CN109663130B

CN109663130B - Use of a combination of a PD-1 antibody and a MEK inhibitor for the preparation of a medicament for the treatment of tumors

Info

Publication number: CN109663130B
Application number: CN201811186529.1A
Authority: CN
Inventors: 邹建军; 徐瑞华; 刘毅
Original assignee: Jiangsu Hengrui Medicine Co Ltd
Current assignee: Jiangsu Hengrui Medicine Co Ltd
Priority date: 2017-10-13
Filing date: 2018-10-12
Publication date: 2021-06-29
Anticipated expiration: 2038-10-12
Also published as: CN109663130A

Abstract

The present invention relates to the use of a PD-1 antibody or antigen-binding fragment thereof in combination with a MEK inhibitor in the preparation of a medicament for the treatment of a tumor. In particular, the invention relates to the use of the combination of a PD-1 antibody or an antigen-binding fragment thereof and a compound (1) or a pharmaceutically acceptable salt thereof for the preparation of a medicament for the treatment of tumors.

Description

Use of a combination of a PD-1 antibody and a MEK inhibitor for the preparation of a medicament for the treatment of tumors

Technical Field

Use of a PD-1 antibody or antigen-binding fragment thereof in combination with a MEK inhibitor in the manufacture of a medicament for the treatment of a tumor.

Background

The occurrence and development of malignant tumors are closely related to abnormal sustained activation of intracellular signaling pathways. The signaling pathway consisting of MAPK (Mitogen-activated protein kinase) controls a number of important physiological processes in cells, with the ERK pathway (Extracellular regulated protein kinase pathway, RAS-RAF-MEK1/2-ERK1/2, Extracellular regulatory protein kinase) being the classical MAPK pathway with abnormal cascade signaling that occurs most frequently in human tumors. Abnormal activation of the ERK pathway is usually caused by acquired mutations in members of the RAS (Rat sarcoma gene, Rat sarcoma protein homologous gene) and RAF (rapid accepted Fibrosarcoma kinase) gene families ([ J ]. Biochimica et Biophysica Acta (BBA) -Molecular Cell Research,2007,1773(8): 1263-. The cancer of bile duct, carcinoma of large intestine, lung cancer, bladder cancer, pancreatic cancer and prostatic cancer all have RAS coding gene, while melanoma, papillary thyroid cancer and low-differentiation ovarian cancer have BRAF gene mutation. BRAF and its downstream target MEK are kinases in the MAPK pathway, playing an important role in cell proliferation ([ J ]. Drug design, maintenance and therapy,2016,10: 43.). The PD-1 (programmed death receptor 1) antibody can specifically recognize and combine with the PD-1 on the surface of the lymphocyte, block a PD-1/PD-L1 signal channel, further activate the immune killing effect of the T cell on the tumor, mobilize the immune system of the organism and eliminate the tumor cells in the body.

With the progress of the study of PD-1/PD-L1 immunotherapy, many patients were found to be insensitive or resistant to immune checkpoint therapy, and meta-analysis by Xia Bu et al showed that immunotherapeutic resistance targeting PD-1/PD-L1 might be associated with upregulation of certain genes, such as IL-10, VEGFA, VEGFC, FLT1, ANGPT2, and monocyte and macrophage driver (J. Trends in molecular menu, 2016,22(6):448 and 451) involved in immune expression. MEK protein kinase is one of the important regulatory targets of normal T cell and RAS signal pathway mutation tumor, and MEK inhibition can block the proliferation of mouse naive T cell, and enhance the anti-tumor activity of previously activated T cell by damaging TCR-driven apoptosis, so that the MEK inhibitor and an antibody with PD-1/PD-L1 target can generate a lasting tumor inhibition effect in combination ([ J ]. Immunity,2016,44(3): 609-621.). WO2016040892 discloses the combination of a PD-1 antibody selected from Pembrolizumab et al and a MEK inhibitor for the treatment of various tumors; WO2016011160 discloses the combination of a PD-1 antibody selected from Nivolumab, Pembrolizumab and the like and a MEK inhibitor for the treatment of a variety of B-RAF antagonist-resistant tumors; WO2014195852 and WO2014193898 disclose the treatment of B-RAF V600 mutant melanoma in combination with Trametinib or Dabrafenib in combination with PD-L1 antibody or PD-1 antibody; WO2013019906 discloses the treatment of B-RAF V600E mutant tumors with various PD-1 antibodies in combination with various MEK inhibitors.

Several clinical studies are currently under development in which PD-1 or PD-L1 is combined with a MEK inhibitor for the treatment of tumors. Clinical studies by Wilson H.Miller et al disclosing Atezolizumab (PD-L1) in combination with cobimetinib for the treatment of metastatic melanoma showed that the combination was comparable to either aloneHas higher DCR (disease control rate) and median progression free survival (mPFS) (Atezolizumab (A)) + cobimetinib (C) in metastic melanoma (mel): Updated safety and clinical activity [ J]2017.); the results of clinical studies reported by Michelle Rohlfs et al on the treatment of metastatic melanoma with a BRAF inhibitor +/-MEK inhibitor in combination with a PD-1 immune checkpoint blocker (pembrolizumab or nivolumab) showed that mPFS was 7.6 months in patients in the group, with the BRAF inhibitor + MEK inhibitor + PD-1 immune checkpoint blocker group at 8.5 months and the BRAF inhibitor + PD-1 immune checkpoint blocker group at 7.2 months, and subjects were well-tolerated (BRAF with the same or with a MEK inhibition plus PD-1 checkpoint blockade for the same treatment of metastatic melanomas [ J]2016.); antoni Ribas et al reported that Pembrolizumab combined with dabrafenib and trametinib treats BRAF^V600E/KThe results of clinical studies of mutant advanced melanoma showed that ORR (Total remission rate) was 60% in 15 subjects that could be evaluated, but severe adverse reactions such as neutropenia occurred in 3 subjects (Pembrolizumab in combination with debrafenib and transmitinib for BRAF-mutated advanced melanoma: Phase 1KEYNOTE-022 study [ J].2016)。

In summary, although studies on PD-1 antibody and MEK inhibitor have been reported, BRAF inhibitor is mainly combined with MEK inhibitor and PD-L1 antibody, and clinical benefit and safety of subjects are not satisfactory, so there is still a need in the field of treatment to further explore suitable PD-1 antibody in combination with MEK inhibitor for treating relapsing refractory tumors.

