Pharmaceutical composition, kit and use thereof

Abstract

Disclosed in the present invention are a pharmaceutical composition, a kit, and the use thereof. The pharmaceutical composition provided by the present invention comprises a first component and a second component, wherein the first component is a compound represented by formula I or a pharmaceutically acceptable salt thereof, and the second component is a CDK4 / 6 inhibitor. By means of combined administration of the first component and the second component, the present invention has significant inhibitory effect and synergistic effect on breast cancer cells, and in particular still has excellent inhibitory effect and strong synergistic effect and reduced drug resistance on mutant breast cancer cells.

Description

A pharmaceutical composition, a pillbox, and its application

[0001] This application claims priority to Chinese patent application 2024116224561, filed on November 14, 2024. The entire contents of the aforementioned Chinese patent application are incorporated herein by reference. Technical Field

[0002] This invention relates to a pharmaceutical composition, a medicine box, and its application. Background Technology

[0003] Breast cancer is the most common cancer among women, the second most common new cancer in women, and the fifth leading cause of cancer death in women. 70% of breast cancers are estrogen receptor-positive (ER+) / HER2-negative. The growth of these ER+ breast cancer cells is mediated by the interaction between estrogen and ERα, a ligand-activated transcription factor encoded by the ESR1 gene. Therefore, endocrine therapy (ET), which aims to block the estrogen-ER signaling pathway, has become the gold standard for the treatment of ER+ breast cancer. ET includes the following in sequence: (1) selective estrogen receptor modulators (SERMs), such as tamoxifen, which competitively bind to ERα, alter the conformation of ERα, prevent its dimerization, thereby inhibiting the signaling pathway and inhibiting the proliferation of breast cancer cells; (2) aromatase inhibitors (AIs), such as anastrozole, letrozole, and exemestane, which reduce the conversion of testosterone to estrogen in the human body, thereby inhibiting the ERα signaling pathway; (3) selective estrogen receptor degraders (SERDs), such as fulvestrant, which bind to ERα, thereby preventing the migration of ERα from the cytoplasm to the nucleus, so that the bound ERα cannot transmit signals, remains in the cytoplasm and is degraded by proteases in the cytoplasm, thereby blocking the ER signaling pathway and inhibiting the proliferation of cancer cells.

[0004] ET can be used independently as a monotherapy or in combination with other treatments or sequentially, depending on the characteristics of the breast cancer and the patient's overall health. Existing ET treatments exhibit primary resistance and can lead to numerous adverse reactions. For example, 30% of ER+ / HER2- breast cancers are unresponsive to tamoxifen (Droog M, et al. 2013). SERM has dual activity as an ER antagonist and agonist, potentially inducing endometrial cell growth and promoting endometrial cancer (Martinkovich S, et al. 2014). Fulvestrant is approved for the treatment of ET-resistant breast cancer. However, intramuscular administration is inconvenient for patients, leading to decreased treatment adherence. Furthermore, the clinical efficacy of fulvestrant is significantly limited due to insufficient receptor occupancy and poor pharmacokinetics (Chen YC, et al. 2022). More importantly, resistance often develops during ET treatment in most patients who respond to the therapy, leading to disease recurrence and progression. One important resistance mechanism is acquired ESR1 mutation. ESR1 mutation is one of the most well-known and widely studied molecular mechanisms of treatment resistance in breast cancer. ESR1 mutations can lead to persistent activation of ligand-independent estrogen pathways, promoting tumor growth and inducing resistance to estradiol (ET). The incidence of these mutations depends on the duration of ET, and is likely present in approximately 20%–40% of patients with metastatic breast cancer who have previously received anti-inflammatory drugs (AIs). ESR1 mutations not only affect the efficacy of subsequent ET, but their incidence is also higher when ET is combined with CDK4 / 6 inhibitors (Chaudhary N, et al. 2024; Jesselohn R, et al. 2014). Therefore, current research focuses on developing safer, more limited, and more precise breast cancer therapies, including strategies to address resistance mechanisms.

