CN118059108A - Pharmaceutical composition and application thereof in preparation of antitumor drugs - Google Patents

Abstract

The present invention discloses a pharmaceutical composition and its application in the preparation of an anti-tumor drug, which is composed of olanzapine and osimertinib, and can significantly improve the therapeutic and/or preventive effects of tumors and reverse the generation of drug resistance. The present invention forms a new pharmaceutical composition with olanzapine and osimertinib, and the two act synergistically, thereby significantly improving the inhibitory effect on lung cancer cell proliferation and lung cancer tumor and organoid growth.

Description

A pharmaceutical composition and its application in preparing anti-tumor drugs

Technical Field

The present invention relates to the technical field of medicine, and in particular to a pharmaceutical composition and application thereof in the preparation of anti-tumor drugs.

Background technique

Lung cancer is one of the most common malignant tumors causing cancer-related deaths worldwide, and its incidence rate ranks second. In recent years, the incidence rate has been increasing year by year, posing a serious threat to human health. Lung cancer is highly heterogeneous and can be divided into multiple subtypes based on pathological and clinical characteristics, of which about 80%-85% are non-small cell lung cancer. In recent years, the rapid development of molecular targeted therapy and immunotherapy has benefited some patients in the advanced stage. However, due to the emergence of recurrence, metastasis and drug resistance, only a small number of lung cancer patients have achieved long-term clinical efficacy. Therefore, developing new drugs and formulating new anti-tumor treatment strategies are the key to controlling the malignant progression of lung cancer.

Olanzapine is a clinically used atypical antipsychotic drug with a molecular formula of C ₁₇ H ₂₀ N ₄ S. It is mainly used to treat schizophrenia and bipolar disorder with few adverse reactions. Olanzapine can inhibit dopamine receptors and bind to 5-hydroxytryptamine receptors. It is currently reported that olanzapine has a certain anti-cancer effect, but its specific anti-tumor mechanism and target are still unclear.

The third-generation epidermal growth factor receptor tyrosine kinase inhibitor (EGFR-TKI) osimertinib is an oral irreversible pyrimidine-based epidermal growth factor receptor tyrosine kinase inhibitor that forms an irreversible covalent bond with EGFR T790M or EGFR mutations through the cysteine-797 residue, and has a selective inhibitory effect on EGFR sensitive activating mutations and T790M resistance mutations. However, similar to patients treated with first- or second-generation EGFR-TKIs, patients treated with osimertinib will eventually develop resistance. Therefore, it is urgent to develop a new generation of treatments and rational combination drug strategies to overcome EGFR-TKI resistance.

Summary of the invention

In order to overcome the defects of the prior art, the technical problem to be solved by the present invention is to provide a pharmaceutical composition and its use in the preparation of anti-tumor drugs. Compared with the individual components, the pharmaceutical composition has higher anti-cancer activity, significantly improves the inhibitory effect on lung cancer cells and lung cancer tumors, significantly inhibits the growth of organoids derived from lung cancer patients, and avoids the drug resistance of the original anti-lung cancer drugs.

The technical solution of the present invention is: a pharmaceutical composition, which comprises: olanzapine and osimertinib.

Preferably, the molar ratio of olanzapine to osimertinib is: 1-400:0.039-5.

Preferably, the molar ratio of olanzapine to osimertinib is: 1.00-400:0.039-5.

Preferably, the molar ratio of olanzapine to osimertinib is: 400:0.039-5.

Furthermore, the present invention provides a pharmaceutical preparation comprising the above-mentioned pharmaceutical active ingredient and a pharmaceutically acceptable carrier.

Preferably, the pharmaceutically acceptable carrier is a filler, a wetting agent, a binder, a disintegrant or a lubricant.

Preferably, the drug is in the form of an oral preparation, and the specific preparation form may be but is not limited to oral tablets, granules, injections, and capsules.

The present invention also provides an application of the pharmaceutical composition in the preparation of an anti-tumor drug, wherein the anti-tumor drug is used for treating lung cancer.

Preferably, the tumor comprises one or more of small cell lung cancer, non-small cell lung squamous cell carcinoma, non-small cell lung adenocarcinoma, non-small cell adenosquamous carcinoma and large cell lung cancer.

Preferably, the anti-tumor drug is used to inhibit the proliferation of tumor cells, and the tumor cells are PC-9 cells, H1975 cells or PC-9 ^{del19-T790M-C797S} cells.

