WO2020020306A1 - 一种羊毛甾醇前药化合物的晶型及其应用 - Google Patents

一种羊毛甾醇前药化合物的晶型及其应用 Download PDF

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WO2020020306A1
WO2020020306A1 PCT/CN2019/097773 CN2019097773W WO2020020306A1 WO 2020020306 A1 WO2020020306 A1 WO 2020020306A1 CN 2019097773 W CN2019097773 W CN 2019097773W WO 2020020306 A1 WO2020020306 A1 WO 2020020306A1
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Prior art keywords
lanosterol
lens
eye drops
compound
eye
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PCT/CN2019/097773
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English (en)
French (fr)
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刘奕志
王延东
李小林
罗志
贺海鹰
黎健
陈曙辉
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中山大学中山眼科中心
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Priority to CN201980049648.9A priority Critical patent/CN112543762B/zh
Priority to KR1020217005334A priority patent/KR20210038596A/ko
Priority to SG11202100665WA priority patent/SG11202100665WA/en
Priority to US17/262,483 priority patent/US11149055B2/en
Application filed by 中山大学中山眼科中心 filed Critical 中山大学中山眼科中心
Priority to AU2019308940A priority patent/AU2019308940B2/en
Priority to EP19841413.8A priority patent/EP3828193A4/en
Priority to MX2021000900A priority patent/MX2021000900A/es
Priority to BR112021001172-2A priority patent/BR112021001172A2/pt
Priority to JP2021509916A priority patent/JP7046266B2/ja
Priority to RU2021103501A priority patent/RU2766088C1/ru
Priority to CA3107361A priority patent/CA3107361C/en
Publication of WO2020020306A1 publication Critical patent/WO2020020306A1/zh
Priority to PH12021550182A priority patent/PH12021550182A1/en
Priority to CONC2021/0000932A priority patent/CO2021000932A2/es
Priority to ZA2021/00923A priority patent/ZA202100923B/en

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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/0012—Galenical forms characterised by the site of application
    • A61K9/0048—Eye, e.g. artificial tears
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/56—Compounds containing cyclopenta[a]hydrophenanthrene ring systems; Derivatives thereof, e.g. steroids
    • A61K31/57—Compounds containing cyclopenta[a]hydrophenanthrene ring systems; Derivatives thereof, e.g. steroids substituted in position 17 beta by a chain of two carbon atoms, e.g. pregnane or progesterone
    • A61K31/573—Compounds containing cyclopenta[a]hydrophenanthrene ring systems; Derivatives thereof, e.g. steroids substituted in position 17 beta by a chain of two carbon atoms, e.g. pregnane or progesterone substituted in position 21, e.g. cortisone, dexamethasone, prednisone or aldosterone
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/08—Solutions
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P27/00—Drugs for disorders of the senses
    • A61P27/02—Ophthalmic agents
    • A61P27/12—Ophthalmic agents for cataracts
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07J—STEROIDS
    • C07J9/00—Normal steroids containing carbon, hydrogen, halogen or oxygen substituted in position 17 beta by a chain of more than two carbon atoms, e.g. cholane, cholestane, coprostane
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07B—GENERAL METHODS OF ORGANIC CHEMISTRY; APPARATUS THEREFOR
    • C07B2200/00—Indexing scheme relating to specific properties of organic compounds
    • C07B2200/13—Crystalline forms, e.g. polymorphs

Definitions

  • the present invention relates to a crystalline form of a compound of formula (I) and the use of the crystalline form in the preparation of a medicament for treating ophthalmic diseases.
  • Cataract is a disease of the eye that occurs on the lens in the eyeball.
  • the opacity of the lens is collectively called cataract.
  • Aging, heredity, metabolic abnormalities, trauma, radiation, poisoning, and local malnutrition can all cause lens capsule damage, increase its permeability, lose its barrier effect, or cause lens metabolism disorders, degeneration of lens proteins, and turbidity.
  • the lens of the eyeball changes from transparent to opaque, which affects the eyes to receive sunlight, it will affect the vision of the eyes.
  • the turbidity of the eyeball is lighter, the impact on vision is lighter.
  • the degree of turbidity gradually deepens the vision will also increase, and severe cases will lead to blindness.
  • Cataract is one of the most common blinding eye diseases and it is the main cause of blindness. Because the mechanism of cataract formation is not clear, no breakthrough has been achieved in drug treatment. Therefore, the only effective treatment currently identified is surgical treatment.
  • the clinical treatment drugs for cataract include: 1 aldose reductase inhibitors, such as catarrhine (Kataling, Carinyo, Bainertin), facrine, benzyl lysine, etc .; 2 Oxidative damage drugs, such as glutathione, taurine, aspirin, etc .; 3 nutrient metabolism drugs, such as vitamins, carotenoids, etc .; 4 Chinese medicine compound prescriptions include Dendrobium Yeguang Pill, Qiju Dihuang Pill, Shi Ju Ming San, etc. These drugs for cataracts have been confirmed by long-term clinical trials. They can only delay the deterioration of cataracts and cannot reverse them, thus treating cataracts.
  • 1 aldose reductase inhibitors such as catarrhine (Kataling, Carinyo, Bainertin), facrine, benzyl lysine, etc .
  • Oxidative damage drugs such as glutathione, taurine, aspirin, etc .
  • Lanosterol is an amphiphilic molecule enriched in the lens. It is synthesized by a key cyclization reaction of lanosterol synthase (LSS) in the cholesterol synthesis pathway, which can reduce the abnormal aggregation of lens proteins and make them re-established. Regular arrangement to restore crystal transparency. Studies have shown that lanosterol synthase can be detected in the lens. In addition, in the Shumiya cataract rat study, a specific combination of homozygous mutations of martosterol synthase and farnesyl diphosphate farnesyl transferase 1 (FDFT1) can reduce cholesterol levels in the lens and cause cataracts.
  • LDS lanosterol synthase
  • FDFT1 farnesyl diphosphate farnesyl transferase 1
  • the present invention provides Form A of the compound of formula (I), whose X-ray powder diffraction (XRPD) pattern has characteristic diffraction peaks at the following 2 ⁇ angles: 8.60 ⁇ 0.2 °, 15.06 ⁇ 0.2 °, and 17.22 ⁇ 0.2 °.
  • XRPD X-ray powder diffraction
  • the X-ray powder diffraction pattern of the above-mentioned Form A has characteristic diffraction peaks at the following 2 ⁇ angles: 8.60 ⁇ 0.2 °, 9.38 ⁇ 0.2 °, 10.57 ⁇ 0.2 °, 12.54 ⁇ 0.2 °, 14.43 ⁇ 0.2 °, 15.06 ⁇ 0.2 °, 17.22 ⁇ 0.2 °, and 25.18 ⁇ 0.2 °.