[0005] To address these challenges, a new generation of oral SERDs has emerged. In 2023, the FDA approved erastrant as the first oral SERD for the treatment of postmenopausal women or adult men with ESR1 mutations who have experienced disease progression after at least one round of ET therapy (Shah M, et al. 2024). Furthermore, the treatment landscape for ER+ / HER2- breast cancer has shifted from sequential monotherapy to combination therapy. ET combined with CDK4 / 6 inhibitors has significantly improved the prognosis of patients with metastatic HR+ breast cancer, significantly prolonging PFS. Currently marketed CDK inhibitors—palbociclib, abecilibi, ribociclib, and dalcilibi—have all shown highly consistent efficacy in prolonging median progression-free survival. Clinical studies at PALOMA, MONALEESA, MONARCH, and DAWNA have demonstrated that, compared to ET alone, combination therapy with ET results in a longer median progression-free survival. However, only the combination of fulvestrant with palbociclib or abecilibi, ribociclib, and nonsteroidal anti-inflammatory drugs (NSAIDs) with ribociclib prolonged overall survival; other combinations failed to achieve the expected benefit in overall survival.

[0006] Overall, developing drugs with key advantages, including greater safety, stronger efficacy, and long-term patient benefits, remains a formidable responsibility for the pharmaceutical industry. Exploring the synergistic effects of novel oral SERDs combined with CDK4 / 6 inhibitors, overcoming the limitations of some ET-CDK4 / 6 inhibitor combinations that have failed to significantly prolong overall survival, and ultimately providing long-term patient benefits remains a new direction for breast cancer treatment. Summary of the Invention

[0007] The technical problem this invention aims to solve is to overcome the limitations of existing breast cancer treatments due to primary or secondary drug resistance, as well as the resulting pain and intramuscular injection side effects, leading to poor patient compliance. This invention provides a pharmaceutical composition, a kit, and its application. The pharmaceutical composition of this invention, through the combined administration of the first and second components, exhibits significant inhibitory and synergistic effects on breast cancer cells, particularly demonstrating excellent inhibitory and strong synergistic effects against mutant breast cancer cells while reducing drug resistance.

[0008] This invention provides a pharmaceutical composition comprising a first component and a second component, wherein the first component is a compound of formula I or a pharmaceutically acceptable salt thereof, and the second component is a CDK4 / 6 inhibitor;

[0009] In one embodiment, the pharmaceutically acceptable salt of the compound represented by Formula I may be a conventionally pharmaceutically acceptable salt in the art, preferably a hydrochloride, hydrobromide, nitrate, carbonate, bicarbonate, phosphate, monohydrogen phosphate, dihydrogen phosphate, sulfate, hydrogen sulfate, hydroiodide, phosphite, acetate, propionate, isobutyrate, maleate, malonate, benzoate, succinate, octanoate, fumarate, lactate, mandelate, phthalate, benzenesulfonate, p-toluenesulfonate, citrate, tartrate, or methanesulfonate.

[0010] In one embodiment, the pharmaceutically acceptable salt of the compound represented by Formula I is a methanesulfonate.

[0011] In one embodiment, the compound represented by formula I is the compound represented by formula Ia:

[0012] In one embodiment, the pharmaceutically acceptable salt of the compound represented by Formula I is the compound represented by Formula Ib:

[0013] In one embodiment, the CDK4 / 6 inhibitor may be one or more of the conventional CDK4 / 6 inhibitors in the art: for example, palbociclib, abemaciclib, ), Ribociclib Dalpicilib ), Kumoshili One or more of the following: Culmerciclib, trilaciclib, lerociclib, bireociclib, FCN-437c, and tebecilib (BPI-16350).

[0014] In one embodiment, the CDK4 / 6 inhibitor is palbociclib, the structure of which is shown in Formula II.

[0015] In one embodiment, the pharmaceutical composition comprises a first component being a compound of formula Ia; and a second component being selected from one or more of palbociclib, abecilib, ribociclib, dalcilib, cumocilib, trilasiclib, lerocilib, piroxicillin, FCN-437c, and tebecilib, preferably the first component being a compound of formula Ia and the second component being palbociclib.

