Use of GW3965 in the manufacture of a medicament for reducing doxorubicin-induced cardiac damage

Abstract

This invention discloses the application of GW3965 in the preparation of drugs that alleviate cardiac damage caused by doxorubicin. Experiments have demonstrated that GW3965 can, while inhibiting tumor growth, alleviate cardiac dysfunction related to doxorubicin cardiotoxicity, reduce doxorubicin-induced cardiac fibrosis, and inhibit doxorubicin-induced cardiomyocyte atrophy and apoptosis. This suggests that GW3965 can alleviate cardiac damage caused by doxorubicin, increase the safety of doxorubicin chemotherapy, and has promising clinical application prospects.

Description

Technical Field

[0001] This invention belongs to the pharmaceutical field and relates to the application of compound GW3965 in the preparation of drugs that alleviate cardiac damage caused by doxorubicin. Background Technology

[0002] Doxorubicin is an anthracycline antitumor drug widely used as a chemotherapy agent to treat various types of cancer. However, the irreversible myocardial damage it causes significantly limits its clinical use. A key characteristic of doxorubicin-induced cardiac injury is cardiomyocyte damage, but currently there are no specific drugs targeting this condition. Investigating the pathogenesis of doxorubicin-induced cardiac injury and identifying drugs and treatments with clinical value will significantly improve the clinical benefits of this condition, improve the long-term prognosis of cancer patients undergoing chemotherapy, and has important clinical translational significance. Summary of the Invention

[0003] Our research group has conducted a systematic study on the specific pathological mechanisms of doxorubicin-induced cardiac injury, explored potential therapeutic targets, and carried out cell and animal experiments using GW3965 to treat doxorubicin-induced cardiac injury. We found that this compound not only significantly reduces doxorubicin-induced cardiac pathological changes and cardiac systolic and diastolic dysfunction, reverses myocardial damage, and effectively prevents heart failure, but also reduces tumor growth. Based on these findings, this invention provides the following technical solution.

[0004] This invention provides the application of GW3965, namely 3-[3-[N-(2-chloro-3-trifluoromethylbenzyl)-(2,2-diphenylethyl)amino]propoxy]phenylacetic acid, in the preparation of a drug to alleviate cardiac damage caused by doxorubicin.

[0005] The aforementioned GW3965 can be the hydrochloride salt, specifically 3-[3-[N-(2-chloro-3-trifluoromethylbenzyl)-(2,2-diphenylethyl)amino]propoxy]phenylacetic acid hydrochloride, CAS number: 405911-17-3, with the following chemical structural formula:

[0006]

[0007] The aforementioned cardiac damage caused by doxorubicin includes, but is not limited to, cardiac systolic and diastolic dysfunction, cardiac pathological changes, heart failure, or dilated cardiomyopathy.

[0008] The term "relief" refers to treatment or prevention, i.e., prevention and treatment. Accordingly, the drug is used to treat or prevent conditions caused by doxorubicin, including: impaired cardiac systolic and diastolic function, cardiac pathological changes, heart failure, or dilated cardiomyopathy.

[0009] In one embodiment, the drug is administered to a tumor host undergoing doxorubicin chemotherapy.

[0010] The aforementioned drugs may contain a therapeutically effective amount of GW3965 as the sole active ingredient.

[0011] In another embodiment, the above-mentioned drug is a pharmaceutical composition that, in addition to containing a therapeutically effective amount of GW3965 as an active ingredient, also contains other pharmaceutical ingredients for preventing and treating heart damage.

[0012] As an example, other pharmaceutical ingredients used to prevent cardiac damage from doxorubicin may be the lysophosphatidylcholine receptor 5 antagonist TC LPA5 4 (CAS No.: 1393814-38-4) reported in CN2023100606133 or the myeloperoxidase inhibitor PF-1355 (also known as PF-06281355, CAS No.: 1435467-38-1) reported in CN 2023103256235.

[0013] Optionally, the above-mentioned drug is a pharmaceutical composition that, in addition to containing a therapeutically effective amount of GW3965, also contains a medically acceptable carrier or excipient.

[0014] For example, the pharmaceutically acceptable excipients mentioned above are selected from one or more combinations of buffers, encapsulating agents, fillers, binders, transdermal absorbents, humectants, disintegrants, absorption enhancers, surfactants, colorants, flavoring agents, and adsorbents.

[0015] The dosage form of the above-mentioned drugs can be oral preparations or injections.

[0016] The oral preparations mentioned above are selected from the following group: tablets, capsules (including but not limited to dispersible capsules and gelatin capsules), granules, powders, solutions, and syrups.

[0017] In the oral formulation, the pharmaceutically acceptable carrier includes one or more of the following: fillers or extenders, binders, humectants, disintegrants, absorbents, lubricants, buffers, complexing agents, and colorants.

[0018] When the drug is in the form of an injection, it is suitable for intravenous injection or intravenous infusion.