The anti-PD-1 antibody provided by the invention, WO201508584 discloses a sequence and a preparation method of the antibody, the PD-1 antibody is currently in the domestic Clinical stage I, the safety is good, and reported Clinical research results show that the antibody has a certain anti-tumor effect ([ J ]. Journal of Clinical Oncology 35(2017): e15572-e 15572); in addition, WO2015058589 discloses a structure and a preparation method of the MEK inhibitor, in vitro research shows that the MEK inhibitor has better tumor inhibition activity, and the specific structure is shown as follows:

disclosure of Invention

The present invention provides the use of an immunotherapeutic agent selected from a PD-1 antibody or an antigen-binding fragment thereof in combination with a MEK inhibitor in the manufacture of a medicament for the treatment of a tumour; the MEK inhibitor is selected from AZD-8330, GDC-0623, CI-1040, WX-554, TAK-733, HL-085, BI-847325, CEP-1347, Binimetib, Pimaseritib, Cobimetinib, PD-0325901, RO-5126766, Trametinib, Refametinib, AS-703988, E-6201, Selumetinib, or a compound (1) shown below or a pharmaceutically acceptable salt thereof,

in a preferred embodiment of the present invention, the MEK inhibitor is preferably selected from Binimetinib, Pimasertib, Cobimetinib, Trametinib, Refametinib, compound (1) or a pharmaceutically acceptable salt thereof.

PD-1 antibodies or antigen-binding fragments thereof are known, preferably the light chain variable region of the PD-1 antibody or antigen-binding fragment thereof comprises LCDR1, LCDR2 and LCDR3 as shown in SEQ ID NO. 4, SEQ ID NO. 5 and SEQ ID NO. 6, respectively.

The heavy chain variable region of the PD-1 antibody or the antigen-binding fragment thereof comprises HCDR1, HCDR2 and HCDR3 shown as SEQ ID NO 1, SEQ ID NO 2 and SEQ ID NO 3, respectively.

Wherein, the CDR sequences are shown in the following table:

name (R)	Sequence of	Numbering
			HCDR1	SYMMS	SEQID NO：1
HCDR2	TISGGGANTYYPDSVKG	SEQID NO：2
			HCDR3	QLYYFDY	SEQID NO：3
LCDR1	LASQTIGTWLT	SEQID NO：4
			LCDR2	TATSLAD	SEQID NO：5
LCDR3	QQVYSIPWT	SEQID NO：6

Preferably, the PD-1 antibody or an antigen-binding fragment thereof is a PD-1 humanized antibody.

Preferably, the humanized antibody light chain variable region sequence is as shown in SEQ ID NO 10 or a variant thereof; the variant preferably has 0-10 amino acid changes in the light chain variable region; more preferably the amino acid change of A43S. The humanized antibody heavy chain variable region sequence is shown as SEQ ID NO. 9 or the variant thereof; the variant preferably has 0-10 amino acid changes in the heavy chain variable region; more preferably the amino acid change of G44R.

The variable region sequences of the heavy and light chains of the humanized antibody are shown as follows:

heavy chain variable region

EVQLVESGGGLVQPGGSLRLSCAASGFTFSSYMMSWVRQAPGKGLEWVATISGGGANTYYPDSVKGRFTISRDNAKNSLYLQMNSLRAEDTAVYYCARQLYYFDYWGQGTTVTVSS

SEQID NO：9

Light chain variable region

DIQMTQSPSSLSASVGDRVTITCLASQTIGTWLTWYQQKPGKAPKLLIYTATSLADGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQVYSIPWTFGGGTKVEIK

SEQID NO：10

Preferably, the humanized antibody light chain sequence is as shown in SEQ ID NO. 8 or a variant thereof; the variant preferably has 0-10 amino acid changes in the light chain variable region; more preferably the amino acid change of A43S. The humanized antibody heavy chain sequence is shown as SEQ ID NO. 7 or the variant thereof; the variant preferably has 0-10 amino acid changes in the heavy chain variable region; more preferably the amino acid change of G44R.

Particularly preferably, the light chain sequence of the PD-1 humanized antibody is shown as SEQ ID NO. 8, and the heavy chain sequence is shown as SEQ ID NO. 7.