[0016] In one embodiment, the pharmaceutical composition comprises a first component being a compound of formula Ib; and a second component being selected from one or more of palbociclib, abecilibiclib, ribociclib, dalcilibiclib, cumocilibiclib, trilasiclib, lerocilibiclib, piroxicillin, FCN-437c, and tebecilibiclib, preferably the first component being a compound of formula Ib and the second component being palbociclib.

[0017] In one embodiment, the first component and the second component are present in the form of active ingredients in the pharmaceutical composition.

[0018] The present invention also provides the use of the pharmaceutical composition described herein in the preparation of a medicament for the prevention and / or treatment of breast cancer.

[0019] In one embodiment, the breast cancer is preferably positive (ER+) / HER2 negative breast cancer, preferably wild-type breast cancer or mutant breast cancer, such as breast cancer with ESR1 D538G mutation.

[0020] The present invention also provides the use of the first component in the preparation of a medicament for the prevention and / or treatment of breast cancer in combination with the second component.

[0021] The present invention also provides the use of the second component in the preparation of a medicament for the prevention and / or treatment of breast cancer in combination with the first component.

[0022] The present invention also provides a combination medicine box, comprising:

[0023] A first container containing the first component; and,

[0024] A second container contains the second component.

[0025] In the uses and combination kits described in this invention, the breast cancer is as described above.

[0026] In the pharmaceutical composition, uses, and combination kits of the present invention, the first component and the second component have a synergistic effect.

[0027] In the pharmaceutical composition, use and combination kit of the present invention, the mass ratio of the first component and the second component can be selected according to conventional art, preferably 1:(5-15), for example 1:8 or 1:10.

[0028] In the pharmaceutical compositions, uses, and combination kits of the present invention, the first component and the second component can be used independently in conventional dosage forms in the art, preferably independently in gastrointestinal or non-gastrointestinal dosage forms, and more preferably independently in gastrointestinal dosage forms. The gastrointestinal dosage forms include, for example, tablets, capsules, granules, powders, or oral liquids.

[0029] In the pharmaceutical composition, uses, and combination kits of the present invention, the first component and the second component are administered independently by oral or injection (e.g., subcutaneous, intradermal, intravenous, intraperitoneal, or intramuscular administration), preferably by oral administration.

[0030] In the pharmaceutical composition, uses, and combination kits described in this invention, the dosage of the first component and the second component is not specifically limited and can be selected according to conventional practices in the art.

[0031] Preferably, the dosage of the first component is 1-1000 mg, preferably 30-400 mg, 60-350 mg, 100-300 mg, or 150-250 mg; for example, 5 mg, 10 mg, 15 mg, 20 mg, 25 mg, 30 mg, 35 mg, 40 mg, 45 mg, 50 mg, 55 mg, 60 mg, 65 mg, 70 mg, 75 mg, 80 mg, 85 mg, 90 mg, 100 mg, 125 mg, 150 mg, 175 mg, 200 mg, 240 mg, 250 mg, 280 mg, 300 mg, 320 mg, 350 mg, 400 mg, 450 mg, 500 mg, 600 mg, 650 mg, 700 mg, 750 mg, 800 mg, 825 mg, 850 mg, 900 mg, 950 mg, or 1000 mg.

[0032] Preferably, the dosage of the second component is 1-1000 mg, more preferably 150-1000 mg, 200-950 mg, 300-900 mg, 400-850 mg, 500-800 mg, or 600-750 mg; for example, 5 mg, 10 mg, 15 mg, 20 mg, 25 mg, 30 mg, 35 mg, 40 mg, 45 mg, 50 mg, 55 mg, 60 mg, 65 mg, 7 mg, etc. 0mg, 75mg, 80mg, 85mg, 90mg, 100mg, 125mg, 150mg, 175mg, 200mg, 240mg, 250mg, 280mg, 300mg, 320mg, 350mg, 400mg, 450mg, 500mg, 550mg, 600mg, 650mg, 700mg, 750mg, 800mg, 900mg, 1000mg.

[0033] In the pharmaceutical composition, uses, and combination kits of the present invention, the frequency of administration of the first component and the second component can be adjusted according to clinical needs, and is preferably once a day, twice a day, three times a day, or once every two days.