Preferably, the anti-tumor drug is used to inhibit tumor growth, and the tumor is a PC-9 subcutaneous transplant tumor, a H1975 subcutaneous transplant tumor, or a PC-9 ^{del19-T790M-C797S} subcutaneous transplant tumor.

Preferably, the anti-tumor drug is used to inhibit the growth of organoids derived from tumor patients.

Compared with the prior art, the beneficial effects of the present invention are as follows:

1. The present invention combines olanzapine and osimertinib into a new pharmaceutical composition, which significantly improves the inhibitory effect on lung cancer cells and lung cancer tumors through the synergistic effect of the two, significantly inhibits the growth of organoids derived from lung cancer patients, and reverses osimertinib resistance.

2. The present invention combines olanzapine and osimertinib into a new pharmaceutical composition, and the effective dose of the combined use of the two is significantly lower than the clinical dose of each, which meets the requirements of clinical drug safety and efficacy.

3. The present invention combines olanzapine and osimertinib into a compound preparation, and both drugs are clinically administered orally. The compound preparation can improve the patient's drug compliance.

BRIEF DESCRIPTION OF THE DRAWINGS

Figure 1 shows the effect of the drug combination on the proliferation of different lung adenocarcinoma cells. (A) Effect of olanzapine alone on the proliferation of PC-9 cells; (B) Effect of olanzapine combined with osimertinib on the proliferation of PC-9 cells; (C) Effect of olanzapine combined with osimertinib on the proliferation of NCI-H1975 cells.

FIG2 shows the effect of the drug composition on the proliferation of osimertinib-resistant PC-9 ^{del19-T790M-C797S} cells.

Figure 3 shows the effect of the drug combination on the growth of organoids from different lung adenocarcinoma patients. (A) The effect of olanzapine monotherapy at different concentration gradients on the growth of organoids from the first lung adenocarcinoma patient; (B) The effect of osimertinib monotherapy at different concentration gradients and the combination of osimertinib and osimertinib on the growth of organoids from the first lung adenocarcinoma patient; (C) The effect of osimertinib monotherapy at different concentration gradients on the growth of organoids from the second lung adenocarcinoma patient; (D) The effect of osimertinib monotherapy at different concentration gradients and the combination of osimertinib and osimertinib on the growth of organoids from the second lung adenocarcinoma patient.

Detailed ways

Various exemplary embodiments of the present invention will now be described in detail. This detailed description should not be considered as limiting the present invention, but should be understood as a more detailed description of certain aspects, features, and embodiments of the present invention.

It should be understood that the terms described in the present invention are only for describing special embodiments and are not intended to limit the present invention. In addition, for the numerical range in the present invention, it should be understood that each intermediate value between the upper and lower limits of the scope is also specifically disclosed. The intermediate value in any stated value or stated range, and each smaller range between any other stated value or intermediate value in the described range is also included in the present invention. The upper and lower limits of these smaller ranges can be independently included or excluded in the scope.

Unless otherwise indicated, all technical and scientific terms used herein have the same meanings as those generally understood by those skilled in the art. Although the present invention describes only preferred methods and materials, any methods and materials similar or equivalent to those described herein may also be used in the implementation or testing of the present invention. All documents mentioned in this specification are incorporated by reference to disclose and describe the methods and/or materials related to the documents. In the event of conflict with any incorporated document, the contents of this specification shall prevail. The experimental methods for which specific conditions are not indicated in the examples are selected according to conventional methods and conditions, or according to the product specifications.

It will be apparent to those skilled in the art that various modifications and variations may be made to the specific embodiments of the present invention description without departing from the scope or spirit of the present invention. Other embodiments derived from the present invention description will be apparent to those skilled in the art. The present invention description and examples are exemplary only.

The words “include,” “including,” “have,” “contain,” etc. used in this document are open-ended terms, meaning including but not limited to.

The room temperature described in the embodiments is the conventional room temperature in the art, preferably 15-30°C.

The experimental results are expressed as mean ± standard error. After comparison with parametric or non-parametric variance test, p < 0.05 is considered to be significantly different, and p < 0.01 is considered to be extremely significantly different.

Example 1

The CCK8 assay was used to conduct a tumor cell proliferation experiment to compare the effects of olanzapine and osimertinib alone and in combination on lung cancer cell proliferation. The experimental steps are as follows:

Cells were plated at 8,000 cells/well in a 96-well plate. After the cells adhered, different concentrations of olanzapine or osimertinib alone or in combination (both solvents were DMSO) were added and incubated at 37°C, 5% carbon dioxide incubator for 24 hours. After incubation, 10 μL of CCK-8 solution was added to each well, and the culture plate was placed in the incubator for incubation for 1-4 hours. The absorbance at 450 nm was measured with a microplate reader, and the IC50 value of each group was calculated.