The sequences of the heavy chain and the light chain of the PD-1 humanized antibody are shown as follows:

heavy chain

EVQLVESGGGLVQPGGSLRLSCAASGFTFSSYMMSWVRQAPGKGLEWVATISGGGANTYYPDSVKGRFTISRDNAKNSLYLQMNSLRAEDTAVYYCARQLYYFDYWGQGTTVTVSSASTKGPSVFPLAPCSRSTSESTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTKTYTCNVDHKPSNTKVDKRVESKYGPPCPPCPAPEFLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSQEDPEVQFNWYVDGVEVHNAKTKPREEQFNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKGLPSSIEKTISKAKGQPREPQVYTLPPSQEEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSRLTVDKSRWQEGNVFSCSVMHEALHNHYTQKSLSLSLGK

SEQID NO：7

Light chain

DIQMTQSPSSLSASVGDRVTITCLASQTIGTWLTWYQQKPGKAPKLLIYTATSLADGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQVYSIPWTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC

SEQID NO：8

In a preferred embodiment of the invention, said tumor is selected from the group consisting of a malignant tumor, a benign tumor; the malignant tumor is selected from malignant epithelial tumor, sarcoma, myeloma, leukemia, lymphoma, melanoma, head and neck tumor, brain tumor, peritoneal cancer, mixed tumor, and children malignant tumor; the malignant epithelial tumor is selected from lung cancer, breast cancer, liver cancer, pancreatic cancer, colorectal cancer, gastric cancer, gastroesophageal adenocarcinoma, esophageal cancer, small intestine cancer, cardiac carcinoma, endometrial cancer, ovarian cancer, fallopian tube cancer, vulval cancer, testicular cancer, prostate cancer, penile cancer, kidney cancer, bladder cancer, anal cancer, gallbladder cancer, bile duct cancer, teratoma and heart tumor; the head and neck tumor is selected from nasopharyngeal carcinoma, laryngeal carcinoma, thyroid carcinoma, tongue cancer, and oral cancer; the sarcoma is selected from Askin tumor, chondrosarcoma, Ewing's sarcoma, malignant vascular endothelioma, malignant nerve sheath tumor, osteosarcoma, and soft tissue sarcoma; the myeloma is selected from isolated myeloma, multiple myeloma, diffuse myeloma, leukemia myeloma, and marrow-type myeloma; the leukemia is selected from acute lymphocytic leukemia, chronic lymphocytic leukemia, acute myelogenous leukemia, chronic myelogenous leukemia, hairy cell leukemia, T cell lymphocytic leukemia, large granular lymphocytic leukemia, adult T cell leukemia; the lymphoma is selected from non-Hodgkin lymphoma and Hodgkin lymphoma; the brain tumor is selected from the group consisting of neuroepithelial tissue tumor, cranial nerve and spinal nerve tumor, meningeal tissue tumor; the malignant tumor of children is selected from nephroblastoma, neuroblastoma, retinoblastoma, and germ cell tumor of children.

In another preferred embodiment of the present invention, said lung cancer is selected from the group consisting of non-small cell lung cancer, small cell lung cancer; the breast cancer is selected from Hormone Receptor (HR) positive breast cancer, human epidermal growth factor receptor-2 (HER2) positive breast cancer and triple negative breast cancer; the renal cancer is selected from clear renal cell carcinoma, papillary renal cell carcinoma, chromophobe renal cell carcinoma, collecting duct carcinoma; the neuroepithelial tissue tumor is selected from preferably astrocytoma, anaplastic astrocytoma, glioblastoma; the liver cancer is selected from primary liver cancer and secondary liver cancer, and the primary liver cancer is selected from hepatocellular carcinoma, cholangiocellular carcinoma and mixed liver cancer; the colorectal cancer is selected from colon cancer and rectal cancer.

In another preferred embodiment of the present invention, the tumor is selected from hodgkin's lymphoma, non-hodgkin's lymphoma, prostate cancer, pancreatic cancer, lung cancer, esophageal cancer, liver cancer, bile duct cancer, breast cancer, colorectal cancer, stomach cancer, kidney cancer, acute myelogenous leukemia, myelodysplastic syndrome, glioma, tumor of unknown primary site.

In a preferred embodiment of the invention, the tumor is mediated by PD-1 and/or expresses PD-L1.

In a preferred embodiment of the invention, the tumor is selected from the group consisting of RAS mutant tumors, RAF mutant tumors, RAS/RAF/MEK pathway-aberrant tumors, RAS/RAF pathway-aberrant tumors.

In a preferred embodiment of the invention, said RAS mutant is selected from the group consisting of an HRas mutant, a KRas mutant, an NRas mutant; the RAF mutant is selected from A-RAF mutant and B-RAF mutant. Wherein the B-RAF mutant is preferably selected from the group consisting of B-RAF V600E mutant, B-RAF V600K mutant, B-RAF V600D mutant and B-RAF V600R mutant.

In the present invention, the mutation is a positive mutation.

In a preferred embodiment of the invention, the tumor is selected from the group consisting of a medium-advanced tumor, a relapsed refractory tumor, a tumor that has failed treatment with a chemotherapeutic drug and/or has relapsed, a tumor that has failed treatment with a radiotherapy, a tumor that has failed treatment with a targeted drug and/or has relapsed, a tumor that has failed treatment with an immunotherapy and/or has relapsed.

In a preferred embodiment of the invention, the tumor is resistant or resistant to an immunotherapeutic agent or immunotherapy, preferably the immunotherapeutic agent targets PD-1 and/or PD-L1 or CTLA-4 (cytotoxic T lymphocyte-associated protein 4); the immunotherapy is selected from Immune Checkpoint Blockade (ICB) therapy, chimeric antigen receptor T cell immunotherapy (CAR-T therapy), autologous cell immunotherapy (CIK therapy).

