Use of anti-PD-1 antibodies in the treatment of nasopharyngeal carcinoma

Abstract

The present invention relates to the use of an anti-PD-1 antibody or its antigen-binding fragment in the manufacture of a medicament for preventing or treating a malignant tumor, and the use of an anti-PD-1 antibody or its antigen-binding fragment in combination with gemcitabine and cisplatin in the manufacture of a medicament for preventing or treating a malignant tumor, preferably nasopharyngeal carcinoma. The present invention also relates to a method for predicting the therapeutic effect of an anti-PD-1 antibody or its antigen-binding fragment in the treatment of nasopharyngeal carcinoma using biomarkers.

Description

[Technical Field]

[0001] The present invention relates to the use of an anti-PD-1 antibody or an antigen-binding fragment thereof in the treatment of malignant tumors. Specifically, the present invention relates to the use of an anti-PD-1 antibody or an antigen-binding fragment thereof in the manufacture of a medicament for treating nasopharyngeal carcinoma, the use of an anti-PD-1 antibody or an antigen-binding fragment thereof in combination with gemcitabine and cisplatin in the manufacture of a medicament for preventing or treating malignant tumors, and a method for predicting the therapeutic effect of an anti-PD-1 antibody or an antigen-binding fragment thereof in the treatment of nasopharyngeal carcinoma using biomarkers. [Background technology]

[0002] Immune escape is one of the hallmarks of cancer. Ahmadzadeh, M. et al., Blood, 114:1537-44, discloses that tumor-specific T lymphocytes are often present in the tumor microenvironment, in draining lymph nodes and peripheral blood, but are usually unable to control tumor progression due to the presence of a network of immunosuppressive mechanisms in the tumor microenvironment. CD8 + Tumor-infiltrating lymphocytes (TILs) typically express activation-induced inhibitory receptors, including CTLA-4 and PD-1, whereas tumor cells often express immunosuppressive ligands, including PD-1 ligand 1 (PD-L1, also known as B7-H1 or CD274), which inhibit T cell activation and effector function. In the inhibitory mechanism, PD-1 and its ligands are important pathways utilized by tumor cells to suppress activated T cells in the tumor microenvironment.

[0003] Programmed death receptor 1 (PD-1) plays an important role in immune regulation and maintaining peripheral tolerance. PD-1 is primarily expressed on activated T cells and B cells and its function is to suppress lymphocyte activation, a normal peripheral tissue tolerance mechanism of the immune system that prevents and treats immune hyperactivity. However, activated T cells infiltrating the tumor microenvironment overexpress PD-1, and inflammatory factors secreted by activated leukocytes induce overexpression of PD-1 ligands, PD-L1 and PD-L2, by tumor cells. This results in persistent activation of the activated T cell PD-1 pathway in the tumor microenvironment, suppressing T cell function and preventing tumor cell killing. Therapeutic anti-PD-1 antibodies block this pathway, partially restoring T cell function and allowing activated T cells to continue killing tumor cells.

[0004] Over the past decade, blockade of the PD-1 / PD-L1 pathway has proven to be an effective route for inducing durable antitumor responses in various cancer indications. Monoclonal antibodies (mAbs) that block the PD / PD-L1 pathway can enhance the activation and effector function of tumor-specific T cells, reduce tumor burden, and improve survival. Between 2014 and 2017, the FDA approved two anti-PD1 monoclonal antibodies (nivolumab and pembrolizumab) and three anti-PD-L1 monoclonal antibodies (atezolizumab, avelumab, and durvalumab) for treating human tumors.

[0005] Nasopharyngeal carcinoma (NPC) is a malignant tumor arising in the apex and lateral walls of the nasopharynx and is the most common malignant tumor of the ear, nose, and throat. According to a World Health Organization study, 80% of NPC cases worldwide are in China. Due to its insidious onset, NPC has a strong tendency to metastasize, with approximately 75% of patients reaching late-stage disease at the time of initial diagnosis, resulting in regional lymph node and / or distant metastasis. While comprehensive treatment primarily consisting of radiation therapy is generally highly effective for early-stage NPC, recurrence or metastasis after treatment carries a poor prognosis, making it a major cause of treatment failure and reduced survival rates. Epstein-Barr virus (EBV) infection is important in the progression of NPC. According to the WHO classification, NPC is classified into three histopathological types: keratinizing (type I), nonkeratinizing (type II), and basal-squamous (type III) cell carcinoma.

[0006] However, some of these commercially available antibodies still have safety issues, such as adverse drug reactions, and therefore there remains a high unmet clinical need for effective therapies to treat malignant tumors (e.g., nasopharyngeal carcinoma). Summary of the Invention

[0007] The present invention provides use of an anti-PD-1 antibody or an antigen-binding fragment thereof in the manufacture of a medicament for preventing or treating malignant tumors, and use of an anti-PD-1 antibody or an antigen-binding fragment thereof in combination with gemcitabine and cisplatin in the manufacture of a medicament for preventing or treating malignant tumors.

[0008] In another aspect, the present invention provides a method for preventing or treating a malignant tumor, comprising administering to an individual in need thereof an effective amount of an anti-PD-1 antibody or antigen-binding fragment thereof described in the present invention, or an anti-PD-1 antibody or antigen-binding fragment thereof in combination with gemcitabine and cisplatin.

[0009] In another aspect, the present invention provides an anti-PD-1 antibody or antigen-binding fragment thereof, or a combination of an anti-PD-1 antibody or antigen-binding fragment thereof with gemcitabine and cisplatin for treating or preventing a malignant tumor. In one or more embodiments, the malignant tumor described in the present invention is nasopharyngeal carcinoma.

[0010] In one or more embodiments, the malignant tumor according to the present invention is recurrent or metastatic nasopharyngeal carcinoma.

[0011] In one or more embodiments, the malignant tumors described herein have PD-L1 expression of >1% in immunohistochemical staining analysis of tumor tissue sections. In a preferred embodiment, the nasopharyngeal carcinomas described herein have PD-L1 expression of >25% in tumor tissue section analysis.

[0012] In one or more embodiments, the malignant tumor according to the present invention is selected from keratinizing nasopharyngeal carcinoma and non-keratinizing nasopharyngeal carcinoma, preferably keratinizing nasopharyngeal carcinoma.

[0013] In one or more embodiments, the nasopharyngeal carcinoma described in the present invention is a nasopharyngeal carcinoma in which genomic amplification of the 11q13 region of chromosome CCND1, FGF14, FGF3, or FGF4 has not been detected in peripheral blood circulating tumor DNA or tumor tissue, or the patient is a nasopharyngeal carcinoma patient in which genomic amplification of the 11q13 region of chromosome CCND1, FGF14, FGF3, or FGF4 has not been detected in peripheral blood circulating tumor DNA or tumor tissue.

[0014] In one or more embodiments, the patient is a patient with nasopharyngeal carcinoma that is incurable after standard systemic treatment or progresses after 6 months of radiochemotherapy.

[0015] In one or more embodiments, the nasopharyngeal carcinoma described in the present invention is a nasopharyngeal carcinoma in which the EBV DNA copy number in peripheral blood on day 28 of treatment is reduced by two-fold or more compared to before administration on day 0.

[0016] In one or more embodiments, the anti-PD-1 antibodies or antigen-binding fragments thereof described herein comprise a light chain complementarity determining region set forth in the amino acid sequences of SEQ ID NOs:1, 2, and 3, and a heavy chain complementarity determining region set forth in the amino acid sequences of SEQ ID NOs:4, 5, and 6.

[0017] In one or more embodiments, the anti-PD-1 antibody or antigen-binding fragment thereof described in the invention comprises a light chain variable region set forth in the amino acid sequence SEQ ID NO:7 and a heavy chain variable region set forth in the amino acid sequence SEQ ID NO:8.

[0018] In one or more embodiments, the anti-PD-1 antibody described in the present invention comprises a light chain having the amino acid sequence set forth in SEQ ID NO:9 and a heavy chain having the amino acid sequence set forth in SEQ ID NO:10.

[0019] In one or more embodiments, the anti-PD-1 antibody described in the present invention is one or more selected from nivolumab, pembrolizumab, toripalimab, sintilimab, camrelizumab, tislelizumab, and cemiplimab, preferably toripalimab.

[0020] In one or more embodiments, the anti-PD-1 antibody described in the present invention is a monoclonal antibody or an antigen-binding fragment thereof.

[0021] In one or more embodiments, the anti-PD-1 antibodies or antigen-binding fragments thereof described in the present invention are administered separately.

[0022] In one or more embodiments, the administration dose of the anti-PD-1 antibody or antigen-binding fragment thereof described in the present invention is selected from about 0.1 mg / kg to about 10.0 mg / kg of individual body weight, for example, about 0.1 mg / kg, about 0.3 mg / kg, about 1 mg / kg, about 2 mg / kg, about 3 mg / kg, about 5 mg / kg, or 10 mg / kg of individual body weight, or a fixed dose of about 120 mg to about 480 mg, for example, a fixed dose of 120 mg, 240 mg, 360 mg, or 480 mg, preferably 3 mg / kg of individual body weight or a fixed dose of 240 mg.

[0023] In one or more embodiments, the administration frequency of the anti-PD-1 antibodies or antigen-binding fragments thereof described in the present invention is about once per week, once per two weeks, once per three weeks, once per four weeks, or once per month, preferably once per two weeks.

[0024] In one or more embodiments, the anti-PD-1 antibodies or antigen-binding fragments thereof described in the present invention are administered at a dose of 1 mg / kg individual body weight, 3 mg / kg individual body weight, 10 mg / kg individual body weight, or a fixed dose of 240 mg or 480 mg, administered once every two or three weeks.

[0025] In one or more embodiments, the anti-PD-1 antibodies or antigen-binding fragments thereof described in the present invention are administered in liquid form, e.g., injection, by a parenteral route, e.g., intravenous infusion.

[0026] In one or more embodiments, the administration cycle of the anti-PD-1 antibodies or antigen-binding fragments thereof described in the present invention is one week, two weeks, three weeks, one month, two months, three months, four months, five months, six months or more, optionally, the duration of each administration cycle is the same or different, and the interval between each administration cycle is the same or different.

[0027] In one or more embodiments, the anti-PD-1 antibodies or antigen-binding fragments thereof described in the present invention are administered in combination with gemcitabine and cisplatin.

[0028] In one or more embodiments, the administration dose of the anti-PD-1 antibody or antigen-binding fragment thereof described in the present invention is selected from about 0.1 mg / kg to about 10.0 mg / kg of individual body weight, for example, about 0.1 mg / kg, about 0.3 mg / kg, about 1 mg / kg, about 2 mg / kg, about 3 mg / kg, about 5 mg / kg, or 10 mg / kg of individual body weight, or a fixed dose of about 120 mg to about 480 mg, for example, a fixed dose of 120 mg, 240 mg, 360 mg, or 480 mg, preferably a fixed dose of 3 mg / kg of individual body weight or 240 mg. The single dose of gemcitabine is about 600 mg / m 2 ~About 1400mg / m 2 Body surface area, e.g., 800 mg / m 2 , 1000 mg / m 2 or 1200 mg / m 2 is the body surface area, The single dose of cisplatin is about 40 mg / m 2 ~about 120mg / m 2 Body surface area, e.g., 60 mg / m 2 , 80 mg / m 2 or 100 mg / m 2 Body surface area.

[0029] In one or more embodiments, the administration frequency of the anti-PD-1 antibody or antigen-binding fragment thereof described in the present invention is about once a week, once every two weeks, once every three weeks, once every four weeks, or once a month, preferably once every three weeks; the administration frequency of the gemcitabine is about once a week, once every two weeks, once every three weeks, twice every three weeks, once every four weeks, or once a month, preferably twice every three weeks; and the administration frequency of the cisplatin is about once a week, once every two weeks, once every three weeks, once every four weeks, or once a month, preferably once every three weeks.

[0030] In one or more embodiments, the anti-PD-1 antibody or antigen-binding fragment thereof described in the present invention is administered at a fixed dose of 240 mg once every three weeks, and the gemcitabine is administered at a single dose of about 1000 mg / m 2body surface area, administered twice every three weeks, and the cisplatin is administered at a single dose of about 80 mg / m 2 body surface area and is administered once every three weeks.

