二萜化合物或其盐在制备防治急性肺损伤的药物中的应用Application of diterpenoids or their salts in the preparation of medicines for preventing and treating acute lung injury
技术领域technical field
本申请涉及生物医药领域,具体的涉及一种二萜化合物或其药学上可接受的盐在制备防治急性肺损伤的药物中的应用。The present application relates to the field of biomedicine, in particular to the application of a diterpene compound or a pharmaceutically acceptable salt thereof in the preparation of a medicine for preventing and treating acute lung injury.
背景技术Background technique
急性肺损伤(Acute lung injury,ALI)是一种常见危重症,病死率极高,严重威胁重症受试者的生命并影响其生存质量。多种因素可诱发ALI,主要包括①直接肺损伤因素:严重肺部感染、胃内容物吸入、肺挫伤、吸入有毒气体、淹溺、氧中毒等;②间接肺损伤因素:严重感染、严重的非胸部创伤、急性重症胰腺炎、大量输血、体外循环、弥漫性血管内凝血等。Acute lung injury (ALI) is a common critical illness with a very high mortality rate, which seriously threatens the lives of critically ill subjects and affects their quality of life. A variety of factors can induce ALI, mainly including ① direct lung injury factors: severe lung infection, inhalation of gastric contents, lung contusion, inhalation of toxic gases, drowning, oxygen poisoning, etc.; ② indirect lung injury factors: severe infection, severe Non-thoracic trauma, acute severe pancreatitis, massive blood transfusion, cardiopulmonary bypass, disseminated intravascular coagulation, etc.
尽管对急性肺损伤的研究逐步深入,但其发病机制仍不完全清楚。目前认为,炎症反应在急性肺损伤的发生发展过程中起到关键作用,急性肺损伤发生时,促炎和抗炎反应平衡被打破,导致肺内皮细胞和肺上皮屏障遭到破坏,肺泡毛细血管膜完整性破坏,大量炎症介质刺激的中性粒细胞过度跨上皮迁移,促进促炎因子和细胞毒性介质的释放,肺组织受损,严重的外溢到全身循环,导致多器官衰竭。促炎细胞因子肿瘤坏死因子-α(TNF-α)和白细胞介素-1β(IL-1β)的高表达是急性肺损伤早期反应的重要事件,可破坏肺泡毛细血管屏障,增加肺水肿。白细胞介素-6的高表达使得的急性肺损伤受试者更易发展为多器官衰竭,循环IL-6已被证明是不同病因所致急性肺损伤疾病的良好预测因子。细胞因子的局部释放可以启动一系列放大和调节作用,导致中性粒细胞激活和有毒介质的释放增加,并产生大量活性氧自由基,加重急性肺损伤肺水肿严重程度。因此,抑制急性肺损伤过程中的炎症因子表达和释放,抑制中性粒细胞浸润及氧化应激损伤,对于防治急性肺损伤具有重要意义。Although the research on acute lung injury has gradually deepened, its pathogenesis is still not fully understood. At present, it is believed that inflammatory response plays a key role in the occurrence and development of acute lung injury. When acute lung injury occurs, the balance of pro-inflammatory and anti-inflammatory responses is disrupted, resulting in the destruction of lung endothelial cells and lung epithelial barriers, and alveolar capillaries. Disruption of membrane integrity, excessive transepithelial migration of neutrophils stimulated by a large number of inflammatory mediators, promoting the release of pro-inflammatory factors and cytotoxic mediators, damage to lung tissue, and severe spillover into the systemic circulation lead to multi-organ failure. High expression of the proinflammatory cytokines tumor necrosis factor-α (TNF-α) and interleukin-1β (IL-1β) are important events in the early response to acute lung injury, which can disrupt the alveolar-capillary barrier and increase pulmonary edema. High expression of interleukin-6 predisposes acute lung injury subjects to develop multiple organ failure, and circulating IL-6 has been shown to be a good predictor of acute lung injury disease of different etiologies. The local release of cytokines can initiate a series of amplification and regulation effects, resulting in increased neutrophil activation and release of toxic mediators, and the production of a large number of reactive oxygen species, aggravating the severity of acute lung injury and pulmonary edema. Therefore, inhibiting the expression and release of inflammatory factors in the process of acute lung injury, inhibiting neutrophil infiltration and oxidative stress injury is of great significance for the prevention and treatment of acute lung injury.
现有技术中关于治疗急性肺损伤的策略还很有限,开发出更多的急性肺损伤的治疗途径是非常有意义的。The strategies for the treatment of acute lung injury in the prior art are still limited, and it is very meaningful to develop more therapeutic approaches for acute lung injury.
发明内容SUMMARY OF THE INVENTION
针对现有技术的不足,本申请的目的在于提供一种二萜化合物或其药学上可接受的盐在制备预防和/或治疗急性肺损伤的药物中的应用。In view of the deficiencies of the prior art, the purpose of the present application is to provide the application of a diterpene compound or a pharmaceutically acceptable salt thereof in the preparation of a medicine for preventing and/or treating acute lung injury.
为达到此目的,本申请采用以下技术方案:In order to achieve this purpose, the application adopts the following technical solutions:
本申请首次发现式Ⅰ所示的二萜化合物具有显著的防治急性肺损伤的作用,其对脂多糖诱导的急性肺损伤小鼠模型显示出良好的防治作用,可改善急性肺损伤小鼠肺组织病理学变化, 减轻肺水肿损伤,减轻肺部血气屏障的破坏,抑制肺损伤肺组织中肿瘤坏死因子α、白细胞介素1β和白细胞介素6的表达,抑制肺损伤肺组织中性粒细胞的浸润和氧化应激反应。上述结果表明该类化合物可用于防治急性肺损伤,为急性肺损伤的治疗提供了一种新思路和药物。The present application finds for the first time that the diterpenoid compound represented by formula I has a significant effect of preventing and treating acute lung injury, it has a good preventive and therapeutic effect on the mouse model of acute lung injury induced by lipopolysaccharide, and can improve the lung tissue of mice with acute lung injury. Pathological changes, reduce pulmonary edema injury, reduce the damage of pulmonary blood gas barrier, inhibit the expression of tumor necrosis factor α, interleukin 1β and interleukin 6 in the lung tissue of lung injury, inhibit the expression of neutrophils in the lung tissue of lung injury. Infiltration and oxidative stress. The above results show that the compounds can be used to prevent and treat acute lung injury, and provide a new idea and drug for the treatment of acute lung injury.
一方面,本申请提供一种二萜化合物或其药学可接受的盐在制备预防和/或治疗急性肺损伤的药物中的应用,其中所述二萜化合物具有式I所示的结构:On the one hand, the application provides the application of a diterpene compound or a pharmaceutically acceptable salt thereof in the preparation of a medicine for preventing and/or treating acute lung injury, wherein the diterpene compound has the structure shown in formula I:
式Ⅰ所示的二萜化合物的分子结构及其合成方法已在专利CN104761460A,WO2016150207A1公开的文本中记载,在此通过全文引入本文。The molecular structure of the diterpene compound represented by formula I and the synthesis method thereof have been described in the published texts of patents CN104761460A and WO2016150207A1, which are incorporated herein in their entirety.
在某些实施方式中,所述药学上可接受的盐包括酒石酸盐、硬脂酸盐、草酸盐、枸橼酸盐、乳酸盐、山梨酸盐、延胡索酸盐、甲酸盐、乙酸盐、苯甲酸盐、苯磺酸盐、乙磺酸盐、树脂酸盐、三氟乙酸盐、马来酸盐、甲磺酸盐、富马酸盐、氨基酸盐、烟酸盐、磷酸盐、硫酸盐、盐酸盐或氢溴酸盐。In certain embodiments, the pharmaceutically acceptable salts include tartrate, stearate, oxalate, citrate, lactate, sorbate, fumarate, formate, acetic acid Salt, benzoate, benzenesulfonate, ethanesulfonate, resinate, trifluoroacetate, maleate, mesylate, fumarate, amino acid salt, nicotinate, phosphoric acid salt, sulfate, hydrochloride or hydrobromide.
在某些实施方式中,所述药物抑制肺组织和肺泡灌洗液中TNF-α,IL-1β和IL-6的表达。In certain embodiments, the medicament inhibits the expression of TNF-α, IL-1β and IL-6 in lung tissue and bronchoalveolar lavage fluid.
在某些实施方式中,所述药物抑制肺组织中髓过氧化物酶的活性。In certain embodiments, the medicament inhibits the activity of myeloperoxidase in lung tissue.
在某些实施方式中,所述药物抑制肺组织中丙二醛的含量。In certain embodiments, the drug inhibits the level of malondialdehyde in lung tissue.
在某些实施方式中,所述药物促进肺组织中超氧化物岐化酶的活性。In certain embodiments, the drug promotes superoxide dismutase activity in lung tissue.
在某些实施方式中,所述药物抑制肺组织湿干重比值的增加。In certain embodiments, the drug inhibits an increase in the ratio of wet to dry weight of lung tissue.
在某些实施方式中,所述药物抑制肺泡灌洗液中的蛋白含量。In certain embodiments, the drug inhibits the protein content of the bronchoalveolar lavage fluid.
在某些实施方式中,所述药物还包括药学上可接受的辅料。In certain embodiments, the medicament further includes pharmaceutically acceptable excipients.
在某些实施方式中,所述辅料包括载体、稀释剂、赋形剂、填充剂、粘合剂、润湿剂、崩解剂、乳化剂、助溶剂、增溶剂、渗透压调节剂、表面活性剂、包衣材料、着色剂、pH调节剂、抗氧剂、抑菌剂或缓冲剂中的任意一种或至少两种的组合。In certain embodiments, the adjuvants include carriers, diluents, excipients, fillers, binders, wetting agents, disintegrants, emulsifiers, cosolvents, solubilizers, osmotic pressure regulators, surface Any one or a combination of at least two of active agents, coatings, colorants, pH adjusters, antioxidants, bacteriostatic agents, or buffers.
所述至少两种的组合例如稀释剂和赋形剂的组合、粘合剂和润湿剂的组合、乳化剂和助溶剂的组合等,其他任意的组合方式均可选择。The combination of the at least two kinds, such as the combination of diluent and excipient, the combination of binder and wetting agent, the combination of emulsifier and cosolvent, etc., can be selected in any other combination manner.
在某些实施方式中,所述药物的剂型为药学上可接受的任意一种剂型。In certain embodiments, the dosage form of the medicament is any pharmaceutically acceptable dosage form.