In a preferred embodiment of the invention, preferably, the immunotherapeutic agent is selected from the group consisting of PD-1 antibodies, PD-L1 antibodies, CTLA-4 antibodies, and PD-1 antibodies include, but are not limited to, Pidilizumab, MEDI-0680, AMP-224, PF-06801591, TSR-042, JS-001, GLS-010, PDR-001, Genolimzumab, Camrelizumab, BGB-A317, IBI-308, REGN-2810, Pembrolizumab, Nivolumab; the PD-L1 antibodies include, but are not limited to, MSB-0011359-C, CA-170, LY-3300054, BMS-936559, Durvalumab, Avelumab, Atezolizumab; the CTLA-4 antibodies include, but are not limited to, ipilimumab, AK-104, JHL-1155, ATOR-1015, AGEN-1884, PRS-010, tremelimumab, IBI-310, MK-1308, BMS-986218, SN-CA21, FPT-155, KN-044, CG-0161, ONC-392, AGEN-2041, PBI-5D3H 5.

In a preferred embodiment of the invention, the dosage of the PD-1 antibody or antigen-binding fragment thereof is selected from 1 to 10mg/kg, preferably from 1mg/kg, 2mg/kg, 3mg/kg, 4mg/kg, 5mg/kg, 6mg/kg, 7mg/kg, 8mg/kg, 9mg/kg, 10mg/kg, more preferably 1mg/kg, 3mg/kg, 4mg/kg, 5mg/kg, 6mg/kg, 10 mg/kg.

In a preferred embodiment of the invention, the dosage of the PD-1 antibody or antigen-binding fragment thereof is selected from the group consisting of 50-600mg, preferably 50mg, 60mg, 70mg, 75mg, 100mg, 125mg, 150mg, 175mg, 200mg, 225mg, 250mg, 375mg, 400mg, 425mg, 450mg, 475mg, 500mg, 600mg, more preferably 60mg, 100mg, 200mg, 400mg, 600 mg.

In a preferred embodiment of the invention, the MEK inhibitor is present in an amount selected from 0.01-500mg, preferably from 0.1mg, 0.25mg, 0.5mg, 0.75mg, 1mg, 2mg, 3mg, 4mg, 5mg, 6mg, 7mg, 8mg, 9mg, 10mg, 11mg, 12mg, 12.5mg, 15mg, 17.5mg, 20mg, 22.5mg, 25mg, 30mg, 45mg, 50mg, 60mg, 70mg, 75mg, 80mg, 90mg, 100mg, 125mg, 150mg, 175mg, 200mg, 225mg, 250mg, 275mg, 300mg, 400mg, 500mg, more preferably 0.25mg, 0.5mg, 1mg, 2mg, 3mg, 4mg, 10mg, 15mg, 20mg, 30mg, 45mg, 50mg, 60mg, 75mg, 100 mg.

In a preferred embodiment of the invention, the MEK inhibitor is selected from compound (1), or a pharmaceutically acceptable salt thereof, in a dose selected from 0.01-500mg, preferably from 0.1mg, 0.25mg, 0.5mg, 0.75mg, 1mg, 2mg, 3mg, 4mg, 5mg, 6mg, 7mg, 8mg, 9mg, 10mg, 11mg, 12mg, 12.5mg, 15mg, 17.5mg, 20mg, 22.5mg, 25mg, 30 mg.

In a preferred embodiment of the present invention, the pharmaceutically acceptable salt of compound (1) is selected from the group consisting of p-toluenesulfonate.

In the present invention, the PD-1 antibody or antigen-binding fragment thereof is administered in combination with a MEK inhibitor for tumor therapy, either in a sequence in which the MEK inhibitor is administered prior to, or concurrently with, administration of the PD-1 antibody or antigen-binding fragment thereof, or the MEK inhibitor is administered after administration of the PD-1 antibody or antigen-binding fragment thereof; preferably, the MEK inhibitor is administered prior to the administration of the PD-1 antibody or antigen-binding fragment thereof.

In the present invention, the PD-1 antibody or antigen-binding fragment thereof is used in combination with a MEK inhibitor for tumor therapy, and both can be administered during the same dosing cycle.

In the present invention, the treatment cycle may be 1 day, 3 days, 1 week, 2 weeks, 3 weeks (21 days), 3-4 weeks (21-28 days), 4 weeks (28 days), preferably 3 weeks or 3-4 weeks or 4 weeks.

In the present invention, the treatment cycle includes, but is not limited to, a chemotherapy cycle or a radiotherapy cycle or other related targeted drug treatment cycle or an immunotherapy cycle.

In the present invention, the MEK inhibitor and the PD-1 antibody or antigen-binding fragment thereof may be used in combination for treating a tumor in the same or different treatment cycles, and during the treatment of a tumor, the MEK inhibitor may be administered in combination with the PD-1 antibody or antigen-binding fragment thereof simultaneously with, before or after, or in combination with a treatment regimen according to a different tumor-preferred chemotherapy regimen or radiotherapy regimen or targeted small molecule drug treatment regimen or immunotherapy regimen, including but not limited to cellular immunotherapy (e.g., CAR-T therapy, tumor vaccine, CIK therapy, etc.); in addition, the combined administration of the MEK inhibitor and the PD-1 antibody or antigen-binding fragment thereof may also be performed alone without combination with other treatment regimens.

In the present invention, the PD-1 antibody or antigen-binding fragment thereof and MEK inhibitor, when used in combination, either before, or after, may be subjected to a regimen of treatment of tumors of different pathological types and progression stages as recommended by various oncological guidelines or guidelines, including, but not limited to, NCCN (national cancer network integration promulgated guidelines for clinical practice of various malignancies) or the malignant oncological guidelines promulgated by the department of health of china.

