Combination therapy of lung cancer using anti-pd-1 antibody and anti-ctl a-4 antibody

Abstract

The present disclosure provides methods for treating a subject having lung cancer, the methods comprising administering to the subject a therapeutically effective amount of: (a) an antibody or antigen-binding portion thereof that specifically binds to the Programmed Death-1 (PD-1) receptor and inhibits PD-1 activity; and (b) an antibody or antigen-binding portion thereof that specifically binds to Cytotoxic T-Lymphocyte Antigen-4 (CTLA-4) and inhibits CTLA-4 activity.

Description

[0001] This application is a divisional application of Chinese patent application No. 201680067708.6 (filed on November 18, 2016, invention title: Treatment of lung cancer using a combination of anti-PD-1 antibody and anti-CTLA-4 antibody). Technical Field

[0002] The present invention relates to a method for treating lung cancer in a subject, comprising administering to the subject a combination of an anti-programmed death-1 (PD-1) antibody and an anti-cytotoxic T-lymphocyte antigen-4 (CTLA-4) antibody. Background Technology Human cancers exhibit numerous genetic and epigenetic alterations, resulting in neoantigens that can potentially be recognized by the immune system (Sjoblom et al. (2006) Science 314:268-74). The adaptive immune system, comprising T and B lymphocytes, possesses potent anti-cancer potential, exhibiting broad capabilities and fine specificity in responding to diverse tumor antigens. Furthermore, the immune system displays considerable plasticity and a memory component. Successful utilization of all these properties of the adaptive immune system will make immunotherapy unique among all cancer treatment modalities.

[0003] PD-1 is a key immune checkpoint receptor expressed by activated T and B cells and mediates immunosuppression. PD-1 is a member of the CD28 receptor family, which includes CD28, CTLA-4, ICOS, PD-1, and BTLA. Two cell surface glycoprotein ligands of PD-1, programmed death-ligand-1 (PD-L1) and programmed death-ligand-2 (PD-L2), have been identified. They are expressed on antigen-presenting cells and in many human cancers, and have been shown to downregulate T cell activation and cytokine secretion upon binding to PD-1.

[0004] Nirumab (formerly known as 5C4, BMS-936558, MDX-1106, or ONO-4538) is a fully human IgG4 (S228P) PD-1 immune checkpoint inhibitor antibody that selectively blocks the interaction with PD-1 ligands (PD-L1 and PD-L2), thereby blocking the downregulation of anti-tumor T cell function (US Patent No. 8,008,449; Wang et al., 2014 Cancer Immunol Res. 2(9):846-56).

[0005] Ipilimumab (YERVOY®) is a fully human IgG1 monoclonal antibody that blocks the binding of CTLA-4 to its B7 ligand, thereby stimulating T cell activation and improving overall survival (OS) in patients with advanced melanoma (Hodi et al. (2010) N Engl J Med 363:711-23). ​​In a phase 1 clinical trial, concomitant therapy with nirushumab and ipilimumab produced rapid and deep tumor regression in a large proportion of patients with advanced melanoma, and was significantly more effective than either antibody alone (Wolchok et al. (2013) N Engl J Med 369(2):122-33; WO 2013 / 173223). However, it remains unknown whether this combination of immunomodulatory antibodies will have similar effects in other tumor types.

[0006] In the United States and worldwide, NSCLC is the leading cause of cancer death (NCCN GUIDELINES®, version 3.2014 – Non-Small Cell Lung Cancer, available at: www.nccn.org / professionals / physician_gls / pdf / nscl.pdf, last accessed May 14, 2014). Patients with stage IV NSCLC who are relatively unresponsive to chemotherapy but have good performance status (PS) benefit from treatment with chemotherapy drugs, including platinum-based agents (e.g., cisplatin, carboplatin), taxanes (e.g., paclitaxel, albumin-bound paclitaxel, docetaxel), vinorelbine, vinblastine, etoposide, pemetrexed, and gemcitabine, and various combinations of these drugs. Summary of the Invention