[0034] The present invention also provides a method for treating tumors, comprising administering a therapeutically effective amount of the pharmaceutical composition as described herein to a subject (e.g., a human or a mouse) in need of such treatment.

[0035] Among the above applications and treatment methods:

[0036] In one embodiment, the pharmaceutical composition exhibits a long-term tumor inhibition rate; the tumor inhibition rate can be 80-150%; preferably 110.5% or 122.83%.

[0037] In one embodiment, the pharmaceutical composition achieves an objective response rate to tumor greater than 80%, for example, 87.5% or 100%.

[0038] In one embodiment, the pharmaceutical composition can increase the tumor growth inhibition rate by, for example, 5% to 35%, such as 9.6% or 23.97%, compared to using the first component alone.

[0039] In one embodiment, the pharmaceutical composition can increase the tumor growth inhibition rate by, for example, 40-100%, such as 51.92% or 67.7%, compared to using the second component alone.

[0040] As used in this article, “treatment” means therapeutic therapy. When a specific condition is involved, treatment means: (1) alleviating one or more biological manifestations of the disease or condition; (2) interfering with (a) one or more points in a biological cascade that causes or precipitates the condition or (b) one or more biological manifestations of the condition; (3) improving one or more symptoms, effects or side effects associated with the condition, or one or more symptoms, effects or side effects associated with the condition or its treatment; or (4) slowing the development of the condition or one or more biological manifestations of the condition.

[0041] As used herein, the term "therapeutic effective amount" refers to an amount of compound sufficient to effectively treat the disease or condition described herein when administered to a subject. The amount of compound constituting a "therapeutic effective amount" will vary depending on the compound, the condition and its severity, and the age of the subject to be treated, but may be adjusted as needed by those skilled in the art.

[0042] As used in this article, the term "container" refers to any container and cap suitable for storing, transporting, dispensing and / or handling pharmaceuticals.

[0043] As used in this article, "pharmaceutical composition" refers to a composition containing a specified active ingredient that can be prepared into the same dosage form.

[0044] As used herein, the term "subject" refers to any animal, preferably a mammal, that is about to be or has already been administered the compound or composition according to embodiments of the invention. As used herein, the term "mammal" includes any mammal. Examples of mammals include, but are not limited to, cattle, horses, sheep, pigs, cats, dogs, mice, rats, rabbits, guinea pigs, monkeys, and humans, with humans being the most preferred.

[0045] The pharmaceutical compositions of this invention may further comprise pharmaceutically acceptable carriers / excipients, such as those widely used in the pharmaceutical manufacturing industry. Excipients primarily serve to provide a safe, stable, and functional pharmaceutical composition, and may also provide methods for dissolving the active ingredient at a desired rate after administration to a subject, or for promoting effective absorption of the active ingredient after administration to a subject. Excipients may be inert fillers or provide a function, such as stabilizing the overall pH of the composition or preventing degradation of the active ingredient. Pharmaceutically acceptable excipients may include one or more of the following: binders, suspending agents, emulsifiers, diluents, fillers, granulators, adhesives, disintegrants, lubricants, anti-adhesion agents, flow aids, wetting agents, gelling agents, absorption delay agents, dissolution inhibitors, enhancers, adsorbents, buffers, chelating agents, preservatives, colorants, flavoring agents, and sweeteners. Pharmaceutically acceptable carriers can take many forms depending on the desired formulation for administration. As an illustrative example, suitable carriers and additives for solid oral dosage forms include starches, sugars, diluents, granulating agents, lubricants, binders, disintegrants, etc. Pharmaceutically acceptable carriers or excipients should generally be non-toxic. Pharmaceutical compositions according to the invention may contain one or more suitable carriers / excipients. The amount and type of carrier / excipient will vary as needed. Those skilled in the art will be able to readily determine appropriate carriers / excipients to be added to the pharmaceutical compositions of the invention based on the current disclosure.

[0046] In this invention, the term "active ingredient" refers to the active ingredient in the pharmaceutical composition of this invention, namely the compound represented by Formula I or a pharmaceutically acceptable salt thereof, and CDK4 / 6 inhibitors.