In the present invention, the PD-1 antibody or antigen-binding fragment thereof is used in combination with a MEK inhibitor for the treatment of a tumor within the same treatment cycle (e.g., one treatment cycle of 28 days), the MEK inhibitor being administered either simultaneously with or before or after the PD-1 antibody or antigen-binding fragment thereof; the MEK inhibitor is administered during the same administration cycle for a continuous period of time, preferably 1-21 days, 1-28 days, which may begin at any one time point of a treatment cycle, preferably at the beginning of the treatment cycle; the PD-1 antibody or antigen-binding fragment thereof can be administered at a frequency of 2 weeks/time, 3 weeks/time, or 4 weeks/time over the same administration period, and can be administered on

days

1 and 15 of a treatment cycle.

The invention relates to a "combination" which is a mode of administration and means that at least one dose of a PD-1 antibody or antigen-binding fragment thereof and at least one dose of a MEK inhibitor, both of which exhibit a pharmacological effect, are administered over a period of time. The time period may be within one administration cycle, preferably within 4 weeks, within 3 weeks, within 2 weeks, within 1 week, or within 24 hours, more preferably within 12 hours. The PD-1 antibody or antigen-binding fragment thereof and the MEK inhibitor may be administered simultaneously or sequentially. Such terms include treatments in which the PD-1 antibody or antigen-binding fragment thereof and the MEK inhibitor are administered by the same route of administration or different routes of administration. The mode of administration of the combinations of the invention is selected from simultaneous administration, separate formulation and co-administration or separate formulation and sequential administration.

In the present invention, the present invention further relates to a use in a medicament, wherein the PD-1 antibody or an antigen-binding fragment thereof is administered once a day, twice a day, three times a day, once a week, once a three week, once a month, and the MEK inhibitor is administered once a day, twice a day, three times a day, once a week, once a two week, once a three week, once a month.

In a particularly preferred embodiment, the PD-1 antibody or antigen-binding fragment thereof is administered once a week and the MEK inhibitor is administered once a day.

In a preferred embodiment of the invention, the PD-1 antibody or antigen-binding fragment thereof is administered by injection, for example, subcutaneously or intravenously, the PD-1 antibody or antigen-binding fragment thereof being formulated in an injectable form prior to injection. A particularly preferred injectable form of the PD-1 antibody or antigen-binding fragment thereof is an injection solution or a lyophilized powder injection comprising the PD-1 antibody or antigen-binding fragment thereof, a buffer, a stabilizer, and optionally further comprising a surfactant. The buffer can be one or more selected from acetate, citrate, succinate and phosphate. The stabilizer may be selected from sugars or amino acids, preferably disaccharides, such as sucrose, lactose, trehalose, maltose. The surfactant is selected from polyoxyethylene hydrogenated castor oil, glycerin fatty acid ester, polyoxyethylene sorbitan fatty acid ester, preferably the polyoxyethylene sorbitan fatty acid ester is polysorbate 20, 40, 60 or 80, most preferably polysorbate 20. The most preferred injectable form of the PD-1 antibody or antigen-binding fragment thereof comprises the PD-1 antibody or antigen-binding fragment thereof, acetate buffer, trehalose, and polysorbate 20.

The mode of administration of the combinations of the invention is selected from simultaneous administration, separate formulation and co-administration or separate formulation and sequential administration.

The administration route of the combination of the present invention is selected from oral administration, parenteral administration, transdermal administration, and the parenteral administration includes, but is not limited to, intravenous injection, subcutaneous injection, and intramuscular injection.

The present invention provides the above immunotherapeutic agent selected from a PD-1 antibody or an antigen-binding fragment thereof, in combination with the above MEK inhibitor as a medicament for treating a tumor.

In the present invention, there is provided a method of treatment comprising administering to a patient an immunotherapeutic agent as described above and a MEK inhibitor as described above, wherein the immunotherapeutic agent is selected from a PD-1 antibody or antigen binding fragment thereof.

The invention also provides a pharmaceutical kit, or a pharmaceutical pack, comprising the aforementioned MEK inhibitor and a PD-1 antibody or antigen-binding fragment thereof.

The invention also provides a pharmaceutical composition comprising an effective amount of a PD-1 antibody, or antigen-binding fragment thereof, and a MEK inhibitor, as described previously, in combination with one or more pharmaceutically acceptable excipients, diluents, or carriers.

Drawings

FIG. 1 evaluation of treatment in Each subject

Detailed Description

Term of

In order that the invention may be more readily understood, certain technical and scientific terms are specifically defined below. Unless clearly defined otherwise herein, all other 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 belongs.

The term "humanized antibody", also known as CDR-grafted antibody (CDR), refers to an antibody produced by grafting mouse CDR sequences into a human antibody variable region framework, i.e., a different type of human germline antibody framework sequence. Can overcome the strong antibody variable antibody reaction induced by the chimeric antibody because of carrying a large amount of mouse protein components. Such framework sequences can be obtained from public DNA databases or published references that include germline antibody gene sequences. Germline DNA Sequences of genes such as the human heavy and light chain variable regions can be found in the "VBase" human germline sequence database (available on the Internet www.mrccpe.com.ac.uk/VBase), as well as in Kabat, E.A. et al, 1991Sequences of Proteins of Immunological Interest, 5 th edition. In a preferred embodiment of the invention, the CDR sequences of the mouse humanized antibody of PD-1 are selected from

SEQ ID NO

1, 2, 3, 4, 5, 6.