在某些实施方式中,所述剂型包括混悬剂、颗粒剂、胶囊剂、散剂、片剂、乳剂、溶液剂、滴丸剂、注射剂、栓剂、灌肠剂、气雾剂、喷雾剂、贴剂或滴剂中的任意一种。In certain embodiments, the dosage forms include suspensions, granules, capsules, powders, tablets, emulsions, solutions, drop pills, injections, suppositories, enemas, aerosols, sprays, patches or any of the drops.
本申请还提供如式Ⅰ所示的二萜化合物或其药学上可接受的盐在制备抑制肺组织和肺泡灌洗液中TNF-α,IL-1β和IL-6表达的药物中的应用。The present application also provides the use of the diterpene compound represented by formula I or a pharmaceutically acceptable salt thereof in the preparation of a medicament for inhibiting the expression of TNF-α, IL-1β and IL-6 in lung tissue and bronchoalveolar lavage fluid.
本申请还提供如式Ⅰ所示的二萜化合物或其药学上可接受的盐在制备抑制肺组织中髓过氧化物酶活性的药物中的应用。The present application also provides the use of the diterpene compound represented by formula I or a pharmaceutically acceptable salt thereof in the preparation of a medicament for inhibiting the activity of myeloperoxidase in lung tissue.
本申请还提供如式Ⅰ所示的二萜化合物或其药学上可接受的盐在制备抑制肺组织中丙二醛含量的药物中的应用。The present application also provides the use of the diterpene compound represented by formula I or a pharmaceutically acceptable salt thereof in the preparation of a medicament for inhibiting the content of malondialdehyde in lung tissue.
本申请还提供如式Ⅰ所示的二萜化合物或其药学上可接受的盐在制备促进肺组织中超氧化物岐化酶活性的药物中的应用。The present application also provides the use of the diterpene compound represented by formula I or a pharmaceutically acceptable salt thereof in the preparation of a medicament for promoting the activity of superoxide dismutase in lung tissue.
本申请还提供如式Ⅰ所示的二萜化合物或其药学上可接受的盐在制备抑制肺组织湿干重比值增加的药物中的应用。The present application also provides the use of the diterpene compound represented by formula I or a pharmaceutically acceptable salt thereof in the preparation of a medicament for inhibiting the increase in the ratio of wet to dry weight of lung tissue.
本申请还提供如式Ⅰ所示的二萜化合物或其药学上可接受的盐在制备抑制肺泡灌洗液中的蛋白含量的药物中的应用。The present application also provides the use of the diterpene compound represented by formula I or a pharmaceutically acceptable salt thereof in the preparation of a medicament for inhibiting the protein content in bronchoalveolar lavage fluid.
在某些实施方式中,其中所述肺组织为急性损伤的肺组织。In certain embodiments, wherein the lung tissue is acutely injured lung tissue.
另一方面,本申请提供一种预防和/或治疗急性肺损伤的方法,其包括向有需要的受试者施用有效量的式I所示的二萜化合物或其药学上可接受的盐。In another aspect, the present application provides a method for preventing and/or treating acute lung injury, comprising administering an effective amount of the diterpene compound represented by formula I or a pharmaceutically acceptable salt thereof to a subject in need thereof.
在某些实施方式中,其中所述施用包括选自口服、肠胃外、经鼻、吸入、气管内、肺内和支气管内的施用途径将治疗有效量的式I所示的二萜化合物或其药学上可接受的盐施用至受试者。In certain embodiments, wherein said administering comprises a route of administration selected from the group consisting of oral, parenteral, nasal, inhalation, intratracheal, intrapulmonary and intrabronchial, a therapeutically effective amount of the diterpene compound of formula I or its A pharmaceutically acceptable salt is administered to a subject.
在某些实施方式中,其中所述式I所示的二萜化合物或其药学上可接受的盐的剂量为约0.01mg至约5000mg。In certain embodiments, the dosage of the diterpene compound represented by Formula I or a pharmaceutically acceptable salt thereof is about 0.01 mg to about 5000 mg.
在某些实施方式中,所述的方法包括将所述式I所示的二萜化合物或其药学上可接受的盐物以每天1至5次或更多次的剂量施用至患受试者。In certain embodiments, the method comprises administering the diterpene compound of formula I or a pharmaceutically acceptable salt thereof to the subject in a dose of 1 to 5 or more times per day .
在某些实施方式中,其进一步包括选自由以下组成的组的至少一种另外的治疗:手术、类固醇疗法、非类固醇疗法、抗病毒疗法、抗真菌疗法、抗微生物疗法、免疫抑制剂疗法、抗感染疗法、抗高血压疗法、营养补充剂及其任意组合。In certain embodiments, it further comprises at least one additional treatment selected from the group consisting of surgery, steroid therapy, non-steroid therapy, antiviral therapy, antifungal therapy, antimicrobial therapy, immunosuppressive therapy, Anti-infective therapy, anti-hypertensive therapy, nutritional supplement, and any combination thereof.
在某些实施方式中,其中所述类固醇疗法包括施用地塞米松、丙酸倍氯米松、布地奈德和/或氟替卡松。In certain embodiments, wherein the steroid therapy comprises administration of dexamethasone, beclomethasone propionate, budesonide, and/or fluticasone.
另一方面,本申请提供一种试剂盒,其包括式I所示的二萜化合物或其药学上可接受的盐。In another aspect, the present application provides a kit comprising the diterpene compound represented by formula I or a pharmaceutically acceptable salt thereof.
在某些实施方式中,所述试剂盒还包括容器。In certain embodiments, the kit further comprises a container.
在某些实施方式中,所述试剂盒还包括使用说明材料,其中所述说明材料包括使用选自经口服、肠胃外、经鼻、吸入、气管内、肺内和支气管内的施用途径治疗或预防受试者中的急性肺损伤的说明书。In certain embodiments, the kit further includes instructional material, wherein the instructional material includes treatment or treatment using a route of administration selected from the group consisting of oral, parenteral, nasal, inhalation, intratracheal, intrapulmonary, and intrabronchial. Instructions for preventing acute lung injury in a subject.
相对于现有技术,本申请具有以下有益效果:Compared with the prior art, the present application has the following beneficial effects:
本申请首次发现式Ⅰ所示的二萜化合物具有显著的防治急性肺损伤的作用,其对脂多糖诱导的急性肺损伤小鼠模型显示出良好的防治作用,可改善急性肺损伤小鼠肺组织病理学变化,减轻肺水肿损伤,减轻肺部血气屏障的破坏,抑制肺损伤肺组织中肿瘤坏死因子α、白细胞介素1β和白细胞介素6的表达,抑制肺损伤肺组织中性粒细胞的浸润和氧化应激反应。上述结果表明该类化合物可用于防治急性肺损伤,为急性肺损伤的治疗提供了一种新思路和药物。The present application finds for the first time that the diterpenoid compound represented by formula I has a significant effect of preventing and treating acute lung injury, it has a good preventive and therapeutic effect on the mouse model of acute lung injury induced by lipopolysaccharide, and can improve the lung tissue of mice with acute lung injury. Pathological changes, reduce pulmonary edema injury, reduce the damage of pulmonary blood-gas barrier, inhibit the expression of tumor necrosis factor α, interleukin 1β and interleukin 6 in lung tissue of lung injury, inhibit the expression of neutrophils in lung tissue of lung injury. Infiltration and oxidative stress. The above results show that the compounds can be used to prevent and treat acute lung injury, and provide a new idea and drug for the treatment of acute lung injury.
本领域技术人员能够从下文的详细描述中容易地洞察到本申请的其它方面和优势。下文的详细描述中仅显示和描述了本申请的示例性实施方式。如本领域技术人员将认识到的,本申请的内容使得本领域技术人员能够对所公开的具体实施方式进行改动而不脱离本申请所涉及发明的精神和范围。相应地,本申请的附图和说明书中的描述仅仅是示例性的,而非为限制性的。Other aspects and advantages of the present application can be readily appreciated by those skilled in the art from the following detailed description. Only exemplary embodiments of the present application are shown and described in the following detailed description. As those skilled in the art will recognize, the content of this application enables those skilled in the art to make changes to the specific embodiments disclosed without departing from the spirit and scope of the invention to which this application relates. Accordingly, the drawings and descriptions in the specification of the present application are only exemplary and not restrictive.
附图说明Description of drawings
本申请所涉及的发明的具体特征如所附权利要求书所显示。通过参考下文中详细描述的示例性实施方式和附图能够更好地理解本申请所涉及发明的特点和优势。对附图简要说明如下:The invention to which this application relates is set forth with particularity characteristic of the appended claims. The features and advantages of the inventions involved in this application can be better understood by reference to the exemplary embodiments described in detail hereinafter and the accompanying drawings. A brief description of the drawings is as follows:
图1显示的是本申请实施例1中各组小鼠的肺组织病理切片图(A-F依次为空白对照组、LPS模型组、LPS+HY1702 5mg/kg组、LPS+HY1702 10mg/kg组、LPS+HY1702 30mg/kg组、LPS+地塞米松(DEX)5mg/kg组);Figure 1 shows the lung histopathological sections of mice in each group in Example 1 of the present application (AF are blank control group, LPS model group, LPS+HY1702 5mg/kg group, LPS+HY1702 10mg/kg group, LPS+HY1702 10mg/kg group in turn +HY1702 30mg/kg group, LPS+dexamethasone (DEX) 5mg/kg group);
图2显示的是本申请实施例1中各组小鼠的肺湿/干重比统计图;Fig. 2 shows the lung wet/dry weight ratio statistics chart of each group of mice in Example 1 of the present application;
图3显示的是本申请实施例1中各组小鼠的BALF中蛋白浓度统计图;Fig. 3 shows the statistic diagram of protein concentration in the BALF of each group of mice in Example 1 of the present application;
图4显示的是本申请实施例1中各组小鼠肺组织中TNF-α,IL-1β,IL-6的蛋白含量统计图(a、b、c依次代表TNF-α,IL-1β,IL-6);Figure 4 shows the statistical graph of the protein content of TNF-α, IL-1β, and IL-6 in the lung tissue of each group of mice in Example 1 of the present application (a, b, and c represent TNF-α, IL-1β, IL-6);
图5显示的是本申请实施例1中各组小鼠BALF中相应mRNA的表达水平统计图(a、b、c依次代表TNF-α,IL-1β,IL-6);Figure 5 shows a statistical graph of the expression levels of corresponding mRNAs in the BALF of each group of mice in Example 1 of the present application (a, b, and c represent TNF-α, IL-1β, and IL-6 in turn);
图6显示的是本申请实施例1中各组小鼠肺组织中MPO活性统计图;Fig. 6 shows the MPO activity statistics diagram in the lung tissue of each group of mice in Example 1 of the present application;
图7显示的是本申请实施例1中各组小鼠肺组织中丙二醛(MDA)含量统计图;Fig. 7 shows the malondialdehyde (MDA) content statistic diagram in the lung tissue of each group of mice in Example 1 of the present application;
图8显示的是本申请实施例1中各组小鼠肺组织中超氧化物歧化酶(SOD)的活性统计图。Figure 8 shows a statistical graph of the activity of superoxide dismutase (SOD) in the lung tissue of each group of mice in Example 1 of the present application.
