Application of daphnetin in preparing medicine for treating viral pneumonia

Abstract

The present invention relates to the use of daphnetin in the preparation of a drug for treating viral pneumonia. When daphnetin or a pharmaceutically acceptable salt or solvate thereof is used for the treatment of viral pneumonia, it has a good therapeutic effect. The present invention also provides the use of daphnetin or a pharmaceutically acceptable salt or solvate thereof and a glucocorticoid in the preparation of a drug for treating viral pneumonia, as well as a pharmaceutical composition containing the two and a preparation method thereof. When daphnetin and a glucocorticoid are used in combination for the treatment of viral pneumonia, the two have a synergistic effect and can significantly improve the efficacy of viral pneumonia.

Description

Technical Field

[0001] The invention relates to the field of medical technology, and in particular to application of daphnetin in preparing medicines for treating viral pneumonia. Background Art

[0002] Daphnetin is an active ingredient extracted from Daphne Korean Nakai, a plant of the genus Daphne. It is also known as Daphnetin. It is the first new drug obtained in my country through chemical synthesis. Its chemical name is 7,8-dihydroxycoumarin.

[0003] Daphne has the effects of dilating coronary blood vessels, increasing coronary blood flow, reducing myocardial oxygen consumption, improving myocardial metabolism and promoting cardiac function recovery, dilating peripheral blood vessels, resisting arterial thrombosis and inhibiting platelet aggregation; it also has the anti-inflammatory effect of stimulating the pituitary-adrenal cortex system.

[0004] After absorption, daphneline is widely distributed in the body, with the highest distribution in the kidney, followed by the lung, spleen, and plasma. The main excretion route is the kidney. Clinically, it is mainly used as an adjuvant treatment for thromboangiitis obliterans and other occlusive vascular diseases and coronary heart disease. Summary of the invention

[0005] The purpose of the present invention is to provide the use of daphnetin in the preparation of a medicine for treating viral pneumonia. When daphnetin is used for the treatment of viral pneumonia, it has a good therapeutic effect.

[0006] To this end, in a first aspect, the present invention provides the use of daphnetin or a pharmaceutically acceptable salt or solvate thereof in the preparation of a medicament for treating viral pneumonia.

[0007] In some embodiments, the medicament uses daphnetin or a pharmaceutically acceptable salt or solvate thereof as an active ingredient.

[0008] In some embodiments, the daphnetin or a pharmaceutically acceptable salt or solvate thereof is present in the medicament in an amount effective for treating the viral pneumonia.

[0009] In some embodiments, in the medicament, the content of daphnetin or a pharmaceutically acceptable salt or solvate thereof as an active ingredient is 0.1-99% (w / w).

[0010] In some embodiments, the viral pneumonia includes pneumonia caused by any one or more viruses in the following group: coronavirus (e.g., coronavirus 229E, coronavirus OC43, coronavirus NL63, coronavirus HKU1, SARS coronavirus, MERS coronavirus, 2019 novel coronavirus), influenza virus (e.g., type A H1N1 virus, etc.), parainfluenza virus, respiratory syncytial virus, human metapneumovirus, rhinovirus, adenovirus, herpes virus (e.g., herpes simplex virus, varicella-zoster virus, cytomegalovirus), etc.

[0011] In some embodiments, the drug is administered orally, by injection, implant, spray or inhalation.

[0012] In some embodiments, the dosage form of the drug includes but is not limited to injection, tablet, granule, capsule, pill, sustained-release agent, oral liquid preparation, powder or gel.

[0013] In some embodiments, the drug further comprises a drug carrier or a pharmaceutically acceptable excipient.

[0014] The second aspect of the present invention provides the use of daphnetin or a pharmaceutically acceptable salt or solvate thereof and a glucocorticoid in the preparation of a medicament for treating viral pneumonia.

[0015] In some embodiments, the glucocorticoid comprises at least one selected from the group consisting of prednisone, methylprednisolone, betamethasone, beclomethasone dipropionate, prednisolone, hydrocortisone, and dexamethasone.

[0016] In some embodiments, in the drug, the mass ratio of daphnetin or a pharmaceutically acceptable salt or solvate thereof to glucocorticoid is 1-99:0.1-99.

[0017] In some embodiments, in the drug, the mass percentage of daphnetin or its pharmaceutically acceptable salt or solvate is 0.1-99%.

[0018] In some embodiments, the viral pneumonia includes pneumonia caused by any one or more viruses in the following group: coronavirus (e.g., coronavirus 229E, coronavirus OC43, coronavirus NL63, coronavirus HKU1, SARS coronavirus, MERS coronavirus, 2019 novel coronavirus), influenza virus (e.g., type A H1N1 virus), parainfluenza virus, respiratory syncytial virus, human metapneumovirus, rhinovirus, adenovirus, herpes virus (e.g., herpes simplex virus, varicella-zoster virus, cytomegalovirus), etc.

