PHARMACEUTICAL COMBINATION FOR TREATING TUMORS AND APPLICATION THEREOF

Abstract

Provided are a pharmaceutical combination for treating tumors and an application thereof. The pharmaceutical combination comprises a compound represented by formula A, a pharmaceutically acceptable salt thereof, a solvate thereof or a solvate of a pharmaceutically acceptable salt thereof, and a “PD-1 inhibitor and/or PD-L1 inhibitor”. The components of the pharmaceutical combination, when used in combination, can significantly increase the inhibition rate of each individual drug on tumor growth, and there were no acute adverse reactions in mice after administration, demonstrating that such a combination therapy has good safety and effectiveness.

##STR00001##

Claims

1. A pharmaceutical combination, comprising: a compound of formula A: ##STR00006## a pharmaceutically acceptable salt thereof, a solvate thereof or a solvate of the pharmaceutically acceptable salt thereof; and a PD-1 inhibitor or a PD-L1 inhibitor, or a combination thereof.

2. The pharmaceutical combination according to claim 1, wherein the pharmaceutically acceptable salt of the compound of formula A is mefuparib hydrochloride; and the PD-1 inhibitor is selected from the group consisting of one or more of a PD-1 antibody, a PD-1 polypeptide inhibitor and a PD-1 micromolecular inhibitor, toripalimab, sintilimab, camrelizumab, pembrolizumab, nivolumab, and Tuoyi; and the PD-L1 inhibitor is selected from the group consisting of one or more of a PD-L1 antibody, a PD-L1 polypeptide inhibitor and a PD-L1 micromolecular inhibitor, atezolizumab, durvalumab and alirocumab.

3. The pharmaceutical combination according to claim 1, wherein the pharmaceutical combination comprises mefuparib hydrochloride and toripalimab.

4. A pharmaceutical composition X, comprising: the compound of formula A, the pharmaceutically acceptable salt thereof, the solvate thereof or the solvate of the pharmaceutically acceptable salt thereof described in claim 1; the PD-1 inhibitor or the PD-L1 inhibitor or a combination thereof described in claim 1; and a pharmaceutically acceptable excipient.

5. The pharmaceutical composition X according to claim 4, wherein the pharmaceutical composition X is in an injectable dosage form or an oral dosage form.

6. A pharmaceutical composition Y, comprising: a first pharmaceutical composition, and a second pharmaceutical composition; wherein the first pharmaceutical composition comprises: a compound of formula A, a pharmaceutically acceptable salt thereof, a solvate thereof or a solvate of the pharmaceutically acceptable salt thereof, ##STR00007## and a pharmaceutically acceptable excipient; the second pharmaceutical composition comprises: a PD-1 inhibitor or a PD-L1 inhibitor or a combination thereof, and a pharmaceutically acceptable excipient.

7. The pharmaceutical composition Y according to claim 6, wherein the first pharmaceutical composition is in an oral dosage form or, the second pharmaceutical composition is in an injectable dosage form; or the first pharmaceutical composition is in an oral dosage form and the second pharmaceutical composition is in an injectable dosage form.

8. A kit, comprising: a first container comprising the first pharmaceutical composition described in claim 6; and a second container comprising the second pharmaceutical composition described in claim 6.

9. A method of preventing or treating a tumor, comprising administering a pharmaceutical combination according to claim 1.

10. The method according to claim 9, wherein the tumor is a solid or hematological tumor or a combination thereof; and the compound of formula A, the pharmaceutically acceptable salt thereof, the solvate thereof or the solvate of the pharmaceutically acceptable salt thereof and the PD-1 inhibitor or the PD-L1 inhibitor, or a combination thereof is administered simultaneously or separately; and the compound of formula A, the pharmaceutically acceptable salt thereof, the solvate thereof or the solvate of the pharmaceutically acceptable salt thereof is administered at a dose of 100-1000 mg; and the compound of formula A, the pharmaceutically acceptable salt thereof, the solvate thereof or the solvate of the pharmaceutically acceptable salt thereof is administered at a frequency of 0.5-2 doses/day; and the compound of formula A, the pharmaceutically acceptable salt thereof, the solvate thereof or the solvate of the pharmaceutically acceptable salt thereof is administered orally; and the PD-1 inhibitor or the PD-L1 inhibitor or a combination thereof is administered at a dose of 50-500 mg; and the PD-1 inhibitor or the PD-L1 inhibitor or a combination thereof is administered at a frequency of once every 7-31 days; and the PD-1 inhibitor or the PD-L1 inhibitor or a combination thereof is administered orally or by injection.

