WO2022217719A1 - 一种促进细胞运动的化合物、包含其的药物组合物及其制备和应用 - Google Patents

一种促进细胞运动的化合物、包含其的药物组合物及其制备和应用 Download PDF

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WO2022217719A1
WO2022217719A1 PCT/CN2021/097808 CN2021097808W WO2022217719A1 WO 2022217719 A1 WO2022217719 A1 WO 2022217719A1 CN 2021097808 W CN2021097808 W CN 2021097808W WO 2022217719 A1 WO2022217719 A1 WO 2022217719A1
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isobutoxymethyl
quinolin
compound
pharmaceutical composition
hydrochloride
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PCT/CN2021/097808
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English (en)
French (fr)
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钱朝南
李长志
陈金东
周红娟
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广州市朝利良生物科技有限公司
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    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D215/00—Heterocyclic compounds containing quinoline or hydrogenated quinoline ring systems
    • C07D215/02—Heterocyclic compounds containing quinoline or hydrogenated quinoline ring systems having no bond between the ring nitrogen atom and a non-ring member or having only hydrogen atoms or carbon atoms directly attached to the ring nitrogen atom
    • C07D215/16—Heterocyclic compounds containing quinoline or hydrogenated quinoline ring systems having no bond between the ring nitrogen atom and a non-ring member or having only hydrogen atoms or carbon atoms directly attached to the ring nitrogen atom with hetero atoms or with carbon atoms having three bonds to hetero atoms with at the most one bond to halogen, e.g. ester or nitrile radicals, directly attached to ring carbon atoms
    • C07D215/20—Oxygen atoms
    • C07D215/24—Oxygen atoms attached in position 8
    • C07D215/26—Alcohols; Ethers thereof
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P17/00—Drugs for dermatological disorders
    • A61P17/02—Drugs for dermatological disorders for treating wounds, ulcers, burns, scars, keloids, or the like

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  • the application belongs to the field of medicinal chemistry, and specifically relates to a compound for promoting cell movement, a pharmaceutical composition containing the same, and its preparation and application.
  • Cell motility is critical in embryonic development, tumor metastasis, wound healing, and immune responses. It has been reported that living cells respond to biochemical signals to move to high concentrations, and cells also have mechanical forces in response to The same molecular network responds to chemical and mechanical signals, combining potentially conflicting signals into unified pathways.
  • 8-Hydroxyquinoline compounds are common intermediates in organic chemical synthesis, which are widely used in medicine, materials, chemical industry and other fields. 8-Hydroxyquinoline and its derivatives are commonly used in the determination and separation of metals, and are also used as pharmaceutical intermediates.
  • the present application provides a compound for promoting cell movement and its preparation and application.
  • the compound can be used to prepare medicines for promoting wound healing.
  • the drug does not cause the spread of tumor cells while accelerating wound healing in patients.
  • the present application provides a compound that promotes the movement of fibroblasts and does not promote the movement of tumor cells, and the compound is 5-(isobutoxymethyl)quinolin-8-ol or 5-(isobutyryl) Oxymethyl) quinolin-8-ol hydrochloride, wherein, the chemical structural formula of 5-(isobutoxymethyl) quinolin-8-ol is shown in formula G06, 5-(isobutoxymethyl) The chemical structural formula of quinolin-8-ol hydrochloride is shown in formula G07:
  • the present application also provides a preparation method of the compound 5-(isobutoxymethyl)quinolin-8-ol hydrochloride, and the preparation steps are as follows:
  • step 1 All the 5-chloromethylquinolin-8-ol obtained in step 1 was added to anhydrous isobutanol, and the consumption of anhydrous isobutanol was 15 mL/g relative to 5-chloromethylquinolin-8-ol, and Add 10% catalyst potassium iodide of 5-chloromethylquinolin-8-ol molar equivalent, under nitrogen protection atmosphere, heat up and reflux for overnight reaction, after the reaction is completed, cool down to room temperature, adjust the pH value with saturated potassium carbonate solution to 6-7, then extracted twice with dichloromethane, the organic layer was dried over anhydrous sodium sulfate, concentrated to dryness to obtain a crude product, and the obtained crude product was separated and purified by column chromatography to obtain 5-(isobutoxymethyl)quinoline -8-ol;
  • the 5-(isobutoxymethyl)quinolin-8-ol obtained in step 2 was dissolved in anhydrous methanol, and cooled to 0°C, and concentrated hydrochloric acid was slowly added dropwise under nitrogen protection to maintain the pH value of the reaction system at 2 After the reaction was completed, methanol and excess concentrated hydrochloric acid were removed by vacuum distillation to obtain a yellow-brown solid crude product.
  • the obtained crude product was stirred and reacted in ethyl acetate and petroleum ether system for 8 hours, filtered, and filtered. The cake was washed with ethyl acetate and petroleum ether mixture until neutral, and the filter cake was vacuum-dried to obtain the pure product 5-(isobutoxymethyl)quinolin-8-ol hydrochloride.
  • the present application also provides a pharmaceutical composition comprising the compound of formula G07 or a pharmaceutically acceptable salt thereof.