[0047] When the pharmaceutical compositions of the present invention are administered to a subject for the treatment or prevention of a disease, condition, or illness, the active ingredients in the pharmaceutical compositions may be administered via the same route or via different routes. Administration may be made via any route described herein, including but not limited to oral or injectable methods. The specific route of administration will depend on the therapeutic agent itself and the formulation, as well as the disease, condition, or illness to be prevented or treated. Based on this disclosure, the skill level of those skilled in the art is sufficient to determine the route of administration for other therapeutic agents. The active ingredients in the pharmaceutical compositions of the present invention may be administered to the subject for a period of time (drug cycle), followed by a period during which no compound is administered (non-drug cycle). The required number of dosing cycles and non-drug cycles may be repeated. The required length and number of dosing cycles or non-drug cycles will depend on the type and / or severity of the disease, condition, or illness being treated or prevented, as well as the subject's sex, age, weight, and other parameters (e.g., the subject's biological, physical, and physiological condition). The active ingredients in the pharmaceutical compositions of the present invention may be administered to the subject simultaneously for a period of time or sequentially for a period of time. Based on the disclosure in this document, the skill level of a person of ordinary skill in the art will be sufficient to determine the appropriate length and number of dosing and / or non-dosing cycles, for example, if weight loss exceeds 15%, discontinue the medication until weight recovers to within 10% before resuming dosing.

[0048] Those skilled in the art will recognize that the therapeutic or preventative effective amount of the pharmaceutical composition of the present invention can vary depending on factors such as age, diet, health, the severity and type of symptoms or diseases, conditions or illnesses, complications and formulations sought for treatment or prevention, etc. Based on the disclosure of the present invention, those skilled in the art will be able to readily determine the therapeutic or preventative effective amount to be administered to a subject in order to elicit the desired biological or medical response in the subject.

[0049] The combined use of the active ingredients in the pharmaceutical composition according to the present invention can have a synergistic effect in the treatment or prevention of breast cancer-related diseases, symptoms or conditions.

[0050] Unless otherwise stated, the word “or” or “and” in this article refers to “and / or”.

[0051] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention pertains. Otherwise, certain terms used herein have the meanings set forth in this specification.

[0052] Based on common knowledge in the field, the above-mentioned preferred conditions can be combined arbitrarily to obtain various preferred embodiments of the present invention.

[0053] BALB / c nude mice: Supplier: Beijing Vital River Laboratory Animal Technology Co., Ltd. (Zhejiang Branch); Animal Qualification Certificate No.: 20210628Abzz0619000201.

[0054] Palbociclib: Purchased from MCE.

[0055] Unless otherwise specified, all reagents and raw materials used in this invention are commercially available.

[0056] The positive and progressive effects of this invention are as follows:

[0057] The pharmaceutical composition of the present invention can be used to treat breast cancer, and has excellent inhibitory effects on both mutated MCF7 breast cancer cells and MCF7 breast cancer cells.

[0058] 1. For mutated MCF7 breast cancer cells: (1) the tumor inhibition rate is greater than 80%, for example, reaching 122.83%, and the objective response rate (ORR) is greater than 80%, for example, reaching 100%;

[0059] (2) Compared with the first component alone (e.g., the compound shown in Formula Ib), the tumor inhibition rate was increased by 5-35%, for example by 23.97%, and the objective response rate (ORR) was increased by at least 80%, for example by 100%; compared with the second component alone (commercially available pebocillin), the tumor inhibition rate was increased by 40-100%, for example by 51.92%, and the objective response rate (ORR) was increased by at least 85%, for example by 100%, with significant inhibitory effect and good tolerability;

[0060] (3) According to calculations, in terms of the effect of inhibiting mutated MCF7 breast cancer cells, the synergistic factor of the first component and the second component combined is greater than 2, and the better synergistic factor is 2.21, which shows excellent strong synergistic effect.

