Use of combination of anti-PD-1 antibody and VEGFR inhibitor in preparation of drug for treating cancers

Abstract

Disclosed is the use of a combination of an anti-PD-1 antibody and a VEGFR inhibitor in the preparation of a drug for treating cancers.

Claims

1. A method of reducing an adverse effect of an anti-PD-1 antibody in a subject in need of a treatment of a cancer, the method comprising administering to the subject, an anti-PD-1 antibody and apatinib or the pharmaceutically acceptable salt thereof, wherein the adverse effect comprises at least hemangiomas, and the anti-PD-1 antibody comprises: an antibody light chain variable region comprising LCDR1, LCDR2 and LCDR3 having the amino acid sequences of SEQ ID NO: 4, SEQ ID NO: 5 and SEQ ID NO: 6, respectively; and an antibody heavy chain variable region comprising HCDR1, HCDR2 and HCDR3 having the amino acid sequences of SEQ ID NO: 1, SEQ ID NO: 2 and SEQ ID NO: 3, respectively.

2. The method of claim 1, comprising administering to the subject a pharmaceutically acceptable salt of apatinib selected from the group consisting of mesylate salt of apatinib and hydrochloride salt of apatinib.

3. The method of claim 1, wherein the anti-PD-1 antibody is a humanized antibody.

4. The method of claim 3, wherein the humanized antibody comprises an antibody light chain variable region having the amino acid sequence of SEQ ID NO:8 and an antibody heavy chain variable region having the amino acid sequence of SEQ ID NO:7.

5. The method of claim 3, wherein the humanized antibody comprises an antibody light chain variable region comprising a mutant sequence of the amino acid sequence of SEQ ID NO:8 having the amino acid substitution A43S, and an antibody heavy chain variable region comprising a mutant sequence of the amino acid sequence of SEQ ID NO: 7 having the amino acid substitution G44R.

6. The method of claim 1, wherein the cancer is selected from the group consisting of breast cancer, lung cancer, liver cancer, gastric cancer, intestinal cancer, renal cancer, melanoma and non-small cell lung cancer.

7. The method of claim 6, wherein the subject has failed at least one chemotherapy prior to the administration of the anti-PD-1 antibody and apatinib or the pharmaceutically acceptable salt thereof.

8. The method of claim 1, wherein the anti-PD-1 antibody is administered at a dose of 2 mg/kg to 6 mg/kg or from 100 mg to 1000 mg per administration, and the apatinib or the pharmaceutically acceptable salt thereof is administered orally at a dose from 100 mg to 500 mg.

9. The method of claim 1, wherein the anti-PD-1 antibody is administered at a dose of 3 mg/kg body weight of the subject or 200 mg once every two weeks, and apatinib or the pharmaceutically acceptable salt thereof is administered orally at a dose of 125 mg, 250 mg, 375 mg, or any dosage in between, once daily.

Description

BRIEF DESCRIPTION OF THE DRAWINGS

(1) FIG. 1. Effect of administration of antibody and compound on the relative volume of MC38 (PD-L1) xenograft in tumor-bearing mice.

(2) FIG. 2. Effect of administration of antibody and compound on body weight of tumor-bearing mice with MC38 (PD-L1) xenograft, wherein*indicates p<0.05, vs blank control group.

(3) FIG. 3. Effect of administration of antibody and compound on MC38 (PD-L1) xenograft in tumor-bearing mice-tumor weight.

DETAILED DESCRIPTION OF THE INVENTION

(4) The present invention is further described below in conjunction with the examples. These examples are not intended to limit the scope of the present invention.

Example 1: Effect of the Anti-PD-1 Antibody and Apatinib Mesylate, Administered Alone or in Combination, on Human PD-1 Transgenic C57 Mice which Bear Mouse Colon Cancer Cell MC-38 (PD-L1) Xenograft Transferred with PD-L1 Gene

(5) 1. Study Purposes

(6) Human PD-1 transgenic mice were used as test animals. The effects of anti-PD-1 antibody in combination with apartinib on human PD-1 expressed in transgenic C57 mice were evaluated, and the transgenic mice bear mouse colon cancer cell MC-38 (PD-L1) xenograft transfected with PD-L1 gene.