  • the pharmaceutical composition is liquid, paste, gel, dressing or spray.
  • composition further comprises one or more pharmaceutically acceptable carriers or pharmaceutical excipients.
  • the present application further provides the use of the pharmaceutical composition in the preparation of a medicament for promoting wound healing.
  • the present application also provides an application of the compound or pharmaceutical composition in the preparation of a medicament for promoting wound healing in surgical patients with tumors complicated by diabetes.
  • the compound 5-(isobutoxymethyl)quinolin-8-ol hydrochloride provided in this application has a remarkable effect on the healing of skin wounds in mice, and can be used to prepare medicines for promoting wound healing. It does not cause the spread of tumor cells while healing, and has extremely high medicinal potential.
  • Fig. 1 is the synthetic route of the compound 5-(isobutoxymethyl)quinolin-8-ol hydrochloride of the application.
  • Figure 2 is the nuclear magnetic spectrum of the compound 5-(isobutoxymethyl)quinolin-8-ol hydrochloride of the present application.
  • Figure 3 shows the effect of the compound 5-(isobutoxymethyl)quinolin-8-ol hydrochloride of the present application on the survival of NIH/3T3 cells and the compound 5-(isobutoxymethyl)quinolin-8- Schematic representation of the results of the alcohol hydrochloride IC50 assay.
  • FIG. 4 is a graph showing the effect of the compound 5-(isobutoxymethyl)quinolin-8-ol of the present application in promoting NIH/3T3 sports scratching experiments.
  • FIG. 5 is a graph showing the effect of the compound 5-(isobutoxymethyl)quinolin-8-ol of the present application on the inhibition of scratching of nasopharyngeal carcinoma cells CNE2.
  • Figure 6 is a graph showing the effect of the compound 5-(isobutoxymethyl)quinolin-8-ol hydrochloride of the present application in promoting NIH/3T3 sports scratching experiment.
  • FIG. 7 is a graph showing the effect of the compound 5-(isobutoxymethyl)quinolin-8-ol hydrochloride of the present application on the inhibition of scratching of nasopharyngeal carcinoma cells CNE2.
  • FIG. 8 is a graph showing the effect of the compound 5-(isobutoxymethyl)quinolin-8-ol of the present application on the promotion of NIH/3T3 movement and migration experiments.
  • FIG. 9 is a graph showing the effect of the compound 5-(isobutoxymethyl)quinolin-8-ol of the present application in inhibiting the migration of nasopharyngeal carcinoma cells CNE2.
  • FIG. 10 is a graph showing the effect of the compound 5-(isobutoxymethyl)quinolin-8-ol hydrochloride of the present application on the promotion of NIH/3T3 movement and migration experiments.
  • FIG. 11 is a graph showing the effect of the compound 5-(isobutoxymethyl)quinolin-8-ol hydrochloride of the present application in inhibiting the migration of nasopharyngeal carcinoma cells CNE2.
  • Figure 12 is a graph showing the experimental effect of the compound 5-(isobutoxymethyl)quinolin-8-ol hydrochloride of the present application on the healing of skin wounds in normoglycemic rats.
  • Figure 13 shows the effect of the compound 5-(isobutoxymethyl)quinolin-8-ol hydrochloride of the present application on the skin wound healing rate of normoglycemic rats.
  • Figure 14 is a graph showing the experimental effect of the compound 5-(isobutoxymethyl)quinolin-8-ol hydrochloride of the present application on the healing of skin wounds in diabetic mice.
  • Figure 15 shows the effect of the compound 5-(isobutoxymethyl)quinolin-8-ol hydrochloride of the present application on the wound healing rate of diabetic rat skin.
  • the reagents used in this application are all commonly used reagents, which can be purchased from conventional reagent production and sales companies.
  • step 1 All the 5-chloromethylquinolin-8-ol obtained in step 1 was added to anhydrous isobutanol, and the consumption of anhydrous isobutanol was 15 mL/g relative to 5-chloromethylquinolin-8-ol, and Add 10% catalyst potassium iodide of 5-chloromethylquinolin-8-ol molar equivalent, under nitrogen protection atmosphere, heat up and reflux for overnight reaction, after the reaction is completed, cool down to room temperature, adjust the pH value with saturated potassium carbonate solution to 7, and then extracted twice with dichloromethane. The organic layer was dried over anhydrous sodium sulfate and concentrated to dryness to obtain a crude product. The obtained crude product was separated and purified by column chromatography to obtain 5-(isobutoxymethyl)quinoline-8 -alcohol;
  • the 5-(isobutoxymethyl)quinolin-8-ol obtained in step 2 was dissolved in anhydrous methanol, and cooled to 0° C. Concentrated hydrochloric acid was slowly added dropwise under nitrogen protection, and the pH value of the reaction system was maintained at 3 , and reacted for 8 hours. After the reaction was completed, methanol and excess concentrated hydrochloric acid were removed by vacuum distillation to obtain a yellow-brown solid crude product. The obtained crude product was stirred and reacted in ethyl acetate and petroleum ether system for 8 hours, filtered, and the filter cake was washed with acetic acid. The mixed solution of ethyl ester and petroleum ether was washed until neutral, and the filter cake was vacuum-dried to obtain the pure product 5-(isobutoxymethyl)quinolin-8-ol hydrochloride.