PC-9 and NCI-H1975 cells were purchased from Wuhan Pronocell Life Science Co., Ltd.; olanzapine was purchased from MedChemExpress (MCE); and osimertinib was purchased from MCE.

The results showed that when olanzapine and osimertinib were used in combination on PC-9 or NCI-H1975 cells, the inhibitory effect on cell proliferation was significantly stronger than that of osimertinib alone. When olanzapine and osimertinib were combined at different concentrations, different combined effects were shown. As shown in Figure 1, the specific results are as follows:

For PC-9 cells, the _IC50 of olanzapine after 48 hours of administration was 483.1 μM, and the _IC50 of olanzapine after 72 hours of administration was 175.7 μM; the _IC50 of osimertinib alone for 24 hours was 0.871 μM, the _IC50 of osimertinib combined with 400 μM olanzapine was 0.016 μM, the _IC50 of osimertinib combined with 200 μM olanzapine was 0.119 μM, and the _IC50 of osimertinib combined with 100 μM olanzapine was 0.286 μM.

For NCI-H1975 cells, the IC ₅₀ of osimertinib alone for 24 hours was 4.122 μM, the IC ₅₀ of osimertinib combined with 400 μM olanzapine was 0.213 μM, the IC ₅₀ of osimertinib combined with 200 μM olanzapine was 3.189 μM, and the IC ₅₀ of osimertinib combined with 100 μM olanzapine was 3.139 μM.

Example 2

The tumor cell proliferation experiment was performed by CCK8 experiment to compare the effects of fixed concentration of olanzapine combined with different concentrations of osimertinib on the proliferation of osimertinib-resistant PC-9 ^{del19-T790M-C797S} lung cancer cells. The experimental method was the same as that in Example 1.

PC-9 ^{del19-T790M-C797S} cells were purchased from Nanjing Kebai Biotechnology Co., Ltd.

Inhibition rate = 1-(OD value of drug-treated group-OD value of blank group)/(OD value of control group-OD value of blank group) × 100%.

The results are shown in Figure 2 and Table 1. The inhibition rate of osimertinib combined with olanzapine on osimertinib-resistant cells was significantly higher than that of the single drug group. The IC ₅₀ of osimertinib alone for 24 hours was 4.882 μM, and the IC ₅₀ of osimertinib combined with olanzapine was 1.708 μM, that is, osimertinib combined with olanzapine had a synergistic inhibitory effect on PC-9 ^{del19-T790M-C797S} cells.

Table 1. The proliferation inhibition rate of PC-9 ^{del19-T790M-C797S} cells treated with fixed concentration of olanzapine combined with different concentrations of osimertinib

The present invention found that the inhibition rate of osimertinib combined with olanzapine on osimertinib-resistant cells was significantly higher than that of the single drug group, and had significant anti-tumor efficacy. The combination regimen has the potential to reduce the effective drug concentration and reduce the side effects caused by high-dose targeted drugs.

Example 3

The effect of the drug composition on the growth of lung cancer tumor cells was investigated by in vivo experiments. Lung cancer cells (1*10 ⁶ /0.1 mL per mouse) were subcutaneously inoculated into the right side of the mouse. Seven days after inoculation, the tumor grew to a volume of 80-100 mm ³ . The mice were randomly divided into 4 groups, with 3 mice in each group: control solvent treatment group (normal saline), olanzapine alone treatment group (0.75 mg/kg, 1.5 mg/kg, 3 mg/kg, 6 mg/kg), osimertinib alone treatment group (1.25 mg/kg, 2.5 mg/kg, 5 mg/kg, 10 mg/kg), and drug combination treatment group (olanzapine (mg/kg): osimertinib (mg/kg): 0.75:1.25, 0.75:2.5, 0.75:5, 0.75:10, 1.5:1.25, 1.5:2.5, 1.5:5, 1.5:10, 3:1.25, 3:2.5, 3:5, 3:10, 6:1.25, 6:2.5, 6:5, 6:10).