(7) 2. Test Antibodies and Compounds

(8) The anti-PD-1 antibody was prepared according to the method disclosed in WO2015085847 in which the corresponding code of the antibody is H005-1, and the sequences of the heavy and light chain are shown in SEQ ID NO: 7 and SEQ ID NO: 8 in the present invention. Lot number: P1512, 200 mg/vial, formulated into 20 mg/ml before use.

(9) Apatinib mesylate was prepared according to the method disclosed in CN101676267A, lot number: 668160401; molecular weight: 493.58; purity: 99.60%.

(10) 3. Experimental Animals

(11) Human PD-1 transgenic C57 mice, specific-pathogen-free (SPF), with different body weights, 50% male and 50% female, were purchased from IsisInnovation Limited, UK.

(12) 4. Drug Preparation

(13) Anti-PD-1 antibody (3 mg/kg): anti-PD-1 antibody stock solution (20 mg/ml) was adjusted to a concentration of 0.3 mg/ml with phosphate buffered saline (PBS), and the intraperitoneal injection volume was 0.2 ml/mouse.

(14) Apatinib (200 mg/kg): 400 mg apatinib was dissolved in 20 ml of 0.5% sodium carboxymethylcellulose (NaCMC), adjusted to 20 mg/ml, and was administered in 0.2 ml per mouse by gavage.

(15) The solvent vehicle (HIgG) control contained human IgG (3 mg/kg dissolved in 0.5% CMC, adjusted to 0.3 mg/ml, and the volume for intraperitoneal injection was 0.2 ml/mouse.

(16) 5. Test Method

(17) 5.1 C57 Mice were Adapted to the Laboratory Environment for >5 Days.

(18) 5.2 Tumor Cells Transplantation

(19) Skin preparation was performed on human PD-1 transgenic C57 mice one day before MC38 (PD-L1) cells (5×10.sup.6/mouse) were inoculated subcutaneously at the right flank on June 12. The tumors were then grown for 8 days. When the tumors reached 142.17±13.30 mm.sup.3, the animals were randomly assigned to 4 groups (d0) with 8 mice in each group (four male mice and four female mice in each group).

(20) 5.3 Dose and Method of Administration

(21) Anti-PD-1 antibody was injected intraperitoneally, Q2D*7 (once every 2 days, 7 times in total), and apatinib was administered by oral gavage, QD*14 (once a day for 14 days). The specific drug administration regimen is shown in Table 1.

(22) 5.4 Determination of Volume of Xenograft and Body Weight of Mice

(23) Tumor volume and body weight were measured twice a week and data were recorded.

(24) 5.5 Statistics

(25) Excel 2003 statistical software was used: the mean was calculated by avg; the SD value was calculated by STDEV; the SEM value was calculated by STDEV/SQRT; the P value indicating the difference between groups is calculated by TTEST.

(26) The formula for calculating tumor volume (V) is:
V=½×L.sub.long×L.sub.short.sup.2
Relative volume(RTV)=V.sub.T/V.sub.0
Tumor inhibition rate(%)=(C.sub.RTV−T.sub.RTV)/C.sub.RTV(%)

(27) Wherein, V.sub.0 and V.sub.T are the tumor volume at the beginning of the experiment and at the end of the experiment, respectively. C.sub.RTV and T.sub.RTV are the relative tumor volumes of the blank control group and the experimental group at the end of the experiment, respectively.