[0007] This disclosure relates to a method of treating a subject with lung cancer, comprising administering to the subject a combination of: (a) an antibody or antigen-binding portion thereof that specifically binds to the programmed death-1 (PD-1) receptor and inhibits PD-1 activity, wherein the PD-1 antibody or antigen-binding portion thereof is administered at a dose ranging from about 0.1 mg / kg to about 5.0 mg / kg body weight, about once every 2 weeks; and (b) an antibody or antigen-binding portion thereof that specifically binds to cytotoxic T-lymphocyte antigen-4 (CTLA-4) and inhibits CTLA-4 activity, wherein the CTLA-4 antibody or antigen-binding portion thereof is administered at a dose ranging from about 1 mg / kg to about 5.0 mg / kg body weight, about once every 6 weeks or 12 weeks. In some embodiments, the lung cancer is non-small cell lung cancer (NSCLC). In other embodiments, the NSCLC has squamous histology. In still other embodiments, the NSCLC has non-squamous histology.

[0008] In some embodiments, the anti-PD-1 antibody or its antigen-binding portion cross-competes with nilumab for binding to human PD-1. In some embodiments, the anti-PD-1 antibody or its antigen-binding portion is a chimeric, humanized, or human monoclonal antibody or a portion thereof. In other embodiments, the anti-PD-1 antibody or its antigen-binding portion comprises a heavy chain constant region having a human IgG1 or IgG4 isotype. In one embodiment, the anti-PD-1 antibody is nilumab. In one embodiment, the anti-PD-1 antibody is pembrolizumab.

[0009] In some embodiments, the anti-CTLA-4 antibody or its antigen-binding portion is a chimeric, humanized, or human monoclonal antibody or a portion thereof. In some embodiments, the anti-CTLA-4 antibody or its antigen-binding portion comprises a heavy chain constant region having a human IgG1 isotype. In some embodiments, the anti-CTLA-4 antibody is ipilimumab. In other embodiments, the anti-CTLA-4 antibody is trimemumab. In some embodiments, the anti-CTLA-4 antibody or its antigen-binding portion cross-competes with ipilimumab for binding to human CTLA-4.

[0010] In some embodiments, the anti-PD-1 antibody or its antigen-binding portion is administered at a dose of about 1 mg / kg or about 3 mg / kg body weight approximately every 2 weeks. In some embodiments, the anti-CTLA-4 antibody or its antigen-binding portion is administered at a dose of about 1 mg / kg body weight. In other embodiments, the anti-PD-1 antibody or its antigen-binding portion is administered at a dose of about 3 mg / kg body weight approximately every 2 weeks, and the anti-CTLA-4 antibody or its antigen-binding portion is administered at a dose of about 1 mg / kg body weight approximately every 12 weeks. In some embodiments, the anti-PD-1 antibody or its antigen-binding portion is administered at a dose of about 1 mg / kg body weight approximately every 2 weeks, and the anti-CTLA-4 antibody or its antigen-binding portion is administered at a dose of about 1 mg / kg body weight approximately every 6 weeks.

[0011] In some embodiments, subjects treated with the disclosed method demonstrate progression-free survival for at least approximately 1 month, at least approximately 2 months, at least approximately 3 months, at least approximately 4 months, at least approximately 5 months, at least approximately 6 months, at least approximately 7 months, at least approximately 8 months, at least approximately 9 months, at least approximately 10 months, at least approximately 11 months, at least approximately 1 year, at least approximately 18 months, at least approximately 2 years, at least approximately 3 years, at least approximately 4 years, or at least approximately 5 years. In some embodiments, subjects demonstrate progression-free survival for at least approximately 8 months after initial dosing.

[0012] In some embodiments, subjects have longer progression-free survival when treated with an anti-PD-1 antibody or its antigen-binding portion every 2 weeks at a dose of 1 mg / kg body weight and an anti-CTLA-4 antibody or its antigen-binding portion every 6 weeks ("Regimen B"), compared to when subjects are treated with an anti-PD-1 antibody or its antigen-binding portion every 2 weeks at a dose of 1 mg / kg body weight and an anti-CTLA-4 antibody or its antigen-binding portion every 12 weeks ("Regimen A"). In some embodiments, subjects receiving Regimen A have a progression-free survival at least about 1 week, at least about 2 weeks, at least about 3 weeks, at least about 1 month, at least about 2 months, at least about 3 months, at least about 4 months, at least about 5 months, or at least about 6 months longer than subjects receiving Regimen B. In some embodiments, subjects receiving Regimen A have a progression-free survival at least about 3 months longer than subjects receiving Regimen B.