The term "antigen-binding fragment" refers to Fab fragments, Fab 'fragments, F (ab') 2 fragments, and Fv fragments sFv fragments that bind to human PD-1, having antigen-binding activity; comprising one or more CDR regions of an antibody of the invention selected from SEQ ID NO 1 to SEQ ID NO 6. The Fv fragment contains the variable regions of the antibody heavy and light chains, but lacks the constant region, and has the smallest antibody fragment with the entire antigen-binding site. Generally, Fv antibodies also comprise a polypeptide linker between the VH and VL domains and are capable of forming the structures required for antigen binding. Two antibody variable regions can also be joined together with different linkers into a single polypeptide chain, known as single chain antibodies (scFv) or single chain fv (sFv). The term "binds to PD-1" in the context of the present invention means capable of interacting with human PD-1. The term "antigen binding site" of the present invention refers to a three-dimensional spatial site that is not antigenically contiguous and is recognized by an antibody or antigen binding fragment of the present invention.

The term "immunotherapy" refers to the immunotherapy of diseases by using the immune system, and in the present invention, mainly refers to the method of stimulating and enhancing the anti-tumor immune response of the body by increasing the immunogenicity of tumor cells and the sensitivity to killing by effector cells, and the method of infusing immune cells and effector molecules into the body of a host to cooperate with the immune system of the body to kill tumors and inhibit the growth of tumors.

Detailed Description

The present invention will be further described with reference to the following examples, which are not intended to limit the scope of the present invention.

Example 1: clinical study on compound (1) p-toluenesulfonate in combination with PD-1 antibody in treatment of advanced solid tumor

1. Test antibodies and compounds

The PD-1 antibody was prepared according to the method disclosed in WO2015085847, corresponding to the code H005-1, and the heavy and light chain sequences are shown in seq id NO:7 and SEQ ID NO: 8. batch number: p1512, 200 mg/count, and preparing 20mg/ml for later use.

Compound (1) tosylate (hereinafter Compound A) may be prepared as disclosed in WO2016155473 and formulated into tablets in a format of 0.125 mg/tablet, 0.5 mg/tablet, 2 mg/tablet.

2. Criteria for Subjects in groups

(1) Age 18-70 years;

(2) diagnosis of advanced solid tumors without corresponding standard treatment or treatment failure;

(3) has RAS or RAF positive mutation;

(4) ECOG score 0-1;

(5) life expectancy of at least 3 months;

(6) sufficient bone marrow, liver, kidney, lung and heart function must be present.

3. Method of administration

The group-entry subject firstly receives single administration of the compound A, and the compound A is eluted for 7-10 days after oral administration, and then enters a continuous administration stage, wherein the compound A tablet is taken once a day, and a cycle is formed by 28 continuous days;

cycles

1 and 15 of the 1 st cycle were administered sequentially with H005-1 at a dose of 200 mg/dose. Combination until disease progression, intolerance of toxicity, subject active withdrawal of informed notice, or investigator's judgment that the subject is no longer eligible to continue receiving treatment.

Up to now, 10 subjects with recurrent or metastatic colorectal cancer participated in a dose escalation phase of clinical trials. The subject received compound a (0.125mg/qd, 0.25mg/qd and 0.5mg/qd) in combination with the PD-1 antibody (200 mg). A total of 5 subjects had withdrawn from the study. Disease progression in 2 subjects, treatment-independent SAE in 1 subject, 1 brain metastasis subject (not meeting inclusion criteria), and 1 subject (0.5mg/qd dose group) experienced dose-limiting toxicity. There were still 5 subjects in the study treatment, 2 of which were PR (11.5 months and 7 months, respectively) and 2 additional subjects with stable disease (9 months and 1 month, respectively). 1 subject will receive tumor assessment. The treatment evaluation of each subject is shown in fig. 1.

The gene mutation in each subject was as follows:

genotype of the subject

Sequence listing

<110> Hengrui pharmaceutical Co., Ltd of Jiangsu

Use of <120> PD-1 antibody in combination with a MEK inhibitor for the preparation of a medicament for the treatment of tumors

<160> 10

<170> SIPOSequenceListing 1.0

<210> 1

<211> 5

<212> PRT

<213> mouse source (Mus musculus)

<400> 1

Ser Tyr Met Met Ser

1 5

<210> 2

<211> 17

<212> PRT

<213> mouse source (Mus musculus)

<400> 2

Thr Ile Ser Gly Gly Gly Ala Asn Thr Tyr Tyr Pro Asp Ser Val Lys

1 5 10 15

Gly

<210> 3

<211> 7

<212> PRT

<213> mouse source (Mus musculus)

<400> 3

Gln Leu Tyr Tyr Phe Asp Tyr

1 5

<210> 4

<211> 11

<212> PRT

<213> mouse source (Mus musculus)

<400> 4

Leu Ala Ser Gln Thr Ile Gly Thr Trp Leu Thr

1 5 10

<210> 5

<211> 7

<212> PRT

<213> mouse source (Mus musculus)

<400> 5

Thr Ala Thr Ser Leu Ala Asp

1 5

<210> 6

<211> 9

<212> PRT

<213> mouse source (Mus musculus)

<400> 6

Gln Gln Val Tyr Ser Ile Pro Trp Thr

1 5

<210> 7

<211> 443

<212> PRT

<213> Artificial Sequence (Artificial Sequence)

<220>

<221> PEPTIDE

<222> (1)..(443)