[0061] 2. For MCF7 breast cancer cells: (1) The tumor inhibition rate is greater than 80%, for example, reaching 110.5%, and the objective response rate (ORR) is greater than 80%, for example, reaching 87.5%;

[0062] (2) Compared with the first component alone (e.g., the compound shown in Formula Ib), the tumor inhibition rate was increased by 5-35%, for example by 9.6%, and the objective response rate (ORR) was increased by at least 80%, for example by 87.5%. Compared with the second component alone (commercially available pebocillin), the tumor inhibition rate was increased by 40-100%, for example by 67.7%, and the objective response rate (ORR) was increased by at least 80%, for example by 87.5%, demonstrating significant inhibitory effects.

[0063] (3) According to calculations, in terms of the effect of inhibiting MCF7 breast cancer cells, the synergistic factor of the first component and the second component in combination is greater than 1.1, for example, 1.44, which shows excellent synergistic effect.

[0064] 3. The pharmaceutical composition of the present invention has broad clinical application prospects. Attached Figure Description

[0065] Figure 1 shows the tumor growth curves of MCF7 cell subcutaneous xenograft tumor model mice carrying the ESR1 D538G mutation after treatment with the compound.

[0066] Figure 2 shows the tumor growth curves of a subcutaneous xenograft model of human breast cancer MCF7 cells in tumor-bearing mice after administration of the test compound. Detailed Implementation

[0067] The present invention is further illustrated below by way of embodiments, but the invention is not limited to the scope of the embodiments described herein. Experimental methods in the following embodiments that do not specify specific conditions were performed according to conventional methods and conditions, or as selected according to the product instructions.

[0068] Formulation A is a formulation containing the compound shown in Formula I. Its preparation method and composition are described in Example 5 of Chinese Patent CN118045052.

[0069] Example 1

[0070] In vivo pharmacodynamics of formulation A in a mouse model of subcutaneous xenografted human breast cancer MCF7 cells carrying the ESR1 D538G mutation.

[0071] 1.1 Cell Information

[0072] Cells: MCF7 ESR1 D538G cells

[0073] Culture conditions: EMEM medium with 10% fetal bovine serum, 1% antibiotic-antifungal agent, 2mM L-glutamine, 1% NEAA, and cultured in a 37℃ 5% CO2 incubator.

[0074] 1.2 Cell Culture Preparation

[0075] Human breast cancer MCF7 cells carrying the ESR1 D538G mutation were cultured in vitro using standard monolayer conditions: EMEM medium supplemented with 10% fetal bovine serum, 1% antibiotic-antifungal agent, 2 mM L-glutamine, and 1% NEAA, incubated at 37°C in a 5% CO2 incubator. Cells were passaged twice weekly using trypsin-EDTA digestion. When cell saturation reached 80%-90% and the desired cell count was achieved, cells were collected, counted, and seeded.

[0076] 1.3 Tumor cell inoculation and grouping

[0077] Cell seeding: 0.2 mL (10 × 10⁻⁶) 6 MCF7 ESR1 D538G cells (PBS + matrix gel, volume ratio 1:1) were subcutaneously inoculated into the right posterior dorsal region of each BALB / c nude mouse, resulting in an average tumor volume of 147 mm². 3 Dosing will begin at the designated time. Experimental groups and dosing regimens are shown in Table 1 below. The first day of dosing is designated as day 0.

[0078] Table 1. Grouping and Dosing Regimens of Experimental Animals

[0079] 1.4 Experimental Results

[0080] This experiment evaluated the efficacy of formulation A in a human breast cancer MCF7 cell xenograft model carrying the ESR1 D538G mutation, with the Vehicle group (10% HP-β-CD in water) as the control. Tumor volumes and statistical analyses at different time points for each group are shown in Figure 1 and Table 2.

[0081] Table 2. Antitumor effects of the test substances on a subcutaneous xenograft model of human breast cancer MCF7 ESR1 D538G cells. Note: a. Mean ± SEM. b. Calculated using relative tumor volume (RTV). c. Calculated using tumor volume (TV). d. p-value obtained by one-way ANOVA analysis of relative tumor volume compared to the solvent therapy group. e. d. Tumor response was assessed using mRECIST. PD, progressive disease; SD, stable disease; PR, partial response; CR, complete response. Objective response rate (ORR) is defined as the proportion of animals with CR or PR.