  • the yield of the 5-chloromethylquinolin-8-ol prepared by the above preparation method is 52%; the yield of the prepared 5-(isobutoxymethyl)quinolin-8-ol is 65%, and the prepared 5-(isobutoxymethyl)quinolin-8-ol hydrochloride has a yield of 95%.
  • Test example 1 Compound cytotoxicity detection (MTS test)
  • the results suggest that the compounds of the present application exhibit dose-dependent cytotoxicity to the above cell lines, and normal cell lines are more sensitive than tumor cell lines. Based on the IC50 results, we used 1 ⁇ g/mL as the highest concentration in subsequent functional experiments to ensure that the compound does not produce obvious cytotoxicity.
  • Alcohol hydrochloride can accelerate NIH/3T3 movement, and the promoting effect increases with the concentration, showing a dose-dependent manner; at the same time, G06 significantly inhibits CNE2 migration of nasopharyngeal carcinoma cells at various concentrations, and G07 can significantly inhibit CNE2 movement at high concentration .
  • mice 28 4-week-old male C57BL/6 mice were randomly selected into the modeling group, and the remaining 14 mice were in the non-modeling group. Measure the basal body weight to confirm that the body weights of mice within and between groups are similar;
  • mice in the model group started to eat high-fat feed as the first day of the experiment, while the mice in the non-model group ate ordinary feed. Record the body weight every week, observe the hair, eating and urination status, remove the feed at night on the 28th day, and fast without food and water overnight;
  • STZ streptozotocin
  • mice in the model group were injected with STZ solution (40 mg/kg/d) intraperitoneally, once a day for 5 consecutive days. High-fat feeding was resumed on the second day; mice in the non-modeling group were intraperitoneally injected with an equal volume of double-distilled water, once a day, for 5 consecutive days. Fasting was continued on the first day of injection, and normal feeding was resumed on the second day of injection. Continue to observe the mice eating, drinking and urinating;
  • mice in the modeling group the mice that stabilized at or above 11.1 mmoL/L for more than 3 consecutive times and appeared "polydipsia, polyuria, polyphagia, and weight loss" were defined as successful diabetic mice and included in the follow-up experiments. , the mice that did not meet the model standards were withdrawn from the experiment, and the success rate of modeling was calculated;
  • mice In the non-modeling group mice, the mice whose stability did not exceed 11.1 mmoL/L for more than 3 consecutive times, and did not show "polydipsia, polyuria, polyphagia, and weight loss" were included in the follow-up experiments. The rat withdrew from the experiment.
  • the successfully modeled diabetes group was randomly divided into a negative control group, a low-dose group, a medium-dose group and a high-dose group, with no less than 3 mice in each group; the mice in the normal blood sugar group were also randomly divided into Negative control group, low-dose group, middle-dose group and high-dose group, each group has no less than 3 mice;
  • mice in each group were moved to the operating room, and gas anesthesia was performed using a gas anesthesia device and isoflurane;
  • mice in each group were determined.
  • the negative control group was given an equal volume of PBS (solvent group)
  • the low-dose group was 3 mg/kg
  • the middle-dose group was 10 mg/kg
  • the high-dose group was given an equal volume of PBS (solvent group).
  • the dose group was 30 mg/kg.
  • the mode of administration is intraperitoneal injection.
  • the day of mouse wounding was daG07 1, and the mice in each group were injected with compound/PBS on daG07 1, 3, 5, 7, and 9, respectively.