11. The method according to claim 10, wherein the compound of formula A, the pharmaceutically acceptable salt thereof, the solvate thereof or the solvate of the pharmaceutically acceptable salt thereof is administered orally at a dose of 100-1000 mg and at a frequency of 0.5-2 doses/day; and the PD-1 inhibitor or the PD-L1 inhibitor or a combination thereof is administered by injection at a dose of 50-500 mg and at a frequency of once every 7-31 days.

12-14. (canceled)

15. The method of claim 9, wherein: the compound of formula A, the pharmaceutically acceptable salt thereof, the solvate thereof or the solvate of the pharmaceutically acceptable salt thereof is administered at a dose of 100-1000 mg and a frequency of 0.5-2 doses/day; the PD-1 inhibitor or the PD-L1 inhibitor or a combination thereof, is administered at a dose of 50-500 mg at a frequency of once every 7-31 days; and the pharmaceutically acceptable salt thereof, the solvate thereof or the solvate of the pharmaceutically acceptable salt thereof and the PD-1 inhibitor or the PD-L1 inhibitor, or a combination thereof, are administered through the same route of administration.

16. The method of claim 10, wherein the tumor is selected from one or more of lung cancer, colon cancer, rectal cancer, breast cancer, prostate cancer, liver cancer, pancreatic cancer, brain cancer, kidney cancer, ovarian cancer, stomach cancer, skin cancer, bone cancer, neuroglioma, glioblastoma, hepatocellular carcinoma, papillary kidney cancer, head and neck cancer, leukemia, lymphoma, myeloma and multiple myeloma.

17. The method of claim 9, wherein the compound of formula A, the pharmaceutically acceptable salt thereof, the solvate thereof or the solvate of the pharmaceutically acceptable salt thereof; and the PD-1 inhibitor, the PD-L1 inhibitor or a combination thereof; are administered simultaneously or separately.

18. The method of claim 9, wherein the compound of formula A, the pharmaceutically acceptable salt thereof, the solvate thereof or the solvate of the pharmaceutically acceptable salt thereof is administered at a dose of 100-1000 mg.

19. The method of claim 9, wherein the compound of formula A, the pharmaceutically acceptable salt thereof, the solvate thereof or the solvate of the pharmaceutically acceptable salt thereof is administered at a frequency of 0.5-2 doses/day.

20. The method of claim 9, wherein the compound of formula A, the pharmaceutically acceptable salt thereof, the solvate thereof or the solvate of the pharmaceutically acceptable salt thereof is administered orally.

21. The method of claim 9, wherein the PD-1 inhibitor, the PD-L1 inhibitor or a combination thereof, is administered at a dose of 50-500 mg.

22. The method of claim 9, wherein the PD-1 inhibitor, the PD-L1 inhibitor or a combination thereof, is administered at a frequency of once every 7-31 days;

23. The method of claim 9, wherein the PD-1 inhibitor, the PD-L1 inhibitor or a combination thereof, is administered orally or by injection.

Description

BRIEF DESCRIPTION OF THE DRAWINGS

[0085] FIG. 1 shows the percentage change in body weight of the experimental animals after start of treatment.

[0086] FIG. 2 shows the tumor growth curve in the experimental animals after start of treatment.

[0087] FIG. 3 shows the tumor volume in the experimental animals after treatment (PG-D24), where “-” indicates the median.

[0088] FIG. 4 shows the FACS assay results of hCD45 in peripheral blood of the experimental animals 3 weeks after PBMC inoculation, where “-” indicates the median.

DETAILED DESCRIPTION

[0089] The present invention is further illustrated by the following examples, which are not intended to limit the present invention. Experimental procedures without specified conditions in the following examples were performed in accordance with conventional procedures and conditions, or in accordance with instructions.

[0090] 1. Test Compounds

[0091] Test compound 1: Tuoyi (toripalimab, PD-1 monoclonal antibody) injection.

[0092] Test compound 2: mefuparib hydrochloride.

[0093] 2. Preparation of Test Compounds

[0094] 2.1 Solvent Selection: Tuoyi PD-1 Monoclonal Antibody Injection: PBS; Mefuparib Hydrochloride: Normal Saline.

[0095] 2.2 Preparation:

TABLE-US-00001 TABLE 1 Preparation and storage of test compounds Final concentration Test compound Preparation (mg/mL) Storage Tuoyi PD-1 A proper volume of sample was measured and added into 1 Prepared monoclonal a glass flask by using a microsyringe. A proper volume of before use antibody PBS was added into the glass flask, and the mixture was injection well mixed. Mefuparib A proper amount of sample was measured using an 4 Once a week hydrochloride electronic balance and added into a glass flask. A proper volume of normal saline was added into the glass flask, and the mixture was well mixed.