[0019] This invention is the first to discover that GW3965 can reduce doxorubicin-induced cardiac damage, prevent or treat doxorubicin-induced cardiac damage and related diseases, improve the prognosis of cancer patients receiving doxorubicin chemotherapy, and has promising clinical application prospects. Attached Figure Description

[0020] Figure 1The statistical results of tumor growth, tumor volume, and weight in mice after GW3965 administration are shown. A shows photographs of tumor tissue obtained from mice euthanized and dissected after 6 weeks of doxorubicin injection and feeding; the top row shows tumors in mice not treated with GW3965, and the bottom row shows tumors in mice treated with GW3965. B shows the statistical curves of the change in mean tumor volume over time in mice in the control diet and GW3965-treated groups. C shows the statistical curves of the mean tumor weight in mice in the control diet and GW3965-treated groups after 6 weeks of doxorubicin injection and feeding (ns: no statistical difference; *: P < 0.05; **: P < 0.01; ***: P < 0.001).

[0021] Figure 2 The results of echocardiography and statistical bar graphs of ejection fraction (EF) and fractional shortening (FS) are shown for mice in the untreated group (Control diet) and the GW3965 treated group (GW3965 diet) after 6 weeks of injection of saline and doxorubicin, respectively (ns: no statistical difference; *: P < 0.05; **: P < 0.01; ***: P < 0.001).

[0022] Figure 3 The results of HE and WGA staining of heart tissues from untreated mice (Control diet) and GW3965-treated mice (GW3965 diet) after 6 weeks of injection of saline and doxorubicin, respectively, are shown. In the images, A is an HE-stained photograph of heart tissue; B is a bar chart of cardiomyocyte area; and C is a WGA-stained photograph of heart tissue.

[0023] Figure 4 The results of masson staining of heart tissues from untreated mice (Control diet) and GW3965-treated mice (GW3965 diet) after 6 weeks of injection of saline and doxorubicin, respectively, are shown. The left image is a photograph of the masson staining of heart tissue; the right image is a bar chart of cardiac fibrosis rate (ns: no statistical difference; *: P < 0.05; **: P < 0.01; ***: P < 0.001).

[0024] Figure 5The results show the levels of myocardial injury markers TnT and CK-MB in plasma of untreated mice (Control diet) and GW3965-treated mice (GW3965 diet) 6 weeks after injection of saline and doxorubicin, respectively. The left panel is a bar chart showing the statistical analysis of CK-MB levels in plasma; the right panel is a bar chart showing the statistical analysis of TnT levels in plasma (ns: no statistical difference; *: P < 0.05; **: P < 0.01; ***: P < 0.001).

[0025] Figure 6 The results of TUNEL staining in heart tissues from untreated (Control diet) and GW3965-treated (GW3965 diet) mice after 6 weeks of doxorubicin injection are shown. The left image is a photograph of the TUNEL staining in the heart tissue; the right image is a bar chart of TUNEL-positive cells (ns: no statistical difference; *: P < 0.05; **: P < 0.01; ***: P < 0.001). Detailed Implementation

[0026] The use of GW3965 for cardiac protection has not been previously reported. Our research group is the first to investigate whether GW3965 has a protective effect against doxorubicin-induced cardiac injury, finding that this compound can effectively prevent / treat doxorubicin-induced cardiomyopathy. Animal experiments showed that GW3965 can significantly alleviate cardiac dysfunction related to doxorubicin cardiotoxicity while inhibiting tumor growth, reducing doxorubicin-induced cardiac fibrosis, and inhibiting doxorubicin-induced cardiomyocyte atrophy and apoptosis. This suggests that GW3965 can improve the condition of doxorubicin-induced cardiac injury, providing a new dosing regimen for the prevention and treatment of doxorubicin-induced cardiac injury, and has significant clinical translational value.

[0027] As a clinical application, GW3965 can be formulated into a cardioprotective drug. In this document, to describe its role in a drug or pharmaceutical composition, GW3965 may be referred to as the "active compound".

[0028] The drug may be a single-component drug containing a therapeutically effective amount of GW3965, or a pharmaceutical composition containing other components such as a pharmaceutically acceptable carrier.

[0029] GW3965 can be used as the sole active ingredient in this drug, or it can be used in combination with other drug ingredients used to prevent cardiac damage. For example, it can be used in combination with the lysophosphatidylcholine receptor 5 antagonist TC LPA54 (CAS No.: 1393814-38-4) reported in CN2023100606133 or the myeloperoxidase inhibitor PF-1355 (also known as PF-06281355, CAS No.: 1435467-38-1) reported in CN 2023103256235 as two active compounds administered to tumors receiving doxorubicin chemotherapy.

[0030] As a specific embodiment of the use of two or more drug components in combination, the drug is a pharmaceutical composition containing a therapeutically effective amount of GW3965 and / or other drug components for preventing cardiac damage.

[0031] The above-mentioned pharmaceutical composition containing GW3965 has the combined effect of the chemotherapy drug doxorubicin and GW3965, which can be used to treat malignant tumors and reduce doxorubicin-induced cardiac damage.