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Abstract

本申请属于药物化学领域，具体涉及一种促进细胞运动的化合物、药物组合物及其制备和应用。本申请提供的化合物为5-(异丁氧基甲基)喹啉-8-醇或5-(异丁氧基甲基)喹啉-8-醇盐酸盐，经试验证实，该化合物可以促进NIH/3T3运动的同时，抑制鼻咽癌细胞CNE2运动，并且具有良好的安全性，对小鼠皮肤伤口愈合效果显著，能够应用于制备促进伤口愈合的药物，尤其适用于肿瘤合并糖尿病的手术患者，可以加速患者伤口愈合而不造成肿瘤细胞扩散，具有极高的药用潜力。

Description

一种促进细胞运动的化合物、包含其的药物组合物及其制备和应用 技术领域
本申请属于药物化学领域，具体涉及一种促进细胞运动的化合物、包含其的药物组合物及其制备和应用。
背景技术
细胞运动在胚胎发育、肿瘤转移、伤口愈合和免疫应答中至关重要，有报道称：活细胞会对生化信号做出应答，向高浓度的地方运动，并且细胞也存在应答机械力，细胞使用同样的分子网络对化学和机械信号做出反应，将可能相互冲突的信号结合起来形成统一的路径。
8-羟基喹啉类化合物是常见的有机化学合成中间体，在医药、材料、化工等领域均有广泛的应用。8-羟基喹啉及其衍生物，常用于金属的测定和分离，还用作医药中间体，作为合成克泻痢宁、氯碘喹啉、扑喘息敏的原料，也是染料、农药中间体。
现有的促进伤口愈合的药物，在促进伤口愈合的同时，也存在有肿瘤扩散的风险，特别是对于肿瘤合并糖尿病患者，有必要提供一种可以加速患者创口愈合而不造成肿瘤细胞扩散的药物。
发明内容
本申请提供了一种促进细胞运动的化合物及其制备和应用。该化合物可用于制备促进伤口愈合的药物。特别是该药物在加速患者创口愈合的同时不会造成肿瘤细胞的扩散。
第一方面，本申请提供了一种促进成纤维细胞运动、不促进肿瘤细胞运动的化合物，所述化合物为5-(异丁氧基甲基)喹啉-8-醇或5-(异丁氧基甲基)喹啉-8-醇盐酸盐，其中，5-(异丁氧基甲基)喹啉-8-醇的化学结构式如式G06所示，5-(异丁氧基甲基)喹啉-8-醇盐酸盐的化学结构式如式G07所示：
Figure PCTCN2021097808-appb-000001
第二方面，本申请还提供了所述化合物5-(异丁氧基甲基)喹啉-8-醇盐酸盐的制备方法，其制备步骤如下：
步骤1：合成5-氯甲基喹啉-8-醇
室温下，将1eq 8-羟基喹啉和20eq多聚甲醛加入到的浓盐酸中，浓盐酸用量相对8-羟基喹啉为20mL/g，氮气保护氛围下，升温到60℃，直至反应结束，待反应完成后，降温至室温，用饱和碳酸钾溶液调节pH值至6～7，用乙酸乙酯萃取两次，用无水硫酸钠干燥有机相，随后将有机相浓缩至干，得粗品，所得粗品经柱层析分离纯化，得5-氯甲基喹啉-8-醇；所述多聚甲醛聚合度为8～100；
步骤2：合成5-(异丁氧基甲基)喹啉-8-醇
将步骤1所得的5-氯甲基喹啉-8-醇全部加入到无水异丁醇中，无水异丁醇用量相对5-氯甲基喹啉-8-醇为15mL/g，并加入5-氯甲基喹啉-8-醇摩尔当量的10％的催化剂碘化钾，在氮气保护氛围下，升温回流反应过夜，待反应完成之后，降温至室温，用饱和碳酸钾溶液调节pH值至6～7，再用二氯甲烷萃取两次，有机层经无水硫酸钠干燥，浓缩至干，得粗品，所得粗品柱析层分离纯化，得5-(异丁氧基甲基)喹啉-8-醇；
步骤3：合成5-(异丁氧基甲基)喹啉-8-醇盐酸盐
将步骤2所得的5-(异丁氧基甲基)喹啉-8-醇溶于无水甲醇，并降温至0℃，氮气保护条件下缓慢滴入浓盐酸，维持反应体系pH值在2～4之间反应8小时，待反应完成之后，真空减压蒸馏去除甲醇和过量的浓盐酸，得到黄棕色固体粗品，所得粗品于乙酸乙酯和石油醚体系中搅拌反应8小时，过滤，滤饼用乙酸乙酯和石油醚混合液洗涤至中性，滤饼真空干燥，得纯产品5-(异丁氧基甲基)喹啉-8-醇盐酸盐。
第三方面，本申请还提供了一种药物组合物，所述药物组合物包含如式G07 所述的化合物或其药学上可接受的盐。
进一步地，所述药物组合物为液体、膏状体、凝胶、敷料或喷雾。
进一步地，所述药物组合物进一步包含一种或多种药学上可接受的载体或药用辅料。
第四方面，本申请进一步提供了所述的药物组合物在制备用于促进伤口愈合的药物中的应用。
第五方面，本申请还提供了所述的化合物或药物组合物在制备用于促进肿瘤合并糖尿病的手术患者伤口愈合的药物中的应用。
与现有技术相比，本申请具有以下优势：
(1)本申请首次获得5-(异丁氧基甲基)喹啉-8-醇及5-(异丁氧基甲基)喹啉-8-醇盐酸盐，5-(异丁氧基甲基)喹啉-8-醇盐酸盐在其安全剂量下可有效促进细胞运动，加快伤口愈合。