[0031] In one or more embodiments, the anti-PD-1 antibody or antigen-binding fragment thereof, gemcitabine, and cisplatin described in the present invention are administered in liquid dosage forms, e.g., injections, by a parenteral route, e.g., intravenous infusion.

[0032] In one or more embodiments, the administration cycle of the anti-PD-1 antibody or antigen-binding fragment thereof, gemcitabine, and cisplatin described in the present invention is one week, two weeks, three weeks, one month, two months, three months, four months, five months, six months, or more, respectively, and optionally, the duration of each administration cycle is the same or different, and the interval between each administration cycle is the same or different.

[0033] In another aspect, the present invention provides a drug combination comprising an anti-PD-1 antibody or an antigen-binding fragment thereof, gemcitabine, and cisplatin.

[0034] In one or more embodiments, the anti-PD-1 antibody or antigen-binding fragment thereof described herein comprises a light chain complementarity determining region represented by the amino acid sequence of SEQ ID NOs:1, 2, and 3 and a heavy chain complementarity determining region represented by the amino acid sequence of SEQ ID NOs:4, 5, and 6; preferably, the anti-PD-1 antibody or antigen-binding fragment thereof comprises a light chain variable region represented by the amino acid sequence of SEQ ID NO:7 and a heavy chain variable region represented by the amino acid sequence of SEQ ID NO:8; preferably, the anti-PD-1 antibody comprises a light chain represented by the amino acid sequence of SEQ ID NO:9 and a heavy chain represented by the amino acid sequence of SEQ ID NO:10; and more preferably, the anti-PD-1 antibody is toripalimab.

[0035] In another aspect, the present invention provides use of a reagent for detecting gene mutations or amplifications in the 11q13 region of CCND1, FGF14, FGF3 or FGF4 chromosome in circulating tumor DNA in an individual's peripheral blood and / or tumor tissue in the manufacture of a kit for predicting the efficacy of treating nasopharyngeal carcinoma with an anti-PD-1 antibody.

[0036] In another aspect, the present invention provides use of a reagent for detecting the EBV DNA copy number in an individual's peripheral blood in the manufacture of a kit for predicting the efficacy of treating nasopharyngeal carcinoma with an anti-PD-1 antibody.

[0037] In another aspect, the present invention provides a method for predicting the efficacy of treating nasopharyngeal carcinoma with an anti-PD-1 antibody, comprising detecting a genetic mutation or amplification in circulating tumor DNA in the peripheral blood of an individual and / or in the 11q13 region of CCND1, FGF14, FGF3, or FGF4 chromosomes in tumor tissue prior to treatment, wherein the presence of a genetic mutation or amplification in the 11q13 region of CCND1, FGF14, FGF3, or FGF4 chromosomes indicates that the malignant tumor subject is unsuitable for use with an anti-PD-1 antibody.

[0038] In another aspect, the present invention provides a method for predicting the efficacy of treating nasopharyngeal carcinoma with an anti-PD-1 antibody, comprising detecting an individual's peripheral blood EBV DNA copy number on day 28 of treatment, wherein a two-fold or greater decrease in peripheral blood EBV DNA copy number indicates that the tumor patient is suitable for treatment with an anti-PD-1 antibody.

[0039] In another aspect, the present invention provides a detection kit comprising reagents for detecting gene mutations or amplifications in the 11q13 region of CCND1, FGF14, FGF3 or FGF4 chromosome in the peripheral blood and / or tumor tissue of an individual.

[0040] In another aspect, the present invention provides a detection kit comprising reagents for detecting the EBV DNA copy number in the peripheral blood of an individual.

[0041] In another aspect, the present invention provides a detection kit comprising a reagent for detecting a gene mutation or amplification in the 11q13 region of CCND1, FGF14, FGF3 or FGF4 chromosome in an individual's peripheral blood and / or tumor tissue, and a reagent for detecting the EBV DNA copy number in an individual's peripheral blood.

[0042] In another aspect, the present invention provides a kit, the kit comprising: one or more single pharmaceutical dosage units of an anti-PD-1 antibody or antigen-binding fragment thereof described in any of the embodiments herein; or one or more single pharmaceutical dosage units of an anti-PD-1 antibody or antigen-binding fragment thereof, one or more single pharmaceutical dosage units of gemcitabine, and one or more single pharmaceutical dosage units of cisplatin, preferably wherein the anti-PD-1 antibody or antigen-binding fragment thereof is as described in any of the embodiments herein; or One or more single drug dosage units comprise a drug combination according to any of the embodiments herein. [Brief explanation of the drawings]

[0043] [Figure 1] Clinical response was assessed based on RECIST v1.1. 1a: Maximum tumor change compared to baseline was assessed for baseline patients with at least one post-treatment imaging assessment (n=190). Column length represents the maximum decrease or minimum increase in target lesions. 1b: Changes in individual tumor burden over time from baseline were assessed (n=190). [Figure 2] 2a: Progression-free survival (PFS) of patients with nasopharyngeal carcinoma in this study, 2b: Overall survival (OS) of patients with nasopharyngeal carcinoma in this study, 2c: Durable response (DR) of patients with nasopharyngeal carcinoma in this study, 2d: Progression-free survival (PFS) of patients with keratinizing and non-keratinizing nasopharyngeal carcinoma in this study, 2e: Overall survival (OS) of patients with keratinizing and non-keratinizing nasopharyngeal carcinoma in this study. [Figure 3]3a: Relationship between clinical response and tumor PD-L1 expression and TMB. PD-L1 positivity was defined as any intensity of membrane staining >1% in tumor or immune cells using SP142 IHC. TMB was calculated by whole-exon sequencing of somatic mutations within the coding region. 3b: Progression-free survival (PFS) for PD-L1+ vs. PD-L1- patients. 3c: Overall survival (OS) for PD-L1+ vs. PD-L1- patients. 3d: Progression-free survival (PFS) for the 10% of patients with the highest TMB values ​​and the 90% of patients with the lowest TMB values. 3e: Overall survival (OS) for the 10% of patients with the highest TMB values ​​and the 90% of patients with the lowest TMB values. [Figure 4] Gene mutations and frequencies were determined in 174 patients by whole exome sequencing (WES). [Figure 5] 5a: Relationship between plasma EBV DNA copy number and disease stabilization (SD) in patients with nasopharyngeal carcinoma (n=35), 5b: Relationship between plasma EBV DNA copy number and disease complete / partial remission (CR / PR) in patients with nasopharyngeal carcinoma (n=34), 5c: Relationship between plasma EBV DNA copy number and disease progression (PD) in patients with nasopharyngeal carcinoma (n=80). [Figure 6] 6a: Independent review committee-assessed PFS (in patients willing to treat) based on RECIST v1.1; 6b: Investigator-assessed PFS (in patients willing to treat) based on RECIST v1.1; 6c: Treatment effect (progression-free survival) for subgroups. [Figure 7] Overall survival (OS) (treatment-seeking patients). [Figure 8] Duration of response was assessed by an independent review committee according to RECIST v1.1. In the figure, "chemotherapy" refers to the administration of gemcitabine and cisplatin. DETAILED DESCRIPTION OF THE INVENTION

[0044] The present invention relates to a method for treating malignant tumors. The method of the present invention comprises administering an anti-PD-1 antibody or an antigen-binding fragment thereof to a patient in need thereof. The malignant tumor described in the present invention is nasopharyngeal carcinoma. The present invention also relates to a method for predicting the therapeutic effect of an anti-PD-1 antibody in treating a patient with a malignant tumor, particularly nasopharyngeal carcinoma, using a biomarker.

[0045] term In order to make the present invention more readily understood, some technical terms are specifically defined below. Unless expressly stated otherwise elsewhere herein, all technical terms used herein have the meanings commonly understood by one of ordinary skill in the art to which this invention belongs.

[0046] "Administration," "giving," and "treating" refer to the introduction of a composition containing a therapeutic agent into a subject using any of a variety of methods or delivery systems known to those of skill in the art. Anti-PD-1 antibody administration routes include intravenous, intramuscular, subcutaneous, peritoneal, spinal, or other parenteral routes such as injection or infusion. "Parenteral administration" refers to a mode of administration other than enteral or topical administration, usually by injection, and includes, but is not limited to, intravenous, intramuscular, intra-arterial, intrathecal, intralymphatic, intralesional, intracapsular, intraorbital, intracardiac, intradermal, intraperitoneal, transtracheal, subcutaneous, subcuticular, intra-articular, subcapsular, subarachnoid, intraspinal, intradural, and intrasternal injection and infusion, and in vivo electroporation.

[0047] As used herein, an "adverse reaction" (AE) refers to any untoward, usually unintended, or undesired sign, symptom, or disorder associated with the use of a medical treatment. For example, an adverse reaction may be related to immune system activation or immune system cell expansion in response to the treatment. A medical treatment may have one or more associated AEs, and each AE may have the same or different levels of severity.

[0048] "Tumor burden" refers to the total amount of tumor material distributed throughout the body. Tumor burden refers to the total number of cancer cells throughout the body or the total size of a tumor. Tumor burden can be assayed by various methods known in the art, for example, measuring the size of a tumor using calipers after it has been removed from a subject, or using imaging techniques while it is in the body (e.g., ultrasound, bone scan, computed tomography (CT), or magnetic resonance imaging (MRI) scan).

[0049] The term "tumor size" refers to the total size of a tumor, which may be measured as the length and width of the tumor. Tumor size can be assayed by various methods known in the art, for example, by measuring the size of a tumor using calipers after it has been removed from a subject, or by measuring the size of the tumor while it is in the body using imaging techniques (e.g., bone scan, ultrasound, CT, or MRI scan).

[0050] The terms "subject," "individual," and "object" include any living organism, preferably an animal, more preferably a mammal (e.g., rat, mouse, dog, cat, rat, etc.), and most preferably a human. The terms "subject" and "patient" are used interchangeably herein.

[0051] As used herein, the term "antibody" refers to any form of antibody capable of achieving the desired biological activity or binding activity. Therefore, it is used in the broadest sense and is not limited to monoclonal antibodies, polyclonal antibodies, multispecific antibodies, humanized, fully human antibodies, chimeric antibodies, and single-domain antibodies derived from camelids. An "antibody" specifically binds to an antigen and comprises at least two heavy (H) chains and two light (L) chains interconnected by disulfide bonds. Each heavy chain comprises a heavy chain variable region (VH) and a heavy chain constant region, which comprises three constant domains: CH1, CH2, and CH3. Each light chain comprises a light chain variable region (VL) and a light chain constant region, which comprises one constant domain: CL. The VH and VL regions may be further subdivided into hypervariable regions called complementarity-determining regions (CDRs), which are interspersed with more conservative regions called framework regions (FRs). Generally, from N-terminus to C-terminus, both light and heavy chain variable domains comprise FR1, CDR1, FR2, CDR2, FR3, CDR3, and FR4. Amino acids are usually assigned to each domain based on the following definitions: Sequences of Proteins of Immunological Interest, Kabat et al., National Institutes of Health, Bethesda, Md.; 5th ed., NIH Publication No. 91-3242 (1991); Kabat (1978) Adv. Prot. Chem. 32:1-75; Kabat et al., (1977) J. Biol. Chem. 252:6609-6616; Chothia et al., (1987) J. Mol. Biol. 196:901-917; or Chothia et al., (1989) Nature 341:878-883.

[0052] The carboxy-terminal portion of the heavy chain may define a constant region primarily responsible for effector function. Human light chains are typically classified as kappa and lambda chains. Human heavy chains are typically classified as mu, delta, gamma, alpha, or epsilon, and the antibody isotype is defined as IgM, IgD, IgG, IgA, and IgE, respectively. IgG subclasses are well known to those skilled in the art and include, but are not limited to, IgG1, IgG2, IgG3, and IgG4.

[0053] The term "antibody" includes natural and non-natural Abs, monoclonal and polyclonal Abs, chimeric and humanized Abs, human or non-human Abs, fully synthetic Abs, and single-chain Abs. Non-human Abs can be humanized by recombinant methods to reduce immunogenicity in humans.