[0013] In some embodiments, subjects have longer progression-free survival when treated with an anti-PD-1 antibody or its antigen-binding portion every 2 weeks at a dose of 3 mg / kg body weight every 2 weeks ("Regimen C") and an anti-CTLA-4 antibody or its antigen-binding portion every 12 weeks ("Regimen A"). In some embodiments, subjects receiving Regimen A have a progression-free survival at least about 1 week, at least about 2 weeks, at least about 3 weeks, at least about 4 weeks, at least about 5 weeks, at least about 6 weeks, at least about 7 weeks, at least about 8 weeks, at least about 9 weeks, at least about 10 weeks, at least about 11 weeks, at least about 12 weeks, at least about 13 weeks, at least about 14 weeks, or at least about 15 weeks longer than subjects receiving Regimen C. In some embodiments, subjects receiving Regimen A have a progression-free survival at least about 3 months longer than subjects receiving Regimen C.

[0014] In some embodiments, the subject has a lung tumor expressing ≥ 1% PD-L1, ≥ 5% PD-L1, ≥ 10% PD-L1, ≥ 25% PD-L1, or ≥ 50% PD-L1. In some embodiments, the combination is administered as long as clinical benefit is observed or until disease progression or uncontrollable toxicity occurs. In one embodiment, anti-PD-1 and anti-CTLA-4 antibodies are formulated for intravenous administration. In some embodiments, an anti-PD-1 antibody or its antigen-binding portion and an anti-CTLA-4 antibody or its antigen-binding portion are administered sequentially to the subject. In some embodiments, anti-PD-1 and anti-CTLA-4 antibodies are administered within 30 minutes of each other. In one embodiment, an anti-PD-1 antibody or its antigen-binding portion is administered prior to an anti-CTLA-4 antibody or its antigen-binding portion. In another embodiment, an anti-CTLA-4 antibody or its antigen-binding portion is administered prior to an anti-PD-1 antibody or its antigen-binding portion. In some embodiments, the anti-PD-1 antibody or its antigen-binding portion and the anti-CTLA-4 antibody or its antigen-binding portion are administered simultaneously in separate compositions. In some embodiments, the anti-PD-1 antibody or its antigen-binding portion and the anti-CTLA-4 antibody or its antigen-binding portion are administered simultaneously as a single composition.

[0015] In one embodiment, the anti-PD-1 antibody or its antigen-binding portion is administered at a subtherapeutic dose. In some embodiments, the anti-CTLA-4 antibody or its antigen-binding portion is administered at a subtherapeutic dose. In some embodiments, both the anti-PD-1 antibody or its antigen-binding portion and the anti-CTLA-4 antibody or its antigen-binding portion are administered at subtherapeutic doses.

[0016] This disclosure also relates to a kit for treating a subject with lung cancer, the kit comprising: (a) an anti-PD-1 antibody or its antigen-binding portion thereof in an amount ranging from about 4 mg to about 500 mg; (b) a CTLA-4 antibody or its antigen-binding portion thereof in an amount ranging from about 40 mg to about 500 mg; and (c) instructions for using the PD-1 antibody or its antigen-binding portion and the CTLA-4 antibody or its antigen-binding portion in any of the disclosed methods.

[0017] Other features and advantages of the invention will become apparent from the following detailed description and embodiments, which should not be considered limiting. All cited references, including scientific papers, newspaper reports, GenBank accessions, patents, and patent applications cited in their entirety herein, are expressly incorporated herein by reference. Attached Figure Description

[0018] Figure 1 This shows the dosing regimens for nilusumab and ipilimumab.