[0019] In some embodiments, the drug is administered orally, by injection, implant, spray or inhalation.

[0020] In some embodiments, the dosage form of the drug includes but is not limited to injection, tablet, granule, capsule, pill, sustained-release agent, oral liquid preparation, powder or gel.

[0021] In some embodiments, the drug further comprises a drug carrier or a pharmaceutically acceptable excipient.

[0022] The third aspect of the present invention provides a pharmaceutical composition, which comprises daphnetin or a pharmaceutically acceptable salt or solvate thereof and a glucocorticoid.

[0023] In some embodiments, the glucocorticoid comprises at least one selected from the group consisting of prednisone, methylprednisolone, betamethasone, beclomethasone dipropionate, prednisolone, hydrocortisone, and dexamethasone.

[0024] In some embodiments, in the pharmaceutical composition, the mass ratio of daphnetin or a pharmaceutically acceptable salt or solvate thereof to glucocorticoid is 1-99:0.1-99.

[0025] In some embodiments, in the pharmaceutical composition, the mass percentage of daphnetin or its pharmaceutically acceptable salt or solvate is 0.1-99%.

[0026] In some embodiments, the pharmaceutical composition is an oral administration agent, an injection administration agent, an implant administration agent, a spray administration agent or an inhalation administration agent.

[0027] In some embodiments, the dosage form of the pharmaceutical composition includes, but is not limited to, injection, tablet, granule, capsule, pill, sustained-release agent, oral liquid preparation, powder or gel.

[0028] In some embodiments, the pharmaceutical composition further comprises a pharmaceutical carrier or a pharmaceutically acceptable excipient.

[0029] The fourth aspect of the present invention provides a method for preparing the pharmaceutical composition described in the third aspect of the present invention, which comprises uniformly mixing daphnetoclax or a pharmaceutically acceptable salt or solvate thereof, a glucocorticoid and an optional drug carrier or a pharmaceutically acceptable excipient.

[0030] The fifth aspect of the present invention provides a method for treating viral pneumonia, comprising administering to a subject an effective amount of a drug comprising daphnetin or a pharmaceutically acceptable salt or solvate thereof.

[0031] In some embodiments, the medicament further comprises a glucocorticoid.

[0032] In some embodiments, the glucocorticoid comprises at least one selected from the group consisting of prednisone, methylprednisolone, betamethasone, beclomethasone dipropionate, prednisolone, hydrocortisone, and dexamethasone.

[0033] In some embodiments, the subject includes a mammal; for example, a human, a rabbit, a pig, a sheep, a cow, a rat, a mouse, a monkey, etc.

[0034] Compared with the prior art, the technical solution of the present invention has the following beneficial effects:

[0035] (1) The present invention provides the use of daphnetin in the preparation of a drug for treating viral pneumonia, which not only expands the application scope of daphnetin, but also provides a new option for the treatment of viral pneumonia.

[0036] (2) The present invention provides the use of daphnetin and glucocorticoid in combination for the preparation of a drug for treating viral pneumonia. The combination of the two has a synergistic effect in the treatment of viral pneumonia and can significantly improve the treatment effect.

[0037] (3) The present invention provides a pharmaceutical composition for treating viral pneumonia. The pharmaceutical composition has a good effect in treating viral pneumonia, and each component in the drug is a known drug that can be used clinically and has a high safety. BRIEF DESCRIPTION OF THE DRAWINGS

[0038] Various other advantages and benefits will become apparent to those skilled in the art by reading the detailed description of the preferred embodiments below. The accompanying drawings are only used for the purpose of illustrating the preferred embodiments and are not to be considered as limiting the present invention. In the accompanying drawings:

[0039] Figure 1 :For viral pneumonia mouse model, the weight changes of mice after treatment with different doses of daphnetoin;

[0040] Figure 2 : For viral pneumonia mouse model, pathological changes of mouse lung tissue after treatment with different doses of daphneline;

[0041] Figure 3 :For viral pneumonia mouse model, changes in mouse lung coefficient after treatment with different doses of daphnetin; * indicates p < 0.05 compared with the model group; ** indicates p < 0.01 compared with the model group;

[0042] Figure 4:For viral pneumonia mouse model, the changes in the mRNA expression levels of inflammatory cytokines in the lung tissue of mice after treatment with different doses of daphnetin; where * indicates p < 0.05 compared with the model group; ** indicates p < 0.01 compared with the model group; *** indicates p < 0.001 compared with the model group; **** indicates p < 0.0001 compared with the model group;

[0043] Figure 5 : For the viral pneumonia mouse model, the changes in the levels of inflammatory cytokines in the mouse serum after treatment with different doses of daphnetin; where * indicates p < 0.05 compared with the model group; ** indicates p < 0.01 compared with the model group; *** indicates p < 0.001 compared with the model group; **** indicates p < 0.0001 compared with the model group. DETAILED DESCRIPTION

[0044] The exemplary embodiments of the present disclosure will be described in detail below. It should be understood that the present disclosure can be implemented in various forms and should not be limited by the embodiments set forth herein. On the contrary, these embodiments are provided to enable a more thorough understanding of the present disclosure and to fully convey the scope of the present disclosure to those skilled in the art.