[0054] Unless otherwise specified, the term "antibody fragment" or "antigen-binding fragment" as used herein refers to an antigen-binding fragment of an antibody, i.e., an antibody fragment that retains the ability of the full-length antibody to specifically bind to an antigen, e.g., a fragment that retains one or more CDR regions. Examples of antigen-binding fragments include, but are not limited to, Fab, Fab', F(ab')2, and Fv fragments, diabodies, linear antibodies, single-chain antibody molecules, nanobodies, and multispecific antibodies formed from antibody fragments.

[0055] "Chimeric antibody" refers to antibodies and fragments thereof in which a portion of the heavy and / or light chain is the same as or homologous to a corresponding sequence in an antibody derived from a particular species (e.g., human) or belonging to a particular antibody class or subclass, while other portions of the chain are the same as or homologous to a corresponding sequence in an antibody derived from another species (e.g., mouse) or belonging to another antibody class or subclass, so long as they exhibit the desired biological activity.

[0056] A "human antibody" refers to an antibody that contains only human immunoglobulin sequences. A human antibody may contain murine carbohydrate chains if produced in a mouse, a mouse cell, or a hybridoma derived from a mouse cell. Similarly, a "mouse antibody" or a "rat antibody" refers to an antibody that contains only mouse or rat immunoglobulin sequences, respectively.

[0057] "Humanized antibody" refers to antibody forms comprising sequences derived from non-human (e.g., murine) and human antibodies. Such antibodies contain minimal sequence derived from non-human immunoglobulin. Typically, a humanized antibody will comprise substantially all of at least one, and usually two, variable domains, in which all or substantially all of the hypervariable loops correspond to those of a non-human immunoglobulin and all or substantially all of the FR regions are those of a human immunoglobulin. The humanized antibody optionally further comprises at least a portion of an immunoglobulin constant region (Fc), usually a human immunoglobulin constant region.

[0058] The term "nasopharyngeal carcinoma" refers to a malignant tumor arising in the nasopharyngeal cavity or nasopharynx. Common clinical symptoms include nasal congestion, blood in the nasal discharge, ear congestion, hearing loss, diplopia, and headache. Epstein-Barr virus (EBV) infection is important for the progression of NPC. According to the WHO classification, there are three histopathological types of nasopharyngeal carcinoma: keratinizing (type I), nonkeratinizing (type II), and basal-squamous (type III). Nonkeratinizing nasopharyngeal carcinoma is closely associated with EBV and is associated with a higher response to radiation therapy and a better overall survival rate.

[0059] The term "immunotherapy" refers to treating a subject having a disease or at risk of infection or recurrence of disease by methods that induce, enhance, suppress, or otherwise modify the immune response. "Treatment" or "therapy" of a subject refers to any type of intervention or process performed on a subject, or administration of an active agent to a subject, with the purpose of reversing, ameliorating, improving, slowing, or preventing the onset, progression, severity, or recurrence of symptoms, complications, or disease, or biochemical markers associated with disease.

[0060] "Programmed death receptor-1 (PD-1)" refers to an immunosuppressive receptor belonging to the CD28 family. PD-1 is primarily expressed on pre-activated T cells in vivo and binds to two ligands, PD-L1 and PD-L2. As used herein, the term "PD-1" includes human PD-1 (hPD-1), hPD-1 variants, isotypes and species homologs, and analogs that share at least one shared epitope with hPD-1.

[0061] A "therapeutically effective amount" or "therapeutically effective dose" of a drug or therapeutic agent refers to any amount of drug that, when used alone or in combination with other therapeutic agents, protects a subject from the onset of disease or promotes regression of disease, as evidenced by a decrease in the severity of disease symptoms, an increase in the frequency and duration of symptom-free periods of disease, or prevention of injury or disability due to pain from the disease. The ability of a therapeutic agent to promote disease regression can be assessed using a variety of methods known to those of skill in the art, for example, assaying the activity of the agent in human subjects during clinical trials, in animal model systems predictive of human efficacy, or in vitro assays.

[0062] A therapeutically effective amount of a drug includes a "prophylactically effective amount," i.e., any amount of a drug that inhibits the onset or recurrence of cancer when administered alone or in combination with an anti-tumor agent to a subject at risk of developing cancer or suffering from the recurrence of cancer.

[0063] "Biotherapeutic agent" refers to a biomolecule, e.g., an antibody or fusion protein, that blocks ligand / receptor signaling in any biological pathway that supports tumor maintenance and / or growth or suppresses anti-tumor immune responses.

[0064] Unless otherwise specified, "CDR" as used herein refers to the complementarity determining regions of an immunoglobulin variable region as defined using the Kabat numbering system.

[0065] A "therapeutic anti-PD-1 monoclonal antibody" refers to a mature form of an antibody that specifically binds to a particular PD-1 expressed on the surface of some mammalian cells. Mature PD-1 lacks the pre-secretion leader sequence, also called the leader peptide. The terms "PD-1" and "mature PD-1" are used interchangeably herein, and unless clearly defined or clearly seen from the context, they should be understood to refer to the same molecule.

[0066] As described herein, a therapeutic anti-human PD-1 antibody or anti-hPD-1 antibody refers to a monoclonal antibody that specifically binds to mature human PD-1.

[0067] As used herein, "framework region" or "FR" refers to an immunoglobulin variable region without the CDR regions.

[0068] An "isolated antibody or antigen-binding fragment thereof" refers to a molecule that is in a purified state, where the designated molecule is substantially free of other biological molecules, e.g., nucleic acids, proteins, lipids, carbohydrates, or other materials (e.g., cellular debris or growth medium).

[0069] "Patient," "subject," or "subject" refers to any single subject requiring a medical procedure or participating in a clinical trial, epidemiological study, or used as a control, and is usually a mammal, including humans and other mammals such as horses, cows, dogs, or cats.

[0070] The "RECIST 1.1 response criteria" described herein refer to the definitions described by Eisenhauver et al., EA et al., Eur. J Cancer 45:228-247 (2009) for target or non-target failure based on the background of the measured response. Prior to immunotherapy, these criteria were the most commonly used to evaluate the therapeutic efficacy of solid tumors. However, with the advent of the immunotherapy era, many challenges not previously encountered in tumor evaluation have emerged. Based on newly emerging phenomena caused by immunotherapy itself, the RECIST Working Group revised the existing "RECIST v.1.1" in 2016 to propose new criteria, i.e., the "irRECIST criteria" described herein, to better evaluate the therapeutic efficacy of immunotherapy drugs.

[0071] The term "ECOG" scoring scale is an index for understanding a patient's general health status and ability to tolerate treatment based on their physical fitness. The ECOG physical fitness status scoring standard scores are 0, 1, 2, 3, 4, and 5. A score of 0 means that the patient's activity capacity is completely normal and there is no difference from the pre-disease activity capacity. A score of 1 indicates the ability to walk around freely and engage in light physical activity, including general housework or office work, but is unable to engage in heavy physical activity.

[0072] "Sustained response" refers to a sustained therapeutic effect following cessation of a therapeutic agent or combination therapy described herein. In some embodiments, the sustained response has a duration that is at least the same as the duration of treatment, or that is at least 1.5, 2.0, 2.5, or 3 times the duration of treatment.

[0073] "Tissue section" refers to a single portion or slice of a tissue sample, such as a thin tissue section cut from a normal tissue or tumor sample.

[0074] "Treatment" of cancer as described herein refers to achieving at least one positive therapeutic effect (e.g., a reduction in the number of cancer cells, a reduction in tumor volume, a reduction in the rate of cancer cell invasion into surrounding organs, or a reduction in the rate of tumor metastasis or tumor growth) in a subject suffering from or diagnosed with cancer using a therapeutic regimen described herein (e.g., administration of an anti-PD-1 antibody). Positive therapeutic effects of cancer can be measured in many ways (see W.A. Weber, J. Nucl. Med., 50:1S-10S (2009)). For example, for tumor growth inhibition, according to NCI standards, a T / C of ≦42% is the minimum level of anti-tumor activity. T / C (%) = median treated tumor volume / median control tumor volume × 100. PFS (also referred to as "time to tumor progression") refers to the length of time during and after treatment during which cancer does not grow, and includes the time a patient experiences CR or PR and the time a patient experiences SD. DFS refers to the length of time a patient remains disease-free during and after treatment. OS refers to an increase in life expectancy compared to an initial or untreated individual or patient. The combination treatment regimen of the present invention that effectively treats cancer patients may vary depending on various factors (e.g., the patient's condition, age, weight, and the ability of the therapy to stimulate the subject's anti-cancer response). Although an embodiment of the present invention may not achieve an effective positive therapeutic effect in all subjects, statistically, it should be effective and achieve a positive therapeutic effect in a significant number of subjects.

[0075] The terms "mode of administration" and "dosing regimen" are used interchangeably and refer to the amount and time of administration of each therapeutic agent in the combination of the present invention.

[0076] The term "immunohistochemistry (IHC)" refers to a method that uses the principle of specific binding between antigens and antibodies to develop coloring agents (fluorescein, enzymes, metal ions, isotopes) marked by antibodies through a chemical reaction to identify antigens (polypeptides and proteins) in tissue cells, thereby conducting localization, qualitative, and relative quantitative studies. In some embodiments of the present invention, PD-L1 detection is performed on tumor tissue samples from subjects before treatment with an anti-PD-1 antibody, using a staining experiment with Roche's anti-human PD-L1 antibody SP142 (Cat No: M4422). In some embodiments, a membrane staining intensity of ≥ 1% of tumor cells is defined as PD-L1 positivity.

[0077] As used herein, the terms "cancer" or "malignant tumor" refer to a wide variety of diseases characterized by the uncontrolled growth of abnormal cells in the body. Unregulated cell division, proliferation, and growth leads to the formation of malignant tumors, which can invade nearby tissues and metastasize to distant parts of the body via the lymphatic system or bloodstream. Cancers suitable for treatment or prevention with the methods, medicaments, and kits of the present invention include, but are not limited to, carcinoma, lymphoma, leukemia, blastoma, and sarcoma. More specific examples of cancer include, but are not limited to, squamous cell carcinoma, myeloma, small cell lung cancer, non-small cell lung cancer, glioma, Hodgkin's lymphoma, non-Hodgkin's lymphoma, acute myeloid leukemia, multiple myeloma, gastrointestinal (tract) cancer, renal cancer, ovarian cancer, liver cancer, lymphoblastic leukemia, lymphocytic leukemia, colorectal cancer, endometrial cancer, kidney cancer, prostate cancer, thyroid cancer, melanoma, chondrosarcoma, neuroblastoma, pancreatic cancer, glioblastoma multiforme, nasopharyngeal cancer, cervical cancer, brain cancer, stomach cancer, bladder cancer, hepatoma, breast cancer, colon cancer, and head and neck cancer.

[0078] As used herein, the term "tumor mutation burden (TMB)" refers to the total number of somatic genetic coding errors, base substitutions, genetic insertions, or deletions detected per million bases. In some embodiments of the present invention, tumor mutation burden (TMB) is estimated by analyzing somatic mutations (coding base substitutions and large base insertions) in a panel of sequences including those studied.

[0079] In the following paragraphs, each aspect of the invention is described in further detail.

[0080] Anti-PD-1 antibody As used herein, "anti-PD-1 antibody" refers to any chemical compound or biological molecule that binds to the PD-1 receptor and blocks the binding of PD-L1 expressed on cancer cells to PD-1 expressed on immune cells (T, B, and NK cells), and preferably also blocks the binding of PD-L2 expressed on cancer cells to PD-1 expressed on immune cells. Alternative nouns or synonyms for PD-1 and its ligands include PDCD1, PD1, CD279, and SLEB2 for PD-1; PDCD1L1, PDL1, B7-H1, B7H1, B7-4, CD274, and B7-H for PD-L1; and PDCD1L2, PDL2, B7-DC, and CD273 for PD-L2. In any of the therapeutic methods, medicaments, and uses of the invention for treating a human individual, the anti-PD-1 antibody blocks the binding of human PD-L1 to human PD-1, and preferably blocks the binding of both human PD-L1 and PD-L2 to human PD1. The human PD-1 amino acid sequence is listed in NCBI locus number NP_005009. The human PD-L1 and PD-L2 amino acid sequences are listed in NCBI locus numbers NP_054862 and NP_079515, respectively.

[0081] As used herein, when referring to an "anti-PD-1 antibody," the term includes antigen-binding fragments thereof, unless otherwise specified or stated.