(2)本申请提供的化合物5-(异丁氧基甲基)喹啉-8-醇盐酸盐经实验证实可在促进NIH/3T3运动同时抑制鼻咽癌细胞CNE2运动，并且具有良好的安全性。
(3)本申请提供的化合物5-(异丁氧基甲基)喹啉-8-醇盐酸盐对小鼠皮肤伤口愈合效果显著，能够用于制备促进伤口愈合的药物，在加速患者创口愈合的同时还不会造成肿瘤细胞的扩散，具有极高的药用潜力。
附图说明
图1为本申请化合物5-(异丁氧基甲基)喹啉-8-醇盐酸盐的合成路线。
图2为本申请化合物5-(异丁氧基甲基)喹啉-8-醇盐酸盐的核磁谱图。
图3为本申请化合物5-(异丁氧基甲基)喹啉-8-醇盐酸盐对NIH/3T3细胞存活的影响及化合物5-(异丁氧基甲基)喹啉-8-醇盐酸盐IC50测定的结果示意图。
图4为本申请化合物5-(异丁氧基甲基)喹啉-8-醇促进NIH/3T3运动划痕实验效果图。
图5为本申请化合物5-(异丁氧基甲基)喹啉-8-醇抑制鼻咽癌细胞CNE2划痕实验效果图。
图6为本申请化合物5-(异丁氧基甲基)喹啉-8-醇盐酸盐促进NIH/3T3运 动划痕实验效果图。
图7为本申请化合物5-(异丁氧基甲基)喹啉-8-醇盐酸盐抑制鼻咽癌细胞CNE2划痕实验效果图。
图8为本申请化合物5-(异丁氧基甲基)喹啉-8-醇促进NIH/3T3运动迁移实验效果图。
图9为本申请化合物5-(异丁氧基甲基)喹啉-8-醇抑制鼻咽癌细胞CNE2迁移实验效果图。
图10为本申请化合物5-(异丁氧基甲基)喹啉-8-醇盐酸盐促进NIH/3T3运动迁移实验效果图。
图11为本申请化合物5-(异丁氧基甲基)喹啉-8-醇盐酸盐抑制鼻咽癌细胞CNE2迁移实验效果图。
图12为本申请化合物5-(异丁氧基甲基)喹啉-8-醇盐酸盐对正常血糖鼠皮肤伤口愈合的影响实验效果图。
图13为本申请化合物5-(异丁氧基甲基)喹啉-8-醇盐酸盐对正常血糖鼠皮肤伤口愈合率的影响。
图14为本申请化合物5-(异丁氧基甲基)喹啉-8-醇盐酸盐对糖尿病鼠皮肤伤口愈合的影响实验效果图。
图15为本申请化合物5-(异丁氧基甲基)喹啉-8-醇盐酸盐对糖尿病鼠皮肤伤口愈合率的影响。
具体实施方式
以下通过试验例的形式，对本申请的所述内容做进一步解释。但本申请上述主题的范围不仅限于以下试验例。
其中，本申请所用试剂均为常用试剂，均可在常规试剂生产销售公司购买。
本申请化合物5-(异丁氧基甲基)喹啉-8-醇盐酸盐的合成路线如图1所示，
本申请化合物5-(异丁氧基甲基)喹啉-8-醇盐酸盐的制备方法为：
步骤1：合成5-氯甲基喹啉-8-醇
室温下，将1eq 8-羟基喹啉和20eq多聚甲醛加入到的浓盐酸中，浓盐酸用量相对8-羟基喹啉为20mL/g，氮气保护氛围下，升温到60℃，直至反应结束，待反应完成后，降温至室温，用饱和碳酸钾溶液调节pH值至6，用乙酸乙酯萃 取两次，用无水硫酸钠干燥有机相，随后将有机相浓缩至干，得粗品，所得粗品经柱层析分离纯化，得5-氯甲基喹啉-8-醇；
步骤2：合成5-(异丁氧基甲基)喹啉-8-醇
将步骤1所得的5-氯甲基喹啉-8-醇全部加入到无水异丁醇中，无水异丁醇用量相对5-氯甲基喹啉-8-醇为15mL/g，并加入5-氯甲基喹啉-8-醇摩尔当量的10％的催化剂碘化钾，在氮气保护氛围下，升温回流反应过夜，待反应完成之后，降温至室温，用饱和碳酸钾溶液调节pH值至7，再用二氯甲烷萃取两次，有机层经无水硫酸钠干燥，浓缩至干，得粗品，所得粗品柱析层分离纯化，得5-(异丁氧基甲基)喹啉-8-醇；
步骤3：合成5-(异丁氧基甲基)喹啉-8-醇盐酸盐
将步骤2所得的5-(异丁氧基甲基)喹啉-8-醇溶于无水甲醇，并降温至0℃，氮气保护条件下缓慢滴入浓盐酸，维持反应体系pH值为3，反应8小时，待反应完成之后，真空减压蒸馏去除甲醇和过量的浓盐酸，得到黄棕色固体粗品，所得粗品于乙酸乙酯和石油醚体系中搅拌反应8小时，过滤，滤饼用乙酸乙酯和石油醚混合液洗涤至中性，滤饼真空干燥，得纯产品5-(异丁氧基甲基)喹啉-8-醇盐酸盐。
本申请化合物5-(异丁氧基甲基)喹啉-8-醇盐酸盐的核磁谱图如图2所示，由图2可知， 1HNMR(400MHz,DMSO)δ11.97(s,1H),9.09(dd,J＝9.5,7.1Hz,3H),8.04(dd,J＝8.6,5.2Hz,2H),7.71(d,J＝7.9Hz,2H),7.46(d,J＝7.9Hz,2H),4.87(s,3H),3.25(d,J＝6.5Hz,3H),1.80(dp,J＝13.3,6.5Hz,2H),0.82(d,J＝6.7Hz,8H)。