[0032] It should be understood that the term “or” as used herein sometimes means “and / or”, and the term “or” sometimes means “and / or”. The term “and / or” as used in phrases such as “A and / or B” is intended to include both A and B; A or B; A (alone); and B (alone). Similarly, the term “and / or” as used in phrases such as “A, B and / or C” is intended to cover each of the following embodiments: A, B and C; A, B or C; A or C; A or B; B or C; A and C; A and B; B and C; A (alone); B (alone); and C (alone).

[0033] As used herein, the phrase "pharmaceutically acceptable" refers to compounds, materials, compositions, and / or dosage forms that, to a reasonable extent of medical judgment, are suitable for use in human and animal tissues without excessive toxicity, irritation, allergic reactions, or other problems or complications, and that are commensurate with a reasonable benefit / risk ratio. Pharmaceutically acceptable carriers are well known in the art and include liquid or solid fillers, diluents, excipients, solvents, or encapsulating materials. Each carrier must be "acceptable" in the sense of compatibility with other components of the formulation and harmlessness to the patient, including, for example, aqueous solutions (such as water or physiologically buffered saline) or other solvents or mediators (such as glycols, glycerin, oils (such as olive oil), or injectable organic esters). Excipients may be selected, for example, to achieve delayed release of the drug or to selectively target one or more cells, tissues, or organs. Pharmaceutical compositions may be in the form of dosage units, such as tablets, capsules (including dispersible capsules and gelatin capsules), granules, powders, solutions, syrups, suppositories, injections, etc.

[0034] As used herein, the term "effective amount" refers to a therapeutic amount required to alleviate at least one or more symptoms of a disease or condition, and involves an adequate amount of medicine to provide the desired effect. Therefore, the term "therapeutic effective amount" refers to a therapeutic amount sufficient to produce a specific effect when administered to a typical subject. In various contexts, effective amount as used herein also includes amounts sufficient to delay the development of a disease condition, alter the course of a disease condition (e.g., but not limited to, slowing the progression of a disease condition), or reverse a disease condition. It should be understood that many methods are known in the art for determining an effective amount for a given application. For example, pharmacological methods for dosage determination can be used in a therapeutic context. In the context of therapeutic or prophylactic application, the amount of composition administered to a subject will depend on the type and severity of the disease and individual characteristics such as general health status, age, sex, weight, and tolerance to the drug. It also depends on the extent, severity, and type of the disease. Those skilled in the art will be able to determine an appropriate dosage based on these and other factors. For example, the therapeutically effective amount of PF-1355 can be determined by referring to its currently safe dosage for use in cancer patients for the treatment of cancer, and by clinical investigation. Appropriate and effective dosage also needs to take into account treatment factors such as drug formulation, individual constitution, weight, age, disease progression, and administration site.

[0035] For reference, when GW3965 is used to prevent and treat a mouse model of doxorubicin-induced cardiomyopathy, the dosage and administration method can be as follows: administer GW3965 in the diet (100 mg / kg) after intravenous injection of doxorubicin (15 mg / kg) into the tail vein of the mouse.

[0036] The drug ingredient GW3965 can also be used in combination with one or more other therapeutic compounds such as TC LPA54 or PF-1355.

[0037] In addition to the main ingredient GW3965, this drug dosage form may also contain a pharmaceutically acceptable carrier. Some examples of materials that can be used as pharmaceutically acceptable carriers include: (1) sugars (such as lactose, glucose, and sucrose); (2) starches (such as corn starch and potato starch); (3) cellulose and its derivatives (such as sodium carboxymethyl cellulose, ethyl cellulose, and cellulose acetate); (4) powdered tragacanth gum; (5) malt; (6) gelatin; (7) talc; (8) excipients (such as cocoa butter and suppository waxes); (9) oils (such as peanut oil, cottonseed oil, safflower oil, sesame oil, olive oil, corn... (10) Diols (such as propylene glycol); (11) Polyols (such as glycerol, sorbitol, mannitol and polyethylene glycol); (12) Esters (such as ethyl oleate and ethyl laurate); (13) Agar; (14) Buffers (such as magnesium hydroxide and aluminum hydroxide); (15) Alginate; (16) Atherless water; (17) Isotonic saline; (18) Ringer's solution; (19) Ethanol; (20) Phosphate buffer solution; and (21) Other non-toxic and compatible substances used in the pharmaceutical preparation.

[0038] The pharmaceutical formulation can be administered to the body via any of a number of routes of administration, including, for example, oral (e.g., as a drenching, tablet, capsule (including dispersible capsules and gelatin capsules), granules, powder, or paste for application to the tongue) in an aqueous or non-aqueous solution or suspension; absorption through the oral mucosa (e.g., sublingual); subcutaneous; transdermal (e.g., as a patch for application to the skin); and topical (e.g., as a cream, ointment, or spray for application to the skin). The GW3965 compound can also be formulated for inhalation. In some embodiments, the GW3965 compound can simply be dissolved or suspended in a sterile solvent.