[0082] Anti-PD-1 antibodies suitable for any of the uses, therapies, medicaments, and kits described herein bind to PD-1 with high specificity and high affinity, blocking the binding of PD-L1 / 2 to PD-1 and transducing PD-1 signals, thereby achieving immunosuppressive effects. In any of the uses, therapies, medicaments, and kits disclosed herein, the anti-PD-1 antibody includes the full-length antibody itself and antigen-binding portions or fragments that bind to the PD-1 receptor and exhibit functional properties similar to those of the full-length Ab in terms of inhibiting ligand binding and upregulating the immune system. In some embodiments, the anti-PD-1 antibody or antigen-binding fragment thereof is an anti-PD-1 antibody or antigen-binding fragment thereof that cross-competes with toripalimab for binding to human PD-1. In other embodiments, the anti-PD-1 antibody or antigen-binding fragment thereof is a chimeric, humanized, or human Ab or antigen-binding fragment thereof. In some embodiments for treating human individuals, the Ab is a humanized Ab.

[0083] In some embodiments, the anti-PD-1 antibody for use in any of the uses, therapies, medicaments, and kits described herein comprises a monoclonal antibody (mAb) or antigen-binding fragment thereof, which specifically binds to PD-1, preferably human PD-1. The mAb may be a human antibody, humanized antibody, or chimeric antibody, and may comprise a human constant region. In some embodiments, the constant region is selected from the group consisting of human IgG1, IgG2, IgG3, and IgG4 constant regions. Preferably, the anti-PD-1 antibody or antigen-binding fragment thereof suitable for any of the uses, therapies, medicaments, and kits described herein comprises a heavy chain constant region of a human IgG1 or IgG4 isotype, more preferably a human IgG4 constant region. In some embodiments, the sequence of the IgG4 heavy chain constant region of the anti-PD-1 antibody or antigen-binding fragment thereof comprises the S228P mutation, which replaces a serine residue in the hinge region with a proline residue typically present at that position in IgG1 isotype antibodies.

[0084] Preferably, in any one embodiment of the uses, therapies, medicaments, and kits described herein, the anti-PD-1 antibody is a monoclonal antibody or antigen-binding fragment thereof, whose light chain CDRs have the amino acid sequences set forth in SEQ ID NOs:1, 2, and 3, and whose heavy chain CDRs have the amino acid sequences set forth in SEQ ID NOs:4, 5, and 6.

[0085] More preferably, in any one embodiment of the uses, therapies, medicaments, and kits described herein, the anti-PD-1 antibody is a monoclonal antibody that specifically binds to human PD-1 and comprises (a) a light chain variable region comprising SEQ ID NO:7, and (b) a heavy chain variable region comprising SEQ ID NO:8.

[0086] More preferably, in any one embodiment of the uses, therapies, medicaments, and kits described herein, the anti-PD-1 antibody is a monoclonal antibody that specifically binds to human PD-1 and comprises (a) a light chain comprising SEQ ID NO:9 and (b) a heavy chain comprising SEQ ID NO:10.

[0087] Table A below provides the amino acid sequence numbers of the light and heavy chain CDRs of exemplary anti-PD-1 antibody mAbs used in the uses, therapies, medicaments and kits described herein:

[0088] [Table A]

[0089] Examples of anti-PD-1 antibodies that bind to human PD-1 and can be used in the uses, therapies, medicaments, and kits described in the present invention are described in WO 2014206107. Human PD-1 mAbs that can be used as the anti-PD-1 antibodies in the uses, therapies, medicaments, and kits described in the present invention include any one of the anti-PD-1 antibodies described in WO 2014206107, including toripalimab (a humanized IgG4 mAb having the structure set forth in WHO Drug Information (Vol. 32, No. 2, pp. 372-373 (2018)) and comprising the light and heavy chain amino acid sequences set forth in SEQ ID NOs: 9 and 10). In a preferred embodiment, the anti-PD-1 antibody that can be used in any one of the uses, therapies, medicaments, and kits described in the present invention is selected from humanized antibodies 38, 39, 41, and 48 described in WO 2014206107. In a particularly preferred embodiment, the anti-PD-1 antibody that can be used in any one of the uses, therapies, medicaments and kits described in the present invention is toripalimab.

[0090] Anti-PD-1 antibodies that can be used in any one of the uses, therapies, medicaments and kits described in the present invention further include FDA-approved Nivolumab and Pembrolizumab.

[0091] In some embodiments, anti-PD-1 antibodies that can be used in any one of the uses, therapies, medicaments, and kits described in the present invention also include anti-PD-L1 monoclonal antibodies that specifically bind to PD-L1 and block the binding of PD-L1 to PD-1, such as nivolumab, pembrolizumab, toripalimab, sintilimab, camrelizumab, tislelizumab, and cemiplimab.

[0092] As used herein, "PD-L1" expression or "PD-L2" expression refers to any detectable expression level of a specific PD-L protein on the cell surface or a specific PD-L mRNA in a cell or tissue. PD-L protein expression can be detected by IHC analysis or flow cytometry of tumor tissue sections using a diagnostic PD-L antibody. Alternatively, PD-L protein expression on tumor cells can be detected by PET imaging using a binding agent that specifically binds to the desired PD-L target (e.g., PD-L1 or PD-L2).

[0093] For methods for quantifying PD-L1 protein expression in IHC analysis of tumor tissue sections, see, but are not limited to, Thompson, RH et al., PNAS 101(49):17174-17179 (2004), Taube, JM et al., Sci Transl Med 4,127ra37 (2012), and Toplian, SL et al., New Eng. J. Med. 366(26):2443-2454 (2012).

[0094] One method uses a simple binary endpoint of positive or negative PD-L1 expression, where a positive result is defined by the percentage of tumor cells showing histological evidence of cell surface membrane staining. A tumor tissue section count of greater than 1% of total tumor cells is defined as positive PD-L1 expression.

[0095] In another method, PD-L1 expression is quantified in tumor tissue sections on tumor cells and infiltrating immune cells. The percentage of tumor cells and infiltrating immune cells showing membrane staining is separately quantified as ≦1%, 1%-50%, and then 50%-100%. For tumor cells, PD-L1 expression is counted as negative if the score is ≦1%, and as positive if the score is >1%.

[0096] In some embodiments, the level of PD-L1 expression by malignant cells and / or infiltrating immune cells within a tumor is assayed as "overexpressed" or "elevated" based on comparison with an appropriately controlled PD-L1 expression level. For example, the PD-L1 controlled protein or mRNA expression level may be the quantified level in a matched section of non-malignant cells or normal tissue of the same type.

[0097] Gemcitabine Gemcitabine is a novel cytidine derivative having the structure shown below:

[0098] [ka]

[0099] Like cytarabine, gemcitabine is activated by deoxycytosine kinase and metabolized by cytidine deaminase after entering the human body. Gemcitabine's main metabolite, difluorodeoxycytidine, is incorporated into intracellular DNA and acts primarily during the G1 / S phase. However, unlike gemcitabine, difluorodeoxycytidine inhibits nucleotide reductase, leading to the reduction of intracellular deoxyribonucleoside triphosphates. Another difference from cytarabine is that gemcitabine can inhibit the reduction of intracellular metabolites by deoxycytosine deaminase, exhibiting self-potentiation.

[0100] In some embodiments of the present invention, gemcitabine may also refer to a composition comprising a therapeutically effective amount of a compound shown in the structural formula above, a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable excipient.

[0101] Cisplatin Cisplatin is a platinum-containing anticancer drug, i.e., cis-diamminedichloroplatinum, which is an orange or yellow crystalline powder that is poorly soluble in water and readily soluble in dimethylformamide. It can be slowly converted to the trans form in aqueous solution and then hydrolyzed. Cisplatin is a compound with the following structure:

[0102] [ka]

[0103] In some embodiments of the present invention, cisplatin may further refer to a composition comprising a therapeutically effective amount of a compound shown in the formula above, a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable excipient.

[0104] Drug combinations The present invention further provides a pharmaceutical combination comprising an anti-PD-1 antibody described herein, gemcitabine, and cisplatin. In this pharmaceutical combination, the anti-PD-1 antibody, gemcitabine, and cisplatin may be provided in the form of a mixture of the three (i.e., in the form of a pharmaceutical composition), or in the form of a mixture of any two and another independent formulation, or each may be provided in the form of an independent formulation. In some embodiments, the pharmaceutical combination contains a three-week dosage regimen comprising one dose of an anti-PD-1 antibody described herein, two doses of gemcitabine, and one dose of cisplatin. When present in the form of independent formulations, each formulation also contains a pharmaceutically acceptable vector in addition to the active ingredients.

[0105] In some embodiments, the anti-PD-1 antibody described in the present invention may be as described in any of the embodiments herein, more preferably an antibody whose light chain CDRs have the amino acid sequences set forth in SEQ ID NOs:1, 2, and 3 and whose heavy chain CDRs have the amino acid sequences set forth in SEQ ID NOs:4, 5, and 6, more preferably a monoclonal antibody comprising a light chain variable region set forth in amino acid sequence SEQ ID NO:7 and a heavy chain variable region set forth in amino acid sequence SEQ ID NO:8, more preferably a monoclonal antibody comprising a light chain set forth in amino acid sequence SEQ ID NO:9 and a heavy chain set forth in amino acid sequence SEQ ID NO:10, more preferably humanized antibodies 38, 39, 41, and 48 described in WO 2014206107, and most preferably toripalimab.

[0106] The drug combinations of the present invention may also include one or more additional therapeutic agents, which may be (for example) chemotherapeutic agents, biotherapeutic agents, immunogenic agents (e.g., attenuated cancer cells, tumor antigens), antigen-presenting cells (e.g., dendritic cells pulsed with tumor-derived antigens or nucleic acids), immunostimulatory cellular factors (e.g., IL-2, IFN-γ, GM-CSF), and cells transfected with a gene encoding an immunostimulatory cellular factor (e.g., but not limited to, GM-CSF).

[0107] Dosage and Administration Regimen The anti-PD-1 antibodies of the present invention may be administered by continuous infusion or interval doses. Single doses may range from about 0.01 to about 20 mg / kg, about 0.1 to about 10 mg / kg of individual body weight, or a fixed dose of about 120 mg to about 480 mg. For example, doses may be about 0.1, about 0.3, about 1, about 2, about 3, about 4, about 5, about 6, about 7, about 8, about 9, or about 10 mg / kg of individual body weight, or a fixed dose of about 120 mg, 240 mg, 360 mg, or 480 mg. Typically, a dosing regimen is designed to achieve this exposure, which results in sustained receptor occupancy (RO) based on the typical pharmacokinetic properties of the Ab. Typical dosing regimens may be administered about once per week, about once every two weeks, about once every three weeks, about once every four weeks, about once per month, or over a longer period. In some embodiments, the individual is administered an anti-PD-1 antibody about once every three weeks, hi some embodiments, the individual is administered an anti-PD-1 antibody about once every two weeks.

[0108] In some embodiments, the anti-PD-1 antibody of the present invention is toripalimab, with a single dose selected from about 1 to about 5 mg / kg of individual body weight. In some embodiments, toripalimab is administered intravenously at a single dose selected from about 1 mg / kg, 2 mg / kg, 3 mg / kg, 4 mg / kg, and 5 mg / kg body weight, or fixed doses of 120 mg, 240 mg, and 360 mg. In some preferred embodiments, toripalimab is administered as a liquid drug, and the selected dose of the drug is administered by intravenous infusion over a 30-60 minute period. In some embodiments, toripalimab is administered at a fixed dose of about 3 mg / kg or about 240 mg once every three weeks (Q3W) over a 30 minute period by intravenous infusion. In some embodiments, toripalimab is administered at a fixed dose of about 3 mg / kg or about 240 mg once every two weeks (Q2W) over a 30 minute period by intravenous infusion.

[0109] The therapeutic agents of the drug combinations of the invention may be administered simultaneously (i.e., in the same drug composition), concurrently (i.e., as separate drug formulations, administered one after the other in any order), or sequentially in any order. It may be particularly useful to use the therapeutic agents in the drug combinations sequentially in different dosage forms (e.g., one drug in a tablet or capsule and another in a sterile liquid formulation) and / or on different administration schedules (e.g., a chemotherapeutic agent administered at least daily and a biotherapeutic agent administered less frequently, e.g., once a week, once every two weeks, or once every three weeks).