  • the X-ray powder diffraction pattern of the above-mentioned Form A has characteristic diffraction peaks at the following 2 ⁇ angles: 4.350 °, 8.598 °, 9.383 °, 10.566 °, 12.542 °, 13.448 °, 14.428 °, 14.591 ° , 15.063 °, 15.453 °, 15.820 °, 16.803 °, 17.216 °, 20.985 °, 21.181 °, 22.225 °, 22.601 °, 22.856 °, 23.726 °, 24.039 °, 24.534 °, 25.185 °, 25.514 °, 25.935 °, 26.570 °, 27.867 °, 28.125 °, 28.416 °, 29.114 °, 29.445 °, 31.914 °, 33.710 °, 34.297 °, 34.329 °, 36.014
  • the XRPD pattern of the above-mentioned Form A is shown in FIG. 1.
  • Table 1 XRPD pattern analysis data of Form A
  • the differential scanning calorimetry (DSC) of the above-mentioned Form A has a starting point of an endothermic peak at 151.75 ⁇ 3 ° C.
  • the DSC spectrum of the above-mentioned Form A is shown in FIG. 2.
  • thermogravimetric analysis curve (TGA) of the above-mentioned Form A has a weight loss of 0.04540% at 151.57 ⁇ 3 ° C.
  • the TGA spectrum of the above-mentioned Form A is shown in FIG. 3.
  • the invention also provides the application of the above-mentioned Form A in the preparation of a medicine for treating ophthalmic diseases.
  • the compound of the present invention has good permeability and is effectively converted into lanosterol in the body, which greatly improves the drug utilization rate of lanosterol; its crystal form has good stability.
  • the intermediate compounds of the present invention can be prepared by a variety of synthetic methods known to those skilled in the art, including the specific embodiments listed below, the embodiments formed by combining them with other chemical synthesis methods, and those skilled in the art.
  • Well-known equivalent alternatives, preferred embodiments include, but are not limited to, the embodiments of the present invention.
  • the solvent used in the present invention is commercially available.
  • DCM stands for dichloromethane
  • DMF stands for N, N-dimethylformamide
  • DMSO dimethyl sulfoxide
  • EtOH stands for ethanol
  • MeOH stands for methanol
  • TFA trifluoroacetic acid
  • TsOH stands for P-toluenesulfonic acid
  • mp melting point
  • EtSO 3 H for ethanesulfonic acid
  • MeSO 3 H for methanesulfonic acid
  • ATP for adenosine triphosphate
  • HEPES for 4-hydroxyethylpiperazineethanesulfonic acid
  • EGTA for ethylene glycol bis (2 -Aminoethyl ether) tetraacetic acid
  • MgCl 2 stands for magnesium dichloride
  • MnCl 2 stands for manganese dichloride
  • DTT stands for dithiothreitol
  • DCC dicyclohex
  • Light tube voltage 40kV
  • light tube current 40mA
  • Anti-scattering slit 7.10mm
  • Test method Take a sample of 10-20mg in a DVS sample tray for testing.
  • the hygroscopicity evaluation is classified as follows:
  • Hygroscopic classification Hygroscopic weight gain * deliquescence Absorb sufficient water to form a liquid
  • Very hygroscopic Moisture gain is not less than 15%
  • Hygroscopic Moisture gain is less than 15% but not less than 2%
  • Slightly hygroscopic Moisture gain is less than 2% but not less than 0.2%
  • Figure 1 XRPD spectrum of Cu-K ⁇ radiation in Form A.
  • Figure 5 Slit lamp observation of the effect of lanosterol and its prodrug 026 (compound of formula (I)) eye drops on sodium selenite-induced neonatal rabbit cataract model.
  • NC Normal control group
  • MC Model control group
  • PC Positive control group
  • LT Lanosterol eye drops treatment group
  • 026 Lanosterol prodrug 026 eye drops treatment group (Lanosterolprodrug 026 eye drops treatment group).
  • Figure 6 Comparison of in vitro lens transparency test results in different groups of New Zealand rabbit cataract models induced by sodium selenite 42 days after administration.
  • NC Normal control group
  • MC Model control group
  • PC Positive control group
  • LT Lanosterol eye drops treatment group
  • 026 Lanosterol prodrug 026 eye drops treatment group (Lanosterolprodrug 026 eye drops treatment group).
  • Grid is 2.12 ⁇ 2.12mm
  • Figure 7 Comparison of the results of glutathione peroxidase (GSH-PX) activity in the lens of each group of newborn New Zealand rabbit cataract models induced by sodium selenite 42 days after administration.
  • NC Normal control group
  • MC Model control group
  • PC Positive control group
  • LT Lanosterol eye drops treatment group
  • 026 Lanosterol prodrug 026 eye drops treatment group (Lanosterolprodrug 026 eye drops treatment group).
  • V.S PC ++ means p ⁇ 0.01, + means p ⁇ 0.05.
  • Figure 8 Slit lamp observes the effect of lanosterol and its prodrug 026 eye drops on a UV-induced cataract model in New Zealand rabbits.
  • NC Normal control group
  • MC Model control group
  • PC Positive control group
  • LT Lanosterol eye drops treatment group
  • 026 Lanosterol prodrug 026 eye drops treatment group (Lanosterolprodrug 026 eye drops treatment group).
  • Figure 9 Comparison of the results of in vitro lens transparency of the New Zealand rabbit cataract model induced by UV in 42 days after administration.
  • NC Normal control group
  • MC Model control group
  • PC Positive control group
  • LT Lanosterol eye drops treatment group
  • 026 Lanosterol prodrug 026 eye drops treatment group (Lanosterolprodrug 026 eye drops treatment group).
  • the grid is 2.12 ⁇ 2.12mm.
  • Figure 10 Comparison of the results of lens glutathione peroxidase (GSH-PX) activity in the UV-induced cataract model of New Zealand rabbits after 42 days of administration in each group.
  • NC Normal control group
  • MC Model control group
  • PC Positive control group
  • LT Lanosterol eye drops treatment group
  • 026 Lanosterol prodrug 026 eye drops treatment group (Lanosterolprodrug 026 eye drops treatment group).
  • V.S PC ++ means p ⁇ 0.01, + means p ⁇ 0.05.
  • Step 1 Synthesis of compound BB-1.
  • the mixture BB-1-1 was subjected to supercritical fluid chromatography (separation conditions Column: Chiralpak AD-3 150 ⁇ 4.6mm ID, 3 ⁇ m; mobile phase: A: CO 2 B: ethanol (0.05% diethanolamine); gradient: 5 minutes Time B (from 5% to 40%, B 40% for 2.5 minutes, then B 5% for 2.5 minutes; flow rate: 2.5 mL / min; column temperature: 35 ° C; wavelength: 220 nm) was isolated to obtain compound BB-1.
  • Step 1 Synthesis of a compound of formula (I).