[0039] The term "subject" as used above refers to a person or an animal. Generally, an animal is a vertebrate, such as a primate, rodent, livestock, or game animal. Primates include chimpanzees, cynomolgus monkeys, spider monkeys, and macaques, such as rhesus monkeys. Rodents include mice, rats, prairie dogs, ferrets, rabbits, and hamsters. Livestock and game animals include cattle, horses, pigs, deer, bison, buffalo, feline species (e.g., domestic cats), and canine species (e.g., dogs, foxes, wolves). In some embodiments, the subject is a mammal, such as a primate like a human. The terms "individual," "patient," and "subject" are used interchangeably herein. Preferably, the subject is a mammal. Mammals can be humans, non-human primates, mice, rats, dogs, cats, horses, or cattle, but are not limited to these examples. The subject can be male or female.

[0040] The pharmaceutical formulation can be conveniently presented in a single dosage form and can be prepared by any method well known in the pharmaceutical field. The amount of active ingredient that can be combined with a carrier material to produce a single dosage form will vary depending on the host being treated and the specific mode of administration. The amount of active ingredient that can be combined with a carrier material to produce a single dosage form will generally be the amount of the compound that produces the therapeutic effect. Typically, this amount, by weight, ranges from about 1% to about 99% of the active ingredient, for example, from about 5% to about 70%.

[0041] Methods for preparing these formulations or compositions include the steps of combining an active compound (such as GW3965) with a carrier and optionally one or more auxiliary components. Typically, formulations are prepared by uniformly and closely combining GW3965 with a liquid carrier or a finely pulverized solid carrier, or both, and then shaping the product (if necessary).

[0042] The formulations of the present invention suitable for oral administration may be in the following forms: capsules (including dispersible capsules and gelatin capsules), flat capsules, pills, tablets, sugar tablets (using a flavoring base, typically sucrose and gum arabic or tragacanth), lyophilic substances, powders, granules, or as solutions or suspensions in aqueous or non-aqueous liquids, or as oil-in-water or water-in-oil liquid emulsions, or as elixirs or syrups, or as soft tablets (using an inert matrix (such as gelatin and glycerin, or sucrose and gum arabic)) and / or as mouthwashes, etc., each containing a predetermined amount of GW3965 as the active ingredient. The compositions or compounds may also be administered as large pills, medicated tablets, or pastes.

[0043] To prepare solid dosage forms (capsules (including dispersible capsules and gelatin capsules), tablets, pills, sugar-coated pills, powders, granules, etc.) for oral administration, the active ingredient is mixed with one or more pharmaceutically acceptable carriers (such as sodium citrate or dicalcium phosphate) and / or any of the following: (1) fillers or extenders (such as starch, lactose, sucrose, glucose, mannitol and / or silicic acid); (2) binders (such as, for example, carboxymethyl cellulose, alginate, gelatin, polyvinylpyrrolidone, sucrose and / or gum arabic); (3) humectants (such as glycerin); (4) Disintegrants (such as agar, calcium carbonate, potato starch or cassava starch, alginate, certain silicates, and sodium carbonate); (5) Solution blockers (such as paraffin); (6) Absorption accelerators (such as quaternary ammonium compounds); (7) Wetting agents (such as, for example, cetyl alcohol and glyceryl monostearate); (8) Absorbents (such as kaolin and bentonite); (9) Lubricants (such as talc, calcium stearate, magnesium stearate, solid polyethylene glycol, sodium lauryl sulfate, and mixtures thereof); (10) Complexing agents (such as modified or unmodified cyclodextrin); and (11) Colorants. In the case of capsules (including dispersible capsules and gelatin capsules), tablets, and pills, the pharmaceutical composition may also contain buffers. Similar types of solid compositions may also be used as fillers in soft-filled and hard-filled gelatin capsules, which use such excipients as lactose and high molecular weight polyethylene glycol.

[0044] Tablets can be manufactured by compression or molding, optionally using one or more excipients. Compressed tablets can be prepared using binders (e.g., gelatin or hydroxypropyl methylcellulose), lubricants, inert diluents, preservatives, disintegrants (e.g., sodium glycolate starch or croscarmellose sodium), surfactants, or dispersants. Molded tablets can be manufactured by molding a mixture of powdered compounds wetted with an inert liquid diluent in a suitable machine.