(28) 6. Test Results

(29) The PD-1 antibody was injected intraperitoneally, Q2D*7, and compound apatinib was administered by oral gavage, QD*14. On day 21 after the initial administration, the tumor inhibition rate of the group administered with the anti-PD-1 antibody (3 mg/kg) alone was 20.40%, and the tumor inhibition rate of the group administered with apatinib (200 mg/kg) alone was 35.67%. The tumor inhibition rate of the combination of the anti-PD-1 antibody (3 mg/kg) and apatinib (200 mg/kg) was 63.07% (significantly different from that in human IgG control group) There were no significant differences between the other administration groups (administration of agent alone) relative to the human IgG control group. From the experimental results, the efficacy of the combination group of the anti-PD-1 antibody (3 mg/kg)+apatinib (200 mg/kg) is superior to that of the anti-PD-1 antibody administrated alone and that of apintinib administrated alone. The body weight of mice in each group was normal, indicating that the drug had no obvious side effects. The specific data are shown in Table 1 and FIGS. 1-3.

Example 2: Clinical Study of Anti-PD-1 Antibody Combined with Apatinib Mesylate in the Treatment of Advanced Malignant Tumor

(30) Inclusion criteria: (1) advanced malignancy; (2) failure in chemotherapy by using first-line, second-line or above; (3) measurable lesions; (4) ECOG score 0-1.

(31) Test drugs: commercially available apatinib mesylate tablet; the anti-PD-1 antibody of Example 1.

(32) Method of administration: Up to Sep. 20, 2017, a total of 31 subjects were screened, 30 subjects were enrolled (14 subjects withdrew from treatment, and 16 subjects were still in the group of administration).

(33) Administration method for subjects No. 001-005 was intravenous infusion of anti-PD-1 antibody, 3 mg/kg, once every 2 weeks; apatinib orally, 500 mg, once a day. Administration method for subjects No. 006-010 was intravenous infusion of anti-PD-1 antibody, 200 mg, once every 2 weeks; apatinib orally, 125 mg, once a day. Administration method for subjects No. 011-031 was intravenous infusion of anti-PD-1 antibody, 200 mg, once every 2 weeks; apatinib orally, 250 mg, once a day.

(34) Clinical Outcome: In terms of effectiveness, in the 6.sup.th week, there were 24 evaluable data for efficacy evaluation, with a disease control rate (DCR) of 83.3% (20/24); in the 12th week, there were 19 evaluable data for efficacy evaluation, with a DCR of 63.2% (12/19); in the 18th week, there were 10 evaluable data for efficacy evaluation, with a DCR of 70% (7/10); in the 24.sup.th week, there were 5 evaluable data for efficacy evaluation, with a DCR of 80% (4/5); up to Sep. 20, 2017, there were 2 hepatocellular carcinoma subjects with their 24-week effect of partial response (PR) and progression-free survival (PFS) of more than 6 months. Among the 24 evaluable data, there were 4 cases showing optimal efficacy with PR, 15 cases of stable disease (SD), and 5 cases of progressive disease (PD). Although the objective response rate (ORR) was only 16.7%, the DCR was high, e.g., as high as 79%, and some subjects had a PFS of more than 6 months. The specific results are shown in Table 2, Table 3 and Table 4. In addition, the dose of apatinib alone in treatment of solid tumor (such as gastric cancer, gastroesophageal junction adenocarcinoma, liver cancer, etc.) was usually up to 850 mg/day (see instructions for apatinib). However, in embodiments of the invention the combination of apatinib and anti-PD-1 antibody makes it possible to reduce the dose of apatinib down to 125 mg/day, and provides improved effectiveness and better safety when compared with apatinib administrated alone. In terms of safety, up to September 20, 11 cases of serious adverse events (SAE) were reported in 8 subjects, and the incidence of SAE was 26.7% (8/30). Seven of the SAEs were observed in subjects No. 001-005 (wherein the dose of apatinib for initial test was high, 500 mg) and accounted for most of the serious adverse events. However, with modified dosage regimen, it was found that good anti-tumor effect could be maintained, and the adverse effects caused by high dose of apintinib could be significantly reduced. In addition, in this clinical study it was surprisingly found that the combination of apatinib and anti-PD-1 antibody showed almost no hemangioma-associated adverse effect in the treatment of malignant tumors when compared with anti-PD-1 antibody alone. Hemangiomas was observed in only one subject who was administrated with PD-1 antibody alone, due to intolerance to combination therapy.