[0110] In some embodiments, at least one therapeutic agent in the drug combination is administered at the same dosage regimen (treatment dose, frequency, and duration) that it would normally be administered if the agents were administered as a monotherapy to treat the same tumor. In other embodiments, the patient receives a lower total amount, e.g., a lower dose, a less frequent dose, and / or a shorter duration of treatment, of at least one therapeutic agent in the combination therapy than when the agents are used as a monotherapy.

[0111] Each therapeutic agent in the drug combination of the present invention may be administered orally or parenterally, including using intravenous, intramuscular, intraperitoneal, subcutaneous, rectal, topical and transdermal routes.

[0112] The gemcitabine of the present invention is administered at its approved or recommended dose for continuous treatment until a clinical response is observed or unacceptable toxicity or disease progression occurs. In some embodiments, a single dose of gemcitabine of the present invention is about 600 mg to about 1400 mg / m 2 In some embodiments, a single dose of gemcitabine is about 800 mg / m 2 , 900 mg / m 2 , 1000 mg / m 2 , 1100 mg / m 2 and 1200 mg / m 2The dose is selected from any one of the following: body surface area. Exemplary dosing regimens may be about once per week, once per two weeks, once per three weeks, twice per three weeks, once per four weeks, or once per month. In some embodiments, gemcitabine is administered to an individual twice per three weeks. In some embodiments, gemcitabine is administered on days 1 and 8 of each treatment cycle, respectively. In some embodiments, gemcitabine is administered at a dose of about 1000 mg / m 2 body surface area and is administered twice every three weeks.

[0113] The cisplatin of the present invention is administered at its approved or recommended dose for continuous treatment until the disease maintenance stage is reached or unacceptable toxicity or disease progression occurs. In some embodiments, a single dose of cisplatin of the present invention is about 40 mg to about 120 mg / m 2 In some embodiments, a single dose of cisplatin is selected from the group consisting of about 60 mg / m2 and about 100 mg / m2 body surface area. 2 , 70 mg / m 2 , 80 mg / m 2 , 90 mg / m 2 and 100 mg / m 2 The dose is selected from any one of the following: body surface area. Exemplary dosing regimens can be about once a week, once every two weeks, once every three weeks, once every four weeks, or once a month. In some embodiments, cisplatin is administered to an individual once every three weeks. In some embodiments, cisplatin is administered at a dose of 80 mg / m 2 body surface area and is administered once every three weeks.

[0114] In some embodiments, toripalimab is administered at a fixed dose of about 240 mg Q3W and gemcitabine is administered at a fixed dose of about 1000 mg / m 2 Cisplatin is administered twice every three weeks at a dose of approximately 80 mg / m² of body surface area. 2 body surface area and administered Q3W.

[0115] In some embodiments, gemcitabine may be administered before or after toripalimab use, and cisplatin may be administered before or after toripalimab use, on the same day as toripalimab use.

[0116] The administration cycles of the anti-PD-1 antibody, gemcitabine, and cisplatin of the present invention may be the same or different, and may be one week, two weeks, three weeks, one month, two months, three months, four months, five months, six months, or more. Optionally, the duration of each administration cycle may be the same or different, and the interval between each administration cycle may be the same or different. For example, in some embodiments, toripalimab is administered at a fixed dose of about 240 mg once every three weeks, and gemcitabine is administered at a fixed dose of about 1000 mg / m 2 Cisplatin is administered twice every three weeks at a dose of approximately 80 mg / m² of body surface area. 2 The three drugs are administered once every three weeks on a body surface area basis, and the administration cycle for all three drugs is three weeks.

[0117] Treatment and Use The present invention provides use of the aforementioned anti-PD-1 antibody or antigen-binding fragment thereof of the present invention, optionally together with gemcitabine and cisplatin, in the manufacture of a medicament for preventing or treating malignant tumors.

[0118] The present invention provides a method for preventing or treating malignant tumors, comprising administering to an individual in need thereof an effective amount of an anti-PD-1 antibody or antigen-binding fragment thereof described in the present invention, optionally together with gemcitabine and cisplatin. The effective amount includes a prophylactically effective amount and a therapeutically effective amount. In a preferred embodiment, the administration regimen (including dosage, frequency of administration, order of administration, etc.) of the preventive or therapeutic method is as described in any of the preceding embodiments.

[0119] The present invention provides the aforementioned anti-PD-1 antibody or antigen-binding fragment thereof of the present invention for preventing or treating malignant tumors.The present invention also provides the aforementioned anti-PD-1 antibody or antigen-binding fragment thereof of the present invention, gemcitabine, and cisplatin in combination for preventing or treating malignant tumors.

[0120] The malignant tumor according to the present invention may be as described in any of the preceding embodiments, preferably the malignant tumor according to the present invention is nasopharyngeal carcinoma, preferably the malignant tumor according to the present invention is recurrent or metastatic nasopharyngeal carcinoma.

[0121] Preferably, the methods, uses, anti-PD-1 antibodies and drug combinations according to any of the embodiments of the present invention are particularly suitable for keratinizing and non-keratinizing nasopharyngeal carcinoma, preferably keratinizing nasopharyngeal carcinoma.

[0122] Preferably, the methods, uses, anti-PD-1 antibodies and drug combinations according to any of the embodiments of the present invention are particularly suitable for malignant tumors that are positive for PD-L1 expression in immunohistochemical staining analysis of tumor tissue sections, preferably malignant tumors that are PD-L1 > 25% in immunohistochemical staining analysis of tumor tissue sections.

[0123] Preferably, the methods, uses, anti-PD-1 antibodies and drug combinations according to any of the embodiments of the present invention are particularly suitable for malignant tumors in which gene amplification of CCND1, FGF14, FGF3 or FGF4 in the 11q13 region of chromosome has not been detected in circulating tumor DNA in peripheral blood or tumor tissue.

[0124] Preferably, the methods, uses, anti-PD-1 antibodies and drug combinations according to any of the embodiments of the present invention are particularly suitable for malignant tumors in which the peripheral blood EBV DNA copy number has decreased by more than two-fold (by day 28 of treatment).

[0125] Preferred anti-PD-1 antibodies for use in malignancies may be as described in any of the embodiments herein, more preferably antibodies whose light chain CDRs have the amino acid sequences set forth in SEQ ID NOs:1, 2, and 3 and whose heavy chain CDRs have the amino acid sequences set forth in SEQ ID NOs:4, 5, and 6, more preferably monoclonal antibodies comprising a light chain variable region set forth in amino acid sequence SEQ ID NO:7 and a heavy chain variable region set forth in amino acid sequence SEQ ID NO:8, more preferably monoclonal antibodies comprising a light chain set forth in amino acid sequence SEQ ID NO:9 and a heavy chain set forth in amino acid sequence SEQ ID NO:10, more preferably humanized antibodies 38, 39, 41, and 48 described in WO 2014206107, and most preferably toripalimab.

[0126] In particularly preferred embodiments, the present invention provides methods for preventing or treating nasopharyngeal carcinoma, comprising administering a therapeutically effective amount of toripalimab or a drug combination described herein to a patient with nasopharyngeal carcinoma. Preferably, the patient has positive PD-L1 expression. In some embodiments, the nasopharyngeal carcinoma is preferably keratinizing nasopharyngeal carcinoma. In some embodiments, the nasopharyngeal carcinoma patient is preferably one in which gene amplification of CCND1, FGF14, FGF3, or FGF4 in the 11q13 region of chromosome 11 is not detected in peripheral blood circulating tumor DNA or tumor tissue. In some embodiments, the patient is preferably a nasopharyngeal carcinoma patient in which the peripheral blood EBV DNA copy number has decreased by more than two-fold after 28 days of treatment.

[0127] In a particularly preferred embodiment, the present invention provides use of an anti-PD-1 antibody or antigen-binding fragment thereof, or a pharmaceutical combination described herein, in the manufacture of a medicament for preventing or treating nasopharyngeal carcinoma. Preferably, PD-L1 expression is positive in immunohistochemical staining analysis of tumor tissue sections of the nasopharyngeal carcinoma. In some embodiments, the nasopharyngeal carcinoma is preferably one in which gene amplification of CCND1, FGF14, FGF3, or FGF4 in the 11q13 region of chromosome 11 is not detected in circulating tumor DNA in peripheral blood or in tumor tissue. In some embodiments, the nasopharyngeal carcinoma preferably exhibits a two-fold or greater reduction in peripheral blood EBV DNA copy number after 28 days of treatment.

[0128] The therapeutic agents described in the present invention can constitute pharmaceutical compositions, for example, pharmaceutical compositions containing an anti-PD-1 antibody described herein and / or an anti-cancer agent other than the anti-PD-1 antibody and another pharmaceutically acceptable vector. As described in the present invention, a "pharmaceutically acceptable vector" includes any and all physiologically compatible solvents, dispersion media, coatings, antibacterial and antifungal agents, isotonic and absorption delaying agents, etc. Preferably, vectors suitable for compositions containing anti-PD-1 antibodies are suitable for intravenous, intramuscular, subcutaneous, parenteral, spinal, or epidermal administration, such as by injection or infusion, while vectors used in compositions containing other anti-cancer agents are suitable for parenteral administration, such as oral administration. Pharmaceutical compositions of the present invention may contain one or more pharmaceutically acceptable salts, antioxidants, water, non-aqueous vectors, and / or adjuvants such as preservatives, wetting agents, emulsifiers, and dispersing agents. In a preferred embodiment, the anti-cancer agent other than the anti-PD-1 antibody comprises gemcitabine and cisplatin.

[0129] Dosing regimens are adjusted to provide the optimal desired response, such as maximum therapeutic response and / or minimal adverse effects. For anti-PD-1 antibodies, including those used in combination with other anti-cancer agents, the dose range may be about 0.01 to about 20 mg / kg, about 0.1 to about 10 mg / kg of individual body weight, or a fixed dose of 120 mg, 240 mg, 360 mg, or 480 mg. For example, the dose may be about 0.1, about 0.3, about 1, about 2, about 3, about 5, or about 10 mg / kg of individual body weight. Typically, dosing regimens are designed to achieve such exposure, which results in sustained receptor occupancy (RO) based on the typical pharmacokinetic properties of the Ab. Typical dosing regimens may be administered about once per week, about once every two weeks, about once every three weeks, about once every four weeks, about once per month, or for longer periods. In some embodiments, an individual receives an anti-PD-1 antibody about once every two weeks.

[0130] Method for predicting therapeutic effect of anti-PD-1 antibody on malignant tumors As used herein, the term "gene amplification" refers to a process in which the copy number of a gene encoding a specific protein is selectively increased while the copy number of other genes is not proportionally increased. In natural conditions, gene amplification is achieved by removing the repeated sequences of the gene from the chromosome, followed by extrachromosomal replication in a plasmid, or by transcribing all repeated sequences of ribosomal RNA into RNA transcripts, which are then transcribed to generate additional copies of the original DNA molecule. In some embodiments of the present invention, gene sequencing analysis is disclosed.

[0131] In some embodiments of the invention, the subjects described in the invention have some unique gene amplification, for example, some subjects have gene amplification in the 11q13 region of chromosome CCND1, FGF14, FGF3, or FGF4. In some embodiments of the invention, the presence of gene amplification in the 11q13 region of chromosome CCND1, FGF14, FGF3, or FGF4 suggests that the patient would not benefit from the treatment of the anti-PD-1 antibody described in the invention alone.

[0132] In some embodiments of the present invention, subjects described in the present invention experienced a reduction in peripheral blood EBV DNA copy number in some subjects on day 28 of treatment. In some embodiments of the present invention, a reduction of more than two-fold in peripheral blood EBV DNA copy number suggests a better therapeutic effect of using an anti-PD-1 antibody described in the present invention in the patient.

[0133] Therefore, the present invention provides a method for predicting the efficacy of treating a malignant tumor in an individual with an anti-PD-1 antibody according to the present invention, in particular toripalimab, comprising detecting a biomarker in the patient's peripheral blood before treatment, wherein the biomarker is selected from, but not limited to, CCND1, FGF14, FGF3 or FGF4 chromosomal 11q13 region mutations, or detecting the EBV DNA copy number in the patient's peripheral blood on day 28 of treatment.