[0045] Tablets and other solid dosage forms of pharmaceutical compositions (such as sugar-coated pills, capsules (including dispersible capsules and gelatin capsules), pellets, and granules) may optionally be scored or prepared with coatings and shells (such as enteric coatings and other coatings well known in the field of pharmaceutical formulation). They may also be formulated to provide slow or controlled release of the active ingredient therein, using, for example, different proportions of hydroxypropyl methylcellulose to provide a desired release profile, other polymer matrices, liposomes, and / or microspheres. They may be sterilized, for example, by filtration through a bacterial-retaining filter, or by a sterilizing agent incorporated into the form of a sterile solid composition, which may be dissolved in sterile water or some other sterile injectable medium just before use. These compositions may also optionally contain an emulsifier and may be compositions that release one or more active ingredients only or preferentially in a portion of the gastrointestinal tract (optionally, in a delayed manner). Examples of encapsulation compositions that may be used include polymeric substances and waxes. The active ingredient may also be in a microencapsulated form, which, where appropriate, has one or more of the excipients described above.

[0046] Liquid dosage forms suitable for oral administration include pharmaceutically acceptable emulsions, reconstituted lyophilic agents, microemulsions, solutions, suspensions, syrups, and elixirs. In addition to the active ingredient, liquid dosage forms may contain inert diluents commonly used in the art (e.g., water or other solvents, cyclodextrins and their derivatives), solubilizers, and emulsifiers (such as ethanol, isopropanol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, propylene glycol, 1,3-butanediol, oils (particularly cottonseed oil, peanut oil, corn oil, germ oil, olive oil, castor oil, and sesame oil), glycerin, tetrahydrofuranol, fatty acid esters of polyethylene glycol and sorbitol, and mixtures thereof).

[0047] In addition to inert diluents, oral compositions may also contain adjuvants such as wetting agents, emulsifiers and suspending agents, sweeteners, flavoring agents, coloring agents, aroma agents and preservatives.

[0048] In addition to the active compound, the suspension may contain suspending agents (such as, for example, ethoxylated isostearyl alcohol, polyoxyethylene sorbitol and dehydrated sorbitol esters, microcrystalline cellulose, aluminum hydroxide, bentonite, agar and tragacanth gum, and mixtures thereof).

[0049] Dosage forms for topical or transdermal application include powders, sprays, ointments, pastes, creams, lotions, gels, solutions, patches, and inhalers. Active compounds can be mixed under sterile conditions with pharmaceutically acceptable carriers and with any preservatives, buffers, or propellants that may be necessary.

[0050] In addition to active compounds, ointments, pastes, creams and gels may contain excipients (such as animal and vegetable fats, oils, waxes, paraffins, starch, tragacanth gum, cellulose derivatives, polyethylene glycol, silicones, bentonite, silicic acid, talc and zinc oxide, or mixtures thereof).

[0051] In addition to the active compound, powders and sprays may contain excipients (such as lactose, talc, silica, aluminum hydroxide, calcium silicate, and polyamide powder, or mixtures of these substances). Sprays may additionally contain conventional propellants (such as chlorofluorocarbons and volatile unsubstituted hydrocarbons such as butane and propane).

[0052] Transdermal patches offer the added advantage of providing controlled delivery of active compounds into the body. Such dosage forms can be manufactured by dissolving or dispersing the active compound in a suitable medium. Absorption enhancers can also be used to increase the flux of the compound across the skin. The rate of such flux can be controlled by providing a rate-controlled membrane or by dispersing the compound in a polymer matrix or gel.

[0053] Examples of suitable aqueous and non-aqueous carriers that can be used in the pharmaceutical compositions of the present invention include water, ethanol, polyols (such as glycerol, propylene glycol, polyethylene glycol, etc.) and suitable mixtures thereof, vegetable oils (such as olive oil), and injectable organic esters (such as ethyl oleate). For example, appropriate flowability can be maintained by using a coating material (such as lecithin), by maintaining the desired particle size in the case of a dispersion, and by using a surfactant.

[0054] These compositions may also contain adjuvants (such as preservatives, wetting agents, emulsifiers, and dispersants). Antimicrobial activity can be ensured by including various antibacterial and antifungal agents (e.g., methylparaben, chlorobutanol, phenolic sorbic acid, etc.). Including isotonic agents (such as sugars, sodium chloride, etc.) in the composition is also desirable. Furthermore, prolonged absorption of injectable drug forms can be achieved by including agents that delay absorption (such as aluminum monostearate and gelatin).

[0055] In some cases, to prolong the effect of a drug, it is desirable to slow the absorption of drugs administered subcutaneously or intramuscularly. This can be achieved by using liquid suspensions of crystalline or amorphous materials with poor water solubility. The absorption rate of the drug then depends on its dissolution rate, which in turn can depend on the crystal size and crystal form. Alternatively, delayed absorption of parenteral drug forms can be achieved by dissolving or suspending the drug in an oil medium.

[0056] Injectable reservoir formulations are created by forming a microencapsulated matrix of the subject compound within a biodegradable polymer, such as poly(lactic-co-glycolic acid). The drug release rate can be controlled depending on the drug-to-polymer ratio and the properties of the specific polymer used. Other examples of biodegradable polymers include poly(orthoesters) and poly(anhydrides). Injectable reservoir formulations are also prepared by trapping the drug within tissue-compatible liposomes or microemulsions.