[0045] The "range" disclosed in the present invention is defined in the form of a lower limit and an upper limit, and a given range is defined by selecting a lower limit and an upper limit, and the selected lower limit and upper limit define the boundaries of a particular range. The range defined in this way can be inclusive or exclusive of the end values, and can be arbitrarily combined, that is, any lower limit can be combined with any upper limit to form a range. For example, if a range of 60 to 120 and 80 to 110 is listed for a particular parameter, it is understood that a range of 60 to 110 and 80 to 120 is also expected. In addition, if the minimum range values ​​1 and 2 are listed, and the maximum range values ​​3, 4 and 5 are listed, the following ranges can all be expected: 1 to 3, 1 to 4, 1 to 5, 2 to 3, 2 to 4 and 2 to 5. In this application, unless otherwise specified, the numerical range "a to b" represents an abbreviation of any real number combination between a and b, where a and b are both real numbers. For example, the numerical range "0-5" means that all real numbers between "0-5" are listed in this document, and "0-5" is just an abbreviation of these numerical combinations. In addition, when a parameter is expressed as an integer ≥ 2, it is equivalent to disclosing that the parameter is, for example, an integer of 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, etc.

[0046] Unless otherwise specified, all embodiments and optional embodiments of the present invention can be combined with each other to form new technical solutions.

[0047] Unless otherwise stated, all technical features and optional technical features of the present invention can be combined with each other to form a new technical solution.

[0048] Unless otherwise specified, all steps of the present invention may be performed sequentially or randomly, preferably sequentially. For example, the method includes steps (a) and (b), which means that the method may include steps (a) and (b) performed sequentially, or may include steps (b) and (a) performed sequentially. For example, mentioning that the method may also include step (c) means that step (c) may be added to the method in any order, for example, the method may include steps (a), (b) and (c), or may include steps (a), (c) and (b), or may include steps (c), (a) and (b), etc.

[0049] Unless otherwise specified, the "include" and "comprising" mentioned in the present invention represent open-ended or closed-ended expressions. For example, the "include" and "comprising" may represent that other components not listed may also be included or comprised, or may only include or comprise the listed components.

[0050] Unless otherwise indicated, the term "effective amount" of the present invention specifies the amount of the compound sufficient to produce the desired response, such as improvement or complete cure of viral pneumonia. For therapeutic purposes, an effective amount is also the amount in which any harmful effects of a compound are offset by the beneficial effects of treatment. The specific effective amount or sufficient amount will vary with factors such as the specific condition being treated, the patient's physical condition (e.g., the patient's weight, age or sex, the type of object being treated, the duration of treatment). The effective amount can be, for example, expressed in grams, milligrams or micrograms or in milligrams per kilogram of body weight (mg / kg). Alternatively, the effective amount can be expressed in terms of the concentration of the active component (e.g., daphneline of the present disclosure), such as molar concentration, mass concentration, volume concentration, molar concentration, molar fraction, mass fraction and mixing ratio. In addition, those skilled in the art can calculate the human equivalent dose of a drug (e.g., a drug comprising daphneline of the present invention) based on the dosage determined from an animal model.

[0051] Unless otherwise indicated, the term "treatment" refers to the effect of administering an effective amount of daphnetoside to a subject or patient, so that a person suffering from viral pneumonia shows improvement in symptoms of viral pneumonia. The improvement is any observable or measurable improvement, and therefore, those skilled in the art will understand that treatment can improve the condition of a subject or patient, but may not be a complete cure of the disease.

[0052] Unless otherwise indicated, the terms "subject" or "patient" are used interchangeably herein to refer to a mammal that can be treated by the medicament or pharmaceutical composition provided by the present invention. The term "mammal" refers to all members of the mammal class, including humans, primates, livestock and domestic animals, such as rabbits, pigs, sheep, cattle, etc. "Subject" or "patient" refers to both male (male) and female (female) genders unless a gender is specifically indicated. In a preferred embodiment, the subject is a human.

[0053] Unless otherwise specified, the term "drug carrier" refers to a system that can change the way a drug enters the human body and its distribution in the body, control the release rate of the drug, and deliver the drug to the target organ. The present invention does not limit the specific types of drug carriers. In some embodiments, the drug carriers that can be used in the drugs or pharmaceutical compositions provided by the present invention include but are not limited to microcapsules, microspheres, nanoparticles, and liposomes.