[0134] The present invention further includes a method for predicting the efficacy of anti-PD-1 antibody treatment in a tumor patient by detecting the presence or absence of gene amplification in the 11q13 region of chromosome CCND1, FGF14, FGF3, or FGF4. Preferably, the presence of gene amplification in the 11q13 region of chromosome CCND1, FGF14, FGF3, or FGF4 indicates that the tumor patient is not suitable for treatment with an anti-PD-1 antibody alone. Preferably, the tumor patient is selected from nasopharyngeal carcinoma patients.

[0135] The present invention further includes a method for predicting the efficacy of anti-PD-1 antibody treatment in a tumor patient by detecting the EBV DNA copy number in the tumor patient's peripheral blood on day 28 after administration. Preferably, a two-fold or greater decrease in the EBV DNA copy number in the peripheral blood (peripheral blood EBV DNA copy number before treatment on day 0 / peripheral blood EBV DNA copy number on day 28≧2) indicates that the tumor patient is suitable for treatment with an anti-PD-1 antibody. Preferably, the tumor patient is selected from nasopharyngeal carcinoma patients.

[0136] The present invention further includes the use of reagents for detecting gene mutations in the 11q13 region of chromosome CCND1, FGF14, FGF3, or FGF4 in the manufacture of a kit for predicting the therapeutic efficacy of an anti-PD-1 antibody against a malignant tumor. Such reagents include, but are not limited to, reagents commonly used in tests, such as primers, probes, and reagents required for PCR.

[0137] The present invention further includes the use of a reagent for detecting EBV DNA copy number in peripheral blood in the manufacture of a kit for predicting the therapeutic efficacy of anti-PD-1 antibodies against malignant tumors. Such reagents include, but are not limited to, reagents commonly used in tests, such as primers, probes, and reagents required for PCR.

[0138] kit The present invention further provides kits, which comprise one or more single pharmaceutical dosage units of an anti-PD-1 antibody or antigen-binding fragment thereof described in any of the embodiments herein, or which comprise one or more single pharmaceutical dosage units of an anti-PD-1 antibody or antigen-binding fragment thereof described in any of the embodiments herein, one or more single pharmaceutical dosage units of gemcitabine described in any of the embodiments herein, and one or more single pharmaceutical dosage units of cisplatin described in any of the embodiments herein.

[0139] In some embodiments, the kit includes one or more single drug dosage units of a drug combination described in any of the embodiments herein. In some embodiments, the kit includes one or more drug formulations, each of which is a 3-week administration dose of one dose of toripalimab, two doses of gemcitabine, and one dose of cisplatin, preferably a fixed dose of about 240 mg of toripalimab and two doses of gemcitabine, each of which is about 1000 mg / m 2 The dose of cisplatin is sufficient for two doses of body surface area, and the dose is about 80 mg / m 2 A single application per body surface area is sufficient.

[0140] The kit of the present invention can be used to treat malignant tumors, such as those described in any of the embodiments herein, particularly nasopharyngeal carcinoma. Preferably, the amount of anticancer active ingredient contained in the kit of the present invention is sufficient to provide one or more treatment courses (e.g., 2-8 treatment courses) according to the treatment method described herein. When the kit includes the drug combination of the present invention, one treatment course includes at least one administration of toripalimab, two administrations of gemcitabine, and one administration of cisplatin.

[0141] Abbreviation The following abbreviations are used throughout the specification and examples of the present invention: BID One dose twice daily CDR Complementarity Determining Region DFS disease-free survival FR Framework Area IgG immunoglobulin G IHC immunohistochemistry OR overall remission rate ORR Objective remission rate OS overall survival PD disease progression PFS Progression-free survival PR partial response CR complete remission SD disease stabilization DLT dose-limiting toxicity MTD maximum tolerated dose AE Adverse Event Q2W One dose every two weeks QD One dose daily CSD long-term solar radiation type non-CSD non-long-term solar radiation type IRC Independent Review Board TRAE Treatment-related adverse reactions SAE serious adverse reaction RO Receptor Occupancy UC urothelial carcinoma RCC renal cell carcinoma MM metastatic melanoma RECIST Response Evaluation Criteria in Solid Tumors irRECIST Response Evaluation Criteria in Immune-Related Solid Tumors Duration of remission MSI microsatellite instability BICR: Double-blind independent center review

[0142] The present invention is further illustrated by the following examples, which should not be construed as limiting the invention. The contents of all references cited throughout the application are expressly incorporated herein by reference. [Example]

[0143] Example 1: Clinical study on the treatment of nasopharyngeal carcinoma with anti-PD-1 antibody Eligibility criteria: Eligible subjects were (1) aged 18 years or older, (2) with recurrent or metastatic nasopharyngeal carcinoma, (3) incurable after standard systemic treatment or disease progression after 6 months of radiochemotherapy, (4) ECOG score of 0 or 1, (5) normal organ function within 10 days of treatment initiation, (6) no history of autoimmune disease or other malignancies, and (7) no prior anti-PD-1 / anti-PD-L1 immunotherapy.

[0144] Subjects were required to have disease evaluable by RECIST v 1.1 standards, not to have received anti-tumor monoclonal drug therapy within 4 weeks prior to treatment, not to have received any anti-tumor drug therapy within 2 weeks prior to treatment, and not to have received systemic steroid drug therapy within 7 days prior to starting treatment.

[0145] Between December 22, 2016, and February 19, 2019, 279 patients with recurrent or metastatic NPC were screened from 17 centers in mainland China, and 190 patients were eligible for this study. The mean age was 46.4 years, and most patients were male (n=158, 83.2%). Among the two histological subtypes, 182 (95.8%) were non-keratinizing NPC and 8 (4.2%) were keratinizing NPC. 116 patients (61.1%) had received at least two prior systemic treatments. The demographic information of eligible subjects is shown in Table 1.

[0146] [Table 1] Note: *PD-L1 positivity is defined as PD-L1 expression ≥1% on tumor cells stained with SP142 IHC.

[0147] Test drug: anti-PD-1 antibody toripalimab (WO 2014206107).

[0148] Enrolled subjects received 3 mg / kg toripalimab intravenously every two weeks (Q2W) until disease progression, intolerable toxicity, the individual agreed to withdraw, the investigator decided to stop treatment, or 24 months of treatment was confirmed.

[0149] Patients were evaluated every 8 weeks during the first year, and every 12 weeks based on RECIST v1.1 and immune-related Response Evaluation Criteria in Solid Tumors (irRECIST). After discontinuation of the drug, patients were evaluated every 3 months.

[0150] Clinical Design: This was a phase II, single-arm, open-label clinical trial to evaluate the safety and antitumor activity of treating patients with recurrent or metastatic nasopharyngeal carcinoma with an anti-PD-1 antibody.

[0151] 1.1 Safety research: As of February 19, 2020, 12 months after the last patient was admitted, patients had received a mean of 8 doses of toripalimab (range: 1-69 doses). One hundred and eighty-one patients (95.3%) experienced a treatment-emergent adverse event (TEAE), of which 141 (74.2%) experienced a treatment-related adverse event (TRAE). The most common (>5%) TRAEs are shown in Table 2. Sixty-three patients (33.2%) experienced grade 3 or higher TEAEs, while 27 patients (14.2%) experienced treatment-related grade 3 or higher TRAEs. Four patients (2.1%) discontinued the drug due to TRAEs, and seven patients (3.7%) discontinued the dose due to TRAEs. Immune-related adverse reactions (AEs) included hypothyroidism in 45 cases (23.7%), hyperthyroidism in 5 cases (2.6%), liver dysfunction in 3 cases (1.6%), interstitial lung disease in 3 cases (1.6%), dermatomyositis in 1 case (0.5%), and autoimmune myocarditis in 1 case (0.5%).

[0152] [Table 2]

[0153] 1.2 Antitumor activity studies: As of February 19, 2020, 94 (49.5%) of the 190 patients had died, 78 (41.1%) had discontinued treatment, and 18 (9.5%) were still on study medication. The median treatment duration was 3.7 months (range, 0.2-34.8 months). Among the 190 patients, the objective response rate (ORR) assessed by IRCIST / RECIST v1.1 was 20.5% (95% CI: 15.0-27.0), with 5 complete responses, 34 partial responses, and 37 stable disease outcomes. The disease control rate (DCR) was 40.0% (95% CI: 33.0-47.3). According to IRC / irRECIST assessment, the ORR was 20.5% (95% CI: 15.0-27.0) and the DCR was 47.9% (95% CI: 40.6-55.2) (Table 3).

[0154] [Table 3] Note: *ORR=(CR+PR) / total number×100% **DCR=(CR+PR+SD) / total number×100%; CR: complete response, PR: partial response, SD: disease stabilization, PD: disease progression, NE: not evaluated, ORR: objective response rate, DCR: disease control rate, CI: confidence interval.

[0155] Seventy-three subjects (38.4%) had a reduction in target lesions compared to baseline, with 48 subjects (25.3%) having a reduction of 30% or more compared to baseline (Figures 1, 1a and 1b). The median duration of response was 1.8 months (95% CI: 1.8-2.1). The median progression-free survival (mPFS) was 1.9 months (95% CI: 1.8-3.5) (Figures 2, 2a). The median overall survival (mOS) was 17.4 months (95% CI: 11.7-22.9) (Figures 2, 2b). Responses were sustained, with a median DOR of 12.8 months (95% CI: 9.4-NE) (Figures 2, 2c). For patients who had received at least two prior lines of therapy (n=116), the ORR was 21.6% (95% CI 14.5-30.1), the median DOR was 21.5 months, the mPFS was 2.0 months, and the mOS was 15.1 months.

[0156] The median OS for subjects who experienced objective remission (n=39) or disease stabilization (n=38) was not reached because only 7 PR / CR subjects and 11 SD subjects died. The median OS for subjects who experienced disease progression (n=113) was 8.4 months.

[0157] 1.3 Immunogenicity Antidrug antibody (ADA) detection was performed in 190 patients. Seven (3.7%) were ADA-positive, including four consecutive positive samples. There were no significant differences in the incidence of AEs, SAEs, grade ≥3 AEs, drug discontinuation or dose delay, or clinical treatment effect between ADA-positive and ADA-negative patients.

[0158] 1.4 Histological subtypes According to IRC evaluation, the ORR for keratinizing NPC (n=8) was significantly better than that for non-keratinizing NPC (n=182), 62.5% vs. 18.7%, respectively, p=0.01. PFS for keratinizing NPC was also significantly better than that for non-keratinizing NPC, 16.6 months vs. 1.9 months, respectively, HR=0.46 (95% CI: 0.25-0.85), p=0.013. OS was not reached and 15.1 months, respectively, a difference that was not statistically significant, HR=0.51 (95% CI: 0.21-1.24), p=0.14 (Figures 2d and 2e).

[0159] Example 2: Study of the association between biomarkers and clinical treatment effects 2.1 PD-L1 expression in tumors PD-L1 expression status was determined in tumor biopsies by SP142 IHC staining, and a tumor proportion score (TPS) >1% was considered positive. Among 190 patients, 48 ​​(25.3%) were PD-L1 positive, and 134 (70.5%) were PD-L1 negative (Figure 3, 3a). The PD-L1 expression status of 8 (4.2%) patients was unknown. Among PD-L1+ patients, 21 (11.1%) were confirmed to have high PD-L1 expression (>25%). According to histological subtype, the PD-L1+ percentage was significantly higher in keratinizing NPC (75.0%) than in nonkeratinizing NPC (24.1%), p = 0.0047. The ORR values ​​for PD-L1+ patients were higher than those for PD-L1- patients, with ORRs of 27.1% and 19.4%, respectively, but the difference was not statistically significant (p=0.31). In patients with PD-L1>25%, the difference in ORR was more significant (38.1% vs. 19.3%, p=0.08) (Figure 3, 3a). PFS and OS were also better in patients with PD-L1 > 25% than in patients with PD-L1 ≤ 25%, with mPFS of 7.2 months and 1.9 months, respectively, HR = 0.64 (95% CI: 0.40-1.02), p = 0.059, and mOS not reached and 15.1 months, respectively, HR = 0.57 (95% CI: 0.31-1.05), p = 0.071 (Figures 3, 3b and 3c), but the differences were not statistically significant.