[0057] For use in the methods of the present invention, the active compound may be administered on its own or as a pharmaceutical composition containing, for example, 0.1% to 99.5% (more preferably, 0.5% to 90%) of an active ingredient in combination with a pharmaceutically acceptable carrier.

[0058] The actual dose level of the active ingredient in a pharmaceutical composition can be varied to obtain an amount of the active ingredient that is effective in achieving the desired therapeutic response for a particular patient, composition, and administration mode, without being toxic to the patient.

[0059] The chosen dose level will depend on a variety of factors, including the specific compound or combination of compounds used, or the activity of its esters, salts or amides, the route of administration, the time of administration, the excretion rate of one or more specific compounds used, the duration of treatment, other drugs, compounds and / or materials used in combination with one or more specific compounds used, the age, sex, weight, condition, general health and prior medical history of the patient being treated, and similar factors well known in the medical field.

[0060] Physicians or veterinarians with ordinary skills in the art can readily determine and prescribe a therapeutically effective amount of the desired pharmaceutical composition. For example, a physician or veterinarian may begin with a dose of the pharmaceutical composition or compound at a level below that required to achieve the desired therapeutic effect and gradually increase the dose until the desired effect is achieved. "Therapeutically effective amount" refers to the concentration of the compound sufficient to cause the desired therapeutic effect. It is generally understood that the effective amount of the compound will vary depending on the subject's weight, sex, age, and medical history. Other factors affecting the effective amount may include, but are not limited to, the severity of the patient's condition, the condition being treated, the stability of the compound, and (if necessary) another type of therapeutic agent administered in conjunction with PF-1355. Larger total doses can be delivered through multiple administrations of the pharmaceutical agent. Methods for determining efficacy and dosage are known to those skilled in the art.

[0061] Generally, the suitable daily dose of the active compound used in the compositions and methods of the present invention will be the amount of the lowest dose of compound that is effective in producing a therapeutic effect. Such an effective dose will generally depend on the factors described above.

[0062] If desired, the effective daily dose of the active compound can be administered as separate sub-dose (one, two, three, four, five, six, or more), optionally in unit dose form, at appropriate time intervals throughout the day. In some embodiments of the invention, the active compound may be administered two or three times daily. In other embodiments, the active compound will be administered once daily.

[0063] Patients receiving this treatment are any animals in need, including primates (especially humans); other mammals (such as horses, cattle, pigs, sheep, cats, and dogs); poultry; and pets.

[0064] The present invention will be further described in detail below with reference to specific embodiments. It should be understood that the following embodiments are for illustrative purposes only and are not intended to limit the scope of the invention.

[0065] In embodiments of the present invention, unless otherwise specified, the experimental operating temperature generally refers to room temperature (10-30°C).

[0066] This article involves the addition amount, content and concentration of various substances. Unless otherwise specified, the percentage content mentioned refers to the mass percentage.

[0067] Statistical analysis: In this study, all numerical variables are expressed as mean ± standard error. Two-tailed Student's t-tests were used for comparisons between two groups, and ANOVA tests were used for comparisons among three groups. A p-value < 0.05 was considered statistically significant.

[0068] Example 1: Grouping of experimental animals and construction of animal models

[0069] Forty 8-week-old male melanoma-bearing A375 cell line mice (1×10⁶ cells per cell line) were randomly selected using a randomized table method. 5 C57BL / 6J mice (purchased from Shanghai Jiesijie Laboratory Animal Co., Ltd.) were administered doxorubicin (2.5 mg / kg) intraperitoneally every other day for a total of 6 injections, with a cumulative dose of 15 mg / kg. They were divided into two groups: a control group (Control diet) and a GW3965 treatment group (GW3965 diet), with 20 mice in each group. GW3965 was purchased from MedChemExpress (catalog number HY-10627A) and prepared as a mouse feed (100 mg / kg), which was continuously fed to the mice for 6 weeks after the first injection of doxorubicin.

[0070] Example 2: Echocardiography assessment of diastolic and systolic cardiac function in mice

[0071] Mice were anesthetized by inhalation with 2% isoflurane, fixed on an ultrasound examination table, and connected to an electrocardiogram (ECG) to maintain a heart rate of 400-500 beats / min. Transthoracic echocardiography was performed on the mice using a Vevo 2100 Doppler ultrasound system to assess cardiac function and structure. Cardiac function parameters were measured and calculated, including ejection fraction (EF), fractional shortening (FS), left ventricular end-diastolic diameter (LVEDD), and left ventricular end-diastolic volume (LVESV). Each measurement was averaged over at least five consecutive cardiac cycles.