具体实施方式detailed description
以下由特定的具体实施例说明本申请发明的实施方式,熟悉此技术的人士可由本说明书所公开的内容容易地了解本申请发明的其他优点及效果。The embodiments of the invention of the present application are described below with specific specific examples, and those skilled in the art can easily understand other advantages and effects of the invention of the present application from the contents disclosed in this specification.
术语定义Definition of Terms
在本申请中,术语“药学上可接受的”通常是指不干扰活性成分生物学活性的有效性的一种或多种无毒物质。这类制剂通常可含有盐、缓冲剂、防腐剂、相容性载体和任选的其它治疗剂。这类药学上可接受的制剂通常也可包含适合给予人的相容性固体或液体填料、稀释剂或包囊材料。用于医药时,盐应该是药学上可接受的盐,但可方便地使用非药学上可接受的盐来制备药学上可接受的盐,不能将它们排除在本申请范围以外。这类药理学和药学上可接受的盐包括但不限于由以下酸制备的盐:氢氯酸、氢溴酸、硫酸、硝酸、磷酸、马来酸、乙酸、水杨酸、柠檬酸、硼酸、甲酸、丙二酸、琥珀酸等。药学上可接受的盐也可制备成碱金属盐或碱土金属盐,如钠盐、钾盐或钙盐。In this application, the term "pharmaceutically acceptable" generally refers to one or more nontoxic substances that do not interfere with the effectiveness of the biological activity of the active ingredient. Such formulations may generally contain salts, buffers, preservatives, compatible carriers and optionally other therapeutic agents. Such pharmaceutically acceptable formulations may also generally contain compatible solid or liquid fillers, diluents or encapsulating materials suitable for administration to humans. When used in medicine, the salts should be pharmaceutically acceptable salts, but non-pharmaceutically acceptable salts can be conveniently used to prepare pharmaceutically acceptable salts, and they cannot be excluded from the scope of this application. Such pharmacologically and pharmaceutically acceptable salts include, but are not limited to, those prepared from the following acids: hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, phosphoric acid, maleic acid, acetic acid, salicylic acid, citric acid, boric acid , formic acid, malonic acid, succinic acid, etc. Pharmaceutically acceptable salts can also be prepared as alkali metal or alkaline earth metal salts, such as sodium, potassium or calcium salts.
在本申请中,术语“药学上可接受的辅料”通常是指对被治疗的受试者的一般健康状况没有持续的不利影响,或者不消除化合物的生物活性或性质,并且相对无毒的物质。这类物质通常可包含盐、缓冲剂、防腐剂、相容性载体和任选的其它制剂。In this application, the term "pharmaceutically acceptable excipient" generally refers to a substance that has no persistent adverse effect on the general health of the subject being treated, or does not eliminate the biological activity or properties of the compound, and is relatively non-toxic . Such materials may generally contain salts, buffers, preservatives, compatible carriers and optionally other formulations.
本申请中,术语“施用”通常是指通过任意引入或递送途径将本申请药物制剂引入受试者的身体中。可以采用本领域技术人员已知的用于使细胞、器官或组织与所述药物接触的任何方法。所述施用可以包括而不限于静脉内、动脉内、鼻内、腹内、肌内、皮下透皮或口服。每日剂量可以划分成一个、两个或更多个合适形式的剂量以在某个时间段期间的一个、两个或更多个时间施用。In this application, the term "administration" generally refers to the introduction of the pharmaceutical formulations of the present application into the body of a subject by any route of introduction or delivery. Any method known to those skilled in the art for contacting cells, organs or tissues with the drug may be employed. The administration may include, without limitation, intravenous, intraarterial, intranasal, intraperitoneal, intramuscular, subcutaneous transdermal, or oral. The daily dose may be divided into one, two or more doses in suitable form to be administered at one, two or more times during a certain period of time.
在本申请中,术语“有效量”或“有效剂量”通常是指足以实现或至少部分实现所需效果的量。药物或治疗剂的“治疗有效量”或“治疗有效剂量”通常是当单独使用或与另一种治疗剂组合使用时促进疾病消退(这通过疾病症状严重程度的降低、疾病无症状期的频度和持续时间的增加、或者由于罹患疾病而引起的损害或残疾的预防来证明)的任何药物量。药物的“预防有效量”或“预防有效剂量”通常是指当单独或与另一种治疗剂组合给有疾病发展或 疾病复发的风险的受试者施用时抑制疾病的发展或复发的药物量。可以使用本领域技术人员已知的多种方法对治疗剂或预防剂促进疾病消退或抑制疾病发展或复发的能力进行评估,比如在处于临床试验期间的人类受试者中、在动物模型系统中预测对人类的功效、或者通过在体外测定中测定药剂的活性。In this application, the term "effective amount" or "effective dose" generally refers to an amount sufficient to achieve, or at least partially achieve, the desired effect. A "therapeutically effective amount" or "therapeutically effective dose" of a drug or therapeutic agent is generally one that, when used alone or in combination with another therapeutic agent, promotes disease regression (this is achieved through a reduction in the severity of disease symptoms, the frequency of asymptomatic periods of the disease) increase in intensity and duration, or as evidenced by the prevention of impairment or disability due to disease). A "prophylactically effective amount" or "prophylactically effective dose" of a drug generally refers to the amount of drug that inhibits the development or recurrence of disease when administered alone or in combination with another therapeutic agent to a subject at risk of disease development or disease recurrence . The ability of a therapeutic or prophylactic agent to promote disease regression or inhibit disease progression or recurrence can be assessed using a variety of methods known to those of skill in the art, such as in human subjects during clinical trials, in animal model systems Efficacy in humans is predicted, or by measuring the activity of an agent in an in vitro assay.
在本申请中,术语“有需要”和类似术语是指已被确定为需要治疗疾病,如急性肺损伤,例如,由脓毒症过程中的腹膜炎引起的急性肺损伤、由脓毒症过程中的静脉菌血症引起的急性肺损伤、由烟尘吸入引起的急性肺损伤、在表面活性物质不足的早产儿中出现的急性肺损伤、由氧中毒引起的急性肺损伤或由来自机械通风的气压伤引起的急性肺损伤的对象。这种确定可以是医疗诊断的结果。此外,“需要”本申请的方法的对象包括已知或被怀疑之前已被诊断为急性肺损伤的那些对象。In this application, the term "in need" and similar terms refers to a condition that has been identified as in need of treatment, such as acute lung injury, eg, acute lung injury caused by peritonitis during sepsis, acute lung injury caused by peritonitis during sepsis acute lung injury from venous bacteremia, acute lung injury from soot inhalation, acute lung injury in preterm infants with insufficient surfactant, acute lung injury from oxygen toxicity, or from air pressure from mechanical ventilation Injury-induced acute lung injury in subjects. This determination may be the result of a medical diagnosis. Furthermore, subjects "in need" of the methods of the present application include those subjects who are known or suspected to have been previously diagnosed with acute lung injury.
在本申请中,术语“受试者”通常是指需要诊断、预后、改善、预防和/或治疗疾病的人或非人动物(包括哺乳动物),诸如人、非人灵长类动物(猿、长臂猿、大猩猩、黑猩猩、猩猩、猕猴)、家畜(狗和猫)、农场动物(家禽如鸡和鸭、马、牛、山羊、绵羊、猪)和实验动物(小鼠、大鼠、兔、豚鼠)。人受试者包括胎儿、新生儿、婴儿、青少年和成人受试者。受试者包括动物疾病模型。In this application, the term "subject" generally refers to a human or non-human animal (including mammals) in need of diagnosis, prognosis, amelioration, prevention and/or treatment of a disease, such as a human, non-human primate (simian) , gibbons, gorillas, chimpanzees, orangutans, macaques), livestock (dogs and cats), farm animals (poultry such as chickens and ducks, horses, cows, goats, sheep, pigs) and laboratory animals (mice, rats, rabbits) , guinea pigs). Human subjects include fetal, neonatal, infant, adolescent and adult subjects. Subjects include animal disease models.
在本申请中,术语“包括”、“包含”、“具有”、“可以”、“含有”及其变体通常旨在是开放式过渡性短语、术语或词语,其不排除额外行为或结构的可能性。术语“由……组成”通常表示不能存在别的组分(或同样地,特征、整数、步骤、等)。除非上下文另有明确规定,单数形式如英文的“a”,“an”,“the”,中文的“一个”、“一种”和“所述/该”一般包括所指代事物的复数形式。In this application, the terms "including", "including", "having", "may", "containing" and variations thereof are generally intended to be open-ended transitional phrases, terms or words that do not preclude additional acts or structures possibility. The term "consisting of" generally means that no other components (or similarly, features, integers, steps, etc.) can be present. Unless the context clearly dictates otherwise, the singular forms such as "a", "an", "the" in English and "a", "an" and "the/the" in Chinese generally include the plural forms of the things they refer to. .
在本申请中,术语“约”通常意指大约(approximately)、在......的附近(intheregionof)、粗略地(roughly)、或左右(around)。当术语“约”当用于指涉数值范围时,截值或特定数值用于指示所载明的数值可与该列举数值有多达10%的差异。因此,术语“约”可用于涵盖自特定值±10%或更少的变异、±5%或更少的变异、±1%或更少的变异、±0.5%或更少的变异、或±0.1%或更少的变异。In this application, the term "about" generally means approximately, in the region of, roughly, or around. When the term "about" is used in reference to a range of values, a cut-off or specific value is used to indicate that the stated value can vary by up to 10% from the recited value. Thus, the term "about" can be used to encompass ±10% or less variation, ±5% or less variation, ±1% or less variation, ±0.5% or less variation, or ±0.5% or less variation from a specified value. 0.1% or less variation.