[0160] 2.2 Tumor mutation burden (TMB) analysis Whole exome sequencing (WES) was performed on tumor biopsies and paired peripheral blood samples from each patient to identify tumor-specific mutations. Valid WES results were obtained in 174 patients. The TMB values ​​in these NPC patients were very low, with a median TMB value of 0.95 mutations per million base pairs (Muts / Mb). Only one patient had MSI-high TMB values, while four patients had TMB values ​​greater than 10 mutations / Mb, and the remaining patients had TMB values ​​less than 5.8 mutations / Mb. This study evaluated clinical response using cutoff values ​​in the first 10% and 20% of TMB values ​​(2.9 and 2.0 Muts / Mb, respectively). The ORRs for the first 10% and first 20% of patients were 17.6% and 14.3%, respectively (Figures 3a and 3b). Four patients with TMB values ​​greater than 10 Muts / Mb, including one MSI-high patient, showed optimal response with disease progression. Furthermore, the 10% of patients with the highest TMB levels and the 90% of patients with the lowest TMB levels had similar PFS at 1.9 months (Figures 3 and 3d). Conversely, the OS values ​​for high TMB patients were lower than those for low TMB patients, at 9.2 months and 17.4 months, respectively, but the difference was not statistically significant (Figures 3 and 3e). As described above, in this study, TMB was not associated with clinical response in patients with advanced NPC treated with toripalimab monotherapy.

[0161] 2.3 Genomic mutation analysis WES identified the most frequently altered genes (≥10%), including CDKN2A (20%), TP53 (13%), NFKB1A (13%), CDKN2B (11%), ETV6 (11%), and MCL1 (10%) (Figure 4). The association between genomic alterations and clinical treatment efficacy was analyzed. According to the study, 11 patients with genomic amplification of CCND1 (n=11) and / or FGF14, FGF3, and FGF4 in the 11q13 region of chromosome 11 had a 0% ORR after treatment with toripalimab. 19 patients with ETV6 mutations had an ORR of only 5.3%.

[0162] 2.4 Plasma EBV DNA copy number Patient plasma was collected before treatment and analyzed for EBV DNA copy number once every 4 weeks using qRT-PCR. Patients with baseline EBV titers <10,000 IU / mL had a higher ORR than patients with EBV titers ≥10,000 IU / mL, 26.7% and 15.4%, respectively (p=0.088). Dynamic plasma EBV DNA copy number data were collected from 149 patients over the course of treatment (Figure 5). The study found that in patients with objective remission (n=34), plasma EBV DNA copy number decreased 31-fold from baseline to the median lowest copy number, whereas in patients with disease stabilization (n=35), it decreased 3-fold, and remained unchanged in patients with disease progression (n=80) (Figure 5). Furthermore, patients who achieved a ≥2-fold decline in plasma EBV DNA copy count on day 28, i.e., 2 weeks before the first radiological assessment of clinical activity (n=60), had a significantly better clinical response rate than patients who achieved a ≤2-fold decline (n=88), with ORRs of 48.3% and 5.7%, respectively (p=0.0001). In contrast, the 14 patients who experienced disease progression experienced at least a ≥2-fold increase in plasma EBV DNA copy count over a median of 3 months before radiologically confirmed disease progression (Figure 5).

[0163] Example 3: Clinical trial of the treatment of nasopharyngeal carcinoma with anti-PD-1 antibody in combination with gemcitabine-cisplatin (GP) Subject Eligibility Criteria: Eligible subjects must be 18-75 years of age, have recurrent or metastatic nasopharyngeal carcinoma (stage IVB according to the Union for International Cancer Control and American Joint Committee on Cancer classification system, NPC, 8th edition), have not received systemic chemotherapy for recurrent or metastatic disease, have at least one measurable lesion according to RECIST 1.1, have a life expectancy of more than 3 months, have an Eastern Cooperative Oncology Group performance status score of 0 or 1, and have normal organ function. For NPC that recurs after systemic treatment, the interval between the recurrence and the last dose of radiation therapy or chemotherapy must be more than 6 months, and toxicity from any prior treatment must have decreased to grade 0 or grade 1 (CTCAE version 5.0) according to the National Cancer Institute's Common Terminology Criteria.

[0164] Exclusion criteria were: a history of severe hypersensitivity reaction to any monoclonal antibody, gemcitabine, cisplatin, or any component of toripalimab; active or untreated central nervous system metastasis or spinal cord compression; potential risk of major bleeding due to necrotic lesions; uncontrolled pleural or pericardial effusion; uncontrolled ascites and uncontrolled tumor-related pain; uncontrolled or symptomatic hypercalcemia; a prior malignancy other than NPC within the past 5 years that excluded those with an extremely low risk of metastasis or death after intended treatment; treatment with anti-PD-1 / PD-L1 / CTLA4 monoclonal antibodies before treatment; use of anti-tumor traditional herbal medicines within the past 4 weeks; and major surgery within the past 28 days. history of or anticipated major surgery during the study period; history of autoimmune disease; use of systemic immunostimulants within 4 weeks or half-life of the drug prior to treatment; use of systemic corticosteroids or immunosuppressive drugs within 2 weeks prior to treatment; history of bone marrow or solid organ transplant; history of non-infectious pneumonia or current pneumonia; use of any live vaccine within the first 4 weeks; active tuberculosis, active hepatitis B virus or hepatitis C virus infection; active human immunodeficiency virus; active significant neurological or psychiatric disorder; grade 2 or higher peripheral neuropathy; pregnant or lactating women; or clinically significant cardiovascular disease.

[0165] Test drug Anti-PD-1 antibody: toripalimab, Suzhou Junmeng Biomedical Technology Co., Ltd. Gemcitabine (Gem), Cisplatin (Cis) Clinical Design This was a randomized, double-blind, placebo-controlled study. Randomization was performed using the Interactive VoiceWeb Remission System (IVRS). Patients were grouped according to ECOG performance status (0 or 1) and disease stage (local recurrence vs. primary metastasis) before inclusion into groups.

[0166] Subjects were randomly assigned 1:1 to Group A and Group B. Group A received toripalimab in combination with gemcitabine (Gem) and cisplatin (Cis) every three weeks (Q3W). Group B received placebo in combination with gemcitabine and cisplatin every three weeks (Q3W). All drugs were administered intravenously. Patients received toripalimab (240 mg) or placebo on day 1 of each 3-week cycle and gemcitabine (1000 mg / m) on days 1 and 8. 2 body surface area) and received cisplatin (80 mg / m 2 Patients received 100 mg of toripalimab (240 mg) (Group A) or placebo (Group B) every 3 weeks as maintenance treatment until disease progression, intolerable toxicity, noncompliance, withdrawal of consent, or up to 6 cycles, based on whichever occurred first during the chemotherapy phase. Patients received either toripalimab (240 mg) (Group A) or placebo (Group B) every 3 weeks as maintenance treatment until disease progression, intolerable toxicity, withdrawal of consent, or at the investigator's discretion, or for a maximum of 2 years. Crossover trials were not permitted as part of the study.

[0167] endpoint The primary endpoint was progression-free survival (PFS) in the intention-to-treat group, defined as the time from randomization to the first documented disease progression or death from any cause (whichever occurred first). Secondary endpoints included overall survival (OS) in the ITT group, objective response rate (ORR), defined as the proportion of patients with confirmed complete or partial response, duration of response (DoR), defined as the time between first documented response and first evidence of progressive disease, disease control rate (DCR), defined as the proportion of patients with an optimal response of complete response (CR), partial response (PR), or disease stabilization (SD), and 1- and 2-year PFS and OS rates.

[0168] evaluation Baseline tumor assessments included nasopharyngeal, neck, chest, and abdominal CT scans (with oral / intravenous contrast unless contraindicated) or MRI or whole-body positron emission tomography (PET) / CT scans. If clinical signs were present, a bone scan should be performed. All known disease sites were recorded at screening and reassessed at each subsequent tumor assessment. The same radiographic procedures were used throughout the study to assess baseline disease sites. Tumor assessments were performed every 6 weeks for the first 12 months, and then every 9 weeks until disease progression, loss of clinical benefit, consent withdrawal, initiation of new anticancer treatment, death, or investigator withdrawal from the study (as determined by disease progression).

[0169] Clinical response was assessed by RECIST v1.1 and irRECIST, investigators, and a double-blind independent review committee (BICR).

[0170] 3.1 Subject Treatment Between November 10, 2018, and October 20, 2019, 408 subjects were screened from 48 sites in China and Singapore (Figure 6). A total of 289 eligible subjects were randomly assigned to either the toripalimab combination group (Group A, n=146) or the placebo combination group (Group B, n=143). Although the proportions of smokers and drinkers were higher in the toripalimab group than in the placebo group, the baseline demographics and disease characteristics of the two groups were generally well balanced (52.1% vs. 41.3%, P=0.077 and 20.5% vs. 12.6%, P=0.082). However, the difference was not statistically significant. PD-L1 expression staining was positive in 74.7% of the toripalimab group and 76.2% of the placebo group, defined as tumor or immune cell positivity of ≥1%. The demographic data of the subjects included in the groups are shown in Table 4.

[0171] During the chemotherapy phase, all patients received at least one dose of study drug; 56 (19.4%) subjects discontinued the study treatment (31 in the toripalimab group and 25 in the placebo group) by the deadline. Both groups received six cycles of chemotherapy. After chemotherapy was completed, 231 (79.9%) patients continued on maintenance treatment (113 in the toripalimab group and 118 in the placebo group). Subjects received an average of 12 courses of toripalimab and 11 courses of placebo.

[0172] [Table 4]

[0173] 3.2 Progression-free survival In a prespecified interim analysis of 128 patients with disease progression or death, the median treatment duration in the toripalimab and placebo groups was 39 and 36 weeks, respectively, according to BICR assessment using RECIST v1.1. The median PFS in the toripalimab group was 11.7 months (95% CI, 11.0-NE) and 8.0 months (95% CI, 7.0-9.5) in the placebo group. Compared with placebo, PFS was significantly improved in the toripalimab group (hazard ratio for progression or death was 0.52; 95% CI, 0.36-0.74; two-sided P = 0.0003) (Figures 6 and 6a). The estimated 1-year PFS in the toripalimab group was 49.4% (95% CI, 36.4–61.1) compared with 27.9% (95% CI, 18.0–38.8) in the placebo group, a difference of 21.4% (95% CI, 5.1–37.8). PFS was superior for toripalimab compared with placebo in all relevant subgroups, including all PD-L1 subgroups (Figures 6 and 6c). For patients with PD-L1-positive tumor cells (TC) ≥ 1% or immune cells (IC) ≥ 1%, the hazard ratio for progression or death between the toripalimab and placebo groups was 0.59 (95% CI, 0.39–0.89), with a median PFS of 11.4 months versus 8.2 months. For patients with PD-L1 positivity on <1% tumor cells (TC) and <1% immune cells (IC), the hazard ratio for progression or death was 0.35 (95% CI, 0.15-0.81), with a median PFS of 11.0 vs. 6.0 months.

[0174] According to investigator assessment using RECIST v1.1, treatment with toripalimab plus chemotherapy reduced the risk of progression or death by 59% (HR = 0.41, 95% CI, 0.28-0.59, P < 0.0001) compared with placebo plus chemotherapy (Figures 6 and 6b). The 1-year PFS rate was 59.5% in the toripalimab group and 20.0% in the placebo group, a difference of 39.5% (95% CI, 25.6-53.5).

[0175] 3.3 Overall survival At the interim analysis deadline on May 30, 2020, 29 deaths were reported: 12 (8.2%) in the toripalimab group and 17 (11.9%) in the placebo group. Neither group reached median survival. The stratified hazard ratio for OS was 0.78 (95% CI, 0.37-1.64, P=0.50). According to the survival update on February 18, 2021, a total of 64 deaths were reported, with 25 deaths in the toripalimab group and 39 deaths in the placebo group. The stratified hazard ratio for OS was 0.60 (95% CI, 0.36-1.00, P=0.0462), indicating a 40% reduction in the risk of immediate death in the toripalimab group compared with the placebo group (Figure 7). The estimated proportion of patients surviving 2 years was 77.8% (95% CI, 68.0-85.0) in the toripalimab group and 63.3% (95% CI, 49.8-74.1) in the placebo group. Median OS for either group was not mature due to the limited number of OS events.