由上述制备方法制备得到的5-氯甲基喹啉-8-醇，其收率为52％；制备得到的5-(异丁氧基甲基)喹啉-8-醇，其收率为65％，制备得到的5-(异丁氧基甲基)喹啉-8-醇盐酸盐，其收率为95％。
以下试验例中，试验例1和试验例4只使用化合物5-(异丁氧基甲基)喹啉-8-醇盐酸盐进行试验，试验例2和试验例3使用5-(异丁氧基甲基)喹啉-8-醇和5-(异丁氧基甲基)喹啉-8-醇盐酸盐两种化合物进行试验，均用二甲基亚砜(DMSO)将化合物溶解为高浓度母液，再用磷酸盐缓冲液(PBS)或培养基稀释配制成一定浓度的溶液。
试验例1、化合物细胞毒性检测(MTS实验)
(1)取适量胰酶消化对数生长期的鼻咽癌CNE2、5-8F细胞株，鼠胚胎成纤维细胞NIH/3T3及角质形成细胞HaCaT，分别离心收集，加DMEM培养基重悬、稀释计数，调整混悬液细胞密度至5000个/孔，以每孔200μL接种于96孔板中，边缘孔加入适量灭菌PBS保持湿润。将96孔板放入37℃、5％CO 2培养箱中过夜。
(2)细胞充分贴壁后，弃去孔内培养基，分别加入含有1.64、4.1、10.24、25.6、64、160、400μg/mL化合物的DMEM培养基，每一浓度设置3个复孔。将96孔板置于37℃、5％CO 2培养箱中培养48h。
(3)避光条件下用DMEM以1:10的比例稀释MTS溶液，弃去96孔板中培养基，每孔加入200μL混合溶液，锡纸包裹避光，于培养箱放置3小时。
(4)将96孔板放入全波长酶标仪(Bio-TekEPOCH2)中，测490nm处各孔吸光度，用ExceL计算各浓度梯度下细胞存活率，用GraphpadPrism7.0绘制药物浓度-反应曲线。
试验结果：
IC50为一定时间药物处理导致半数细胞死亡的药物浓度。如图3所示，本申请化合物在NIH/3T3细胞中48h的IC50＝10.05μg/mL，在HaCaT细胞中为3.837μg/mL，在5-8F细胞中为21.79μg/mL，在CNE2细胞中为61.3μg/mL。该结果提示，本申请化合物对上述细胞株呈现剂量依赖性细胞毒性，且正常细胞株较肿瘤细胞株更为敏感。基于各IC50结果，我们在后续功能实验中以1μg/mL为最高浓度，保证本化合物不产生明显细胞毒性。
试验例2、划痕实验
(1)分别接种适量CNE2和NIH/3T3细胞到六孔板中，待达到一定密度后，根据分组分别加入不含化合物、含0.1μg/mL及1μg/mL化合物的完全培养基，48h后细胞铺满孔板底面。
(2)使用200μL微量移液器的枪头对细胞做划痕。
(3)使用PBS洗细胞2次，洗去因划痕从板底脱离的细胞。
(4)分别于划痕后0h，12h，24h，36h和48h，使用研究级倒置荧光显微镜(OLG07MPUSIX73)对细胞划痕进行观察及拍照。
如图4、图5、图6、图7所示，0.1μg/mL及1μg/mL浓度的化合物5-(异丁氧基甲基)喹啉-8-醇和5-(异丁氧基甲基)喹啉-8-醇盐酸盐均加速了NIH/3T3 运动，同时抑制了鼻咽癌细胞CNE2的迁移。
试验例3、迁移实验
(1)分别接种适量CNE2和NIH/3T3细胞到12孔板中，待达到一定密度后，根据分组分别加入不含化合物、含0.1μg/mL、0.5μg/mL及1μg/mL化合物的完全培养基处理48h，其中NIH/3T3细胞培养基中不含0.5μg/mL的化合物5-(异丁氧基甲基)喹啉-8-醇；CNE2细胞培养基中不含0.5μg/mL的化合物5-(异丁氧基甲基)喹啉-8-醇盐酸盐。
(2)适量胰酶消化加药48h后的CNE2和NIH/3T3细胞，分别离心收集，加入不含血清的基础培养基重悬、稀释计数，调整混悬液细胞密度至80000个/孔(3T3)及50000个/孔(CNE2)。
(3)取直径8mm的Transwell小室放入适配24孔板中，下室加入800μL完全培养基，上室每孔接种200μL细胞悬液，放入37℃、5％CO 2培养箱中过夜。
(4)培养23h后取出孔板，弃去上室、下室培养液，分别加入200μL、800μL甲醛溶液固定细胞。
(5)半小时后弃去上下室中甲醛溶液，分别加入200μL、800μL0.1％结晶紫溶液进行细胞染色。
(6)1小时后，回收上、下室结晶紫溶液，在清水中轻柔洗净TransweLL小室，用棉签擦去小室内面细胞，过夜自然晾干。
(7)在高级研究型显微镜(NIKONECLIPSE80i)下观察小室外侧细胞情况，拍照记录。
如图8、图9、图10、图11所示，各个浓度的化合物5-(异丁氧基甲基)喹啉-8-醇和5-(异丁氧基甲基)喹啉-8-醇盐酸盐均能加速NIH/3T3运动，且促进作用随浓度提高而增加，呈现剂量依赖性；同时G06在各浓度均显著抑制鼻咽癌细胞CNE2迁移，G07在高浓度能显著抑制CNE2运动。