  • Substrate 35.2g of compound BB-1 was dissolved in 550mL of anhydrous dichloromethane, 22.8g of salicylic acid was added, followed by 39.2g of DCC, and 23.2g of DMAP was finally added to obtain a white suspension, which was heated to 35 ° C to react 96 hour.
  • TLC showed a small amount of raw material remaining, stopped the reaction, filtered, and washed the filter residue with 150 mL of dichloromethane.
  • the filtrates were combined and spin-dried to obtain a crude residue.
  • 500 mL of methanol was added to the crude residue, and the mixture was refluxed for 16 hours to obtain a white suspension. The temperature was lowered to 15 ° C., and a white solid was obtained by filtration, and XRPD detection was performed to obtain the Form A of the compound of the formula (I).
  • the hygroscopic weight gain of the compound A of the formula (I) at 25 ° C and 80% RH is 0.747%, which is slightly hygroscopic.
  • Test item X includes: appearance, XRPD, content and related substances. Plan 10-day time point analysis;
  • ICH Represents the guiding principles for photostability testing.
  • New Zealand white rabbits (weighing more than 2 kg in weight and more than 12 weeks of age) were used as experimental animals.
  • 2 New Zealand white rabbits were used.
  • 50 ⁇ L of eye drops were added to the left and right eyes of each rabbit. Eyes were used to collect aqueous humor samples and the other eye was used as a spare. Eye drops are formulated with 1.2% hydroxypropyl methylcellulose (E5 size), 20.5% Poloxamer (P407 size), 1.6% Poloxamer (P188 size), the compound is formulated at a concentration of 5mM, and the eye drops It is a homogeneous suspension.
  • anterior aqueous humor was collected at 0.5, 2, 4, and 6 hours after administration, and the volume of each sample did not exceed 50 ⁇ L.
  • Each animal was given a light anesthesia before each sample was collected. The time point is 3 samples.
  • the collected aqueous humor samples are stored in dry ice immediately after collection or stored in a refrigerator at -80 ⁇ 10 °C. After the samples were collected, the animals were euthanized. The samples were analyzed for compound concentration using a triple quadrupole mass spectrometer (API4000).
  • Tables 3 and 4 show the in vivo DMPK analysis and detection methods; Tables 5 and 6 show the concentration of the drug in the aqueous humor after the compound lanosterol (parent drug) and the prodrug compound eye drops (250 nM per eye).
  • Table 4 Liquid phase method gradients of DMPK lanosterol and compounds of formula (I) in vivo
  • Table 5 New Zealand white rabbit drops, the average concentration of samples in aqueous humor (nM) after instilling 250nmol lanosterol into each eye
  • Table 6 New Zealand white rabbit drops The average concentration (nM) of the aqueous humor sample after instilling 250nmol of the compound of formula (I) into each eye
  • Newborn New Zealand rabbits are P7 days old, normal grade, with 5 young rabbits per litter and a mother rabbit suckling.
  • the experimental young rabbits were randomly divided into 5 groups, 5 young rabbits in each group.
  • NC Normal control group
  • Model control group at 10 days, the young rabbit's neck was subcutaneously injected with sodium selenite solution (in physiological saline) at 20 ⁇ mol / kg body weight, and after 15 days of P 15 days, blank drops without drugs were used. Eye drops in the right eye 3 times a day.
  • Positive control group PC: On day 10 of the young rabbits, a subcutaneous injection of sodium selenite solution (in physiological saline) at 20 ⁇ mol / kg body weight was performed. (Japan Tower Pharmaceuticals) Drop the right eye 3 times a day.
  • Lanosterol eye drops treatment group at 10 days, the young rabbit's neck was injected subcutaneously with sodium selenite solution (in physiological saline) at 20 ⁇ mol / kg body weight, and after 15 days, it was used for 42 consecutive days Lanolinol eye drops are applied to the right eye 3 times a day.
  • Lanosterol prodrug 026 eye drops treatment group (Lanosterolprodrug 026 eye drops treatment group, 026, compound of formula (I)): at day 10, the rabbit's neck was injected subcutaneously with sodium selenite solution (in physiological saline) 20 ⁇ mol / kg of body weight, 42 days after P15, right eye with lanosterol prodrug 026 eye drops 3 times a day.
  • Photograph of slit lamp each group of newborn New Zealand rabbits induced by sodium selenite was observed by slit lamp before, 7 days, 14 days, 21 days and 42 days after administration;
  • Glutathione peroxidase (GSH-PX) activity detection refer to the method provided in the instructions of the GSH-PX activity detection kit (Nanjing Jiancheng Biotechnology Research Institute) to detect the isolated rabbit lens GSH-PX activity.
  • the experimental data were analyzed by One-Way ANOVA using SPSS statistical software, and the comparison between groups was performed by LSD method. The statistical difference was p ⁇ 0.05.
  • Figure 6 compares the lens transparency of each group of New Zealand rabbit cataract models induced by sodium selenite after 42 days of administration.
  • the left lens of each picture is the left eye lens (the left eye is not administered, as a self control), and the right lens is the right eye lens (the right eye is administered according to different groups).
  • 42 days after the administration of lanosterol prodrug 026 eye drops the transparency of the lens of the right eye was significantly higher than that of the left eye, and also significantly higher than that of the MC group, but still lower than that of the NC group. There was no significant change in lens transparency in the right eye of the LT group.
  • GSH-PX activity test The lens GSH-PX activity test results of each group after 42 days of administration showed (see Figure 7). After subcutaneous injection of sodium selenite, the GSH-PX activity of rabbit eye lens was significantly reduced, compared with the NC group. Compared with statistically significant (p (0.01). Lanolinol prodrug 026 eye drops and the positive control drug Carlinyo eye drops can increase the lens GSH-PX activity, which is statistically different from the MC group (p ⁇ 0.01), and the improvement effect of 026 is better than carlin It is more obvious (p ⁇ 0.01). The effect of lanosterol eye drops on the activity of lens GSH-PX is obviously inferior to that of 026 and carlin, and there is no statistical difference compared with MC group (p> 0.05).
  • lanosterol prodrug 026 eye drops can alleviate cataract symptoms, increase lens transparency and lens GSH-PX activity in newborn New Zealand rabbits induced by sodium selenite.
  • the experimental animals were randomly divided into 5 groups of 5 animals each.
  • Normal control group Normal control group, NC: Normal feeding and no administration.
  • Models were irradiated with 313 nm ultraviolet rays for 24 hours, and then the right eye was dropped with blank eye drops without drugs 3 times a day for 42 consecutive days.
  • Positive control group (Positive control group, PC): Models were irradiated with 313 nm ultraviolet rays for 24 hours, and then the right eye was dropped with Carinol Eye Drops (Japan Tower Pharmaceuticals) for 42 consecutive days, 3 times a day.