发明详述Detailed description of the invention
用途use
一方面,本申请提供一种二萜化合物或其药学可接受的盐在制备预防和/或治疗急性肺损伤的药物中的应用,其中所述二萜化合物具有式I所示的结构:On the one hand, the application provides the application of a diterpene compound or a pharmaceutically acceptable salt thereof in the preparation of a medicine for preventing and/or treating acute lung injury, wherein the diterpene compound has the structure shown in formula I:
在某些实施方式中,所述药学上可接受的盐包括酒石酸盐、硬脂酸盐、草酸盐、枸橼酸盐、乳酸盐、山梨酸盐、延胡索酸盐、甲酸盐、乙酸盐、苯甲酸盐、苯磺酸盐、乙磺酸盐、树脂酸盐、三氟乙酸盐、马来酸盐、甲磺酸盐、富马酸盐、氨基酸盐、烟酸盐、磷酸盐、硫酸盐、盐酸盐或氢溴酸盐。In certain embodiments, the pharmaceutically acceptable salts include tartrate, stearate, oxalate, citrate, lactate, sorbate, fumarate, formate, acetic acid Salt, benzoate, benzenesulfonate, ethanesulfonate, resinate, trifluoroacetate, maleate, mesylate, fumarate, amino acid salt, nicotinate, phosphoric acid salt, sulfate, hydrochloride or hydrobromide.
在某些实施方式中,所述药物具有以下效果中的至少一种:In certain embodiments, the medicament has at least one of the following effects:
1)能够抑制肺组织和肺泡灌洗液中TNF-α,IL-1β和IL-6的表达;1) Can inhibit the expression of TNF-α, IL-1β and IL-6 in lung tissue and bronchoalveolar lavage fluid;
2)能够抑制肺组织中髓过氧化物酶的活性;2) Can inhibit the activity of myeloperoxidase in lung tissue;
3)能够抑制肺组织中丙二醛的含量;3) can inhibit the content of malondialdehyde in lung tissue;
4)能够促进肺组织中超氧化物岐化酶的活性;4) Can promote the activity of superoxide dismutase in lung tissue;
5)能够抑制肺组织湿干重比值的增加;和5) can inhibit the increase of the ratio of wet to dry weight of lung tissue; and
6)能够抑制肺泡灌洗液中的蛋白含量。6) Can inhibit the protein content in bronchoalveolar lavage fluid.
在某些实施方式中,所述药物还包括药学上可接受的辅料。In certain embodiments, the medicament further includes pharmaceutically acceptable excipients.
在某些实施方式中,所述辅料包括载体、稀释剂、赋形剂、填充剂、粘合剂、润湿剂、崩解剂、乳化剂、助溶剂、增溶剂、渗透压调节剂、表面活性剂、包衣材料、着色剂、pH调节剂、抗氧剂、抑菌剂或缓冲剂中的任意一种或至少两种的组合。In certain embodiments, the adjuvants include carriers, diluents, excipients, fillers, binders, wetting agents, disintegrants, emulsifiers, cosolvents, solubilizers, osmotic pressure regulators, surface Any one or a combination of at least two of active agents, coatings, colorants, pH adjusters, antioxidants, bacteriostatic agents, or buffers.
例如,所述至少两种的组合可以是稀释剂和赋形剂的组合、粘合剂和润湿剂的组合、乳化剂和助溶剂的组合等,其他任意的组合方式均可选择。For example, the combination of the at least two kinds can be a combination of a diluent and an excipient, a combination of a binder and a wetting agent, a combination of an emulsifier and a cosolvent, etc., and any other combination can be selected.
在某些实施方式中,所述药物的剂型为药学上可接受的任意一种剂型。In certain embodiments, the dosage form of the medicament is any pharmaceutically acceptable dosage form.
在某些实施方式中,所述剂型包括混悬剂、颗粒剂、胶囊剂、散剂、片剂、乳剂、溶液剂、滴丸剂、注射剂、栓剂、灌肠剂、气雾剂、喷雾剂、贴剂或滴剂中的任意一种。In certain embodiments, the dosage forms include suspensions, granules, capsules, powders, tablets, emulsions, solutions, drop pills, injections, suppositories, enemas, aerosols, sprays, patches or any of the drops.
在某些实施方式中,所述药物选自以方式施用:经口服、肠胃外、经鼻、吸入、气管内、肺内和支气管内的施用途径In certain embodiments, the drug is administered selected from the group consisting of oral, parenteral, nasal, inhalation, intratracheal, intrapulmonary and intrabronchial routes of administration
含有本申请的式Ⅰ所示的二萜化合物或其药学上可接受的盐的口服制剂可以包含任何传 统上使用的口服形式,包括片剂、胶囊、含服剂型(buccal forms)、圆锭剂(troches)、菱形锭剂(lozenges)和口服液体、悬浮液或溶液。胶囊可以含有该活性化合物与惰性填料和/或稀释剂的混合物,所述惰性填料和/或稀释剂例如可药用淀粉(例如玉米、马铃薯或木薯淀粉)、糖、人造甜味剂、粉状纤维素(如结晶和微晶纤维素)、面粉、明胶、树胶等等。可用的片剂制剂可以通过传统的压实、湿法造粒或干法造粒法制得,并使用可药用稀释剂、粘合剂、润滑剂、崩解剂、表面改性剂(包括表面活性剂)、助悬或稳定剂,包括但不限于硬脂酸镁、硬脂酸、滑石、十二烷基硫酸钠、微晶纤维素、羧甲基纤维素钙、聚乙烯基吡咯烷酮、明胶、褐藻酸、阿拉伯树胶、黄原胶、柠檬酸钠、复合硅酸盐、碳酸钙、甘氨酸、糊精、蔗糖、山梨糖醇、磷酸二钙、硫酸钙、乳糖、高岭土、甘露醇、氯化钠、滑石、干淀粉和糖粉。表面改性剂包括非离子型和阴离子型表面改性剂。表面改性剂的代表性实例包括但不限于泊洛沙姆188、苯扎氯铵、硬脂酸钙、鲸蜡硬脂醇(Cetostearl alcohol)、Cetomacrogol乳化蜡、失水山梨糖醇酯、胶态二氧化硅、磷酸盐、十二烷基硫酸钠、硅酸镁铝和三乙醇胺。本文中口服制剂可以使用标准延释或缓释制剂以改变活性化合物的吸收。该口服制剂还可以包括在按需要含有适当增溶剂或乳化剂的水或果汁中施用该活性成分。Oral formulations containing the diterpene compound represented by formula I of the present application or a pharmaceutically acceptable salt thereof may comprise any conventionally used oral forms, including tablets, capsules, buccal forms, lozenges (troches), lozenges (lozenges) and oral liquids, suspensions or solutions. Capsules may contain the active compounds in admixture with inert fillers and/or diluents such as pharmaceutically acceptable starches (eg corn, potato or tapioca), sugar, artificial sweeteners, powdered Cellulose (eg crystalline and microcrystalline cellulose), flour, gelatin, gums, and the like. Useful tablet formulations may be prepared by conventional compaction, wet granulation or dry granulation methods using pharmaceutically acceptable diluents, binders, lubricants, disintegrants, surface modifiers (including surface active agent), suspending or stabilizing agents, including but not limited to magnesium stearate, stearic acid, talc, sodium lauryl sulfate, microcrystalline cellulose, calcium carboxymethyl cellulose, polyvinylpyrrolidone, gelatin , alginic acid, gum arabic, xanthan gum, sodium citrate, complex silicate, calcium carbonate, glycine, dextrin, sucrose, sorbitol, dicalcium phosphate, calcium sulfate, lactose, kaolin, mannitol, chloride Sodium, talc, dry starch and powdered sugar. Surface modifiers include nonionic and anionic surface modifiers. Representative examples of surface modifiers include, but are not limited to, poloxamer 188, benzalkonium chloride, calcium stearate, cetostearyl alcohol, cetomacrogol emulsifying wax, sorbitan esters, gums Silica, Phosphate, Sodium Lauryl Sulfate, Magnesium Aluminum Silicate and Triethanolamine. Oral formulations herein can use standard extended or sustained release formulations to alter the absorption of the active compound. The oral formulation may also include administering the active ingredient in water or fruit juice with suitable solubilizers or emulsifiers as required.
在一些情况下,可能合意的是将该化合物以气溶胶形式直接施加到气道中。In some cases, it may be desirable to apply the compound directly into the airways in an aerosol form.
本申请的化合物也可以肠道外给药或腹膜内给药。可以在合适地与表面活性剂(如羟丙基纤维素)混合的水中制备游离碱或可药用盐形式的这些活性化合物的溶液或悬浮液。也可以制备在甘油、液态聚乙二醇和它们在油中的混合物中的分散体。在普通储存和使用条件下,这些制剂含有防腐剂以防止微生物生长。The compounds of the present application may also be administered parenterally or intraperitoneally. Solutions or suspensions of these active compounds in free base or pharmaceutically acceptable salt form can be prepared in water suitably mixed with a surfactant such as hydroxypropylcellulose. Dispersions in glycerol, liquid polyethylene glycols, and mixtures thereof in oils can also be prepared. Under ordinary conditions of storage and use, these preparations contain a preservative to prevent the growth of microorganisms.
适合注射用的药物形式包括无菌水溶液或分散体和用于临时制备无菌可注射溶液或分散体的无菌粉末。在所有情况下,该剂型必须无菌且必须在存在易注射性的程度上流动。其必须在制造和储存条件下稳定且必须防止微生物(如细菌和真菌)的污染作用。载体可以是溶剂或分散介质,含有例如水、乙醇、多元醇(例如甘油、丙二醇和液态聚乙二醇)、它们的合适混合物和植物油。The pharmaceutical forms suitable for injectable use include sterile aqueous solutions or dispersions and sterile powders for the extemporaneous preparation of sterile injectable solutions or dispersion. In all cases, the dosage form must be sterile and must be fluid to the extent that ease of syringability exists. It must be stable under the conditions of manufacture and storage and must be protected against the contaminating action of microorganisms such as bacteria and fungi. The carrier can be a solvent or dispersion medium containing, for example, water, ethanol, polyol (for example, glycerol, propylene glycol, and liquid polyethylene glycol), suitable mixtures thereof, and vegetable oils.
在本申请中,经皮给药可以被理解为包括在身体皮肤和身体通路内衬(包括上皮和粘膜组织)表面上的所有给药。这类给药可以采用本化合物或其可药用盐,以洗液、乳霜、泡沫、贴剂、悬浮液、溶液和栓剂(直肠和阴道)形式进行。In the present application, transdermal administration is understood to include all administrations on the surface of the body skin and body passage linings, including epithelial and mucosal tissues. Such administration can take the form of lotions, creams, foams, patches, suspensions, solutions and suppositories (rectal and vaginal) using the present compound or a pharmaceutically acceptable salt thereof.