试验例4、化合物对小鼠皮肤伤口愈合的影响
一、实验材料
1.1实验动物：
来源：广东省医学实验动物中心(许可证号：SCXK(粤)2018-0002)
品种品系：野生型C57BL/6小鼠
等级：SPF级
规格：只
数量：42只
性别：雄性
周龄：4
1.2实验试剂如表1所示：
表1
Figure PCTCN2021097808-appb-000002
1.3实验仪器如表2所示：
表2
Figure PCTCN2021097808-appb-000003
二、实验方法
2.1高脂饮食联合STZ构建糖尿病小鼠模型
1、分组：随机抽取28只4周龄雄性C57BL/6小鼠进入造模组，余14只为非造模组。测量基础体重，确认组内及组间小鼠体重相近；
2、喂养：以造模组小鼠开始进食高脂饲料为实验第一天，同时非造模组小鼠进食普通饲料。每周记录体重，观察毛发、进食及排尿状况，第28天晚上撤去饲料，禁食不禁水过夜；
3、配制链脲佐菌素(STZ)注射液：将20mgSTZ粉末溶于4mL柠檬酸缓冲液中，配制成5mg/mL的注射液。所有操作在冰上进行，注意避光。STZ溶液现配现用。
4、注射STZ：第二天测量小鼠体重和空腹血糖：造模组小鼠腹腔注射STZ溶液(40mg/kg/d)，每天一次，连续5天，注射第一日继续禁食，注射第二日开始恢复高脂喂养；非造模组小鼠腹腔注射等体积双蒸水，每天一次，连续5天，注射第一日继续禁食，注射第二日开始恢复普通喂养。继续观察小鼠进食、饮水、排尿情况；
5、给药结束2周后，分别检测两组小鼠空腹血糖：
1)造模组小鼠中，定义以连续3次以上稳定在11.1mmoL/L以上并出现“多饮、多尿、多食、消瘦”的小鼠为造模成功的糖尿病鼠，纳入后续实验，不符合模型标准的小鼠退出实验，统计造模成功率；
2)非造模组小鼠中，连续3次以上稳定不超过11.1mmoL/L，并无“多饮、多尿、多食、消瘦”表现的小鼠纳入后续实验，不符合模型标准的小鼠退出实验。
6、实验期间注意造模小鼠的卫生护理，若出现多饮多尿表现，必须注意及时补充饮水，缩短更换垫料间隔。
2.2小鼠背部皮肤全层伤口构建及化合物对皮肤伤口愈合影响的观察
1、分组：将建模成功的糖尿病组随机分为阴性对照组、低剂量组、中剂量组及高剂量组，各组小鼠不少于3只；同样将正常血糖组小鼠随机分为阴性对照组、低剂量组、中剂量组及高剂量组，各组小鼠不少于3只；
2、于建模成功2周后，将各组小鼠移至手术室，利用气麻装置及异氟烷对进行气体麻醉；
3、将完全麻醉的小鼠以俯卧位置于垫巾上，用动物电动剃毛器剃除背部毛发；
4、用75％酒精消毒背部皮肤，用直径为8mm的环钻于背部中线最高处两侧分别打出2个直径约8mm的圆形皮肤全层伤口，用手术剪及镊子剃除皮下组织，暴露肌肉表面筋膜；
5、伤口止血、消毒，开放伤口；以游标卡尺作为对照，拍照记录伤口形态大小。
6、给药：根据预先完成的急性毒性试验结果，确定各组小鼠给药剂量，阴性对照组给予等体积PBS(溶剂组)，低剂量组3mg/kg，中剂量组10mg/kg，高剂量组30mg/kg。给药方式为腹腔注射。小鼠创伤当天为daG07 1，分别于daG07 1、3、5、7、9对各组小鼠进行化合物/PBS注射。
7、温暖手术灯下复苏麻醉小鼠，运回洁净饲育室IVC笼架。
三、试验结果
实验结果如图12-15所示，经各剂量G07处理后的糖尿病鼠及正常血糖鼠，其伤口愈合率均在给药后3～9天内显著提高，提示G07具有明显促进小鼠伤口愈合的作用。
上述实施例仅示例性说明本申请的原理及其功效，而非用于限制本申请。任何熟悉此技术的人士皆可在不违背本申请的精神及范畴下，对上述实施例进行修饰或改变。因此，举凡所属技术领域中具有通常知识者在未脱离本申请所揭示的精神与技术思想下所完成的一切等效修饰或改变，仍应由本申请的权利要求所涵盖。

Claims (7)

  1. 一种促进成纤维细胞运动的化合物，其为5-(异丁氧基甲基)喹啉-8-醇或5-(异丁氧基甲基)喹啉-8-醇盐酸盐，其中，5-(异丁氧基甲基)喹啉-8-醇的化学结构式如式G06所示，5-(异丁氧基甲基)喹啉-8-醇盐酸盐的化学结构式如式G07所示：
    Figure PCTCN2021097808-appb-100001
  2. 5-(异丁氧基甲基)喹啉-8-醇盐酸盐的制备方法，包括：
    步骤1：合成5-氯甲基喹啉-8-醇
    室温下，将1eq 8-羟基喹啉和20eq多聚甲醛加入到浓盐酸中，浓盐酸用量相对8-羟基喹啉为20mL/g，氮气保护氛围下，升温到60℃，直至反应结束，待反应完成后，降温至室温，用饱和碳酸钾溶液调节pH值至6～7，用乙酸乙酯萃取两次，用无水硫酸钠干燥有机相，随后将有机相浓缩至干，得粗品，所得粗品经柱层析分离纯化，得5-氯甲基喹啉-8-醇；