[0019] Figures 2A-2D Show the percentage change from baseline for the target lesion in each of the following treatment groups: Nirutumab 1 mg / kg + Ipilimumab 1 mg / kg Q3W ( Figure 2A Nirumumab 1 mg / kg Q2W + Ipilimumab 1 mg / kg Q6W Figure 2B Nirumab 3 mg / kg Q2W + Ipilimumab 1 mg / kg Q12W Figure 2C ); and nilumumab 3 mg / kg Q2W + ipilimumab 1 mg / kg Q6W ( Figure 2D ).

[0020] Figure 3 This shows the optimal percentage change in tumor burden at target injury relative to baseline tumor PD-L1 expression.

[0021] Figure 4 This displays the response duration of nivo + ipi in first-line NSCLC.

[0022] Figure 5 The efficacy of nivo+ipi was demonstrated at all tumor PD-L1 expression levels.

[0023] Figure 6A and 6B Showing through smoking ( Figure 6A ) and EGFR mutations ( Figure 6B In the first-line NSCLC, the effects of nivo +ipi are displayed.

[0024] Figures 7A-7D This shows a case with pathological CR. Invention Details This invention relates to a method for treating lung cancer patients, comprising administering a combination of an anti-PD-1 antibody and another anticancer agent to the patient. In some embodiments, the other anticancer agent is an anti-CTLA-4 antibody.

[0025] the term To facilitate a clearer understanding of this invention, certain terms are first defined. As used herein, unless otherwise expressly provided, each of the following terms shall have the following meaning. Further definitions are set forth in this application.

[0026] "Administration" means the physical introduction of a composition comprising a therapeutic agent into a subject using any of a variety of methods and delivery systems known to those skilled in the art. Routes of administration for anti-PD-1 antibodies include intravenous, intramuscular, subcutaneous, intraperitoneal, spinal, or other parenteral administration routes, such as by injection or infusion. As used herein, the phrase "parenteral administration" refers to administration modalities other than enteral and local administration, typically performed by injection, and includes, but is not limited to, intravenous, intramuscular, intra-articular, intrathecal, intralymphatic, intralesional, intracystic, intraorbital, intracardiac, intradermal, intraperitoneal, tracheal, subcutaneous, subepidermal, intra-articular, subcystic, subarachnoid, spinal, epidural, and intrasternal injections and infusions, as well as in vivo electroporation. In some embodiments, the composition is administered via a non-parenteral route; in other embodiments, it is administered orally. Other non-parenteral routes include local, epidermal, or mucosal administration routes, such as intranasal, vaginal, rectal, sublingual, or local administration. It can also be applied, for example, once, multiple times, and / or over one or more extended time periods.

[0027] As used herein, an “adverse event” (AE) is any adverse and generally unintentional or undesirable indication (including abnormal laboratory findings), symptom, or illness associated with the application of a medical treatment. For example, an adverse event may be associated with activation of the immune system in response to treatment or expansion of immune system cells (e.g., T cells). Medical treatments may have one or more associated AEs, and each AE may have the same or different levels of severity. References to methods capable of “modifying adverse events” refer to treatment regimens that reduce the incidence and / or severity of one or more AEs associated with the application of different treatment regimens.

[0028] An "antibody" (Ab) should include, but is not limited to, glycoprotein immunoglobulins that specifically bind to an antigen and contain at least two heavy (H) chains and two light (L) chains linked by disulfide bonds, or their antigen-binding portions. Each H chain contains a heavy chain variable region (abbreviated as V in this document). H The heavy chain constant region contains three constant structural domains C. H1 C H2 and C H3 Each light chain contains a light chain variable region (abbreviated as V in this article). L ) and the light chain constant region. The light chain constant region contains a constant structural domain C L V H and V L The region can be further subdivided into highly variable regions known as complementarity-determining regions (CDRs), which are interspersed within more conservative regions known as framing regions (FRs). Each V H and V LIt contains three CDRs and four FRs, arranged in the following order from the amino terminus to the carboxyl terminus: FR1, CDR1, FR2, CDR2, FR3, CDR3, FR4. The variable regions of the heavy and light chains contain binding domains that interact with the antigen. The constant regions of the antibody can mediate the binding of the immunoglobulin to host tissues or factors, including various cells of the immune system (e.g., effector cells) and the first component (C1q) of the classical complement system.