另一方面,本申请还提供如式Ⅰ所示的二萜化合物或其药学上可接受的盐在制备抑制肺组织和肺泡灌洗液中TNF-α,IL-1β和IL-6表达的药物中的应用。On the other hand, the present application also provides the diterpene compound represented by formula I or a pharmaceutically acceptable salt thereof in the preparation of a drug for inhibiting the expression of TNF-α, IL-1β and IL-6 in lung tissue and bronchoalveolar lavage fluid applications in .
另一方面,本申请还提供如式Ⅰ所示的二萜化合物或其药学上可接受的盐在制备抑制肺组 织中髓过氧化物酶活性的药物中的应用。On the other hand, the present application also provides the use of the diterpene compound represented by formula I or a pharmaceutically acceptable salt thereof in the preparation of a medicament for inhibiting the activity of myeloperoxidase in lung tissue.
另一方面,本申请还提供如式Ⅰ所示的二萜化合物或其药学上可接受的盐在制备抑制肺组织中丙二醛含量的药物中的应用。On the other hand, the present application also provides the use of the diterpene compound represented by formula I or a pharmaceutically acceptable salt thereof in the preparation of a medicament for inhibiting the content of malondialdehyde in lung tissue.
另一方面,本申请还提供如式Ⅰ所示的二萜化合物或其药学上可接受的盐在制备促进肺组织中超氧化物岐化酶活性的药物中的应用。On the other hand, the present application also provides the use of the diterpene compound represented by formula I or a pharmaceutically acceptable salt thereof in the preparation of a medicament for promoting the activity of superoxide dismutase in lung tissue.
另一方面,本申请还提供如式Ⅰ所示的二萜化合物或其药学上可接受的盐在制备抑制肺组织湿干重比值增加的药物中的应用。On the other hand, the present application also provides the use of the diterpene compound represented by formula I or a pharmaceutically acceptable salt thereof in the preparation of a medicament for inhibiting the increase in the ratio of wet to dry weight of lung tissue.
另一方面,本申请还提供如式Ⅰ所示的二萜化合物或其药学上可接受的盐在制备抑制肺泡灌洗液中的蛋白含量的药物中的应用。On the other hand, the present application also provides the use of the diterpene compound represented by formula I or a pharmaceutically acceptable salt thereof in the preparation of a medicament for inhibiting the protein content in bronchoalveolar lavage fluid.
在某些实施方式中,所述肺组织为急性损伤的肺组织。In certain embodiments, the lung tissue is acutely injured lung tissue.
治疗方法treatment method
另一方面,本申请还提供一种预防和/或治疗急性肺损伤的方法,其包括向有需要的受试者施用有效量的式I所示的二萜化合物或其药学上可接受的盐。On the other hand, the present application also provides a method for preventing and/or treating acute lung injury, comprising administering to a subject in need an effective amount of the diterpene compound shown in formula I or a pharmaceutically acceptable salt thereof .
在某些实施方式中,其中所述施用包括选自口服、肠胃外、经鼻、吸入、气管内、肺内和支气管内的施用途径将治疗有效量的式I所示的二萜化合物或其药学上可接受的盐施用至受试者。In certain embodiments, wherein said administering comprises a route of administration selected from the group consisting of oral, parenteral, nasal, inhalation, intratracheal, intrapulmonary and intrabronchial, a therapeutically effective amount of the diterpene compound of formula I or its A pharmaceutically acceptable salt is administered to a subject.
施用方案可能会影响有效量的构成。可以在本申请所考虑的疾病或病症发作之前或之后将治疗制剂施用至受试者。此外,可以每天或顺序施用若干分开的剂量以及交错剂量,或者可以连续输注该剂量,或者可以是推注。此外,治疗制剂的剂量可以通过治疗或预防情况的紧急程度所指示的成比例地增加或减少。The regimen of administration may affect what constitutes an effective amount. The therapeutic formulation can be administered to a subject before or after the onset of the disease or condition contemplated herein. Furthermore, several divided and staggered doses may be administered daily or sequentially, or the doses may be infused continuously, or they may be bolus injections. Furthermore, the dosage of the therapeutic agent can be proportionally increased or decreased as indicated by the exigencies of the therapeutic or prophylactic situation.
可以使用已知的方法,以有效治疗本申请所考虑的疾病或病症的剂量和时间段,将本申请的药物施用至受试者,优选地为哺乳动物,更优选地为人。实现治疗效果所必需的治疗化合物的有效量可以根据以下因素而变化,比如受试者的疾病或病症的状态;受试者的年龄、性别和体重;以及治疗化合物治疗本申请中考虑的疾病或病症的能力。可以调整剂量方案以提供最佳的治疗反应。例如,如由治疗情况的紧急程度所指示,可以每天施用若干分开的剂量,或者可以成比例地减少剂量。用于治疗本申请的化合物的有效剂量范围的非限制性实例为约0.01mg/kg体重/每天和5000mg/kg体重/每天。本领域普通技术人员将能够研究相关因素,并做出治疗化合物的有效量的决定,而无需过度的实验。The medicaments of the present application can be administered to a subject, preferably a mammal, more preferably a human, using known methods, at doses and for periods of time effective to treat the disease or condition contemplated herein. The effective amount of the therapeutic compound necessary to achieve a therapeutic effect may vary depending on factors such as the state of the disease or disorder of the subject; the age, sex, and weight of the subject; disease capacity. Dosage regimens can be adjusted to provide the best therapeutic response. For example, several divided doses may be administered daily, or the dose may be proportionally reduced, as indicated by the exigencies of the therapeutic situation. Non-limiting examples of effective dosage ranges for the treatment of the compounds of the present application are about 0.01 mg/kg body weight/day and 5000 mg/kg body weight/day. One of ordinary skill in the art will be able to study the relevant factors and make a determination of an effective amount of a therapeutic compound without undue experimentation.
可以改变本申请的药物中活性成分的实际剂量水平,以便获得对于实现特定受试者、药物和施用方式的期望的治疗反应有效的活性成分的量,而对受试者是无毒的。The actual dosage level of the active ingredient in the medicaments of the present application can be varied in order to obtain an amount of the active ingredient effective to achieve the desired therapeutic response for a particular subject, drug and mode of administration, while being non-toxic to the subject.
本申请化合物的治疗有效量或剂量取决于受试者的年龄、性别和体重,受试者当前的医疗状况以及本申请中考虑的疾病或病症的进展。The therapeutically effective amount or dosage of a compound of the present application will depend on the age, sex and weight of the subject, the subject's current medical condition and the progression of the disease or disorder contemplated in the present application.
具有本领域普通技术的医生,例如内科医生或兽医,可以容易地确定所需药物的有效量并对其开处方。例如,内科医生或兽医可以以低于获得所需水平开始在药物中使用的本申请的化合物的剂量,并逐渐增加剂量直到实现期望的效果。A physician of ordinary skill in the art, such as a physician or veterinarian, can readily determine and prescribe the effective amount of the drug required. For example, a physician or veterinarian may start a dose of a compound of the present application for use in medicine below the level required to achieve the desired effect and gradually increase the dose until the desired effect is achieved.
本申请的式I所示的二萜化合物或其药学上可接受的盐的合适的剂量可以在每天约0.01mg至约5,000mg的范围中,比如每天约0.1mg至约1,000mg,例如约1mg至约A suitable dosage of a diterpene compound of formula I of the present application or a pharmaceutically acceptable salt thereof may be in the range of about 0.01 mg to about 5,000 mg per day, such as about 0.1 mg to about 1,000 mg per day, such as about 1 mg per day to about
500mg,比如约5mg至约250mg。该剂量可以以单剂量或多剂量施用,例如每天1至4次或更多次。当使用多剂量时,每个剂量的量可以相同或不同。例如,每天1mg的剂量可以以两个0.5mg的剂量施用,在给药之间具有约12小时的间隔。500 mg, such as about 5 mg to about 250 mg. The dose may be administered in a single dose or in multiple doses, eg, 1 to 4 or more times per day. When multiple doses are used, the amount of each dose can be the same or different. For example, a dose of 1 mg per day may be administered in two doses of 0.5 mg with an interval of about 12 hours between doses.
用于施用的本申请的式I所示的二萜化合物或其药学上可接受的盐可以在约1μg至约10,000mg、约20μg至约9,500mg、约40μg至约9,000mg、约75μg至约8,500mg、约150μg至约7,500mg、约200μg至约7,000mg、约3050μg至约6,000mg、约500μg至约5,000mg、约750μg至约4,000mg、约1mg至约3,000mg、约10mg至约2,500mg、约20mg至约2,000mg、约25mg至约1,500mg、约30mg至约1,000mg、约40mg至约900mg、约50mg至约800mg、约60mg至约750mg、约70mg至约600mg、约80mg至约500mg的范围中,和在其之间的任何和所有完整增量或局部增量。The diterpene compound represented by formula I of the present application or a pharmaceutically acceptable salt thereof for administration may be in the range of about 1 μg to about 10,000 mg, about 20 μg to about 9,500 mg, about 40 μg to about 9,000 mg, about 75 μg to about 8,500 mg, about 150 μg to about 7,500 mg, about 200 μg to about 7,000 mg, about 3050 μg to about 6,000 mg, about 500 μg to about 5,000 mg, about 750 μg to about 4,000 mg, about 1 mg to about 3,000 mg, about 10 mg to about 2,500 mg, about 20 mg to about 2,000 mg, about 25 mg to about 1,500 mg, about 30 mg to about 1,000 mg, about 40 mg to about 900 mg, about 50 mg to about 800 mg, about 60 mg to about 750 mg, about 70 mg to about 600 mg, about 80 mg to In the range of about 500 mg, and any and all full or partial increments therebetween.
在一些实施方式中,本申请的化合物的剂量为约1mg和约2,500mg。在一些实施方式中,用于本文描述的药物的本申请的化合物的剂量小于约10,000mg、或小于约8,000mg、或小于约6,000mg、或小于约5,000mg、或小于约3,000mg、或小于约2,000mg、或小于约1,000mg、或小于约500mg、或小于约200mg、或小于约50mg。In some embodiments, the doses of the compounds of the present application are between about 1 mg and about 2,500 mg. In some embodiments, the dose of a compound of the present application for use in the medicaments described herein is less than about 10,000 mg, or less than about 8,000 mg, or less than about 6,000 mg, or less than about 5,000 mg, or less than about 3,000 mg, or less than About 2,000 mg, or less than about 1,000 mg, or less than about 500 mg, or less than about 200 mg, or less than about 50 mg.