    步骤2：合成5-(异丁氧基甲基)喹啉-8-醇
    将步骤1所得的5-氯甲基喹啉-8-醇全部加入到无水异丁醇中，无水异丁醇用量相对5-氯甲基喹啉-8-醇为15mL/g，并加入5-氯甲基喹啉-8-醇摩尔当量的10％的催化剂碘化钾，在氮气保护氛围下，升温回流反应过夜，待反应完成之后，降温至室温，用饱和碳酸钾溶液调节pH值至6～7，再用二氯甲烷萃取两次，有机层经无水硫酸钠干燥，浓缩至干，得粗品，所得粗品柱析层分离纯化，得5-(异丁氧基甲基)喹啉-8-醇；
    步骤3：合成5-(异丁氧基甲基)喹啉-8-醇盐酸盐
    将步骤2所得的5-(异丁氧基甲基)喹啉-8-醇溶于无水甲醇，并降温至0℃，氮气保护条件下缓慢滴入浓盐酸，维持反应体系pH值在2～4之间反应8小时，待反应完成之后，真空减压蒸馏去除甲醇和过量的浓盐酸，得到黄棕色固体粗 品，所得粗品于乙酸乙酯和石油醚体系中搅拌反应8小时，过滤，滤饼用乙酸乙酯和石油醚混合液洗涤至中性，滤饼真空干燥，得纯产品5-(异丁氧基甲基)喹啉-8-醇盐酸盐。
  3. 一种药物组合物，其包含权利要求1所述的化合物或其药学上可接受的盐。
  4. 如权利要求3所述药物组合物，其中，所述药物组合物为液体、膏状体、凝胶、敷料或喷雾。
  5. 如权利要求4所述药物组合物，其中，所述药物组合物进一步包含一种或多种药学上可接受的载体或药用辅料。
  6. 如权利要求1所述的化合物或权利要求3-5任一项所述的药物组合物在制备用于促进伤口愈合的药物中的应用。
  7. 如权利要求1所述的化合物或权利要求3-5任一项所述的药物组合物在制备用于促进肿瘤合并糖尿病的手术患者伤口愈合的药物中的应用。
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WO2009137597A1 (en) * 2008-05-06 2009-11-12 The Trustees Of Columbia University In The City Of New York COMPOUNDS THAT INHIBIT PRODUCTION OF sAPPβ AND Aβ AND USES THEREOF
CN113087665A (zh) * 2021-04-14 2021-07-09 广州市朝利良生物科技有限公司 一种促进细胞运动的化合物、药物组合物及其制备和应用

Patent Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2009137597A1 (en) * 2008-05-06 2009-11-12 The Trustees Of Columbia University In The City Of New York COMPOUNDS THAT INHIBIT PRODUCTION OF sAPPβ AND Aβ AND USES THEREOF
CN113087665A (zh) * 2021-04-14 2021-07-09 广州市朝利良生物科技有限公司 一种促进细胞运动的化合物、药物组合物及其制备和应用

Non-Patent Citations (1)

* Cited by examiner, † Cited by third party
Title
XING JING; ZHANG RUKANG; JIANG XIANGRUI; HU TIANWEN; WANG XINJUN; QIAO GANG; WANG JUNJIAN; YANG FENGLING; LUO XIAOMIN; CHEN KAIXIA: "Rational design of 5-((1H-imidazol-1-yl)methyl)quinolin-8-ol derivatives as novel bromodomain-containing protein 4 inhibitors", EUROPEAN JOURNAL OF MEDICINAL CHEMISTRY, ELSEVIER, AMSTERDAM, NL, vol. 163, 1 January 1900 (1900-01-01), AMSTERDAM, NL , pages 281 - 294, XP085576191, ISSN: 0223-5234, DOI: 10.1016/j.ejmech.2018.11.018 *

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