  • Lanosterol eye drops treatment group (LT): Models were irradiated with 313 nm ultraviolet rays for 24 hours, and then the right eye was treated with lanosterol eye drops 3 times a day for 42 consecutive days.
  • Lanosterol prodrug 026 eye drops treatment group (026): 313nm ultraviolet irradiation for 24 hours to build a model, and then for 42 consecutive days with lanosterol prodrug 026 eye drops in the right eye, 3 days a day Times.
  • Photograph of slit lamp each group was observed with slit lamp before the administration of the model, 7 days, 14 days, 21 days and 42 days after the administration;
  • Glutathione peroxidase (GSH-PX) activity detection refer to the method provided in the instructions of the GSH-PX activity detection kit (Nanjing Jiancheng Biotechnology Research Institute) to detect the isolated rabbit lens GSH-PX activity.
  • the experimental data were analyzed by One-Way ANOVA using SPSS statistical software, and the comparison between groups was performed by LSD method. The statistical difference was p ⁇ 0.05.
  • Figure 8 shows that ultraviolet rays can induce cataract in the lens of New Zealand rabbits. Slit lamp observation observed that lanosterol prodrug 026 eye drops were administered for 42 days (Fig. 8-I) and the cataract symptoms were significantly reduced compared with that before administration (Fig. 8-J). However, the changes of cataract symptoms were not obvious before and after administration of carinol eye drops ( Figure 8-E, 8-F) and lanosterol eye drops ( Figure 8-G, 8-H).
  • Figure 9 compares the lens transparency of each group after 42 days of UV-induced cataract model in New Zealand rabbits.
  • the left lens of each picture is the left eye lens (the left eye is not administered, as a self control), and the right lens is the right eye lens (the right eye is administered according to different groups).
  • 42 days after the administration of lanosterol prodrug 026 eye drops the transparency of the lens of the right eye was significantly higher than that of the left eye, and also significantly higher than that of the MC group, but still lower than that of the NC group. There was no significant change in lens transparency in the right eye of the LT group.
  • GSH-PX activity test The lens GSH-PX activity test results of each group after 42 days of administration showed (see Figure 10) that after ultraviolet irradiation, the rabbit eye lens GSH-PX activity was significantly reduced, compared with the NC group. (P ⁇ 0.01 or p ⁇ 0.05).
  • Lanolinol prodrug 026 eye drops and the positive control drug Carlinyo eye drops can increase the lens GSH-PX activity, which is statistically different from the MC group (p ⁇ 0.01), and the improvement effect of 026 is better than carlin More obvious (p (0.05).
  • the effect of lanosterol eye drops on the activity of lens GSH-PX is obviously inferior to that of 026 and carlin, and there is no statistical difference compared with MC group (p> 0.05).
  • lanosterol prodrug 026 eye drops can reduce UV-induced cataract symptoms in New Zealand rabbits, improve lens transparency, and lens GSH-PX activity.

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Abstract

涉及式(Ⅰ)化合物的晶型，以及该晶型在制备治疗眼科疾病药物中的应用。

Description

一种羊毛甾醇前药化合物的晶型及其应用
本申请主张如下优先权：
CN201810826425.6，申请日2018.07.25。
技术领域