[0029] Immunoglobulins can be derived from any of the commonly known isotypes, including but not limited to IgA, secretory IgA, IgG, and IgM. IgG subclasses are also well known to those skilled in the art and include, but are not limited to, human IgG1, IgG2, IgG3, and IgG4. "Isotype" refers to an antibody class or subclass (e.g., IgM or IgG1) encoded by a heavy chain constant region gene. The term "antibody" includes, for example, naturally occurring and non-naturally occurring antibodies; monoclonal and polyclonal antibodies; chimeric and humanized antibodies; human or non-human antibodies; fully synthetic antibodies; and single-chain antibodies. Non-human antibodies can be humanized by recombinant methods to reduce their immunogenicity in humans. Unless explicitly stated otherwise, and unless the context otherwise indicates, the term "antibody" also includes antigen-binding fragments or antigen-binding portions of any of the aforementioned immunoglobulins, and includes monovalent and bivalent fragments or portions, and single-chain antibodies.

[0030] "Isolated antibody" refers to an antibody that substantially does not contain other antibodies with different antigen specificities (e.g., isolated antibodies that specifically bind to PD-1 substantially do not contain antibodies that specifically bind to antigens other than PD-1). However, isolated antibodies that specifically bind to PD-1 may have cross-reactivity with other antigens, such as PD-1 molecules from different species. Furthermore, isolated antibodies may substantially not contain other cellular material and / or chemicals.

[0031] The term "monoclonal antibody" ("mAb") refers to a non-natural preparation of an antibody molecule consisting of a single molecule (i.e., an antibody molecule whose basic sequence is substantially identical and which exhibits single binding specificity and affinity for a specific epitope). mAb is an example of an isolated antibody. Monoclonal antibodies can be produced using hybridoma techniques, recombinant techniques, transgenic techniques, or other techniques known to those skilled in the art.

[0032] A "human" antibody (HuMAb) indicates an antibody having a variable region, wherein both the frame region and the CDR region are derived from human germline immunoglobulin sequences. Furthermore, if the antibody contains a constant region, then that constant region is also derived from a human germline immunoglobulin sequence. The human antibodies of this invention may include amino acid residues not encoded by human germline immunoglobulin sequences (e.g., mutations introduced through random or site-specific in vitro mutagenesis or in vivo somatic mutation). However, the term "human antibody" as used herein is not intended to include antibodies in which a germline-derived CDR sequence from another mammalian species (such as a mouse) has been grafted onto a human frame sequence. The terms "human" antibody and "fully human" antibody are used synonymously.

[0033] "Humanized antibody" refers to an antibody in which some, most, or all of the amino acids outside the CDR domain of a non-human antibody are replaced by corresponding amino acids derived from human immunoglobulins. In one embodiment of the humanized form of the antibody, some, most, or all of the amino acids outside the CDR domain have been replaced by amino acids from human immunoglobulins, while some, most, or all of the amino acids within one or more CDR regions remain unchanged. Minor additions, deletions, insertions, substitutions, or modifications of amino acids are permitted, as long as they do not abolish the antibody's ability to bind a specific antigen. "Humanized" antibodies retain antigen specificity similar to that of the original antibody.

[0034] "Chimeric antibody" refers to an antibody in which the variable region is derived from one species and the constant region is derived from another species, such as an antibody in which the variable region is derived from a mouse antibody and the constant region is derived from a human antibody.

[0035] "Anti-antigen" antibodies are antibodies that specifically bind to antigens. For example, anti-PD-1 antibodies specifically bind to PD-1, and anti-CTLA-4 antibodies specifically bind to CTLA-4.

[0036] An antibody's "antigen-binding portion" (also known as an "antigen-binding fragment") refers to one or more fragments of an antibody that retain the ability to specifically bind to antigens bound by the intact antibody.

[0037] "Cancer" refers to a broad group of diseases characterized by the uncontrolled growth of abnormal cells in the body. Disordered cell division and growth lead to the formation of malignant tumors, which can invade adjacent tissues and can also metastasize to distant parts of the body via the lymphatic system or bloodstream.