Example 3: Phase II Clinical Study of Anti-PD-1 Antibody Combined with Apatinib Mesylate in the Treatment of Advanced Non-Small Cell Lung Cancer

(35) Inclusion criteria: (1) advanced non-small cell lung cancer; (2) failure in chemotherapy by using first-line or second-line or above; (3) measurable lesions; (4) ECOG score 0-1.

(36) Test drugs: commercially available apatinib mesylate tablet; the anti-PD-1 antibody of Example 1.

(37) Method of administration: Anti-PD-1 antibody, once every 2 weeks, intravenous infusion, 200 mg each time; apatinib mesylate orally, once daily, 250 mg or 375 mg or 500 mg each time.

(38) Clinical results: up to July 28, a total of 15 subjects were screened, of which 12 were enrolled. A total of 12 subjects completed at least 1 cycle of administration observation, 10 patients (10/12) had disease in stable condition, and 1 patient had partial remission. See Table 5 for details. Surprisingly, it was found that the combination of apatinib mesylate and anti-PD-1 antibody enhanced the efficacy and reduced the adverse effects when compared with anti-PD-1 antibody administered alone. In this study, the common adverse effects were usually grade I to II, and the incidence of anti-PD-1 antibody-associated or immune-associated adverse effects (such as capillary hemangioma) was only 8% (1 case), the incidence of hypothyroidism was only 8% (1 case), and gastrointestinal adverse effects (such as diarrhea) and skin adverse effect (such as pruritus) were not observed. In an ASCO report published in 2017, anti-PD-1 antibody administrated alone for the treatment of solid tumors in phase I clinical trial exhibited an incidence of capillary hemangioma as high as 79.3%, and the incidence of hypothyroidism was 29.3%, the incidence of pruritus was 19.0%, the incidence of diarrhea was 10.3% (Phase I study of the anti-PD-1 antibody SHR-1210 in patients with advanced solid tumors. (2017): e15572-e15572). Therefore, the combination of apatinib mesylate and anti-PD-1 antibody can not only alleviate or control the tumor proliferation of non-small cell lung cancer (which has experienced chemotherapy failure), but also reduce the anti-PD-1 antibody-associated or immune-mediated adverse effects and improve the life quality of patients.

(39) TABLE-US-00003 TABLE 1 Number Mean tumor volume Mean tumor volume Relative tumor % Tumor of (mm.sup.3) (mm.sup.3) volume inhibition P animals/ Group Administration Route D 0 SEM D 21 SEM D 21 SEM rate D 21 (vs blank) group HIgG (3 mg/kg) Q2D*7 ip 141.46 13.23 1983.55 292.09 14.41 2.07 — — 8 Anti-PD-1 antibody Q2D*7 ip 146.40 12.68 1652.93 309.61 11.47 2.49 20.40% 0.379164 8 (3 mg/kg) Anti-PD-1 antibody Q2D*7/ ip/po 146.11 11.69 771.95 73.42 5.32 0.73 63.07%** 0.001007 8 (3 mg/kg) + apatinib QD(14 D) (200 mg/kg) apatinib QD(14 D) po 139.70 7.59 1263.86 206.54 9.27 1.58 25.67% 0.068923 8 **p < 0.01, vs control group

(40) TABLE-US-00004 TABLE 2 Administration methods: PD-1 antibody 3 mg/kg + apatinib 500 mg Previous Treatment 6 weeks 12 weeks 18 weeks 24 weeks 32 weeks Optimal No. Diagnosis therapy cycle evaluation evaluation evaluation evaluation evaluation efficacy 001 gastric cancer Second-line 1 NA NA NA NA NA Not evaluated therapy 002 gastric cancer Fouth-line 6 SD reduced PD NA NA NA SD therapy 003 gastric cancer Fifth-line 9 SD reduced SD reduced PD PD NA SD therapy 004 hepatocellular First-line 2 SD increased NA NA NA NA SD carcinoma therapy 005 hepatocellular Second-line 1 NA NA NA NA NA Not evaluated carcinoma therapy