本发明涉及式(Ⅰ)化合物的晶型，以及该晶型在制备治疗眼科疾病药物中的应用。
背景技术
白内障属于眼睛的疾病，发生在眼球内的晶状体上，晶状体的浑浊统称为白内障。老化、遗传、代谢异常、外伤、辐射、中毒和局部营养不良等都可引起晶状体囊膜损伤，使其渗透性增加，丧失屏障作用，或导致晶状体代谢紊乱，使晶状体蛋白发生变性，形成混浊。如果眼球的晶状体从透明变成不透明、影响到眼睛接收阳光，那么就会影响眼睛的视力情况。在眼球浑浊较轻时对视力的影响较轻，随着浑浊的程度逐渐加深，视力也会随之加大，严重者会导致失明。白内障是最常见的致盲眼病之一，它是导致失明的主要因素。由于白内障形成的机制尚不明确，药物治疗至今未取得突破性进展。因此，目前唯一确定有效的治疗方法就是手术治疗。
尽管白内障手术方式的不断进步为白内障的治疗提供了巨大的帮助，但手术治疗的治愈率仍然远远低于发生率，存在发生严重并发症的可能；另一方面，白内障的手术治疗成本十分高昂，即使是发达国家，白内障也给医疗保险体系带来了巨大的负担。因此药物的防治起到举足轻重的作用。目前，临床上针对白内障的治疗药物包括：①醛糖还原酶抑制剂，如卡他林(卡他灵、卡林优、白内停)、法可林、苄达赖氨酸等；②抗氧化损伤药物，如谷胱甘肽、牛磺酸、阿司匹林等；③营养代谢类药物，如维生素类、类胡萝卜素等；④中药复方包括石斛夜光丸、杞菊地黄丸、石决明散等。而这些治疗白内障的药物经长期临床试验证实，只能延缓白内障的病情恶化，不能使病情逆转，从而治疗白内障。同时，随着我国开始步入老龄化社会，白内障患者日益增多，对于白内障药物的需求将更为迫切。因此，临床上非常需要安全、疗效好、眼内穿透力强、性质稳定的新品种眼科外用抗白内障药物。
羊毛甾醇是富集于晶状体内的两亲性分子，它是由羊毛甾醇合酶(LSS)在胆固醇合成途径中的一个关键的环化反应来合成，能够降低晶状体蛋白的异常聚集，使其重新规则排列从而恢复晶体透明。已有研究表明，在晶状体中可以检测到羊毛甾醇合酶。此外，在Shumiya白内障大鼠研究中，毛甾醇合酶和法尼基二磷酸法尼基转移酶1(FDFT1)的纯合突变特定组合可以减轻晶状体中胆固醇的水平，并导致白内障。同时我们最近的研究发现，羊毛甾醇在体外和细胞水平可以显著降低预形成的晶状体蛋白质聚集体。在在体水平也证实，羊毛甾醇可以使白内障的病情逆转，晶状体变的澄清透明，此结果已近期发表在Nature杂志，引起全世界广泛注意，是预防和治疗白内障的新分子。
发明内容
本发明提供了式(Ⅰ)化合物的A晶型，其X射线粉末衍射(XRPD)图谱在下列2θ角处具有特 征衍射峰：8.60±0.2°、15.06±0.2°和17.22±0.2°。
Figure PCTCN2019097773-appb-000001
本发明的一些方案中，上述A晶型的X射线粉末衍射图谱在下列2θ角处具有特征衍射峰：8.60±0.2°、9.38±0.2°、10.57±0.2°、12.54±0.2°、14.43±0.2°、15.06±0.2°、17.22±0.2°和25.18±0.2°。
本发明的一些方案中，上述A晶型的X射线粉末衍射图谱在下列2θ角处具有特征衍射峰：4.350°、8.598°、9.383°、10.566°、12.542°、13.448°、14.428°、14.591°、15.063°、15.453°、15.820°、16.803°、17.216°、20.985°、21.181°、22.225°、22.601°、22.856°、23.726°、24.039°、24.534°、25.185°、25.514°、25.935°、26.570°、27.867°、28.125°、28.416°、29.114°、29.445°、31.914°、33.710°、34.297°、34.329°、36.014°、36.108°和38.196°。
本发明的一些方案中，上述A晶型的XRPD图谱如图1所示。
本发明的一些方案中，上述A晶型的XRPD图谱解析数据如表1所示：
表1：A晶型的XRPD图谱解析数据
Figure PCTCN2019097773-appb-000002
本发明的一些方案中，上述A晶型的差示扫描量热曲线(DSC)在151.75±3℃处有一个吸热峰的起始点。
本发明的一些方案中，上述A晶型的DSC图谱如图2所示。
本发明的一些方案中，上述A晶型的热重分析曲线(TGA)在151.57±3℃处失重达0.04540％。
本发明的一些方案中，上述A晶型的TGA图谱如图3所示。
本发明还提供了上述A晶型在制备治疗眼科疾病药物中的应用。
技术效果
作为羊毛甾醇的新型前药，本发明的化合物具有良好的渗透性，并在体内有效的转化为羊毛甾醇，大大提高了羊毛甾醇的药物利用率；其晶型有良好的稳定性。
定义和说明
除非另有说明，本文所用的下列术语和短语旨在含有下列含义。一个特定的短语或术语在没有特别定义的情况下不应该被认为是不确定的或不清楚的，而应该按照普通的含义去理解。当本文出现商品名时，旨在指代其对应的商品或其活性成分。
本发明的中间体化合物可以通过本领域技术人员所熟知的多种合成方法来制备，包括下面列举的具体实施方式、其与其他化学合成方法的结合所形成的实施方式以及本领域技术上人员所熟知的等同替换方式，优选的实施方式包括但不限于本发明的实施例。
本发明具体实施方式的化学反应是在合适的溶剂中完成的，所述的溶剂须适合于本发明的化学变化及其所需的试剂和物料。为了获得本发明的化合物，有时需要本领域技术人员在已有实施方式的基础上对合成步骤或者反应流程进行修改或选择。
下面会通过实施例具体描述本发明，这些实施例并不意味着对本发明的任何限制。
本发明所使用的所有溶剂是市售的，无需进一步纯化即可使用。
本发明所使用的溶剂可经市售获得。本发明采用下述缩略词：DCM代表二氯甲烷；DMF代表N，N-二甲基甲酰胺；DMSO代表二甲亚砜；EtOH代表乙醇；MeOH代表甲醇；TFA代表三氟乙酸；TsOH代表对甲苯磺酸；mp代表熔点；EtSO 3H代表乙磺酸；MeSO 3H代表甲磺酸；ATP代表三磷酸腺苷；HEPES代表4-羟乙基哌嗪乙磺酸；EGTA代表乙二醇双(2-氨基乙基醚)四乙酸；MgCl 2代表二氯化镁；MnCl 2代表二氯化锰；DTT代表二硫苏糖醇；DCC代表二环己基碳二亚胺；DMAP代表4-二甲氨基吡啶；羊毛甾醇前药026代表本发明的式(Ⅰ)化合物。
X射线粉末衍射仪(X-ray powder diffractometer,XRPD)
大约10～20mg样品用于XRPD检测。
详细的XRPD参数如下：
光管：Cu,kα,
Figure PCTCN2019097773-appb-000003
光管电压：40kV，光管电流：40mA
发散狭缝：0.60mm
探测器狭缝：10.50mm
防散射狭缝：7.10mm
扫描范围：4-40deg
步径：0.02deg
步长：0.12秒
样品盘转速：15rpm
动态蒸汽吸附分析(Dynamic Vapor Sorption,DVS)
测试方法：取样品10-20mg样品置于DVS样品盘进行检测。
温度：25℃
平衡dm/dt：0.01％/min：(时间：10min最大180min)
干燥：0％RH，120min
RH(％)测量梯度：10％
RH(％)测量梯度范围：0％～90％～0％
引湿性评价分类如下：
吸湿性分类 吸湿增重*
潮解 吸收足量水分形成液体
极具引湿性 引湿增重不小于15％
有引湿性 引湿增重小于15％但不小于2％
略有引湿性 引湿增重小于2％但不小于0.2％
无或几乎无引湿性 引湿增重小于0.2％
*在25±1℃/80±2％RH下的吸湿增重。
附图说明
图1：A晶型的Cu-Kα辐射的XRPD谱图。
图2：A晶型的DSC谱图。
图3：A晶型的TGA谱图。
图4：A晶型的DVS等温线。