[0082] All tumor-bearing mice in the treatment groups showed good tolerance to the test compounds, and their body weight did not decrease significantly; on the contrary, the weight gain rate of mice in each treatment group showed a certain degree of increase.

[0083] Palbociclib had a TGI of 70.91% (4SD, 4PD) at a dose of 50 mg / kg, while formulation A had a TGI of 98.86% (4SD, 4PD) at a dose of 5 mg / kg. The TGI of the combination therapy of formulation A (5 mg / kg) and Palbociclib (50 mg / kg) was 122.83% (1CR, 7PR). In the combination therapy group, the objective response rate (ORR) reached 100%. Compared with the 0% ORR in the Palbociclib and formulation A monotherapy groups, the combination therapy clearly had a more significant tumor-suppressive effect than monotherapy.

[0084] MCF7 ESR1 D538G cells are mutant human breast cancer cells that develop drug resistance during treatment. Based on a synergistic factor of 2.21 (synergistic factor = ((formulation A / Vehicle) × (Palbociclib / Vehicle)) / (combination group / Vehicle)), it indicates that combination therapy still has significant synergistic activity in a mouse model of drug-resistant ESR1 D538G mutant human breast cancer MCF7 cell xenograft tumors.

[0085] Example 2

[0086] In vivo pharmacodynamic study of formulation A on BALB / c nude mouse model of subcutaneous xenografted human breast cancer MCF-7 cells

[0087] 2.1 Cell Information

[0088] Human breast cancer MCF-7 cells (ATCC, catalog number: ECACC-86012803) were cultured in the same manner as in Example 1.

[0089] 2.2 Tumor cell inoculation and grouping

[0090] Cell inoculation: Estrogen tablets (0.36 mg / tablet; IRA) were subcutaneously injected into the left hindquarters of each BALB / c nude mouse. Three days later, 0.2 ml (1 × 10⁻⁶) of the estrogen solution was injected into the cell line. 7 MCF-7 cells (with matrix gel; corning, volume ratio 1:1) were subcutaneously inoculated into the right posterior dorsal region of each mouse, resulting in an average tumor volume of 188 mmHg. 3 Dosing should begin in groups according to the dosing regimen in Table 3.

[0091] Table 3. Grouping and Dosing Regimens of Experimental Animals

[0092] 2.3 Experimental Results

[0093] This experiment evaluated the efficacy of test formulation A in a human breast cancer xenograft model, with the solvent control group (Vehicle group) as a reference. The tumor volume of each group at different time points and the corresponding statistical analysis results are shown in Table 4 and Figure 2.

[0094] Table 4. Antitumor effects of the tested compounds on a subcutaneous xenograft model of breast cancer MCF7 cells. Note: a. Mean ± SEM. b. Calculated using relative tumor volume (RTV). c: Calculated using tumor volume (TV). d: Tumor response was assessed using mRECIST. PD, progressive disease; SD, stable disease; PR, partial response; CR, complete response. Objective response rate (ORR) is defined as the proportion of animals with CR or PR.

[0095] At a dose of 3 mg / kg, formulation A had a TGI of 100.9% and a disease control rate (DCR) of 100%. The TGI of the Palbociclib (25 mg / kg) monotherapy group was 42.8%, with a P value of 0.219 compared to the control group; therefore, Palbociclib monotherapy did not show antitumor activity. However, when formulation A (3 mg / kg) was used in combination with Palbociclib (25 mg / kg), the TGI was 110.5%; the P values ​​compared with the control group, Palbociclib group, and formulation A (3 mg / kg) group were 0.001, 0.002, and 0.012, respectively; the objective response rate (ORR) reached 87.5% (7 PR, 1 SD); the synergistic factor of the combination therapy was 1.44 (synergistic factor = ((formulation A / Vehicle) × (Palbociclib / Vehicle)) / (combination group / Vehicle)). Therefore, the combination therapy showed a stronger antitumor effect than formulation A alone, which is a synergistic effect.

[0096] While specific embodiments of the present invention have been described above, those skilled in the art should understand that these are merely illustrative examples, and various changes or modifications can be made to these embodiments without departing from the principles and essence of the present invention. Therefore, the scope of protection of the present invention is defined by the appended claims.