[0176] 3.4 Tumor response According to the BICR assessment, 28 patients (19.2%) in the toripalimab group and 16 patients (11.2%) in the placebo group achieved a confirmed complete response, and 85 patients (58.2%) in the toripalimab group and 79 patients (55.2%) in the placebo group achieved a confirmed partial response. The ORR in the toripalimab group was 77.4% (95% CI, 69.8-83.9) and 66.4% (95% CI, 58.1-74.1) in the placebo group (P = 0.0335). The DCR in the toripalimab group was 87.7% (95% CI, 81.2-92.5) and 79.7% (95% CI, 72.2-86.0) in the placebo group (P = 0.0650) (Table 5). The investigator-assessed ORR in the toripalimab group was 80.8% (95% CI, 73.5 to 86.9) and 74.8% (95% CI, 66.9 to 81.7) in the placebo group.

[0177] According to the BICR evaluation in the ITT population, 114 patients in the toripalimab group and 95 patients in the placebo group were responders. The median DoR in the toripalimab group was 10.0 (95% CI, 8.8-NE) months and 5.7 (95% CI, 5.4-6.8) months in the placebo group (HR: 0.50, 95% CI, 0.33-0.78) (Table 5). As of May 30, 2020, the ongoing response rates in the toripalimab and placebo groups were 66% (75 / 114) and 43% (41 / 95), respectively (Figure 8). Investigator assessment of the intention-to-treat population revealed that the median DoR was not reached with toripalimab (95% CI, 9.7 to NE) and was 5.8 (95% CI, 5.7 to 6.9) months in the placebo group (HR: 0.37; 95% CI, 0.24 to 0.56).

[0178] [Table 5]

[0179] 3.5 Adverse Reactions As of May 30, 2020, the median treatment duration in the toripalimab group was 38.7 weeks and in the placebo group was 36.0 weeks. The median exposure duration to cisplatin in the toripalimab and placebo groups was 18.3 and 18.4 weeks, respectively, while the median exposure duration to gemcitabine was 19.3 and 19.6 weeks, respectively. All patients experienced at least one treatment-emergent adverse event (TEAE). The incidence of ≥3 TEAEs in the toripalimab and placebo groups was 89.0% and 89.5%, respectively. The rates of discontinuation of toripalimab / placebo due to TEAEs were 7.5% and 4.9%, respectively. The rates of serious adverse events (SAEs) (41.1% vs. 43.4%) and fatal TEAEs (2.7% vs. 2.8%) were similar between the two groups.

[0180] The most common TEAEs included leukopenia (91.1% in the toripalimab group vs. 94.4% in the placebo group), anemia (88.4% vs. 94.4%), neutropenia (85.6% vs. 93.0%), nausea (69.2% vs. 83.2%), vomiting (67.1% vs. 65.7%), thrombocytopenia (63.0% vs. 62.2%), and decreased appetite (53.4% ​​vs. 58.7%).

[0181] The incidence of grade 3 or higher TEAEs in the two groups was similar and included leukopenia (61.6% vs 58.0%), neutropenia (57.5% vs 63.6%), anemia (47.3% vs 39.9%), thrombocytopenia (32.9% vs 28.7%), pneumonia (10.3% vs 3.5%), lymphopenia (8.9% vs 7.0%), hyponatremia (8.9% vs 4.2%), and hypokalemia (6.8% vs 7.0%).

[0182] 3.6 Conclusion This randomized phase III trial compared the therapeutic efficacy and toxicity of toripalimab in combination with GP (gemcitabine and cisplatin) versus placebo chemotherapy for the treatment of recurrent or metastatic NPC. The results showed that compared with the addition of placebo to chemotherapy, the addition of toripalimab to chemotherapy can provide a higher overall remission rate and longer overall survival, with easily controlled safety.

Claims

1. Use of an anti-PD-1 antibody or an antigen-binding fragment thereof in the manufacture of a medicament for preventing or treating a malignant tumor; or 1. Use of a combination of an anti-PD-1 antibody or an antigen-binding fragment thereof with gemcitabine and cisplatin in the manufacture of a medicament for preventing or treating a malignant tumor, comprising: the malignant tumor is a keratinizing nasopharyngeal carcinoma with PD-L1 expression >1% in immunohistochemical staining analysis of tumor tissue sections; The anti-PD-1 antibody is toripalimab. use.

2. The use described in claim 1, characterized in that the malignant tumor is recurrent or metastatic nasopharyngeal carcinoma.

3. The use according to claim 1, characterized in that the malignant tumor is a keratinizing nasopharyngeal carcinoma with PD-L1 > 25% in immunohistochemical staining analysis of tumor tissue sections.

4. The keratinizing nasopharyngeal carcinoma is a keratinizing nasopharyngeal carcinoma in which genomic amplification of the 11q13 region of CCND1, FGF14, FGF3 or FGF4 chromosomes has not been detected in peripheral blood circulating tumor DNA or tumor tissue, or The use according to claim 1, characterized in that the keratinizing nasopharyngeal carcinoma is a keratinizing nasopharyngeal carcinoma in which the EBV DNA copy number in peripheral blood on day 28 of treatment has decreased by more than two-fold compared to before administration on day 0.

5. Use of an anti-PD-1 antibody or antigen-binding fragment thereof in the manufacture of a medicament for preventing or treating a malignant tumor, wherein the anti-PD-1 antibody or antigen-binding fragment thereof is administered alone, and the dose of the anti-PD-1 antibody or antigen-binding fragment thereof is selected from 0.1 mg / kg to 10.0 mg / kg of individual body weight, or a fixed dose of 120 mg to 480 mg, or In the use of a combination of an anti-PD-1 antibody or an antigen-binding fragment thereof with gemcitabine and cisplatin in the manufacture of a medicament for preventing or treating a malignant tumor, the anti-PD-1 antibody or antigen-binding fragment thereof is administered in combination with gemcitabine and cisplatin, wherein the dose of the anti-PD-1 antibody or antigen-binding fragment thereof is selected from 0.1 mg / kg to 10.0 mg / kg of individual body weight, or a fixed dose of 120 mg to 480 mg, and the single dose of gemcitabine is 600 mg / m ２ ~1400 mg / m ２ body surface area, and the single dose of cisplatin is 40 mg / m ２ ~120 mg / m ２ The use according to any one of claims 1 to 4, characterized in that it is body surface area.

6. Use of an anti-PD-1 antibody or an antigen-binding fragment thereof in the manufacture of a medicament for preventing or treating a malignant tumor, wherein the anti-PD-1 antibody or antigen-binding fragment thereof is administered alone, and wherein the dose of the anti-PD-1 antibody or antigen-binding fragment thereof is selected from 0.1 mg / kg, 0.3 mg / kg, 1 mg / kg, 2 mg / kg, 3 mg / kg, 5 mg / kg, or 10 mg / kg of individual body weight, or a fixed dose of 120 mg, 240 mg, 360 mg, or 480 mg; or 6. The use of an anti-PD-1 antibody or antigen-binding fragment thereof in combination with gemcitabine and cisplatin in the manufacture of a medicament for preventing or treating a malignant tumor, wherein the anti-PD-1 antibody or antigen-binding fragment thereof is administered in combination with gemcitabine and cisplatin, wherein the dose of the anti-PD-1 antibody or antigen-binding fragment thereof is selected from 0.1 mg / kg, 0.3 mg / kg, 1 mg / kg, 2 mg / kg, 3 mg / kg, 5 mg / kg, or 10 mg / kg of individual body weight, or a fixed dose of 120 mg, 240 mg, 360 mg, or 480 mg, wherein the single dose of gemcitabine is 800 mg / m 2 , 1000 mg / m 2 , or 1200 mg / m 2 of body surface area, and the single dose of cisplatin is 60 mg / m 2 , 80 mg / m 2 , or 100 mg / m 2 of body surface area.

7. Use of an anti-PD-1 antibody or an antigen-binding fragment thereof in the manufacture of a medicament for preventing or treating a malignant tumor, wherein the anti-PD-1 antibody or antigen-binding fragment thereof is administered alone; wherein the administration frequency of the anti-PD-1 antibody or antigen-binding fragment thereof is once a week, once every two weeks, once every three weeks, once every four weeks, or once a month; or the use of a combination of an anti-PD-1 antibody or antigen-binding fragment thereof with gemcitabine and cisplatin in the manufacture of a medicament for preventing or treating a malignant tumor, wherein the anti-PD-1 antibody or antigen-binding fragment thereof is administered in combination with gemcitabine and cisplatin; The use according to claim 5, wherein the administration frequency of the anti-PD-1 antibody or antigen-binding fragment thereof is once a week, once every two weeks, once every three weeks, once every four weeks, or once a month; the administration frequency of the gemcitabine is once a week, once every two weeks, once every three weeks, twice every three weeks, once every four weeks, or once a month; and the administration frequency of the cisplatin is once a week, once every two weeks, once every three weeks, once every four weeks, or once a month.

8. Use of an anti-PD-1 antibody or an antigen-binding fragment thereof in the manufacture of a medicament for preventing or treating a malignant tumor, wherein the anti-PD-1 antibody or antigen-binding fragment thereof is administered alone, and the administration frequency of the anti-PD-1 antibody or antigen-binding fragment thereof is once every two weeks, or 8. The use of an anti-PD-1 antibody or antigen-binding fragment thereof in combination with gemcitabine and cisplatin in the manufacture of a medicament for preventing or treating a malignant tumor, wherein the anti-PD-1 antibody or antigen-binding fragment thereof is administered in combination with gemcitabine and cisplatin, wherein the administration frequency of the anti-PD-1 antibody or antigen-binding fragment thereof is once every three weeks, the administration frequency of the gemcitabine is twice every three weeks, and the administration frequency of the cisplatin is once every three weeks.

9. In the use of an anti-PD-1 antibody or an antigen-binding fragment thereof in the manufacture of a medicament for preventing or treating a malignant tumor, the anti-PD-1 antibody or antigen-binding fragment thereof is administered at a dose of 1 mg / kg individual body weight, 3 mg / kg individual body weight, or 10 mg / kg individual body weight, or at a fixed dose of 240 mg, or at a fixed dose of 480 mg, at a frequency of once every two or three weeks, or In the use of a combination of an anti-PD-1 antibody or an antigen-binding fragment thereof with gemcitabine and cisplatin in the manufacture of a medicament for preventing or treating a malignant tumor, the anti-PD-1 antibody or antigen-binding fragment thereof is administered at a fixed dose of 240 mg once every three weeks, and the gemcitabine is administered at a single dose of 1000 mg / m ２ body surface area, and is administered twice every three weeks, and the cisplatin is administered at a single dose of 80 mg / m ２ 2. The use according to claim 1, characterized in that the administration frequency is once every three weeks.

10. The use according to claim 1, wherein the anti-PD-1 antibody or antigen-binding fragment thereof, gemcitabine, and cisplatin are administered parenterally in liquid dosage forms.

11. The use of claim 10, wherein the anti-PD-1 antibody or antigen-binding fragment thereof, gemcitabine, and cisplatin are administered by intravenous infusion.

12. The use according to claim 10, wherein the liquid dosage form is an injection.

13. A drug combination for use in a method for preventing or treating keratinizing nasopharyngeal carcinoma in which PD-L1 expression is >1% in immunohistochemical staining analysis of tumor tissue sections, the drug combination comprising an anti-PD-1 antibody or an antigen-binding fragment thereof, gemcitabine, and cisplatin, wherein the anti-PD-1 antibody is toripalimab.

14. A kit for use in a method for preventing or treating keratinizing nasopharyngeal carcinoma in which PD-L1 expression is greater than 1% in immunohistochemical staining analysis of tumor tissue sections, comprising: one or more single-dose units of an anti-PD-1 antibody or antigen-binding fragment thereof, wherein the anti-PD-1 antibody is toripalimab; or one or more single pharmaceutical dosage units of an anti-PD-1 antibody or antigen-binding fragment thereof, one or more single pharmaceutical dosage units of gemcitabine, and one or more single pharmaceutical dosage units of cisplatin, wherein the anti-PD-1 antibody is toripalimab; or 14. A kit comprising one or more single drug dose units of the drug combination of claim 13.

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