图5：裂隙灯观察羊毛甾醇及其前药026(式(Ⅰ)化合物)滴眼液对亚硒酸钠诱导的新生新西兰兔白内障模型的作用。NC：正常对照组(Normal control group)；MC：模型对照组(Model control group)； PC：阳性对照组(Positive control group)；LT：羊毛甾醇滴眼液处理组(Lanosterol eye drops treatment group)；026：羊毛甾醇前药026滴眼液处理组(Lanosterolprodrug 026 eye drops treatment group)。
图6：亚硒酸钠诱导的新生新西兰兔白内障模型在给药42天后各组体外晶状体透明度检测结果比较。NC：正常对照组(Normal control group)；MC：模型对照组(Model control group)；PC：阳性对照组(Positive control group)；LT：羊毛甾醇滴眼液处理组(Lanosterol eye drops treatment group)；026：羊毛甾醇前药026滴眼液处理组(Lanosterolprodrug 026 eye drops treatment group)。网格为2.12×2.12mm
图7：亚硒酸钠诱导的新生新西兰兔白内障模型在给药42天后各组的晶状体谷胱甘肽过氧化物酶(Glutathione peroxidase,GSH-PX)活性检测结果比较。NC：正常对照组(Normal control group)；MC：模型对照组(Model control group)；PC：阳性对照组(Positive control group)；LT：羊毛甾醇滴眼液处理组(Lanosterol eye drops treatment group)；026：羊毛甾醇前药026滴眼液处理组(Lanosterolprodrug 026 eye drops treatment group)。V.S NC：﹡﹡表示p﹤0.01,﹡表示p﹤0.05；V.S MC：##表示p﹤0.01,#表示p﹤0.05；V.S PC：++表示p﹤0.01,+表示p﹤0.05。
图8：裂隙灯观察羊毛甾醇及其前药026滴眼液对紫外线诱导的新西兰兔白内障模型的作用。NC：正常对照组(Normal control group)；MC：模型对照组(Model control group)；PC：阳性对照组(Positive control group)；LT：羊毛甾醇滴眼液处理组(Lanosterol eye drops treatment group)；026：羊毛甾醇前药026滴眼液处理组(Lanosterolprodrug 026 eye drops treatment group)。
图9：紫外线诱导的新西兰兔白内障模型在给药42天后各组体外晶状体透明度检测结果比较。NC：正常对照组(Normal control group)；MC：模型对照组(Model control group)；PC：阳性对照组(Positive control group)；LT：羊毛甾醇滴眼液处理组(Lanosterol eye drops treatment group)；026：羊毛甾醇前药026滴眼液处理组(Lanosterolprodrug 026 eye drops treatment group)。网格为2.12×2.12mm。
图10：紫外线诱导的新西兰兔白内障模型在给药42天后各组的晶状体谷胱甘肽过氧化物酶(Glutathione peroxidase,GSH-PX)活性检测结果比较。NC：正常对照组(Normal control group)；MC：模型对照组(Model control group)；PC：阳性对照组(Positive control group)；LT：羊毛甾醇滴眼液处理组(Lanosterol eye drops treatment group)；026：羊毛甾醇前药026滴眼液处理组(Lanosterolprodrug 026 eye drops treatment group)。V.S NC：﹡﹡表示p﹤0.01,﹡表示p﹤0.05；V.S MC：##表示p﹤0.01,#表示p﹤0.05；V.S PC：++表示p﹤0.01,+表示p﹤0.05。
具体实施方式
为了更好的理解本发明的内容，下面结合具体实施例来做进一步的说明，但具体的实施方式并不是对本发明的内容所做的限制。
参考例1：片段BB-1
Figure PCTCN2019097773-appb-000004
合成路线：
Figure PCTCN2019097773-appb-000005
步骤1：化合物BB-1的合成。
将混合物BB-1-1经过超临界流体色谱(分离条件Column:Chiralpak AD-3 150×4.6mm I.D.,3μm；流动相:A:CO 2B:乙醇(0.05％二乙醇胺)；梯度:5分钟时间B从5％到40％，B 40％走2.5分钟，然后B 5％走2.5分钟；流速:2.5mL/min；柱温:35℃；波长:220nm)分离得到化合物BB-1。 1H NMR(CDCl 3400MHz):δ＝5.06-5.15(m,1H),5.10(br t,J＝7.2Hz,1H),3.20-3.22(m,1H),3.24(dd,J＝11.5,4.5Hz,1H),1.64-2.09(m,15H),0.77-1.57(m,29H),0.65-0.72ppm(m,3H)。
实施例1：式(Ⅰ)化合物的制备
Figure PCTCN2019097773-appb-000006
合成路线：
Figure PCTCN2019097773-appb-000007
步骤1：式(Ⅰ)化合物的合成.
底物35.2g化合物BB-1溶解在无水二氯甲烷550mL中，加入水杨酸22.8g，随后加入DCC 39.2g，最后加入DMAP 23.2g，得到一白色混悬液，加热至35℃反应96小时。TLC显示原料少量剩余，停止反 应，过滤，用150mL二氯甲烷洗涤滤渣，滤液合并旋干得到粗品残渣。粗品残渣中加入500mL甲醇，回流16小时，得到白色混悬液，降温到15℃，过滤得到白色固体，进行XRPD检测，得到式(Ⅰ)化合物的A晶型。
1H NMR(400MHz,CDCl 3)δ10.89(s,1H),7.77(dd,J＝1.76,8.03Hz,1H),7.38(ddd,J＝1.76,7.09,8.47Hz,1H),6.91(dd,J＝0.88,8.41Hz,1H),6.78-6.85(m,1H),5.00-5.07(m,1H),4.68-4.74(m,1H),1.64-2.04(m,12H),1.62(s,3H),1.57(br s,2H),1.54(s,3H),1.04-1.51(m,9H),0.98(s,3H),0.98(s,3H),0.89(s,3H),0.85(d,J＝6.27Hz,3H),0.82(s,3H),0.63(s,3H)。
实施例2：式(Ⅰ)化合物A晶型的吸湿性研究
实验材料：
SMS DVS Advantage动态蒸汽吸附仪
实验方法：
取式(Ⅰ)化合物A晶型10～15mg置于DVS样品盘内进行测试。
实验结果：
式(Ⅰ)化合物A晶型的DVS谱图如图4所示，△W＝0.747％。
实验结论：
式(Ⅰ)化合物A晶型在25℃和80％RH下的吸湿增重为0.747％，略有吸湿性。
实施例3：式(Ⅰ)化合物的A晶型的预稳定性实验
考察式(Ⅰ)化合物的A晶型在以下3个条件放置并在不同的时间点取样检测性状，XRPD，含量和有关物质。研究条件和检测项目如下表2。
表2：研究条件和检测项目
Figure PCTCN2019097773-appb-000008
注：
*测试项目X包括：外观，XRPD，含量及有关物质。计划10天时间点分析；
ICH：表示光稳定性试验指导原则。
实验步骤：