Claims

1. A pharmaceutical composition, characterized in that, It comprises a first component and a second component, wherein the first component is a compound of Formula I or a pharmaceutically acceptable salt thereof, and the second component is a CDK4 / 6 inhibitor; 2. The pharmaceutical composition according to claim 1, characterized in that, Pharmaceutically acceptable salts of the compounds represented by Formula I are hydrochloride, hydrobromide, nitrate, carbonate, bicarbonate, phosphate, monohydrogen phosphate, dihydrogen phosphate, sulfate, hydrogen sulfate, hydroiodide, phosphite, acetate, propionate, isobutyrate, maleate, malonate, benzoate, succinate, octanoate, fumarate, lactate, mandelate, phthalate, benzenesulfonate, p-toluenesulfonate, citrate, tartrate, or methanesulfonate, such as methanesulfonate.

3. The pharmaceutical composition according to claim 1, characterized in that, It satisfies one or two of the following conditions: (1) The compound represented by formula I is the compound represented by formula Ia. Preferably, it is a compound represented by the following formula Ib. (2) The CDK4 / 6 inhibitor is selected from one or more of palbociclib, abeciclib, ribociclib, dalcilib, cumociclib, trilasiclib, lerocilib, pyrosinib, FCN-437c and tebeciclib, such as palbociclib.

4. The pharmaceutical composition according to claim 1, characterized in that, It satisfies one or more of the following conditions: (1) The first component is a compound represented by formula Ia; the second component is selected from one or more of palbociclib, abecilib, ribociclib, dalcilib, cumocilib, trilasiclib, lerocilib, piroxicillin, FCN-437c and tebecilib, preferably the first component is a compound represented by formula Ia and the second component is palbociclib; (2) The first component is a compound represented by formula Ib; The second component is selected from one or more of palbociclib, abecilibi, ribociclib, dalcilibi, cumocilibi, trilasiclib, lerocilibi, piroxicillin, FCN-437c and tebecilibi, preferably the first component is a compound represented by formula Ib and the second component is palbociclib; (3) The first component and the second component exist in the form of active ingredients; (4) The first component and the second component have a synergistic effect; (5) The mass ratio of the first component to the second component is 1:(5-15), for example 1:8 or 1:10; (6) The first component and the second component are used independently in a gastrointestinal or non-gastrointestinal dosage form; wherein the gastrointestinal dosage form is a tablet, capsule, granule, powder or oral liquid. (7) The administration methods of the first component and the second component are independently oral or injectable, preferably oral; (8) The dosage of the first component is 1-1000 mg, preferably 30-400 mg, 60-350 mg, 100-300 mg, or 150-250 mg; (9) The dosage of the second component is 1-1000 mg, preferably 150-1000 mg, 200-950 mg, 300-900 mg, 400-850 mg, 500-800 mg or 600-750 mg; (10) The application frequency of the first component and the second component is independently once a day, twice a day, three times a day or once every two days.

5. Use of a pharmaceutical composition according to any one of claims 1-4 in the preparation of a medicament for the prevention and / or treatment of breast cancer.

6. The use as described in claim 5, characterized in that, The breast cancer is positive (ER+) / HER2 negative breast cancer, preferably wild-type breast cancer or mutant breast cancer, such as breast cancer with ESR1 D538G mutation.

7. Use of the first component as claimed in any one of claims 1-4 in the preparation of a medicament for the prevention and / or treatment of breast cancer in combination with the second component as claimed in any one of claims 1-4, wherein the use is as described in claim 5 or 6.

8. Use of the second component as claimed in any one of claims 1-4 in the preparation of a medicament for the prevention and / or treatment of breast cancer in combination with the first component as claimed in any one of claims 1-4, wherein the use is as described in claim 5 or 6.

9. A combination pillbox, comprising: A first container comprising the first component as claimed in any one of claims 1-4; and, The second container contains the second component as claimed in any one of claims 1-4.

10. The combination medicine box as described in claim 9, characterized in that, The mass ratio of the first component to the second component, the dosage form, the administration method, the dosage and the frequency of administration are as described in claim 4.

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