在某些实施方式中,将本申请的药物以每天1至5次或更多次的剂量施用至受试者。在另一个实施方式中,将本申请的药物以包括但不限于每天一次、每两天一次、每三天一次、每周一次、每两周一次的剂量范围施用至受试者。本领域技术人员容易显而易见,本申请的药物的各种组合的施用频率在个体与个体之间不同,这取决于许多因素,包括但不限于年龄、待治疗的疾病或病症、性别、整体健康和其他因素。因此,本申请不应被解释为限于任何具体的剂量方案,并且要施用至任何受试者的精确剂量和药物是通过主治医生考虑有关受试者的所有其他因素来确定的。In certain embodiments, a medicament of the present application is administered to a subject in a dose of 1 to 5 or more times per day. In another embodiment, a medicament of the present application is administered to a subject in a dose range including, but not limited to, once daily, once every two days, once every three days, once a week, once every two weeks. It will be readily apparent to those skilled in the art that the frequency of administration of the various combinations of the medicaments of the present application will vary from individual to individual, depending on a number of factors including, but not limited to, age, disease or condition being treated, gender, overall health and other factors. Accordingly, this application should not be construed as limited to any particular dosage regimen, and the precise dosage and drug to be administered to any subject is determined by the attending physician taking into account all other factors relevant to the subject.
应当理解,在非限制性实例中,可以每天、每隔一天、每2天、每3天、每4天或每5天施用每天给药的化合物的量。例如,每隔一天施用一次,则可以在周一开始每天5mg的剂量,在周三施用第一个随后每天5mg的剂量,在周五施用第二个每天5mg的剂量等。It will be appreciated that, in non-limiting examples, the amount of the compound to be administered daily may be administered every day, every other day, every 2 days, every 3 days, every 4 days, or every 5 days. For example, with administration every other day, a dose of 5 mg per day may be started on Monday, the first subsequent dose of 5 mg per day may be administered on Wednesday, the second dose of 5 mg per day may be administered on Friday, etc.
在某些实施方式中,其进一步包括选自由以下组成的组的至少一种另外的治疗:手术、类固醇疗法、非类固醇疗法、抗病毒疗法、抗真菌疗法、抗微生物疗法、免疫抑制剂疗法、抗感染疗法、抗高血压疗法、营养补充剂及其任意组合。例如,所述类固醇疗法可以包括施用地塞米松、丙酸倍氯米松、布地奈德和/或氟替卡松。In certain embodiments, it further comprises at least one additional treatment selected from the group consisting of surgery, steroid therapy, non-steroid therapy, antiviral therapy, antifungal therapy, antimicrobial therapy, immunosuppressive therapy, Anti-infective therapy, anti-hypertensive therapy, nutritional supplement, and any combination thereof. For example, the steroid therapy may include administration of dexamethasone, beclomethasone propionate, budesonide, and/or fluticasone.
试剂盒Reagent test kit
另一方面,本申请提供一种试剂盒,其包括式I所示的二萜化合物或其药学上可接受的盐。In another aspect, the present application provides a kit comprising the diterpene compound represented by formula I or a pharmaceutically acceptable salt thereof.
在某些实施方案中,该试剂盒包含容器和在该容器上或与该容器相连的标签或包装插页。合适的容器包括例如瓶子、管瓶、注射器等。该容器可以由各种材料如玻璃或塑料形成。该容器容纳或含有能有效治疗所选疾病或失调症的药物并且可具有无菌获取口(例如该容器可以是具有可被皮下注射针刺入的塞子的静脉注射液袋或管瓶)。In certain embodiments, the kit comprises a container and a label or package insert on or associated with the container. Suitable containers include, for example, bottles, vials, syringes, and the like. The container can be formed from various materials such as glass or plastic. The container holds or contains a drug effective to treat the selected disease or disorder and may have a sterile access port (eg, the container may be an IV bag or vial with a stopper that can be penetrated by a hypodermic needle).
所述试剂盒可以进一步包括具有可药用稀释缓冲剂(如注射用抑菌水(BWFI)、磷酸盐缓冲盐水、林格氏溶液和葡萄糖溶液)的第二容器。其可以进一步包括从商业和使用者角度看合意的其它材料,包括其它缓冲剂、稀释剂、过滤器、针和注射器。The kit may further comprise a second container with a pharmaceutically acceptable dilution buffer such as bacteriostatic water for injection (BWFI), phosphate buffered saline, Ringer's solution and dextrose solution. It may further include other materials desirable from a commercial and user standpoint, including other buffers, diluents, filters, needles and syringes.
在其他实施方式中,试剂盒进一步包括疗、预防或减轻急性肺损伤的症状的至少一种另外的药剂,例如地塞米松、丙酸倍氯米松、布地奈德或氟替卡松。In other embodiments, the kit further comprises at least one additional agent that treats, prevents or alleviates symptoms of acute lung injury, such as dexamethasone, beclomethasone propionate, budesonide, or fluticasone.
在某些实施方式中,所述试剂盒还包括施用器和其使用说明材料。所述说明材料可以包括用于预防或治疗急性肺损伤,或本申请中考虑的任何其他疾病或病症的说明书。说明材料叙述了应该施用至受试者的式I所示的二萜化合物或其盐的量和频率。In certain embodiments, the kit further includes an applicator and instructional materials for its use. The instructional material may include instructions for preventing or treating acute lung injury, or any other disease or disorder contemplated in this application. The instructional material recites the amount and frequency of the diterpene compound of formula I, or a salt thereof, that should be administered to the subject.
具体地,本申请提供了以下具体实施方式:Specifically, the present application provides the following specific embodiments:
1、一种小分子二萜化合物或其盐在制备防治急性肺损伤的药物中的应用,其特征在于,所述小分子二萜化合物的结构如式Ⅰ所示。1. Use of a small molecule diterpene compound or a salt thereof in the preparation of a medicine for preventing and treating acute lung injury, characterized in that the structure of the small molecule diterpene compound is shown in formula I.
2、如实施方式1所述的应用,其特征在于,所述盐包括酒石酸盐、硬脂酸盐、草酸盐、枸橼酸盐、乳酸盐、山梨酸盐、延胡索酸盐、甲酸盐、乙酸盐、苯甲酸盐、苯磺酸盐、乙磺酸盐、树脂酸盐、三氟乙酸盐、马来酸盐、甲磺酸盐、富马酸盐、氨基酸盐、烟酸盐、磷酸盐、硫酸盐、盐酸盐或氢溴酸盐。2. The use according to Embodiment 1, wherein the salts include tartrate, stearate, oxalate, citrate, lactate, sorbate, fumarate, and formate , acetate, benzoate, benzenesulfonate, ethanesulfonate, resinate, trifluoroacetate, maleate, mesylate, fumarate, amino acid salt, niacin salt, phosphate, sulfate, hydrochloride or hydrobromide.
3、如实施方式1或2所述的应用,其特征在于,所述药物抑制肺组织和肺泡灌洗液中TNF-α,IL-1β和IL-6的表达。3. The use according to Embodiment 1 or 2, wherein the drug inhibits the expression of TNF-α, IL-1β and IL-6 in lung tissue and bronchoalveolar lavage fluid.
4、如实施方式1-3中任一项所述的应用,其特征在于,所述药物抑制髓过氧化物酶的活性。4. The use according to any one of Embodiments 1-3, wherein the drug inhibits the activity of myeloperoxidase.
5、如实施方式1-4中任一项所述的应用,其特征在于,所述药物抑制肺组织中丙二醛的含量。5. The use according to any one of Embodiments 1 to 4, wherein the drug inhibits the content of malondialdehyde in lung tissue.
6、如实施方式1-5中任一项所述的应用,其特征在于,所述药物促进肺组织中超氧化物岐化酶的活性。6. The use according to any one of embodiments 1-5, wherein the drug promotes the activity of superoxide dismutase in lung tissue.
7、如实施方式1-6中任一项所述的应用,其特征在于,所述药物抑制肺组织湿干重比值的增加。7. The use according to any one of Embodiments 1-6, wherein the drug inhibits the increase of the ratio of wet to dry weight of lung tissue.
8、如实施方式1-7中任一项所述的应用,其特征在于,所述药物抑制肺泡灌洗液中的蛋白含量。8. The use according to any one of embodiments 1-7, wherein the drug inhibits the protein content in bronchoalveolar lavage fluid.
9、如实施方式1-8中任一项所述的应用,其特征在于,所述药物的剂型为药学上可接受的任意一种剂型;9. The use according to any one of Embodiments 1-8, wherein the dosage form of the medicine is any pharmaceutically acceptable dosage form;
优选地,所述剂型包括混悬剂、颗粒剂、胶囊剂、散剂、片剂、乳剂、溶液剂、滴丸剂、注射剂、栓剂、灌肠剂、气雾剂、喷雾剂、贴剂或滴剂中的任意一种。Preferably, the dosage forms include suspensions, granules, capsules, powders, tablets, emulsions, solutions, drop pills, injections, suppositories, enemas, aerosols, sprays, patches or drops any of the .
10、如实施方式1-8中任一项所述的应用,其特征在于,所述药物还包括药学上可接受的辅料;10. The use according to any one of Embodiments 1-8, wherein the medicine further comprises a pharmaceutically acceptable adjuvant;
优选地,所述辅料包括载体、稀释剂、赋形剂、填充剂、粘合剂、润湿剂、崩解剂、乳化 剂、助溶剂、增溶剂、渗透压调节剂、表面活性剂、包衣材料、着色剂、pH调节剂、抗氧剂、抑菌剂或缓冲剂中的任意一种或至少两种的组合。Preferably, the adjuvants include carriers, diluents, excipients, fillers, binders, wetting agents, disintegrants, emulsifiers, cosolvents, solubilizers, osmotic pressure regulators, surfactants, Any one or a combination of at least two of coating materials, colorants, pH adjusters, antioxidants, bacteriostatic agents or buffers.
11、式Ⅰ所示的小分子二萜化合物或其盐在制备抑制肺组织和肺泡灌洗液中TNF-α,IL-1β和IL-6表达的药物中的应用。11. Use of the small molecule diterpene compound represented by formula I or its salt in the preparation of a drug for inhibiting the expression of TNF-α, IL-1β and IL-6 in lung tissue and bronchoalveolar lavage fluid.
12、式Ⅰ所示的小分子二萜化合物或其盐在制备抑制髓过氧化物酶活性的药物中的应用。12. Use of the small-molecule diterpene compound represented by formula I or a salt thereof in the preparation of a medicament for inhibiting the activity of myeloperoxidase.