准确称量式(Ⅰ)化合物的A晶型10mg，置于样品瓶中，摊成薄薄一层。60℃、92.5％RH条件下样品，直接用铝箔纸包好瓶口并在铝箔纸上扎些小孔，保证样品能与环境空气充分接触，分别放置于干燥箱和含有饱和硝酸钾溶液的玻璃缸中。光照样品(敞口，不用铝箔纸覆盖)及光照对照品(敞口，整个样 品瓶用铝箔纸覆盖)放置于光照箱中。每个时间点分别称量2份，作为正式供试样品。另外称取式(Ⅰ)化合物的A晶型大约5mg，用于XRPD测试，样品瓶用铝箔纸包好并扎小孔，同样置于对应的干燥箱和含有饱和硝酸钾溶液的玻璃缸中。
含量的分析方法：
柱子型号：Ultimate XB-C18 3.0*50mm,3um；流动相：0.5ml TFA在1L水中(溶剂A)的和0.4ml TFA在1L乙腈中(溶剂B)，使用洗脱梯度95％-100％(溶剂B)超过2分钟并在100％下保持13分钟。流速为1.5毫升/分钟；柱温：30℃
实验结果及结论：实验后晶型都没发生改变，其稳定性良好。
生物活性测试
实验例一：体内眼部药物渗透以及药物转化为羊毛甾醇的研究
本研究采用新西兰大白兔(体重大于2公斤，周龄大于12周)为实验动物，每一个化合物研究使用2只新西兰大白兔，每只兔子左右两眼均滴入50μL的滴眼液，3只眼睛用于采集房水样品，另外1只眼睛作为备用。滴眼液配方为1.2％羟丙基甲基纤维素(E5规格)，20.5％波洛沙姆(P407规格)，1.6％波洛沙姆(P188规格)，化合物配制浓度为5mM，滴眼液为均一混悬液。滴眼液滴入兔子眼睛后，分别于给药后0.5、2、4和6小时采集前房水，每次采样品体积不超过50μL，每次动物采集样品前将给予轻度麻醉，每个时间点为3个样品。采集的房水样品在采集后即刻放入干冰保存或者于-80±10℃冰箱进行保存。样品采集结束后，动物进行安乐死。样品使用三重四极杆质谱仪(API4000)分析化合物浓度。表3及表4表示了体内DMPK分析检测方法；表5和表6展示了化合物羊毛甾醇(母药)以及前药化合物滴眼(250nM每只眼睛)后房水中的药物浓度。
实验结果及结论：羊毛甾醇本身及其本发明前药化合物均能够从角膜或者通过其他途径渗透入房水；并且前药化合物能够在渗透过程中转化为母药羊毛甾醇，并在房水中表现出更高的羊毛甾醇浓度和暴露量。
表3：体内DMPK分析检测方法
Figure PCTCN2019097773-appb-000009
Figure PCTCN2019097773-appb-000010
表4：体内DMPK羊毛甾醇以及式(Ⅰ)化合物液相方法梯度
Figure PCTCN2019097773-appb-000011
表5：新西兰大白兔滴每只眼睛眼液滴入250nmol羊毛甾醇后，房水中的样品平均浓度(nM)
滴眼液化合物名称 羊毛甾醇(母药)
测试化合物名称 羊毛甾醇(母药)
时间(小时) 平均浓度(nM)
0.5 106*
2 496
4 300
6 225
AUC(nM.h) 1779
*BQL:低于检测限,AUC：暴露量。
表6：新西兰大白兔滴每只眼睛眼液滴入250nmol式(Ⅰ)化合物后，房水中的样品平均浓度(nM)
Figure PCTCN2019097773-appb-000012
*BQL:低于检测限，AUC：暴露量。
实验例二：羊毛甾醇滴眼液及其前药对亚硒酸钠诱导的新生新西兰兔白内障模型的药效学研究
1、实验动物
新生新西兰兔P7天龄，普通级，每窝5只幼兔配一只妈妈兔哺乳。
2、分组及处理
实验幼兔随机分为5组，每组5只幼兔。
1)正常对照组(Normal control group,NC)：P10天时幼兔颈部皮下注射生理盐水0.25ml，P15天后不给药。
2)模型对照组(Model control group,MC)：P10天时幼兔颈部皮下注射亚硒酸钠溶液(于生理盐水中)20μmol/kg体重，P15天之后连续42天用不含药物的空白滴眼液滴右眼，每天3次。
3)阳性对照组(Positive control group,PC)：P10天时幼兔颈部皮下注射亚硒酸钠溶液(于生理盐水中)20μmol/kg体重，P15天之后连续42天用卡林优滴眼液(日本参天制药)滴右眼，每天3次。
4)羊毛甾醇滴眼液处理组(Lanosterol eye drops treatment group,LT)：P10天时幼兔颈部皮下注射亚硒酸钠溶液(于生理盐水中)20μmol/kg体重，P15天之后连续42天用羊毛甾醇滴眼液滴右眼，每天3次。
5)羊毛甾醇前药026滴眼液处理组(Lanosterolprodrug 026 eye drops treatment group,026,式(Ⅰ)化合物)：P10天时幼兔颈部皮下注射亚硒酸钠溶液(于生理盐水中)20μmol/kg体重，P15天之后连续42天用羊毛甾醇前药026滴眼液滴右眼，每天3次。
3、实验检测
1)裂隙灯照像：亚硒酸钠诱导的新生新西兰兔每组分别于给药前、给药后7天、14天、21天和42天进行裂隙灯观察；
2)体外晶状体透明度检测：最后一天，解剖出动物眼球，完整分离出包含囊膜的晶状体，将晶状体至于方格纸上(2.12×2.12mm)，拍照显示透过晶状体所拍摄的方格的清晰度。
3)谷胱甘肽过氧化物酶(Glutathione peroxidase,GSH-PX)活性检测：参照GSH-PX活性检测试剂盒(南京建成生物工程研究所)说明书提供的方法检测分离出的各组兔晶状体的GSH-PX活性。实验数据用SPSS统计软件进行One-Way ANOVA分析，用LSD法做各组间比较，统学计差异水平为p﹤0.05。
4、实验结果
1)裂隙灯观察：图5显示亚硒酸钠可诱导新生新西兰兔晶状体发生白内障。裂隙灯观察羊毛甾醇前药026滴眼液给药42天(图5-I)与给药前(图5-J)相比较白内障症状明显减轻。而卡林优滴眼液给药前后(图5-E、5-F)和羊毛甾醇滴眼液给药前后(图5-G、5-H)白内障症状变化不明显。
2)体外晶状体透明度检测：图6比较了亚硒酸钠诱导的新生新西兰兔白内障模型在给药42天后各组的晶状体透明度。每张照片左侧的是左眼晶状体(左眼未给药，作为自身对照)，右侧的是右眼晶状体(右眼按不同分组相应给药)。羊毛甾醇前药026滴眼液给药42天后右眼晶状体透明度明显高于自身左眼的透明度，也明显高于MC组晶状体的透明度，但尚低于NC组的透明度。LT组右眼给药后晶状体透明度未见明显改变。
3)GSH-PX活性检测：给药42天后各组的晶状体GSH-PX活性检测结果显示(见图7)，亚硒酸钠皮下注射后，兔眼晶状体GSH-PX活性显著降低，与NC组相比具有统计学差异(p﹤0.01)。羊毛甾醇前药026滴眼液和阳性对照药卡林优滴眼液可提高晶状体GSH-PX活性，与MC组相比具有统计学差异(p﹤0.01)，且026的提高效果比卡林优更明显(p﹤0.01)。羊毛甾醇滴眼液对晶状体GSH-PX活性的影响明显不如026和卡林优，与MC组相比没有统计学差异(p＞0.05)。
5、结论
上述结果提示，羊毛甾醇前药026滴眼液可减轻亚硒酸钠诱导的新生新西兰兔白内障症状、提高晶状体透明度以及晶状体GSH-PX活性。
实验三、羊毛甾醇滴眼液及其前药对紫外线诱导的新西兰兔白内障模型的药效学研究
1、实验动物
成年新西兰兔2.0-2.5kg，普通级，雌雄不拘，共25只。
2、分组及处理
实验动物随机分为5组，每组5只。
1)正常对照组(Normal control group,NC)：正常饲养，不给药。
2)模型对照组(Model control group,MC)：313nm紫外线照射24小时造模，之后连续42天用不含药物的空白滴眼液滴右眼，每天3次。
3)阳性对照组(Positive control group,PC)：313nm紫外线照射24小时造模，之后连续42天用卡林优滴眼液(日本参天制药)滴右眼，每天3次。
4)羊毛甾醇滴眼液处理组(Lanosterol eye drops treatment group,LT)：313nm紫外线照射24小时造模，之后连续42天用羊毛甾醇滴眼液滴右眼，每天3次。