[0038] “Cytotoxic T-lymphocyte antigen-4” (CTLA-4) represents an immunosuppressive receptor belonging to the CD28 family. CTLA-4 is exclusively expressed on T cells in vivo and binds to two ligands, CD80 and CD86 (also known as B7-1 and B7-2, respectively). As used herein, the term “CTLA-4” includes human CTLA-4 (hCTLA-4), variants of hCTLA-4, isotypes and species homologs of hCTLA-4, and analogs that share at least one common epitope with hCTLA-4. The complete hCTLA-4 sequence can be found in GenBank under accession number AAB59385.

[0039] The term "immunotherapy" refers to the treatment of a subject who has a disease or is at risk of infection or disease recurrence by means of methods including inducing, enhancing, suppressing, or otherwise altering an immune response. "Treatment" or "therapy" for a subject refers to any type of intervention or procedure performed on a subject, or the administration of an active agent to a subject with the aim of reversing, alleviating, improving, suppressing, slowing, or preventing the onset, progression, development, severity, or recurrence of symptoms, complications, or conditions, or disease-related biochemical markers.

[0040] The term "PD-L1 positive" as used herein may be used interchangeably with "at least about 1% PD-L1 expression". In one embodiment, PD-L1 expression may be used by any method known in the art. In another embodiment, PD-L1 expression is measured by automated IHC. PD-L1 positive tumors may thus have at least about 1%, at least about 2%, at least about 5%, at least about 10%, at least about 20%, at least about 25%, at least about 30%, at least about 40%, at least about 50%, at least about 60%, at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, or about 100% of tumor cells expressing PD-L1, as determined by automated IHC. In some embodiments, "PD-L1 positive" means the presence of at least 100 cells expressing PD-L1 on their cell surface.

[0041] “Programmed death-1 (PD-1)” refers to an immunosuppressive receptor belonging to the CD28 family. PD-1 is primarily expressed on previously 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), variants, isotypes, and species homologs of hPD-1, as well as analogs sharing at least one common epitope with hPD-1. The complete hPD-1 sequence can be found in GenBank under GenBank accession number U64863.

[0042] Programmed death-ligand-1 (PD-L1) is one of two cell surface glycoprotein ligands (the other being PD-L2) that target PD-1, and it downregulates T cell activation and cytokine secretion upon binding to PD-1. As used in this article, the term "PD-L1" includes human PD-L1 (hPD-L1), variants, isotypes, and species homologs of hPD-L1, as well as analogs that share at least one epitope with hPD-L1. The complete hPD-L1 sequence can be found in GenBank under accession number Q9NZQ7.

[0043] "Subject" includes any human or non-human animal. The term "non-human animal" includes, but is not limited to, vertebrates such as non-human primates, sheep, dogs, and rodents such as mice, rats, and guinea pigs. In some embodiments, the subject is a human. The terms "subject" and "patient" are used interchangeably herein.

[0044] A “therapeuticly effective amount” or “therapeutic dose” of a drug or therapeutic agent is any amount of the drug, when used alone or in combination with another therapeutic agent, that protects a subject from the onset of disease or promotes disease remission, demonstrated by a reduction in the severity of disease symptoms, an increase in the frequency and duration of asymptomatic phases, or prevention of damage or disability caused by disease suffering. The ability of a therapeutic agent to promote disease remission can be evaluated using a variety of methods known to skilled practitioners, such as in human subjects during clinical trials, in animal model systems predicting efficacy for humans, or by measuring the activity of the drug in in vitro assays.

[0045] As used in this article, “subtherapeutic dose” refers to a dose of a therapeutic compound (e.g., an antibody) that is below the usual or typical dose of the therapeutic compound when administered alone for the treatment of hyperproliferative diseases (e.g., cancer).

[0046] As an example, "anticancer drugs" promote cancer regression or prevent further tumor growth in subjects. In some implementations, a therapeutically effective amount of the drug promotes cancer regression to the point of cancer elimination. "Promoting cancer regression" means that, alone or in combination with an antitumor agent, an effective amount of the drug results in a reduction in tumor growth or size, tumor necrosis, a decrease in the severity of at least one disease symptom, an increase in the frequency and duration of asymptomatic phases, or prevention of damage or disability caused by disease suffering. Furthermore, the terms "effective" and "efficacy" in relation to treatment include both pharmacological efficacy and physiological safety. Pharmacological efficacy refers to the ability of a drug to promote cancer regression in a patient. Physiological safety refers to the level of toxicity or other adverse physiological effects (adverse effects) at the cellular, organ, and / or organismal level caused by drug administration.