The mouse strain used in the above tumor formation experiment was BALB/C-Nude, and the tumor cell line used was human lung adenocarcinoma cell line PC-9. The drug was administered orally by gavage every day, and the tumor volume was measured every 2 days. After 25 days, the tumor growth curve was drawn, the tumor weight was weighed, and the weight loss rate of the mouse was calculated. The weight loss rate of the mouse = (1-mouse weight on the day/mouse weight on the first day) × 100%; the tumor inhibition rate was calculated according to the tumor volume. (The calculation method is: tumor inhibition rate = (1-tumor volume of the combined drug group/tumor volume of the control group) * 100%.

As shown in Table 1, considering the tumor inhibition rate and weight loss rate of mice, the mass ratio of olanzapine to osimertinib is preferably (0.75-1.5): (1.25-5). The dosage of the composition in the present invention is much less than the commonly used dosage of anti-tumor drugs alone, and the side effect of olanzapine in increasing body weight compensates for the effect of osimertinib on reducing the body weight of mice, thereby improving the quality of life. The pharmaceutical composition of the present invention reduces the toxic side effects of targeted drugs on tumor patients and the cost of patient treatment.

Table 2. Dosage of the drug composition and its effect on tumor inhibition rate and body weight of tumor-bearing mice (n=3)

Example 4

The effect of the drug combination on the growth of lung cancer cells was investigated by 3D organoid culture experiment of lung cancer organoids. The tumor tissue of lung cancer patients was minced, digested by enzymatic method to obtain cell suspension, and Matrigel was further added to form a 3D culture system. The IC50 value after adding different gradient concentrations of olanzapine, the IC50 value of osimertinib, and the IC50 value of olanzapine (concentration of its IC30 value) combined with osimertinib combination were detected, and the results are shown in Figure 3. The results showed that the anti-tumor activity of olanzapine alone could be observed in the organoids of 2 patients with lung adenocarcinoma (Figure 3A, 3C); the administration of olanzapine at IC ₃₀ concentration combined with osimertinib can significantly increase the sensitivity of lung cancer organoids to osimertinib and significantly inhibit the growth of lung cancer organoids (Figure 3B, 3D).

The results of Examples 1 to 4 show that the combination of olanzapine and osimertinib can significantly inhibit the proliferation of lung cancer cells and the growth of tumors and organoids, reduce the effective drug concentration, and improve the quality of life.

The above description is only a preferred embodiment of the present invention and does not limit the present invention in any form. Any simple modification, equivalent changes and modifications made to the above embodiments based on the technical essence of the present invention are still within the protection scope of the technical solution of the present invention.

Claims (11)

1. A pharmaceutical composition characterized by: it comprises the following steps: olanzapine and octreotide.

2. The pharmaceutical composition according to claim 1, wherein: the mol ratio of olanzapine to octenib is as follows: 1. -400: 0.039 to 5.

3. The pharmaceutical composition according to claim 1, wherein: the mol ratio of olanzapine to octenib is as follows: 100-400: 0.039 to 5.

4. The pharmaceutical composition according to claim 1, wherein: the mol ratio of olanzapine to octenib is as follows: 400:0.039 to 5.

5. The pharmaceutical composition according to claim 2, wherein: the pharmaceutical composition further comprises a pharmaceutically acceptable carrier.

6. The pharmaceutical composition according to claim 5, wherein: the pharmaceutically acceptable carrier is filler, wetting agent, adhesive, disintegrating agent or lubricant.

7. The pharmaceutical composition according to claim 5, wherein: the preparation form of the medicine is oral tablet, granule, injection or capsule.

8. The use of the pharmaceutical composition according to claim 5 for the preparation of an antitumor drug, characterized in that: the tumor comprises one or more of small cell lung cancer, non-small cell lung squamous carcinoma, non-small cell lung adenocarcinoma, non-small cell adenosquamous carcinoma and large cell lung carcinoma.

9. The use of the pharmaceutical composition according to claim 5 for the preparation of an antitumor drug, characterized in that: the antitumor drug is used for inhibiting proliferation of tumor cells, wherein the tumor cells are PC-9 cells, H1975 cells or PC-9 ^{del19 -T790M-C797S} cells.

10. The use of the pharmaceutical composition according to claim 5 for the preparation of an antitumor drug, characterized in that: the antitumor drug is used for inhibiting tumor growth, and the tumor is PC-9 subcutaneous transplantation tumor, H1975 subcutaneous transplantation tumor or PC-9 ^{del19-T790M-C797S} subcutaneous transplantation tumor.

11. The use of the pharmaceutical composition according to claim 5 for the preparation of an antitumor drug, characterized in that: the antitumor drug is used for inhibiting the growth of the organoid derived from the lung cancer patient.