[0054] Unless otherwise specified, the term "pharmaceutically acceptable excipient" refers to an excipient that, in addition to the active ingredient, has no significant irritation to the organism and does not damage the biological activity and performance of the active ingredient. The use of pharmaceutically acceptable excipients to prepare pharmaceutical preparations is well known to those of ordinary skill in the art. In some embodiments, the pharmaceutically acceptable excipients that can be used in the medicament or pharmaceutical composition provided by the present invention include, but are not limited to: propellants, solubilizers, cosolvents, emulsifiers, colorants, adhesives, disintegrants, fillers, lubricants, wetting agents, osmotic pressure regulators, stabilizers, glidants, flavoring agents, preservatives, suspending agents, coating materials, fragrances, anti-adhesives, integrators, penetration enhancers, pH regulators, buffers, plasticizers, surfactants, foaming agents, defoamers, thickeners, inclusion agents, humectants, absorbents, diluents, flocculants and deflocculating agents, filter aids, and release retardants.

[0055] In the prior art, daphnetin, as a safe clinical drug, has been widely reported for its use, but has not been used for viral pneumonia. The present invention provides the use of daphnetin or a pharmaceutically acceptable salt or solvate thereof in the preparation of a drug for treating viral pneumonia through research.

[0056] Daphnetin, also known as daphnetolide, ancestral dapoxetine, and daphnetin, is the active ingredient of Daphne Korean Nakai. It is the first new drug obtained by chemical synthesis in my country. Its chemical name is 7,8-dihydroxycoumarin, and its molecular formula is C9H6O4. Its chemical structure is shown in formula (I).

[0057]

[0058] The first aspect of the present invention provides the use of daphnetin or a pharmaceutically acceptable salt or solvate thereof in the preparation of a medicament for treating viral pneumonia.

[0059] The present invention has found through research that the experimental animal BALb / c mice are used as research objects to explore the effects of daphnetin at different doses on the viral pneumonia mouse model. By observing the lungs of mice after different doses of treatment (including appearance and pathological sections), detecting the expression of lung index and related markers, the therapeutic effect of daphnetin on viral pneumonia mice is studied. The experimental results show that daphnetin has a significant therapeutic effect on the viral pneumonia mouse model. Further mechanism studies have found that daphnetin may have the effect of inhibiting the activity of cyclooxygenase and 5-ester oxidase. Daphnetin may also produce anti-inflammatory and anti-allergic reactions by inhibiting the synthesis of prostaglandins and leukotrienes. The target of daphnetin is NSP10 / NSP16 methyltransferase, which has an inhibitory effect on a variety of viruses that cause pneumonia, such as the 2019 novel coronavirus, influenza A (H1N1) virus, etc.

[0060] In some embodiments, the drug uses daphnetin or a pharmaceutically acceptable salt or solvate thereof as an active ingredient. For example, daphnetin or a pharmaceutically acceptable salt or solvate thereof can be the only active ingredient of the drug, or one of the active ingredients.

[0061] In some embodiments, the daphnetin or a pharmaceutically acceptable salt or solvate thereof is present in the medicament in an amount effective for treating the viral pneumonia.

[0062] In some embodiments, in the drug, the mass percentage of daphnetin or its pharmaceutically acceptable salt or solvate as the active ingredient is 0.1-99%; for example, it can be about 0.1%, 0.5%, 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20%, 21%, 22%, 23%, 24%, 25%, 26%, 27%, 28%, 29%, 30%, 31%, 32%, 33%, 34%, 35%, 36%, 37%, 38%, 39%, 40%, 41%, 42%, 43%, 44%, 45%, 46%, 47%, 48%, 49%, 50%, 51%, 52%, 53%, 54%, 55%, 56%, 57%, 58%, 59%, 60%, 61%, 62%, 63%, 64%, 65%, 66%, 67%, 68%, 69%, 70%, 71%, 72%, 73%, 74%, 75%, 76%, 77%, 78%, 79%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, 100%, 101%, 102%, 103%, 104%, 105%, 106%, 107%, 108%, 109%, 110%, 111%, 112%, %, 43%, 44%, 45%, 46%, 47%, 48%, 49%, 50%, 51%, 52%, 53%, 54%, 55%, 56%, 57%, 58%, 59%, 60%, 61%, 62%, 63%, 64%, 65%, 66%, 67%, 68%, 69%, 70%, 71%, 72%, 73%, 74%, 75%, 76%, 77%, 78%, 79%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99% etc.

[0063] In some embodiments, the viral pneumonia includes pneumonia caused by any one or more viruses in the following group: coronavirus, such as HCoV-229E, HCoV-OC43, HCoV-NL63, HCoV-HKU1, SARS coronavirus (SARS-CoV), MERS coronavirus (MERS-CoV), 2019 novel coronavirus (COVID-19), etc.; influenza virus, such as type A (H1N1) virus; parainfluenza virus (PIV); respiratory syncytial virus (RSV); human metapneumovirus (HMPV); rhinovirus (RV); adenovirus (AdV); herpes virus, such as herpes simplex virus (HSV), varicella-zoster virus (VZV), cytomegalovirus (CMV), etc.

[0064] In some embodiments, the drug is administered orally, by injection, implant, spray or inhalation.