[0096] 3. Materials and Instruments

TABLE-US-00002 TABLE 2 Reagents Name Unit size Manufacturer Fetal bovine serum (FBS) 500 mL/bottle Gibco Leibovitz's L-15 medium 500 mL/bottle Gibco Penicillin-streptomycin mixed 100 mL/bottle Solarbio solution Trypsin-EDTA digestive juice 100 mL/bottle Solarbio (0.25%) Matrigel 10 mL/bottle Corning PBS (phosphate-buffered saline) 500 mL/bottle Gibco Normal saline (0.9% sodium 500 mL/bottle Shijiazhuang No. chloride injection) 4 Pharmaceutical Co., Ltd. PE-Cy7 anti-Hu CD45 100 Tests Invitrogen PE-Cy7 mouse IgG1 κ Iso Control 20 Tests Invitrogen

TABLE-US-00003 TABLE 3 Instruments and equipment Name Model No. Manufacturer CO.sub.2 incubator 3111 Thermo Microscope IX53 Olympus Electronic balance EJ-1201C Beijing Langke Xingye Weighing Equipment Co., Ltd. SF2000 three-button electronic 111N-101B Guilin Guanglu digital display caliper Measuring Instrument Co., Ltd. ACEA NovoCyte flow cytometer 3130 ACEA

[0097] 4. Experimental Animals and Accommodation Management

[0098] 4.1 Experimental Animals

[0099] Species: Mus Musculus, NCG (NOD-Prkdc.sup.em26Cd52II2rg.sup.em26Cd22/N ju), female; weight: 18-22 g; quantity: 55; supplier: GemPharmatech.

[0100] 4.2 Accommodation Management

[0101] Accommodation management: the experimental animals were housed in individually ventilated cages in an SPF-grade clean laminar flow room of constant temperature and humidity, with 5 mice in each cage.

[0102] The temperature and humidity were controlled in ranges of 23±3° C. and 40%-70%, respectively. The cages were made of polycarbonate. The dimensions were 370 mm×155 mm×135 mm. The cages were paved with autoclaved and clean soft corn cob padding, which was refreshed twice a week. The cages were labeled with the number of animals, sex, species, time of receipt, grouping and start of treatment.

[0103] Feed and water: SPF grade murine diet, disinfected by cobalt 60 radiation. The drinking water was ultrafiltered and autoclaved water. The animals had free access to sterile food and water. The animals were numbered by ear tags.

[0104] 5. Methodology

[0105] 5.1 Cell Culture

[0106] MDA-MB-436 tumor cells (YK-CL-075) were purchased from ATCC. The tumor cells were cultured in the Leibovitz's L-15 medium containing inactivated 10% fetal bovine serum, 100 U/mL of penicillin and 100 μg/mL of streptomycin in a 37° C./5% CO.sub.2 incubator, and were passaged every 3 to 4 days when the cells were overgrown. The tumor cells in logarithmic phase were used for in vivo tumor grafting.

[0107] PBMCs (human peripheral blood mononuclear cells) were purchased from ALLCELLS (Donor#: A0075).

[0108] 5.2 Tumor Cell Grafting and Grouping

[0109] MDA-MB-436 tumor cells were resuspended in a 1:1 (v:v) mixture of PBS and Matrigel at a concentration of 1×10.sup.8 cells/mL, and were grafted subcutaneously at the right flank of mice at 100 μL/mouse. When the tumor grew to about 1000 mm.sup.3, the tumors were aseptically removed and cut into small pieces having dimensions of about 2 mm×2 mm×2 mm, which were grafted subcutaneously at the right flank of experimental animals. After 2 weeks, PBMCs were resuspended in PBS and grafted into mice in an amount of 2×10.sup.6 cells/mouse. The animals were divided into 4 groups of 10 and treated (the day was recorded as PG-D0) when the tumor grew to about 93 mm.sup.3. The specific dosing regimen is shown in Table 4.

[0110] 5.3 Measurement and Experimental Parameters of Mouse Body Weight

[0111] The long and short diameters of the tumor were measured twice every week using a vernier caliper for calculating the tumor volume, and the calculation formula was: volume=0.5×long diameter×short diameter.

[0112] The T/C value was calculated from the tumor volume, where T is the mean relative tumor volume (RTV) of each treatment group and C is the mean RTV of the control group. RTV is the ratio of tumor volume after the treatment to that before the treatment. Tumor growth inhibition (TGI, %)=(1−T/C)×100%.

[0113] Three weeks after PBMC grafting, peripheral blood of the mice was collected, and the proportion of human-derived CD45.sup.+ cells (hCD45 cells) therein was determined by flow cytometry (FACS). After the end of study, the tumors was removed, weighed and photographed.