[0072] Example 3: TUNEL staining test

[0073] The apoptosis level of cardiomyocytes in cardiac tissue was assessed using the TUNEL apoptosis detection kit from Beyotime Biotechnology Co., Ltd. The specific steps were as follows: Tissue sections were fixed with fixative at room temperature for 30 minutes. The fixative was removed, and punching buffer was added, followed by incubation at room temperature for 15 minutes. The punching buffer was removed, and 5% BSA was added for blocking at room temperature for 1 hour. 50 μL of TUNEL detection solution was added, and the sections were incubated at 37°C in the dark for 60 minutes. The blocking buffer was removed, and the prepared primary antibody was added to the tissue, ensuring sufficient coverage for each sample. The slides were placed in a humidified chamber and incubated overnight at 4°C. The primary antibody was removed, and after washing, secondary antibody was added and incubated at room temperature in the dark for 1 hour. The secondary antibody was removed, and the slides were washed three times. The slides were mounted with DAPI anti-quenching mounting medium, ensuring a tight fit between the coverslip and the tissue slide without air bubbles. The slides were then photographed using a confocal microscope.

[0074] Six weeks after the first injection of doxorubicin, cardiac function in mice was assessed by echocardiography, fibrosis was detected by Masson staining, cardiomyocyte area was detected by HE and WGA staining, myocardial damage was detected by plasma TnT and CK-MB, and cardiomyocyte apoptosis in mice in different treatment groups was detected by TUNEL staining.

[0075] Experimental results

[0076] 1. See Figure 1 In mice treated with doxorubicin, tumor volume continued to increase over time after the first injection, while tumor volume increased slowly in mice treated with GW3965 (GW3965 diet), indicating that GW3965 can inhibit tumor growth.

[0077] See Figure 1 In Figure A, the top row shows tumors in mice that were not treated with GW3965, and the bottom row shows tumors in mice that were treated with GW3965. The difference in tumor size between the untreated group (Control diet) and the GW3965-treated group (GW3965 diet) suggests that GW3965 can significantly inhibit tumor growth.

[0078] See Figure 1 In the study, after 6 weeks of doxorubicin injection and feeding, the average tumor weight in the untreated group (Control diet) was about 1.0g, while that in the GW3965 treated group (GW3965 diet) was about 0.43g, indicating that GW3965 inhibited tumor growth.

[0079] 2. See Figure 2 Echocardiographic results in mice showed that GW3965 could improve doxorubicin-induced cardiac dysfunction.

[0080] See Figure 2 In mice in the control diet (B group), after 6 weeks of saline injection, the ejection fraction (EF) was approximately 80%, while after 6 weeks of doxorubicin injection, the EF was approximately 52%. In the GW3965-treated mouse group (GW3965 diet), after 6 weeks of saline injection, the EF was approximately 78%, while after 6 weeks of doxorubicin injection, the EF was approximately 63%. These results indicate that GW3965 can improve the ejection fraction (EF%) in a mouse model of doxorubicin-induced cardiac injury.

[0081] See Figure 2In the control group (C), mice injected with saline for 6 weeks showed a fractional shortening (FS) of approximately 48%, while mice injected with doxorubicin for 6 weeks showed a FS of approximately 26%. Mice treated with GW3965 (GW3965 diet) for 6 weeks showed a FS of approximately 52%, while mice injected with doxorubicin for 6 weeks showed a FS of approximately 32%. These results indicate that GW3965 can improve the fractional shortening (FS%) in a mouse model of doxorubicin-induced cardiac injury.

[0082] By comparing with injected saline solution Figure 2 It also indicates that GW3965 does not affect heart function.

[0083] Improved cardiac function, with increased ejection fraction (EF%) and fractional shortening (FS%), suggests improved survival in the doxorubicin-induced cardiac injury mouse model.

[0084] 3. See also Figure 3 HE and WGA staining results of cardiac tissue in the central anterior chamber (AC) showed that GW3965 alleviated doxorubicin-induced cardiomyocyte atrophy. WGA staining showed that, compared to saline treatment, DOX treatment caused a significant reduction in cardiomyocyte cross-sectional area, while GW3965 treatment significantly alleviated the DOX-induced reduction in myocardial cross-sectional area. HE staining of longitudinal sections of the heart revealed that, compared to saline treatment, DOX treatment caused cardiac atrophy, while GW3965 treatment significantly alleviated DOX-induced cardiac atrophy.

[0085] See Figure 3 In mice in the AB control group (control diet), the cardiomyocyte area was 373 μm² after 6 weeks of injection with saline. 2 Around 272 μm², while after 6 weeks of doxorubicin injection, the cardiomyocyte area was 272 μm². 2 Approximately; after 6 weeks of injection of physiological saline, the cardiomyocyte area of ​​mice in the GW3965 treatment group (GW3965 diet) was 380 μm². 2 Around 291 μm², while after 6 weeks of doxorubicin injection, the cardiomyocyte area was 291 μm². 2 The results further indicate that GW3965 alleviated doxorubicin-induced cardiomyocyte atrophy.

[0086] 4. See Figure 4 The left-middle image shows the results of Masson staining of cardiac tissue. It indicates that DOX treatment increased the area of ​​cardiac fibrosis compared to saline treatment, while GW3965 treatment significantly reduced the area of ​​DOX-induced cardiac fibrosis. GW3965 also alleviated doxorubicin-induced cardiac fibrosis in mice.