[0065] In some embodiments, the dosage form of the drug includes but is not limited to injection, tablet, granule, capsule, pill, sustained-release agent, oral liquid preparation, powder or gel.

[0066] In some embodiments, the drug further comprises a drug carrier or a pharmaceutically acceptable excipient.

[0067] In some embodiments, the dosage form of the drug is a capsule, and the raw materials thereof include, by mass percentage: 80-90% daphnetin, 10-20% filler, 1-5% binder, and 1-5% lubricant.

[0068] In some embodiments, the method for preparing the drug comprises:

[0069] S1. Weighing: Weigh the raw materials according to the following mass percentages: 80-90% daphne, 10-20% filler, 1-5% binder, 1-5% lubricant;

[0070] S2. Prepare the adhesive: dissolve the adhesive in water;

[0071] S3 wet granulation: The daphne is added to the wet granulator, the binder prepared in step S2 is added to the wet granulator, stirred evenly, and the wet granules are formed by a wet granulator;

[0072] S4. Wet granulation: adding the wet granules to a square vibrating screening machine, passing through a sieve of a first mesh number (eg, 20 meshes), to obtain first granules;

[0073] S5. Drying: putting the first particles into a hot air circulation oven to obtain second particles after drying;

[0074] S6. Particle selection: adding the second particles to an oscillating sieve, passing through a second mesh number (eg, 30 mesh) sieve to obtain third particles;

[0075] S7. Granulation: The particles that have not passed the second mesh sieve are added to a multifunctional granulator, and passed through a third mesh sieve (eg, Φ1 sieve) to obtain fourth particles;

[0076] S8 mixing: the steps S6, S7 obtained particles and lubricants, fillers into a three-dimensional mixer and mixed evenly to obtain daphne granules;

[0077] S9. Filling the mixed particles to obtain the drug.

[0078] The second aspect of the present invention provides the use of daphnetin or a pharmaceutically acceptable salt or solvate thereof and a glucocorticoid in the preparation of a medicament for treating viral pneumonia.

[0079] Glucocorticoids are one of the common clinical drugs, but the efficacy of glucocorticoids on viral pneumonia is controversial, and it may also lead to increased mortality, prolonged mechanical ventilation and hospitalization, increased incidence of secondary infections, etc. The present invention finds that by combining the use of daphnetin and glucocorticoids, the two can play a synergistic role in the treatment of viral pneumonia, which has significantly better technical effects than the use of either alone, and can avoid the above-mentioned adverse reactions caused by the use of glucocorticoids alone; this may be related to the lower dosage of glucocorticoids when the two are used in combination, and may also be related to the interaction between the two.

[0080] In some embodiments, the glucocorticoid comprises at least one selected from the group consisting of prednisone, methylprednisolone, betamethasone, beclomethasone dipropionate, prednisolone, hydrocortisone, and dexamethasone.

[0081] In some embodiments, in the drug, the mass ratio of daphnetin or its pharmaceutically acceptable salt or solvate to glucocorticoid is 1-99:0.1-99; for example, 1-90:0.1-90, 1-10:50-90, 10-35:40-80, 50-70:20-30, 60-90:0.1-10, 75-99:1-5, 55-80:10-30, 10:0.1, etc.; for example, in some embodiments, by weight, the weight ratio of daphnetin or its pharmaceutically acceptable salt or solvate in the drug can be selected from the following group: 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18 ,19,20,21,22,23,24,25,26,27,28,29,30,31,32,33,34,35,36,37,38,39,40,41,42,43,44,45,46,47,48,49,50,51,52,53,54,55,56,57,58,5 9, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, etc.;

[0082] The weight percentage of glucocorticoid can be selected from the following group: 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65 3, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, etc.

[0083] In some embodiments, in the drug, the mass percentage of daphnetin or its pharmaceutically acceptable salt or solvate is 0.1-99%; for example, it can be about 0.1%, 0.5%, 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20%, 21%, 22%, 23%, 24%, 25%, 26%, 27%, 28%, 29%, 30%, 31%, 32%, 33%, 34%, 35%, 36%, 37%, 38%, 39%, 40%, 41%, 42%, 43%, 44%, 45%, 46%, 47%, 48%, 49%, 50%, 51%, 52%, 53%, 54%, 55%, 56%, 57%, 58%, 59%, 60%, 61%, 62%, 63%, 64%, 65%, 66%, 67%, 68%, 69%, 70%, 71%, 72%, 73%, 74%, 75%, 76%, 77%, 78%, 79%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, 100%, 101%, 102%, 103%, 104%, 105%, 106%, 107%, 108%, 109%, 110%, 111%, 112%, 3%, 44%, 45%, 46%, 47%, 48%, 49%, 50%, 51%, 52%, 53%, 54%, 55%, 56%, 57%, 58%, 59%, 60%, 61%, 62%, 63%, 64%, 65%, 66%, 67%, 68%, 69%, 70%, 71%, 72%, 73%, 74%, 75%, 76%, 77%, 78%, 79%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, etc.