[0114] 6. Dosing Regimen

TABLE-US-00004 TABLE 4 Dosing regimen Number of Route of Frequency of animals Treatment Dose (mg/kg)* administration administration.sup.# 10 Vehicle (normal saline + PBS) — p.o. + i.p. qod × 13 + q4d × 7 10 Mefuparib hydrochloride 40 p.o. qod × 13 10 Tuoyi PD-1 monoclonal antibody injection 10 i.p. q4d × 7 10 Mefuparib hydrochloride 40 p.o. qod × 13 Tuoyi PD-1 monoclonal antibody injection 10 i.p. q4d × 7 Note: “*” the administration volume was determined as per 10 μL/g body weight, and the treatment can be discontinued until the body weight returned to normal level if body weight was reduced by 15-20%; i.p.: intraperitoneal injection; p.o.: per os; qod × 13: once every other day for 13 doses; q4d × 7: once every four days for 7 doses.

[0115] 7. Statistical Analysis

[0116] Inter-group statistical analysis was performed on tumor volume using IBM SPSS Statistics 22.0 software and mixed linear model analysis. Inter-group statistical analysis was performed on tumor weight using One-Way ANOVA analysis, with p<0.05 indicating significant differences.

EXAMPLE 1. TEST RESULTS

[0117] 1. Reaction and Change in Body Weight of Experimental Animals After Treatment

[0118] During the treatment, no significant acute adverse events were observed after treatment in all groups. In the middle and later stages of the study, mice in all groups showed weight loss, as shown in FIG. 1. The study demonstrated the good safety of the combined use of the mefuparib hydrochloride and the Tuoyi PD-1 monoclonal antibody injection.

[0119] 2. Results of Tumor Growth Inhibition

[0120] The change in tumor volume of the treated experimental animals is shown in Table 5, FIG. 2 and FIG. 3.

TABLE-US-00005 TABLE 5 Tumor (tumor volume) inhibition of test compounds on MDA-MB-436 model Number of Tumor volume (mm.sup.3).sup.a animals Grouping Administration Tumor growth Group (mouse) (PG-D0) (PG-D24) inhibition (%) P Vehicle (normal saline + PBS) 10 93 ± 7 1632 ± 292 — — Tuoyi PD-1 monoclonal 10 94 ± 7 1256 ± 252 29% 0.010 antibody injection Mefuparib hydrochloride 10 93 ± 6  761 ± 130 54% 0.000 Mefuparib hydrochloride + 10 93 ± 6  687 ± 160 60% 0.000 Tuoyi PD-1 monoclonal antibody injection

[0121] Conclusion: on day 24 after grouping (PG-D24), the tumor growth inhibitions were 54%, 29% and 60% for the mefuparib hydrochloride group, the Tuoyi PD-1 monoclonal antibody injection group and the mefuparib hydrochloride+Tuoyi PD-1 monoclonal antibody injection group, respectively. The tumor volumes in all treatment groups were significantly smaller than that of the control group (p<0.05); the tumor volume in the mefuparib hydrochloride+Tuoyi PD-1 monoclonal antibody injection group was significantly smaller than that of the Tuoyi PD-1 monoclonal antibody injection group (p<0.05). These indicated that the combination produced efficacy significantly better than that of PD-1 monotherapy and superior therapeutic trend to that of mefuparib hydrochloride.

[0122] 3. FACS Assay on Peripheral Blood

[0123] FACS assay results of hCD45 in peripheral blood of mice 3 weeks after PBMC grafting are shown in FIG. 4.

[0124] As can be seen from FIG. 4, FACS assay results of hCD45 cells in peripheral blood of the mice were positive, indicating that the humanized immune reconstruction modeling succeeded, and the percentage medians of hCD45 cells in the mefuparib hydrochloride group, the Tuoyi PD-1 monoclonal antibody injection group, and the mefuparib hydrochloride group+the Tuoyi PD-1 monoclonal antibody injection group were all higher than that in the control group, wherein the percentage median of the mefuparib hydrochloride group+the Tuoyi PD-1 monoclonal antibody injection group was higher than those in the mefuparib hydrochloride group and the Tuoyi PD-1 monoclonal antibody injection group.

[0125] In conclusion, in the PBMC humanized MDA-MB-436 subcutaneous tumor grafting model, the combination of the Tuoyi PD-1 monoclonal antibody injection and the mefuparib hydrochloride generates a statistically significant anti-tumor effect, and can effectively inhibit the tumor growth. It demonstrates a tumor growth inhibition significantly higher than that of the Tuoyi PD-1 monoclonal antibody injection and an increased proportion of hCD45 cells in peripheral blood of mice.

[0126] Although specific embodiments of the present invention have been described above, it will be appreciated by those skilled in the art that these embodiments are merely illustrative and that many changes or modifications can be made to these embodiments without departing from the principles and spirit of the present invention. The scope of protection of the present invention is therefore defined by the appended claims.