<223> heavy chain sequence

<400> 7

Glu Val Gln Leu Val Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly

1 5 10 15

Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Ser Tyr

20 25 30

Met Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val

35 40 45

Ala Thr Ile Ser Gly Gly Gly Ala Asn Thr Tyr Tyr Pro Asp Ser Val

50 55 60

Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Ser Leu Tyr

65 70 75 80

Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys

85 90 95

Ala Arg Gln Leu Tyr Tyr Phe Asp Tyr Trp Gly Gln Gly Thr Thr Val

100 105 110

Thr Val Ser Ser Ala Ser Thr Lys Gly Pro Ser Val Phe Pro Leu Ala

115 120 125

Pro Cys Ser Arg Ser Thr Ser Glu Ser Thr Ala Ala Leu Gly Cys Leu

130 135 140

Val Lys Asp Tyr Phe Pro Glu Pro Val Thr Val Ser Trp Asn Ser Gly

145 150 155 160

Ala Leu Thr Ser Gly Val His Thr Phe Pro Ala Val Leu Gln Ser Ser

165 170 175

Gly Leu Tyr Ser Leu Ser Ser Val Val Thr Val Pro Ser Ser Ser Leu

180 185 190

Gly Thr Lys Thr Tyr Thr Cys Asn Val Asp His Lys Pro Ser Asn Thr

195 200 205

Lys Val Asp Lys Arg Val Glu Ser Lys Tyr Gly Pro Pro Cys Pro Pro

210 215 220

Cys Pro Ala Pro Glu Phe Leu Gly Gly Pro Ser Val Phe Leu Phe Pro

225 230 235 240

Pro Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro Glu Val Thr

245 250 255

Cys Val Val Val Asp Val Ser Gln Glu Asp Pro Glu Val Gln Phe Asn

260 265 270

Trp Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr Lys Pro Arg

275 280 285

Glu Glu Gln Phe Asn Ser Thr Tyr Arg Val Val Ser Val Leu Thr Val

290 295 300

Leu His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser

305 310 315 320

Asn Lys Gly Leu Pro Ser Ser Ile Glu Lys Thr Ile Ser Lys Ala Lys

325 330 335

Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Pro Ser Gln Glu

340 345 350

Glu Met Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val Lys Gly Phe

355 360 365

Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly Gln Pro Glu

370 375 380

Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly Ser Phe

385 390 395 400

Phe Leu Tyr Ser Arg Leu Thr Val Asp Lys Ser Arg Trp Gln Glu Gly

405 410 415

Asn Val Phe Ser Cys Ser Val Met His Glu Ala Leu His Asn His Tyr

420 425 430

Thr Gln Lys Ser Leu Ser Leu Ser Leu Gly Lys

435 440

<210> 8

<211> 214

<212> PRT

<213> Artificial Sequence (Artificial Sequence)

<220>

<221> PEPTIDE

<222> (1)..(214)

<223> light chain sequence

<400> 8

Asp Ile Gln Met Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly

1 5 10 15

Asp Arg Val Thr Ile Thr Cys Leu Ala Ser Gln Thr Ile Gly Thr Trp

20 25 30

Leu Thr Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys Leu Leu Ile

35 40 45

Tyr Thr Ala Thr Ser Leu Ala Asp Gly Val Pro Ser Arg Phe Ser Gly

50 55 60

Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Ser Leu Gln Pro

65 70 75 80

Glu Asp Phe Ala Thr Tyr Tyr Cys Gln Gln Val Tyr Ser Ile Pro Trp

85 90 95

Thr Phe Gly Gly Gly Thr Lys Val Glu Ile Lys Arg Thr Val Ala Ala

100 105 110

Pro Ser Val Phe Ile Phe Pro Pro Ser Asp Glu Gln Leu Lys Ser Gly

115 120 125

Thr Ala Ser Val Val Cys Leu Leu Asn Asn Phe Tyr Pro Arg Glu Ala

130 135 140

Lys Val Gln Trp Lys Val Asp Asn Ala Leu Gln Ser Gly Asn Ser Gln

145 150 155 160

Glu Ser Val Thr Glu Gln Asp Ser Lys Asp Ser Thr Tyr Ser Leu Ser

165 170 175

Ser Thr Leu Thr Leu Ser Lys Ala Asp Tyr Glu Lys His Lys Val Tyr

180 185 190

Ala Cys Glu Val Thr His Gln Gly Leu Ser Ser Pro Val Thr Lys Ser

195 200 205

Phe Asn Arg Gly Glu Cys

210

<210> 9

<211> 116

<212> PRT

<213> Artificial Sequence (Artificial Sequence)

<220>

<221> PEPTIDE

<222> (1)..(116)

<223> heavy chain variable region

<400> 9

Glu Val Gln Leu Val Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly

1 5 10 15

Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Ser Tyr

20 25 30

Met Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val

35 40 45

Ala Thr Ile Ser Gly Gly Gly Ala Asn Thr Tyr Tyr Pro Asp Ser Val

50 55 60

Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Ser Leu Tyr

65 70 75 80

Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys

85 90 95

Ala Arg Gln Leu Tyr Tyr Phe Asp Tyr Trp Gly Gln Gly Thr Thr Val

100 105 110

Thr Val Ser Ser

115

<210> 10

<211> 107

<212> PRT

<213> Artificial Sequence (Artificial Sequence)

<220>

<221> PEPTIDE

<222> (1)..(10)

<223> light chain variable region

<400> 10

Asp Ile Gln Met Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly

1 5 10 15

Asp Arg Val Thr Ile Thr Cys Leu Ala Ser Gln Thr Ile Gly Thr Trp

20 25 30

Leu Thr Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys Leu Leu Ile

35 40 45

Tyr Thr Ala Thr Ser Leu Ala Asp Gly Val Pro Ser Arg Phe Ser Gly

50 55 60

Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Ser Leu Gln Pro

65 70 75 80

Glu Asp Phe Ala Thr Tyr Tyr Cys Gln Gln Val Tyr Ser Ile Pro Trp

85 90 95

Thr Phe Gly Gly Gly Thr Lys Val Glu Ile Lys

100 105

Claims

1. Use of an immunotherapeutic agent selected from a PD-1 antibody or an antigen-binding fragment thereof, the light chain variable region of which comprises LCDR1, LCDR2 and LCDR3 as shown in SEQ ID No. 4, SEQ ID No. 5 and SEQ ID No. 6, respectively, in combination with a MEK inhibitor in the manufacture of a medicament for the treatment of a tumor; the heavy chain variable region of the PD-1 antibody or antigen-binding fragment thereof comprises HCDR1, HCDR2 and HCDR3 as shown in SEQ ID NO 1, SEQ ID NO 2 and SEQ ID NO 3, respectively; the MEK inhibitor is compound (1) shown below or a pharmaceutically acceptable salt thereof,