13、Ⅰ所示的小分子二萜化合物或其盐在制备抑制肺组织中丙二醛含量的药物中的应用。13. Use of the small-molecule diterpene compound shown in I or its salt in the preparation of a medicament for inhibiting the content of malondialdehyde in lung tissue.
14、式Ⅰ所示的小分子二萜化合物或其盐在制备促进肺组织中超氧化物岐化酶活性的药物中的应用。14. Use of the small-molecule diterpene compound represented by formula I or a salt thereof in the preparation of a medicament for promoting the activity of superoxide dismutase in lung tissue.
15、式Ⅰ所示的小分子二萜化合物或其盐在制备抑制肺组织湿干重比值增加的药物中的应用。15. Use of the small-molecule diterpene compound represented by formula I or a salt thereof in the preparation of a medicament for inhibiting the increase in the wet-dry weight ratio of lung tissue.
16、式Ⅰ所示的小分子二萜化合物或其盐在制备抑制肺泡灌洗液中的蛋白含量的药物中的应用。16. Use of the small-molecule diterpene compound represented by formula I or a salt thereof in the preparation of a medicament for inhibiting protein content in bronchoalveolar lavage fluid.
不欲被任何理论所限,下文中的实施例仅仅是为了阐释本申请的二萜化合物或其盐的用途等,而不用于限制本申请发明的范围。Without intending to be limited by any theory, the following examples are only to illustrate the use of the diterpene compounds or their salts of the present application, etc., and are not intended to limit the scope of the invention of the present application.
实施例Example
实施本申请的过程、条件、试剂、试验方法等,除以下专门提及的内容之外,均为本领域的普通知识和公知常识,本申请没有特别限制内容。各实施例中未注明具体条件的试验方法,通常按照常规条件或按照制造厂商所建议的条件。Except for the contents specifically mentioned below, the process, conditions, reagents, test methods, etc. for implementing the present application are all common knowledge and common knowledge in the field, and the present application has no special limited contents. The test methods for which specific conditions are not indicated in each embodiment are usually in accordance with conventional conditions or in accordance with the conditions suggested by the manufacturer.
除非另有说明,本说明书中使用的全部专业术语和科学用语的含义均与本申请所属技术领域的技术人员一般理解的含义相同,但如有冲突,以包含定义的本说明书为准。Unless otherwise specified, all technical and scientific terms used in this specification have the same meanings as commonly understood by those skilled in the art to which this application belongs, but in case of conflict, this specification including definitions shall prevail.
下述实施例所涉及的试验动物为SPF级C57BL/6小鼠(8-10周龄,雄性),购自北京昭衍新药研发技术有限公司,实验动物许可证号:SCXK(苏)2018-0006)。The experimental animals involved in the following examples are SPF grade C57BL/6 mice (8-10 weeks old, male), purchased from Beijing Zhaoyan New Drug Research and Development Technology Co., Ltd., experimental animal license number: SCXK (Su) 2018- 0006).
下述实施例所涉及的试剂包括:The reagents involved in the following examples include:
HY1702溶液:由苏州沪云新药研发股份有限公司提供(分子量:413.98,纯度:98.77%,批号:150818),其分子结构如下所示:HY1702 solution: provided by Suzhou Huyun New Drug Research and Development Co., Ltd. (molecular weight: 413.98, purity: 98.77%, batch number: 150818), its molecular structure is as follows:
地塞米松:购自生工生物工程股份有限公司(分子量:392.5,纯度:≥97%,货号:A601187-0005);Dexamethasone: purchased from Sangon Bioengineering Co., Ltd. (molecular weight: 392.5, purity: ≥97%, article number: A601187-0005);
MDA、MPO、SOD检测试剂盒:购自南京建成生物工程研究所;MDA, MPO, SOD detection kits: purchased from Nanjing Jiancheng Bioengineering Institute;
TNF-α,IL-1β,IL-6的ELISA检测试剂盒:购自武汉博士德生物工程有限公司;ELISA detection kits for TNF-α, IL-1β, and IL-6: purchased from Wuhan Boster Biological Engineering Co., Ltd.;
微量样本总RNA提取试剂盒:购自北京天根生化科技有限公司;Micro-sample total RNA extraction kit: purchased from Beijing Tiangen Biochemical Technology Co., Ltd.;
RNA逆转试剂盒和RT-PCR检测试剂盒:购自日本Takara生物技术有限公司。RNA reversal kit and RT-PCR detection kit: purchased from Takara Biotechnology Co., Ltd., Japan.
下述实施例所涉及的数据处理方式为:The data processing methods involved in the following embodiments are:
用GraphPad Prism 5.0软件进行作图并处理实验数据,数据以均值±标准差(Mean±SEM)表示,两样本均数及样本均数与总体均数之间的比较采用t检验,两个及两个以上样本均数差别的显著性检验采用方差分析;###P<0.001,##P<0.01,#P<0.05与对照组相比;***P<0.001,**P<0.01,*P<0.05与LPS模型组相比。GraphPad Prism 5.0 software was used to draw and process experimental data. The data were expressed as mean ± standard deviation (Mean ± SEM). The comparison between the two-sample mean and the sample mean and the overall mean was performed by t test. ANOVA was used to test the significance of differences in means of more than one sample; ###P<0.001, ##P<0.01, #P<0.05 compared with the control group; ***P<0.001, **P<0.01, *P<0.05 compared to LPS model group.
实施例1Example 1
本实施例探究本申请所涉及的小分子二萜化合物对急性肺损伤小鼠模型的防治效果,包括以下内容:This example explores the prevention and treatment effect of the small molecule diterpene compounds involved in the application on the acute lung injury mouse model, including the following content:
(1)LPS诱导急性肺损伤小鼠模型的建立、分组和给药(1) Establishment, grouping and administration of LPS-induced acute lung injury mouse model
将SPF级C57BL/6小鼠分成6组,每组6只,设置空白对照组、LPS模型组、LPS+HY17025mg/kg组、LPS+HY1702 10mg/kg组、LPS+HY1702 30mg/kg组、LPS+地塞米松(DEX)5mg/kg组(灌胃给药,造模前1h给药)。麻醉小鼠,并手术暴露气管,用胰岛素针将5mg/kg LPS或生理盐水沿小鼠会厌软骨下的空隙缓慢打入,随后缝合伤口。造模6h后处死小鼠,获得小鼠肺泡灌洗液(BALF)及肺组织,进行后续指标检测。The SPF grade C57BL/6 mice were divided into 6 groups with 6 mice in each group. The blank control group, the LPS model group, the LPS+HY17025mg/kg group, the LPS+HY1702 10mg/kg group, the LPS+HY1702 30mg/kg group, and the LPS+HY1702 30mg/kg group were set up. Dexamethasone (DEX) 5 mg/kg group (gavage administration, administration 1 h before modeling). The mice were anesthetized, and the trachea was surgically exposed, and 5 mg/kg LPS or normal saline was slowly injected along the subchondral space of the mouse epiglottis with an insulin needle, and then the wound was sutured. The mice were sacrificed 6 hours after modeling, and the mouse bronchoalveolar lavage fluid (BALF) and lung tissue were obtained for subsequent index detection.
(2)HE染色(2) HE staining
取小鼠肺组织,放入4%甲醛溶液中固定48h后,进行脱水、透明和石蜡包埋并作4μm厚切片,然后进行苏木素-伊红(HE)染色,光镜下观察小鼠肺组织病理变化(炎性细胞浸润、肺泡腔破坏及肺泡间隔增厚等)。The mouse lung tissue was taken and fixed in 4% formaldehyde solution for 48 hours, then dehydrated, transparent and paraffin-embedded, and sliced into 4 μm thick sections. Then, hematoxylin-eosin (HE) staining was performed, and the mouse lung tissue was observed under a light microscope. Pathological changes (infiltration of inflammatory cells, destruction of alveolar spaces and thickening of alveolar septa, etc.).
(3)ELISA检测(3) ELISA detection
采集小鼠支气管肺泡灌洗液,按照试剂盒说明书要求检测支气管肺泡灌洗液中的炎性细胞因子TNF-α,IL-1β和IL-6的含量。The bronchoalveolar lavage fluid from mice was collected, and the contents of inflammatory cytokines TNF-α, IL-1β and IL-6 in the bronchoalveolar lavage fluid were detected according to the instructions of the kit.
(4)RNA提取和RT-PCR检测(4) RNA extraction and RT-PCR detection
采集小鼠肺组织,按照试剂盒说明书要求提取肺组织中RNA,并进行RT-PCR实验,检测肺组织中炎性细胞因子TNF-α,IL-1β和IL-6的mRNA表达。Lung tissue was collected from mice, RNA was extracted from lung tissue according to the kit instructions, and RT-PCR experiment was performed to detect the mRNA expression of inflammatory cytokines TNF-α, IL-1β and IL-6 in lung tissue.
(5)小鼠肺泡灌洗液中蛋白含量测定(5) Determination of protein content in mouse bronchoalveolar lavage fluid
采集小鼠支气管肺泡灌洗液,按照BCA蛋白测定试剂盒实验步骤进行操作,酶标仪检测562nm处的吸光度值,计算小鼠肺泡灌洗液中蛋白浓度。The bronchoalveolar lavage fluid of mice was collected and operated according to the experimental steps of the BCA protein assay kit. The absorbance value at 562 nm was detected by a microplate reader, and the protein concentration in the bronchoalveolar lavage fluid of mice was calculated.
(6)小鼠肺组织湿干重比值测定(6) Determination of wet and dry weight ratio of mouse lung tissue
采集小鼠肺组织,取小鼠右肺上叶组织称重记录湿重数值,70℃烘箱中干燥72h,称重获得肺组织干重数值,计算湿干重比值。The lung tissue of mice was collected, and the right upper lobe tissue of the mouse was weighed to record the wet weight value, dried in an oven at 70 °C for 72 h, weighed to obtain the dry weight value of the lung tissue, and the wet to dry weight ratio was calculated.
(7)MPO活性、MDA含量和SOD活性检测(7) Detection of MPO activity, MDA content and SOD activity
采集小鼠肺组织,取左肺制成组织匀浆液,按照试剂盒说明书要求检测肺组织匀浆液中MPO活性、MDA含量和SOD活性。Lung tissue was collected from mice, and the left lung was taken to make tissue homogenate. The MPO activity, MDA content and SOD activity in the lung tissue homogenate were detected according to the kit instructions.