5)羊毛甾醇前药026滴眼液处理组(Lanosterolprodrug 026 eye drops treatment group,026)：313nm紫外线照射24小时造模，之后连续42天用羊毛甾醇前药026滴眼液滴右眼，每天3次。
3、实验检测
1)裂隙灯照像：每组分别于造模后给药前、给药后7天、14天、21天和42天进行裂隙灯观察；
2)体外晶状体透明度检测：最后一天，解剖出动物眼球，完整分离出包含囊膜的晶状体，将晶状体至 于方格纸上(2.12×2.12mm)，拍照显示透过晶状体所拍摄的方格的清晰度。
3)谷胱甘肽过氧化物酶(Glutathione peroxidase,GSH-PX)活性检测：参照GSH-PX活性检测试剂盒(南京建成生物工程研究所)说明书提供的方法检测分离出的各组兔晶状体的GSH-PX活性。实验数据用SPSS统计软件进行One-Way ANOVA分析，用LSD法做各组间比较，统学计差异水平为p﹤0.05。
4、实验结果
1)裂隙灯观察：图8显示紫外线可诱导新西兰兔晶状体发生白内障。裂隙灯观察羊毛甾醇前药026滴眼液给药42天(图8-I)与给药前(图8-J)相比较白内障症状明显减轻。而卡林优滴眼液给药前后(图8-E、8-F)和羊毛甾醇滴眼液给药前后(图8-G、8-H)白内障症状变化不明显。
2)体外晶状体透明度检测：图9比较了紫外线诱导的新西兰兔白内障模型在给药42天后各组的晶状体透明度。每张照片左侧的是左眼晶状体(左眼未给药，作为自身对照)，右侧的是右眼晶状体(右眼按不同分组相应给药)。羊毛甾醇前药026滴眼液给药42天后右眼晶状体透明度明显高于自身左眼的透明度，也明显高于MC组晶状体的透明度，但尚低于NC组的透明度。LT组右眼给药后晶状体透明度未见明显改变。
3)GSH-PX活性检测：给药42天后各组的晶状体GSH-PX活性检测结果显示(见图10)，紫外线照射后，兔眼晶状体GSH-PX活性显著降低，与NC组相比具有统计学差异(p﹤0.01或p﹤0.05)。羊毛甾醇前药026滴眼液和阳性对照药卡林优滴眼液可提高晶状体GSH-PX活性，与MC组相比具有统计学差异(p﹤0.01)，且026的提高效果比卡林优更明显(p﹤0.05)。羊毛甾醇滴眼液对晶状体GSH-PX活性的影响明显不如026和卡林优，与MC组相比没有统计学差异(p＞0.05)。
5、结论
上述结果提示，羊毛甾醇前药026滴眼液可减轻紫外线诱导的新西兰兔白内障症状、提高晶状体透明度以及晶状体GSH-PX活性。

Claims (9)

  1. 式(Ⅰ)化合物的A晶型，其X射线粉末衍射(XRPD)图谱在下列2θ角处具有特征衍射峰：8.60±0.2°、15.06±0.2°和17.22±0.2°。
    Figure PCTCN2019097773-appb-100001
  2. 根据权利要求1所述的A晶型，其X射线粉末衍射图谱在下列2θ角处具有特征衍射峰：8.60±0.2°、9.38±0.2°、10.57±0.2°、12.54±0.2°、14.43±0.2°、15.06±0.2°、17.22±0.2°和25.18±0.2°。
  3. 权利要求2所述的A晶型，其X射线粉末衍射图谱在下列2θ角处具有特征衍射峰：4.350°、8.598°、9.383°、10.566°、12.542°、13.448°、14.428°、14.591°、15.063°、15.453°、15.820°、16.803°、17.216°、20.985°、21.181°、22.225°、22.601°、22.856°、23.726°、24.039°、24.534°、25.185°、25.514°、25.935°、26.570°、27.867°、28.125°、28.416°、29.114°、29.445°、31.914°、33.710°、34.297°、34.329°、36.014°、36.108°和38.196°。
  4. 根据权利要求3所述的A晶型，其XRPD图谱如图1所示。
  5. 根据权利要求1～4任意一项所述的A晶型，其差示扫描量热曲线(DSC)在151.75±3℃处有一个吸热峰的起始点。
  6. 根据权利要求5所述的A晶型，其DSC图谱如图2所示。
  7. 根据权利要求1～4任意一项所述的A晶型，其热重分析曲线(TGA)在151.57±3℃处失重达0.04540％。
  8. 根据权利要求7所述的A晶型，其TGA图谱如图3所示。
  9. 根据权利要求1～8任意一项所述的A晶型在制备治疗眼科疾病药物中的应用。
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Cited By (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2020177714A1 (zh) * 2019-03-04 2020-09-10 中山大学中山眼科中心 一种羊毛甾醇前药化合物的组合物及其制备方法和应用
WO2023020535A1 (zh) * 2021-08-18 2023-02-23 广州润尔眼科生物科技有限公司 甾体化合物在制备预防和/或治疗飞蚊症的药物中的应用
WO2023020536A1 (zh) * 2021-08-18 2023-02-23 广州润尔眼科生物科技有限公司 一种药物组合物及其制备方法和应用
WO2023143452A1 (zh) * 2022-01-28 2023-08-03 广州润尔眼科生物科技有限公司 甾体化合物在预防和/或治疗白内障中的应用
WO2023147640A1 (en) * 2022-02-07 2023-08-10 GERVÁSIO ALVES DA SILVA, Társis Preparation of novel triterpene alcohol derivatives with enhanced bioavailability for cancer, inflammation and pain treatment

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PH12021550182A1 (en) 2022-02-21
CA3107361C (en) 2022-03-22
ZA202100923B (en) 2021-10-27
MA53397A (fr) 2021-06-02
BR112021001172A2 (pt) 2021-04-27
CN112543762A (zh) 2021-03-23
US20210246158A1 (en) 2021-08-12
EP3828193A1 (en) 2021-06-02
MX2021000900A (es) 2021-06-15
CO2021000932A2 (es) 2021-02-17
CA3107361A1 (en) 2020-01-30
KR20210038596A (ko) 2021-04-07
JP2021524499A (ja) 2021-09-13
RU2766088C1 (ru) 2022-02-07
AU2019308940A1 (en) 2021-03-11
CN112543762B (zh) 2022-11-08
US11149055B2 (en) 2021-10-19
AU2019308940B2 (en) 2024-02-08
EP3828193A4 (en) 2022-06-29
SG11202100665WA (en) 2021-02-25
JP7046266B2 (ja) 2022-04-01

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