[0047] As an example of cancer treatment, a therapeutically effective dose of an anticancer drug can inhibit cell or tumor growth by at least about 20%, at least about 40%, at least about 60%, or at least about 80% relative to an untreated subject. In other embodiments of the invention, tumor regression can be observed and persist for at least about 20 days, at least about 40 days, or at least about 60 days. Despite these ultimate measures of treatment effectiveness, the evaluation of immunotherapeutic drugs must also consider “immune-related” response patterns.

[0048] The “immune-related” response pattern refers to a clinical response pattern frequently observed in cancer patients treated with immunotherapies that exert their antitumor effects by inducing cancer-specific immune responses or by altering innate immune processes. This response pattern is characterized by a beneficial therapeutic effect following an initial increase in tumor burden or the appearance of new lesions, which would be classified as disease progression and synonymous with drug failure in the evaluation of conventional chemotherapy. Therefore, appropriate evaluation of immunotherapies may require long-term monitoring of the effects of these agents on the target disease.

[0049] Therapeutic effective amounts of a drug include “preventative effective amounts,” which are any amounts of drug that, when administered alone or in combination with an antitumor agent to a subject at risk of developing cancer (e.g., a subject with a pre-existing condition that has worsened) or a subject at risk of cancer recurrence, inhibit the occurrence or recurrence of cancer. In some embodiments, preventative effective amounts completely prevent the occurrence or recurrence of cancer. “Inhibiting” the occurrence or recurrence of cancer means reducing the likelihood of cancer occurring or recurring, or completely preventing the occurrence or recurrence of cancer.

[0050] The use of alternatives (e.g., "or") should be understood to mean any one, two, or any combination of the alternatives. The indefinite articles "a" or "an" as used herein should be understood to mean "one or more / a combination of" any enumerated or listed components.

[0051] The terms “about” or “substantially comprise” mean a value or composition within an acceptable margin of error for a particular value or composition as determined by one of ordinary skill in the art, which will depend in part on how the value or composition is measured or determined, i.e., the limitations of the measuring system. For example, “about” or “substantially comprise” can mean within one or more standard deviations, as practiced in the art. Alternatively, “about” or “substantially comprise” can mean a range of up to 10% or 20% (i.e., ±10% or ±20%). For example, about 3 mg can include any number between 2.7 mg and 3.3 mg (for 10%) or between 2.4 mg and 3.6 mg (for 20%). Furthermore, particularly with respect to biological systems or processes, the term can mean up to an order of magnitude or up to a maximum of 5 times the numerical value. When a particular value or composition is provided in this application and claims, unless otherwise stated, the meaning of “about” or “substantially comprise” should be assumed to be within an acceptable margin of error for that particular value or composition.

[0052] The terms “about once a week,” “about once every two weeks,” or any other similar dosing interval terms used herein refer to approximate values. “About once a week” can include every 7 days ± 1 day, i.e., every 6 to 8 days. “About once every two weeks” can include every 14 days ± 3 days, i.e., every 11 to 17 days. Similar approximations apply, for example, about once every 3 weeks, about once every 4 weeks, about once every 5 weeks, about once every 6 weeks, and about once every 12 weeks. In some embodiments, a dosing interval of about once every 6 weeks or about once every 12 weeks means that the first dose can be administered on any day of the first week, and then the second dose can be administered on any day of the sixth or twelfth week, respectively. In other embodiments, a dosing interval of about once every 6 weeks or about once every 12 weeks means that the first dose is administered on a specific day of the first week (e.g., Monday), and then the second dose is administered on the same day of the sixth or twelfth week (i.e., Monday), respectively.