[0084] In some embodiments, the viral pneumonia includes pneumonia caused by any one or more viruses in the following group: coronavirus, such as HCoV-229E, HCoV-OC43, HCoV-NL63, HCoV-HKU1, SARS coronavirus (SARS-CoV), MERS coronavirus (MERS-CoV), 2019 novel coronavirus (COVID-19), etc.; influenza virus, such as type A (H1N1) virus; parainfluenza virus (PIV); respiratory syncytial virus (RSV); human metapneumovirus (HMPV); rhinovirus (RV); adenovirus (AdV); herpes virus, such as herpes simplex virus (HSV), varicella-zoster virus (VZV), cytomegalovirus (CMV), etc.

[0085] In some embodiments, the drug is administered orally, by injection, implant, spray or inhalation.

[0086] In some embodiments, the dosage form of the drug includes but is not limited to injection, tablet, granule, capsule, pill, sustained-release agent, oral liquid preparation, powder or gel.

[0087] In some embodiments, the drug further comprises a drug carrier or a pharmaceutically acceptable excipient.

[0088] The third aspect of the present invention provides a pharmaceutical composition, which comprises daphnetin or a pharmaceutically acceptable salt or solvate thereof and a glucocorticoid.

[0089] According to the technical solution of the present invention, by combining the use of daphnetoclax and glucocorticoids in a pharmaceutical composition, the two can play a synergistic role in the treatment of viral pneumonia, which has significantly better technical effects than the use of either drug alone, and can avoid the adverse reactions caused by the use of glucocorticoids alone (such as increased mortality, prolonged mechanical ventilation and hospitalization, increased incidence of superinfection, etc.); this may be related to the lower dosage of glucocorticoids when the two are used in combination, and may also be related to the interaction between the two.

[0090] In some embodiments, the glucocorticoid comprises at least one selected from the group consisting of prednisone, methylprednisolone, betamethasone, beclomethasone dipropionate, prednisolone, hydrocortisone, and dexamethasone.

[0091] In some embodiments, in the pharmaceutical composition, the mass ratio of daphnetin or its pharmaceutically acceptable salt or solvate to glucocorticoid is 1-99:0.1-99; for example, 1-90:1-90, 1-10:50-90, 10-35:40-80, 50-70:20-30, 60-90:0.1-10, 75-99:1-5, 55-80:10-30, 10:0.1, etc.; for example, in some embodiments, by weight, the weight ratio of daphnetin or its pharmaceutically acceptable salt or solvate in the drug can be selected from the following group: 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18 ,19,20,21,22,23,24,25,26,27,28,29,30,31,32,33,34,35,36,37,38,39,40,41,42,43,44,45,46,47,48,49,50,51,52,53,54,55,56,57,58,5 9, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, etc.;

[0092] The weight percentage of glucocorticoid can be selected from the following group: 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65 3, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, etc.

[0093] In some embodiments, in the pharmaceutical composition, the mass percentage of daphnetin or its pharmaceutically acceptable salt or solvate is 0.1-99%; for example, it can be about 0.1%, 0.5%, 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20%, 21%, 22%, 23%, 24%, 25%, 26%, 27%, 28%, 29%, 30%, 31%, 32%, 33%, 34%, 35%, 36%, 37%, 38%, 39%, 40%, 41%, 42%, 43%, 44%, 45%, 46%, 47%, 48%, 49%, 50%, 51%, 52%, 53%, 54%, 55%, 56%, 57%, 58%, 59%, 60%, 61%, 62%, 63%, 64%, 65%, 66%, 67%, 68%, 69%, 70%, 71%, 72%, 73%, 74%, 75%, 76%, 77%, 78%, 79%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, etc.

[0094] In some embodiments, the pharmaceutical composition is administered orally, by injection, implant, spray or inhalation.

[0095] In some embodiments, the dosage form of the pharmaceutical composition includes, but is not limited to, injection, tablet, granule, capsule, pill, sustained-release agent, oral liquid preparation, powder or gel.

[0096] In some embodiments, the pharmaceutical composition further comprises a pharmaceutical carrier or a pharmaceutically acceptable excipient.

[0097] Example 1

[0098] The 8-week-old male BALb / c mice were used as the research objects to explore the effects of different doses of daphnetoclax on the viral pneumonia mouse model and analyze the therapeutic effect of daphnetoclax on viral pneumonia mice.

[0099] (I) Establishment of mouse model of viral pneumonia

[0100] The viral pneumonia mouse model was established using the following method, and the model was established using influenza A (H1N1) virus FM1 strain:

[0101] Eight-week-old BALB / c male mice were randomly divided into a control group (Control), a model group (Model), a low-dose Daphnetin treatment group (Daph-Low), a medium-dose treatment group (Daph-Med), a high-dose treatment group (Daph-High) and a dexamethasone treatment group (DEX), with 12 mice in each group. The mouse viral pneumonia model was established after the experimental mice were adaptively fed for seven days in the Class II Biosafety Laboratory of the School of Basic Medical Sciences of Jilin University.