the tumor is selected from malignant tumor and benign tumor; wherein the malignant tumor is selected from malignant epithelial tumor, sarcoma, myeloma, leukemia, lymphoma, melanoma, head and neck tumor, brain tumor, peritoneal cancer, mixed tumor, and children malignant tumor.

2. The use of claim 1, wherein the PD-1 antibody or antigen-binding fragment thereof is a PD-1 humanized antibody.

3. The use according to claim 2, wherein the light chain variable region sequence of the PD-1 humanized antibody is the sequence shown in SEQ ID No. 10 or a variant thereof having 0-10 amino acid changes in the light chain variable region; the heavy chain variable region sequence is shown as SEQ ID NO. 9 or its variant, and the variant has amino acid change of 0-10 in the heavy chain variable region.

4. The use according to claim 3, wherein the light chain variable region sequence of the PD-1 humanized antibody is the sequence shown in SEQ ID NO 10 or a variant thereof having an amino acid change of A43S in the light chain variable region; the heavy chain variable region sequence is as shown in SEQ ID NO. 9 or a variant thereof having an amino acid change in the heavy chain variable region of G44R.

5. The use of claim 1, wherein the light chain sequence of the PD-1 humanized antibody is represented by SEQ ID NO. 8 and the heavy chain sequence is represented by SEQ ID NO. 7.

6. Use according to any one of claims 1 to 5, wherein the tumour is selected from Hodgkin's lymphoma, non-Hodgkin's lymphoma, prostate cancer, pancreatic cancer, lung cancer, oesophageal cancer, liver cancer, bile duct cancer, breast cancer, colorectal cancer, gastric cancer, renal cancer, acute myeloid leukaemia, myelodysplastic syndrome, glioma, tumours of unknown primary site.

7. The use according to any one of claims 1 to 5, wherein the tumour is mediated by PD-1 and/or expresses PD-L1.

8. Use according to any one of claims 1 to 5, wherein the tumour is selected from RAS mutant tumours or RAF mutant tumours.

9. The use according to any one of claims 1 to 5, wherein the tumour is a RAS/RAF/MEK pathway dysregulated tumour.

10. The use according to any one of claims 1 to 5, wherein the tumour is a tumour of RAS/RAF pathway abnormality type.

11. The use according to any one of claims 1 to 5, wherein the tumour is selected from a tumour of intermediate or advanced stage or a relapsed refractory tumour.

12. The use according to any one of claims 1 to 5, wherein the tumor is selected from the group consisting of a tumor that has failed treatment with a chemotherapeutic drug and/or has recurred, a tumor that has failed treatment with a radiotherapy and/or has recurred, a tumor that has failed treatment with a targeted drug and/or has recurred, a tumor that has failed treatment with an immunotherapy and/or has recurred.

13. The use according to any one of claims 1 to 5, wherein the dose of the PD-1 antibody or antigen-binding fragment thereof is 1 to 10 mg/kg.

14. The use according to any one of claims 1 to 5, wherein the dose of the PD-1 antibody or antigen-binding fragment thereof is selected from the group consisting of 3mg/kg, 4mg/kg and 5 mg/kg.

15. The use according to any one of claims 1 to 5, wherein the dose of PD-1 antibody or antigen-binding fragment thereof is 50 to 600 mg.

16. The use according to any one of claims 1 to 5, wherein the dose of the PD-1 antibody or antigen-binding fragment thereof is selected from the group consisting of 200mg, 400mg, and 600 mg.

17. The use of any one of claims 1 to 5 wherein the MEK inhibitor dose is 0.01 to 500 mg.

18. The use of any one of claims 1-5, wherein the MEK inhibitor dose is selected from 0.1mg, 0.25mg, 0.5mg, 0.75mg, 1mg, 2mg, 3mg, 4mg, 5mg, 6mg, 7mg, 8mg, 9mg, 10mg, 11mg, 12mg, 12.5mg, 15mg, 17.5mg, 20mg, 22.5mg, 25mg, 30mg, 45mg, 50mg, 60mg, 70mg, 75mg, 80mg, 90mg, 100mg, 125mg, 150mg, 175mg, 200mg, 225mg, 250mg, 275mg, 300mg, 400mg, 500 mg.

19. The use of any one of claims 1 to 5, wherein the MEK inhibitor dose is selected from 0.25mg, 0.5mg, 1mg, 2mg, 3mg, 4mg, 10mg, 15mg, 20mg, 30mg, 45mg, 50mg, 60mg, 75mg, 100 mg.

20. Use according to any one of claims 1 to 5, characterized in that the pharmaceutically acceptable salt of compound (1) is p-toluenesulfonate.

21. A pharmaceutical pack comprising a MEK inhibitor according to any one of claims 1-20 and a PD-1 antibody or antigen-binding fragment thereof.

22. A pharmaceutical composition comprising an effective amount of a PD-1 antibody or antigen-binding fragment thereof according to any one of claims 1-20 and a MEK inhibitor, in combination with one or more pharmaceutically acceptable excipients, diluents or carriers.