(41) TABLE-US-00005 TABLE 3 Administration methods: PD-1 antibody 200 mg + apatinib 125 mg Previous Treatment 6 weeks 12 weeks 18 weeks 24 weeks 32 weeks Optimal No. Diagnosis therapy cycle evaluation evaluation evaluation evaluation evaluation efficacy 006 hepatocellular Second-line 18 SD increased SD increased PD PR SD PR carcinoma therapy 007 hepatocellular Second-line 18 SD SD reduced SD reduced SD Performed, SD carcinoma therapy Not evaluated 009 hepatocellular Second-line 18 SD SD reduced SD reduced SD reduced NA SD carcinoma therapy 008 hepatocellular First-line 4 PD NA NA NA NA PD carcinoma therapy 010 gastric cancer Third-line 2 NA NA NA NA NA Not evaluated therapy

(42) TABLE-US-00006 TABLE 4 Administration methods: PD-1 antibody 200 mg + apatinib 250 mg Treatment 6 weeks 12 weeks 18 weeks 24 weeks 32 weeks Optimal No. Diagnosis Therapy cycle evaluation evaluation evaluation evaluation evaluation efficacy 011 hepatocellular Second-line 15 SD reduced PR PR PR NA PR carcinoma therapy 014 hepatocellular Third-line 14 SD reduced SD reduced SD Performed, NA SD carcinoma therapy Not evaluated 019 hepatocellular Second-line 11 SD SD reduced SD NA NA SD carcinoma therapy 021 hepatocellular First-line 9 SD increased SD Performed, NA NA SD carcinoma therapy Not evaluated 027 hepatocellular Second-line 3 SD reduced Not performed NA NA NA SD carcinoma therapy 018 hepatocellular Second-line 4 SD increased PD NA NA NA PD carcinoma therapy 016 gastric cancer Fouth-line 9 PR PD PD NA NA PR therapy 025 gastric cancer Second-line 8 PR PR NA NA NA PR therapy 012 gastric cancer Multi-line 9 SD reduced SD reduced SD increased NA NA SD therapy 013 gastric cancer Third-line 5 SD NA NA NA NA SD therapy 022 gastric cancer Second-line 10 SD reduced SD SD NA NA SD therapy 024 gastric cancer Third-line 6 SD increased PD NA NA NA SD therapy 026 gastric cancer Third-line 8 SD reduced SD NA NA NA SD therapy 028 gastric cancer Third-line 7 SD Performed, NA NA NA SD therapy Not evaluated 015 gastric cancer Second-line 5 9 weeks PD PD NA NA NA PD therapy 017 gastric cancer Fifth-line 4 PD PD NA NA NA PD therapy 023 gastric cancer Second-line 5 PD PD NA NA NA PD therapy 029 gastric cancer Multi-line 5 Performed, NA NA NA NA To be therapy Not evaluated evaluated 030 gastric cancer Third-line 5 Performed, NA NA NA NA To be therapy Not evaluated evaluated 031 gastric cancer Second-line 3 Not NA NA NA NA Not evaluated therapy Performed

(43) TABLE-US-00007 TABLE 5 Efficacy evaluation of enrolled patients Efficacy evaluation-Diameter Screening (mm)/Baseline ratio (%) General No. Baseline 2 cycles 4 cycles evaluation 01001 28 27/−3.6%  15.5/−44.6% PR 01002 20.6 1 cycle, PD hydrothorax increased 01003 76.7 New onset of 74.7/−2.6% SD hydrothorax 01005 122.9 liver metastases PD increased, enlarged 01006 137 118/−13.9% 107/−22% SD 01007 134.2 117.7/−12.2%   107/−20.3 SD 01008 58.3  52/−10.8% SD SD 01010 11  9/−20% SD 01011 36 SD SD 01013 87 SD SD 01014 85.5 SD SD 01015 SD SD