[0102] After the mice were anesthetized with isoflurane, they were fixed on the endotracheal intubation table and intubated by chest light intubation. The specific operation was as follows: the tongue of the mouse was pulled outward with forceps, and a 22G intravenous cannula needle was used as the endotracheal cannula. With the help of the endotracheal cannula lighting, the endotracheal cannula was gently inserted into the mouse trachea, and then 40 μL of PBS-diluted influenza A (H1N1) virus suspension was drawn with a pipette (sterile PBS was used in the control group) and injected along the upper edge of the cannula. After the liquid completely flowed into the lungs, the mouse was placed vertically for 30 seconds to ensure that the liquid completely entered the lungs. After the experiment was completed, the mouse was placed in a cage and waited for it to wake up naturally.

[0103] (II) Treatment with different doses of daphneline

[0104] On the second day after the modeling was completed, the mice in the control group and the model group were given drugs once a day at a fixed time for 8 consecutive days. The mice in the dexamethasone treatment group were given drugs by intraperitoneal injection, and the mice in the other groups were given drugs by gavage. The specific dosages are shown in Table 1. During the period, diet and drinking water were supplied normally, and the general condition of the mice was observed daily and the weight changes were recorded. After three days of treatment (recorded as Day 4), 6 mice were randomly selected from each group, and the peripheral blood of the mice was collected by orbital blood sampling, and the lungs of the mice were dissected to detect the lung index. The pathological changes of the lung tissue were detected after fixation with paraformaldehyde solution; the ELISA and qPCR methods were used to detect the changes in the levels of inflammatory cytokines in the serum and lung tissue of each group of mice. The remaining mice in each group continued to receive treatment, and the sampling experiment was carried out in the same way as above 8 days after the treatment (recorded as Day 9).

[0105] Table 1

[0106] Grouping dose Model Group Same volume of saline Low dose group 25 mg / kg / day Medium dose group 50 mg / kg / day High dose group 100 mg / kg / day Dexamethasone group 10 mg / kg / day

[0107] 3. Evaluation of treatment effect

[0108] 1. Mouse weight

[0109] Viral pneumonia is an infectious disease that is often accompanied by loss of body fluids and a large amount of nutrient consumption, coupled with the impact on the digestive system, which often leads to weight loss in patients. The weight monitoring results of each group of mice are as follows: Figure 1As shown in the figure, compared with the control group, the weight of mice in the model group was significantly reduced. The weight of mice in all treatment groups also decreased, but the decrease level was lower than that in the model group. Among them, the weight of mice in the low-dose daphneline treatment group decreased the least, and the weight loss of mice in the medium-dose and high-dose daphneline treatment groups and the dexamethasone group was more obvious than that in the low-dose daphneline treatment group.

[0110] 2. Pathological changes of lung tissue

[0111] The pathological changes of viral pneumonia caused by influenza virus include: alveolar epithelial damage, alveolar capillary dilation, congestion, widening of alveolar septa, lymphocyte infiltration, proliferation of type II alveolar epithelial cells, and increased inflammatory exudation in the alveolar cavity, and even local or diffuse hyaline membrane formation. HE staining of mouse lung tissue was used to evaluate the lung damage of mice. Figure 2 As shown in the figure, the alveolar structure of the mice in the control group was intact, the thickness of the lung septum was normal, and there was no inflammatory cell infiltration; the alveolar septum of the mice in the model group was significantly thickened, and a large number of inflammatory cells infiltrated the alveoli and lung interstitium, and the structure was chaotic. After 3 days of treatment with daphneline and dexamethasone, the thickening of the alveolar septum of the mice in each treatment group was significantly reduced compared with the mice in the model group, and the infiltration of inflammatory cells was reduced. Among them, except for the positive control dexamethasone, the inflammation reduction in the medium-dose group was the most obvious, followed by the high-dose and low-dose groups. The lung tissue damage of the mice in the model group continued to worsen over time. After 8 days of treatment with low, medium, and high doses of daphneline, the thickening of the alveolar septum of the mice in each treatment group continued to further reduce, and the infiltration of inflammatory cells in the alveoli and lung interstitium was further reduced. Among them, the lung tissue damage of the mice in the medium and high-dose groups was significantly reduced, indicating a good therapeutic effect.

[0112] 3. Lung coefficient

[0113] When pneumonia occurs, due to increased pulmonary vascular permeability, increased tissue fluid exudation, or accompanied by pulmonary edema and fibrosis, the lung wet weight often increases. The lung coefficient (lung coefficient = lung wet weight / body weight × 100%) can be used to assess the degree of lung tissue damage. Figure 3 As shown in the figure, compared with the control group, the lung coefficient of mice in the model group increased significantly, indicating that lung damage was aggravated. After 3 days of treatment with daphneline, the lung coefficients of mice in the low, medium, and high dose groups and the positive control dexamethasone group were lower than those in the model group; after 8 days of treatment with daphneline, the lung coefficients of mice in each group were further reduced, indicating that daphneline can reduce lung damage in mice caused by viral pneumonia.