[0087] See Figure 4 In the right-middle figure, the cardiac fibrosis rate in mice in the untreated group (Control diet) after 6 weeks of saline injection was approximately 2.2%, while the rate after 6 weeks of doxorubicin injection was approximately 8.0%. In the GW3965-treated group (GW3965 diet), the cardiac fibrosis rate after 6 weeks of saline injection was approximately 2.2%, while the rate after 6 weeks of doxorubicin injection was approximately 4.0%. These results further indicate that GW3965 alleviates doxorubicin-induced cardiac fibrosis.

[0088] 5. See also Figure 5 In the untreated group (Control diet), the plasma CK-MB level was approximately 7 ng / mL after 6 weeks of injection with saline, while it was approximately 31 ng / mL after 6 weeks of injection with doxorubicin. In the GW3965 treated group (GW3965 diet), the plasma CK-MB level was approximately 6 ng / mL after 6 weeks of injection with saline, while it was approximately 17 ng / mL after 6 weeks of injection with doxorubicin.

[0089] In the untreated group (Control diet), the plasma TnT level was approximately 6 ng / mL after 6 weeks of injection with saline, while it was approximately 320 ng / mL after 6 weeks of injection with doxorubicin. In the GW3965 treated group (GW3965 diet), the plasma TnT level was approximately 6 ng / mL after 6 weeks of injection with saline, while it was approximately 172 ng / mL after 6 weeks of injection with doxorubicin.

[0090] The results of two myocardial injury markers showed that GW3965 reduced the levels of TnT and CK-MB in the plasma of mice with doxorubicin-induced cardiac injury.

[0091] 6. See also Figure 6 TUNEL staining results of cardiac tissue from mice in the untreated group (Control diet) and the GW3965 treated group (GW3965 diet) 6 weeks after injection of doxorubicin showed that GW3965 could reduce the number of DOX-induced TUNEL-positive cardiomyocytes.

[0092] After 6 weeks of doxorubicin injection, the TUNEL-positive cell rate in untreated mice (Control diet-DOX) was approximately 12.6%; after 6 weeks of doxorubicin injection, the TUNEL-positive cell rate in GW3965-treated mice (GW3965 diet-DOX) was approximately 7.6%, further indicating that GW3965 can alleviate doxorubicin-induced cardiomyocyte apoptosis in mice.

[0093] The above experimental results confirm that GW3965 can reduce cardiac dysfunction related to doxorubicin cardiotoxicity while inhibiting tumor growth, reduce doxorubicin-induced cardiac fibrosis, and inhibit doxorubicin-induced cardiomyocyte atrophy and apoptosis. This suggests that GW3965 can reduce doxorubicin-induced cardiac damage, increase the safety of doxorubicin chemotherapy, and provide a new option for the treatment of malignant tumors.

[0094] It should be understood that the above embodiments are merely preferred embodiments of the present invention and are not intended to limit the present invention in any form or substance. It should be noted that those skilled in the art can make several improvements and additions without departing from the present invention, and these improvements and additions should also be considered within the scope of protection of the present invention.

Claims

1. The use of GW3965 in the preparation of a drug for reducing doxorubicin-induced cardiac damage, wherein the doxorubicin-induced cardiac damage is a condition caused by doxorubicin such as: cardiac fibrosis, cardiac systolic and diastolic dysfunction, heart failure, or dilated cardiomyopathy.

2. The application as described in claim 1, characterized in that, The GW3965 is a hydrochloride salt, specifically 3-[3-[N-(2-chloro-3-trifluoromethylbenzyl)-(2,2-diphenylethyl)amino]propoxy]phenylacetic acid hydrochloride, CAS number: 405911-17-3, with the following chemical structural formula: 。 3. The application as described in claim 1, characterized in that, The drug is used to treat or prevent conditions caused by doxorubicin, including: cardiac systolic and diastolic dysfunction, heart failure, or dilated cardiomyopathy.

4. The application as described in claim 1, characterized in that, The drug is intended for cancer patients receiving doxorubicin chemotherapy.

5. The application as described in claim 1, characterized in that, The drug contains a therapeutically effective amount of GW3965 as its sole active ingredient.

6. The application as described in claim 1, characterized in that, The drug is a pharmaceutical composition that, in addition to containing a therapeutically effective amount of GW3965, also contains other pharmaceutical ingredients for the prevention and treatment of cardiac damage.

7. The application as described in claim 5 or 6, characterized in that, The drug is a pharmaceutical composition that, in addition to containing a therapeutically effective amount of GW3965, also contains a medically acceptable carrier.

8. The application as described in claim 1, characterized in that, The drug is available in oral or injectable form.

9. The application as described in claim 8, characterized in that, The oral preparations are selected from the group consisting of tablets, capsules, granules, powders, and syrups.

10. The application as described in claim 8, characterized in that, The injectable is suitable for intravenous injection.

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