 shown.

[0214] like Figure 4As shown in the figure, it was found that the preferred compounds all had agonist effects on EP2 receptors in vitro, among which compounds LO7, LO10 and LO12 had better agonist activities on EP2 receptors than OMDI, EC 50 The results were 1.688nM, 0.8223nM, and 3.69nM, respectively (Table 1). A compound with good in vitro EP2 receptor agonist activity was screened from three different types of NO donors. Combined with the results of in vitro NO release in rabbit aqueous humor and the metabolism of the original drug after eye drop administration, LO10, LO16, and LO19 were selected as the preferred compounds for further in vivo evaluation of their biological activity.

[0215] Table 1 shows the agonistic effects of preferred compounds on EP2 receptors

[0216] CompoundID <![CDATA[EC 50 (nM)]]> Bottom Top PGE2 0.01412 0.64 100.20 LO7 1.688 17.86 101.70 LO10 0.8223 4.76 100.90 LO12 3.69 5.63 101.80 LO16 11.57 13.29 ＝100.0 LO17 14.65 12.45 ＝100.0 LO19 9.511 8.04 ＝100.0 OMDI 4.86 8.19 100.00

[0217] 5. Intraocular pressure-lowering effect of NO-donating omidatepag derivatives in normal intraocular pressure rabbits

[0218] Experimental Methods: New Zealand white rabbits (1-2 kg, half male and half female) were used. Before the experiment, the rabbits were adapted to the environment for 3 days (temperature 25°C, humidity 40%-70%), and their eyes were examined to ensure normal eye condition. Before administration, normal intraocular pressure was measured in both eyes using a rebound tonometer. Five measurements were taken for each eye, and the maximum and minimum values ​​were removed. The average value was taken as the 0-hour intraocular pressure (control group). The left eye of each rabbit was uniformly selected for administration, and the right eye was not treated. The corneal surface and conjunctival sac were rinsed with normal saline to keep the eye moist. First, we used OMDI as a positive control (the concentration was set at 0.002%. Unless otherwise specified, all preparations were made with normal saline) to evaluate the intraocular pressure-lowering activity of compounds LO10 (furazan N-oxide), LO16 (nitrate), and LO19 (azodiol salt) (equimolar to OMDI). After administration, the intraocular pressure was measured at 1, 2, 3, 4, 5, 6, 7, 8, and 24 hours, and the data were recorded. The experimental results are shown in the figure below. Figure 5 shown.

[0219] like Figure 5 As shown in the results, we found that all three NO-donating omidatepag derivatives were able to reduce intraocular pressure in normal rabbits after administration. LO10 demonstrated superior IOP-lowering activity at 3 and 24 hours after administration, with significant differences compared to LO16, LO19, and OMDI. The IOP-lowering effect was sustained for a longer period, with no rebound effect at 24 hours.

[0220] The present invention discloses a nitric oxide donor type Omidenepag derivative or a pharmacologically acceptable salt thereof represented by general formula I. Ocular metabolism experiments show that all compounds can produce the active substance Omidenepag (OMD) and release NO in aqueous humor, thereby synergistically producing an intraocular pressure lowering effect with OMD. In vitro EP2 agonist activity shows that the three nitric oxide donor type Omidenepag derivatives all retain good EP2 receptor agonist activity, among which compound LO10 has an in vitro EC2 agonist activity on EP2 receptor. 50 The concentration of 0.8223 nM is superior to OMDI (4.86 nM), retaining a strong EP2 receptor agonist effect. In in vitro stability experiments in solvents, all three nitric oxide donor-type omidatepag derivatives retained high stability. Among them, compound LO10 still had 87% remaining on the seventh day, which is comparable to OMDI (87%). It retains its original stability while improving its efficacy. In the intraocular pressure lowering experiment in normal intraocular pressure rabbits, all three nitric oxide donor-type omidatepag derivatives showed good intraocular pressure lowering activity. At a dose equimolar to the marketed concentration of OMDI, compound LO10's intraocular pressure lowering activity was superior to the positive control OMDI at 3 and 24 hours after administration. The intraocular pressure lowering lasted longer, the eye irritation was less, and there was no rebound phenomenon after 24 hours. Therefore, according to the present invention, it is possible to provide a new compound with excellent properties for the prevention and treatment of ophthalmic diseases (particularly glaucoma or ocular hypertension).

Claims

1. A nitric oxide donor type Omidenepag derivative, characterized in that: Its structural formula is: Wherein, R1 is selected from Or -H; R2 is selected from methylene, ethylene, propylene, butylene, pentylene, or isosorbide R3 is selected from or -ONO2.

2. The nitric oxide donor type Omidenepag derivative according to claim 1, characterized in that The R1 is H.

3. The nitric oxide donor type Omidenepag derivative according to claim 1, characterized in that R1 is H, R2 is selected from methylene, ethylene, propylene, butylene or pentylene, and R3 is selected from 4. The nitric oxide donor type Omidenepag derivative according to claim 1, characterized in that The structural formula of the derivative is:

5. A method for preparing the nitric oxide donor type Omidenepag derivative according to any one of claims 1 to 4, characterized in that: The following steps are involved: (1) Compound III Omidenepag and 9-fluorenylmethyl chloroformate undergo amide condensation reaction to obtain intermediate IV; (2) Intermediate IV undergoes ester condensation with compound R3-R2-OH or nucleophilic substitution with compound R3-R2-Cl to obtain compound II; (3) Compound II is deprotected from the 9-fluorenylmethoxycarbonyl protecting group to obtain the target compound I; The synthetic route is as follows: Wherein, R1 is selected from Or -H; R2 is selected from methylene, ethylene, propylene, butylene, pentylene, or isosorbide R3 is selected from Or -ONO2; when R1 is When , step (3) is not performed.

6. The method for preparing the nitric oxide donor type Omidenepag derivative according to claim 5, characterized in that: In step (1), the reaction temperature is -20 to 25° C., and the reaction solvent is one or more of anhydrous dichloromethane, chloroform, ethyl acetate, redistilled acetone, anhydrous tetrahydrofuran, anhydrous N,N-dimethylformamide, dimethyl sulfoxide or dioxane.

7. The method for preparing the nitric oxide donor type Omidenepag derivative according to claim 5, characterized in that: In step (2), the reaction temperature is -20 to 25° C., and the solvent is selected from one or more of anhydrous acetonitrile, anhydrous dichloromethane, chloroform, ethyl acetate, redistilled acetone, anhydrous tetrahydrofuran, anhydrous N,N-dimethylformamide, dimethyl sulfoxide or dioxane.

8. The method for preparing the nitric oxide donor type Omidenepag derivative according to claim 5, characterized in that: In step (3), the reaction temperature is -20 to 25° C., and the solvent is selected from one or more of anhydrous piperidine, anhydrous acetonitrile, anhydrous dichloromethane, chloroform, ethyl acetate, redistilled acetone, anhydrous tetrahydrofuran, anhydrous N,N-dimethylformamide, dimethyl sulfoxide or dioxane.

9. Use of the nitric oxide donor Omidenepag derivative according to any one of claims 1 to 4 in the preparation of a medicament for preventing and / or treating diseases associated with increased intraocular pressure.

10. The use according to claim 9, characterized in that The diseases associated with increased intraocular pressure include ocular hypertension, glaucoma and diabetic eye complications.

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