[0114] 4. Expression levels of inflammatory cytokines mRNA in lung tissue

[0115] Extensive and uncontrollable cytokine storm is one of the important causes of acute respiratory distress syndrome and multiple organ failure during the progression of viral pneumonia. IL-6 and IL-1β are the main participants of inflammatory cytokine storm. Therefore, the mRNA expression of IL-6 and IL-1β in mouse lung tissue homogenate was detected. Figure 4 As shown in the figure, the mRNA expression levels of inflammatory cytokines IL-6 and IL-1β in the lung tissue of mice in the model group were significantly increased, and increased over time. After 3 days of treatment with daphneline, the mRNA expression levels of IL-6 and IL-1β in lung tissue decreased. After 8 days of treatment with daphneline, the mRNA expression levels of the above-mentioned related inflammatory factors in the low, medium and high dose groups were further reduced, and there were significant differences compared with the model group.

[0116] 5. Serum inflammatory cytokine levels

[0117] In addition to causing excessive inflammatory responses in the lungs, inflammatory cytokines can also be released into the blood circulation, leading to severe systemic inflammatory responses. Therefore, the levels of inflammatory cytokines IL-6, IL-1β, TNF-α, and IFN-γ in mouse serum were detected. Figure 5 As shown in the figure, the levels of inflammatory factors in the serum of mice in the model group were significantly increased and gradually worsened over time; while the levels of inflammatory factors in the serum of mice in the positive control dexamethasone group were significantly decreased. It is worth noting that although the levels of inflammatory factors in the serum of mice in the low-dose daphneline treatment group increased significantly after 3 days of daphneline treatment, they were significantly reduced after 8 days of daphneline treatment. Overall, after 8 days of daphneline treatment, the levels of inflammatory factors in the serum of mice in the low-, medium-, and high-dose daphneline treatment groups were significantly reduced, indicating that daphneline can reduce the levels of inflammatory cytokines in the serum of mice with viral pneumonia, showing a good anti-inflammatory effect.

[0118] Example 2

[0119] Based on Example 1, the effect of the combination of daphnetoclax and glucocorticoids on the viral pneumonia mouse model was further investigated.

[0120] The modeling method of viral pneumonia mouse model, as well as the settings of the control group (Control) and the model group (Model) are the same as in Example 1. Each experimental group is intragastrically administered according to the following doses, namely: Group 1, daphnetin 50 mg / kg / d; Group 2: dexamethasone 0.5 mg / kg / d; Group 3: methylprednisolone 30 mg / kg / d; Group 4: daphnetin 25 mg / kg / d, dexamethasone 0.25 mg / kg / d; Group 5, daphnetin 25 mg / kg / d, methylprednisolone 15 mg / kg / d. There were 10 mice in each group, and the drug was administered once a day at a fixed time. After 5 consecutive days of administration, the lung coefficient was calculated and the results are shown in Table 2.

[0121] Table 2

[0122]

[0123] Among them, each experimental group had significant differences compared with the model group (p < 0.05). When low-dose daphnetin was used in combination with dexamethasone or methylprednisolone, a synergistic effect was shown, indicating that the combination of daphnetin and glucocorticoids can produce synergistic effects in the treatment of viral pneumonia. In particular, although the calculated lung coefficient of group 4 was higher than that of the control group, the difference between the two was not significant.

[0124] The above is only a preferred specific embodiment of the present invention, but the protection scope of the present invention is not limited thereto. Any changes or substitutions that can be easily thought of by a person skilled in the art within the technical scope disclosed by the present invention should be included in the protection scope of the present invention. Therefore, the protection scope of the present invention shall be based on the protection scope of the claims.

Claims

1. The use of daphnetin and glucocorticoid in combination for preparing a drug for treating viral pneumonia, wherein: The glucocorticoid is dexamethasone; the mass ratio of daphnetin to glucocorticoid is 10:0.1; and the viral pneumonia is pneumonia caused by the influenza A (H1N1) virus.

2. The use according to claim 1, characterized in that In the medicine, the mass percentage of daphnetin is 0.1-99%.

3. The use according to claim 1 or 2, characterized in that: The medicine is an oral administration agent, an injection administration agent, an implant administration agent, a spray administration agent or an inhalation administration agent.

4. The use according to claim 1 or 2, characterized in that: The dosage form of the medicine is tablet, granule, capsule, pill, sustained-release preparation, oral liquid preparation, powder or gel.

5. The use according to claim 1 or 2, characterized in that: The medicine also includes a drug carrier or a pharmaceutically acceptable excipient.

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