试验结果如下所示:The test results are as follows:
(1)HY1702减轻LPS诱导的急性肺损伤小鼠(ALI小鼠)肺组织病理学改变,如图1所示(control代表对照组):对照组小鼠肺组织结构完整,肺泡腔清晰,肺泡间隔内无明显炎性细胞的浸润(图1的A);LPS模型组小鼠肺组织结构遭到破坏,肺泡的间隔增厚,且有大量的炎性细胞浸润(图1的B)。经HY1702干预后,小鼠肺部炎症细胞浸润减少,肺组织结构破坏现象和程度得到不同程度缓解(图1的C-E),其中HY1702 30mg/kg组的改善程度最大;DEX给药组小鼠肺组织也同样得到了改善(图1的F)。综上表明HY1702能够通过改善肺泡壁的损坏,减少肺组织炎症细胞的浸润从而起到保护急性肺损伤的作用。(1) HY1702 alleviates the pathological changes of lung tissue in LPS-induced acute lung injury mice (ALI mice), as shown in Figure 1 (control represents the control group): the lung tissue structure of the control group mice is complete, the alveolar cavity is clear, and the alveoli There was no obvious inflammatory cell infiltration in the septum (Figure 1A); the lung tissue structure of the LPS model group was destroyed, the alveolar septa were thickened, and a large number of inflammatory cells infiltrated (Figure 1B). After the intervention of HY1702, the infiltration of inflammatory cells in the lungs of mice was reduced, and the damage to the lung tissue structure was alleviated to varying degrees (CE in Figure 1). Among them, the HY1702 30 mg/kg group had the greatest improvement; Organization also improved (Fig. 1F). In conclusion, HY1702 can protect the acute lung injury by improving the damage of alveolar wall and reducing the infiltration of inflammatory cells in lung tissue.
(2)HY1702减轻LPS诱导的ALI小鼠肺水肿及肺部血气屏障的破坏(2) HY1702 attenuates LPS-induced pulmonary edema and pulmonary blood-gas barrier damage in ALI mice
水肿是炎症的典型症状,肺湿/干重比能够反应肺水肿的程度。如图2所示(control代表 对照组),与对照组相比,LPS诱导后小鼠肺组织湿干重比值显著增加(P<0.001),小鼠肺组织出现肺水肿。HY1702组较LPS组呈剂量依赖性显著降低湿干重比,缓解小鼠肺水肿程度;DEX组小鼠肺水肿程度也得到显著缓解。综上表明HY1702通过减轻急性肺损伤肺水肿,在一定程度上对ALI小鼠起到保护作用。Edema is a typical symptom of inflammation, and the lung wet/dry weight ratio can reflect the degree of pulmonary edema. As shown in Figure 2 (control represents the control group), compared with the control group, the ratio of wet to dry weight in the lung tissue of the mice after LPS induction was significantly increased (P<0.001), and the lung tissue of the mice appeared pulmonary edema. Compared with the LPS group, the HY1702 group significantly reduced the wet-dry weight ratio in a dose-dependent manner, and relieved the degree of pulmonary edema in the mice; the degree of pulmonary edema in the DEX group was also significantly relieved. In conclusion, HY1702 can protect ALI mice to a certain extent by alleviating pulmonary edema in acute lung injury.
BALF中蛋白浓度可反映肺部血气屏障的完整性。如图3所示(control代表对照组),与正常对照组相比,LPS组的BALF中蛋白浓度显著增加。HY1702组较LPS组呈剂量依赖性抑制BALF中蛋白浓度,同时DEX组小鼠BALF中蛋白浓度也得到了显著的抑制(P<0.001)。提示HY1702能够缓解肺部富蛋白液渗出情况,减轻ALI小鼠肺部血气屏障的破坏。The protein concentration in BALF can reflect the integrity of the blood-gas barrier in the lung. As shown in Figure 3 (control represents the control group), the protein concentration in the BALF of the LPS group was significantly increased compared with the normal control group. Compared with the LPS group, the HY1702 group inhibited the protein concentration in BALF in a dose-dependent manner, and the protein concentration in the BALF of the mice in the DEX group was also significantly inhibited (P<0.001). It is suggested that HY1702 can alleviate the exudation of protein-rich fluid in the lungs and reduce the damage of the blood-gas barrier in the lungs of ALI mice.
(3)HY1702抑制ALI小鼠肺组织炎症因子的表达(3) HY1702 inhibits the expression of inflammatory factors in the lung tissue of ALI mice
在炎症发生过程中,大量促炎因子如TNF-α,IL-1β和IL-6的释放,促进机体内环境中炎性因子反应的发生。ELISA及RT-PCR结果表明,与对照组相比,LPS诱导小鼠肺组织中TNF-α,IL-1β,IL-6的蛋白含量(分别如图4中的a、b、c,control代表对照组)及BALF中相应mRNA(分别如图5中的a、b、c,control代表对照组)表达水平均显著增加;HY1702组较LPS组呈剂量依赖性显著抑制TNF-α,IL-1β,IL-6在蛋白及mRNA转录水平上的表达。DEX组也呈现出显著的抑制作用。表明HY1702可以通过抑制ALI小鼠肺组织中炎症细胞因子TNF-α,IL-1β,IL-6的表达,进一步抑制炎症反应的加剧来防治急性肺损伤。In the process of inflammation, a large number of pro-inflammatory factors such as TNF-α, IL-1β and IL-6 are released to promote the occurrence of inflammatory factor responses in the body's internal environment. The results of ELISA and RT-PCR showed that compared with the control group, LPS induced the protein content of TNF-α, IL-1β, and IL-6 in the lung tissue of mice (respectively, a, b, c in Figure 4, control represents control group) and the corresponding mRNA in BALF (a, b, c in Figure 5, respectively, control represents the control group), the expression levels were significantly increased; the HY1702 group significantly inhibited TNF-α and IL-1β in a dose-dependent manner compared with the LPS group , IL-6 expression at the protein and mRNA transcription levels. The DEX group also exhibited a significant inhibitory effect. It indicated that HY1702 could prevent and treat acute lung injury by inhibiting the expression of inflammatory cytokines TNF-α, IL-1β and IL-6 in the lung tissue of ALI mice, and further inhibiting the aggravation of inflammatory response.
(4)HY1702抑制ALI小鼠肺组织中髓过氧化物酶(Myeloperoxidase,以下简称MPO)活性(4) HY1702 inhibited the activity of myeloperoxidase (MPO) in lung tissue of ALI mice
中性粒细胞的积聚是LPS诱导的急性肺损伤的特征之一,MPO的活性是中性粒细胞功能和激活的标志。如图6所示(control代表对照组):与对照组相比,HY1702组的MPO活性均较LPS组明显下降,且呈剂量性抑制;DEX组也呈现出显著的抑制MPO活性作用。表明HY1702可减轻肺组织中性粒细胞浸润的急性炎症反应。Neutrophil accumulation is one of the hallmarks of LPS-induced acute lung injury, and MPO activity is a hallmark of neutrophil function and activation. As shown in Figure 6 (control represents the control group): compared with the control group, the MPO activity of the HY1702 group was significantly lower than that of the LPS group, and it was dose-dependently inhibited; the DEX group also showed a significant inhibitory effect on MPO activity. It was shown that HY1702 can alleviate the acute inflammatory response of neutrophil infiltration in lung tissue.
(5)HY1702抑制ALI小鼠肺部氧化应激反应(5) HY1702 inhibits lung oxidative stress in ALI mice
急性肺损伤发生时体内产生大量的活性氧,机体防御系统处于负荷状态,无法及时清除从而导致过氧化产物在体内堆积,产生一些列氧化产物损伤机体。检测肺组织中氧化相关产物丙二醛(MDA)及超氧化物歧化酶(SOD)可间接反映出机体抗氧化能力。如图7和图8所示(control代表对照组),与对照组相比,LPS组MDA含量显著增加,SOD活性显著下降;HY1702组及DEX组均能较LPS组一定程度上显著抑制MDA含量,增强SOD的活性。表明HY1702可以通过调节氧化应激反应来调控炎症反应的发生,从而防护LPS诱导的急性 肺损伤。When acute lung injury occurs, a large amount of reactive oxygen species is produced in the body, and the body's defense system is in a state of load, which cannot be cleared in time, resulting in the accumulation of peroxidation products in the body, resulting in a series of oxidation products that damage the body. Detection of oxidation-related products malondialdehyde (MDA) and superoxide dismutase (SOD) in lung tissue can indirectly reflect the body's antioxidant capacity. As shown in Figure 7 and Figure 8 (control represents the control group), compared with the control group, the MDA content in the LPS group was significantly increased, and the SOD activity was significantly decreased; both the HY1702 group and the DEX group could significantly inhibit the MDA content to a certain extent compared with the LPS group. , enhance the activity of SOD. It indicated that HY1702 could regulate the occurrence of inflammatory response by regulating oxidative stress response, thereby protecting against LPS-induced acute lung injury.
申请人声明,本申请通过上述实施例来说明本申请的一种小分子二萜化合物或其盐在制备防治急性肺损伤的药物中的应用,但本申请并不局限于上述实施例,即不意味着本申请必须依赖上述实施例才能实施。所属技术领域的技术人员应该明了,对本申请的任何改进,对本申请产品各原料的等效替换及辅助成分的添加、具体方式的选择等,均落在本申请的保护范围和公开范围之内。The applicant declares that this application illustrates the application of a small molecule diterpene compound or its salt in the preparation of medicines for preventing and treating acute lung injury through the above-mentioned examples, but the application is not limited to the above-mentioned examples, that is, it is not limited to the above-mentioned examples. It means that the present application must rely on the above-mentioned embodiments to be implemented. Those skilled in the art should understand that any improvement to the application, the equivalent replacement of each raw material of the product of the application, the addition of auxiliary components, the selection of specific methods, etc., all fall within the scope of protection and disclosure of the application.
以上详细描述了本申请的优选实施方式,但是,本申请并不限于上述实施方式中的具体细节,在本申请的技术构思范围内,可以对本申请的技术方案进行多种简单变型,这些简单变型均属于本申请的保护范围。The preferred embodiments of the present application have been described in detail above. However, the present application is not limited to the specific details of the above-mentioned embodiments. Within the scope of the technical concept of the present application, various simple modifications can be made to the technical solutions of the present application. These simple modifications All belong to the protection scope of this application.
另外需要说明的是,在上述具体实施方式中所描述的各个具体技术特征,在不矛盾的情况下,可以通过任何合适的方式进行组合,为了避免不必要的重复,本申请对各种可能的组合方式不再另行说明。In addition, it should be noted that the specific technical features described in the above-mentioned specific embodiments can be combined in any suitable manner unless they are inconsistent. The combination method will not be specified otherwise.