[0053] The term "weight-based dosage" as used in this article refers to a dosage administered to a patient that is calculated based on the patient's weight. For example, when a patient weighing 60 kg requires 3 mg / kg of anti-PD-1 antibody, an appropriate amount of anti-PD-1 antibody (i.e., 180 mg) can be calculated and used for administration.

[0054] Regarding the method of the present invention, the term "fixed dose" refers to the presence of two or more different antibodies (e.g., anti-PD-1 antibody and a second antibody, such as anti-CTLA-4 antibody) in a specific (fixed) ratio to each other in a single composition. In some embodiments, the fixed dose is based on the weight of the antibody (e.g., mg). In some embodiments, the fixed dose is based on the concentration of the antibody (e.g., mg / ml). In some embodiments, the ratio is at least about 1:1, about 1:2, about 1:3, about 1:4, about 1:5, about 1:6, about 1:7, about 1:8, about 1:9, about 1:10, about 1:15, about 1:20, about 1:30, about 1:40, about 1:50, about 1:60, about 1:70, about 1:80, about 1:90, about 1:100, about 1:120, about 1:140, about 1:160, about 1:1 80, approximately 1:200, approximately 200:1, approximately 180:1, approximately 160:1, approximately 140:1, approximately 120:1, approximately 100:1, approximately 90:1, approximately 80:1, approximately 70:1, approximately 60:1, approximately 50:1, approximately 40:1, approximately 30:1, approximately 20:1, approximately 15:1, approximately 10:1, approximately 9:1, approximately 8:1, approximately 7:1, approximately 6:1, approximately 5:1, approximately 4:1, approximately 3:1, or approximately 2:1 mg primary antibody (e.g., anti-PD-1 antibody): mg secondary antibody (e.g., anti-CTLA-4 antibody). For example, a 3:1 ratio of anti-PD-1 antibody to anti-CTLA-4 antibody could mean that a vial could contain approximately 240 mg of anti-PD-1 antibody and 80 mg of anti-CTLA-4 antibody, or approximately 3 mg / ml of anti-PD-1 antibody and 1 mg / ml of anti-CTLA-4 antibody.

[0055] Regarding the methods and dosages of this invention, the term "uniform dose" is used to refer to the dose administered to a patient regardless of the patient's weight or body surface area (BSA). The uniform dose is therefore not provided as a mg / kg dose, but rather as an absolute amount of the agent (e.g., anti-PD-1 antibody). For example, a 60 kg person and a 100 kg person may receive the same dose of antibody (e.g., 240 mg of anti-PD-1 antibody).

[0056] As described herein, any concentration range, percentage range, ratio range, or integer range shall be understood to include the value of any integer within the enumerated range, and, where appropriate, to include fractions (such as one-tenth and one-hundredth of an integer), unless otherwise indicated.

[0057] The different aspects of the invention are described in more detail in the following sections.

[0058] The method of the present invention This disclosure provides a method of treating a subject with lung cancer, the method comprising administering to the subject a therapeutically effective amount of a combination of the following agents: (a) an anticancer agent, which is an antibody that specifically binds to the PD-1 receptor and inhibits PD-1 activity or an antigen-binding portion thereof; and (b) an anti-CTLA-4 antibody. Because NSCLC accounts for more than 85% of lung tumors, in this embodiment, lung cancer is NSCLC. In other embodiments, the subject is a human patient. In some embodiments, the subject is a chemotherapy-naïve patient (e.g., a patient who has not previously received any chemotherapy). In other embodiments, the subject of the combination therapy of the present invention has received another cancer therapy (e.g., chemotherapy) but is resistant to or tolerant to said other cancer therapy. In some specific embodiments, the subject of the combination therapy of the present invention has cancer cells expressing a mutated form of the EGFR or KRAS gene. In some embodiments, the subject has PD-L1+ cancer cells. In some embodiments, the subject has PD-L1- cancer cells. In some embodiments, the subject has never smoked. In some embodiments, the subject previously smoked. In one embodiment, the subject currently smokes. In some embodiments, the subject has squamous cell carcinoma. In some implementations, the subject has non-squamous cancer cells.

[0059] In some embodiments, the therapies of the present invention (e.g., administration of anti-PD-1 antibodies and CTLA-4 antibodies) effectively increase the duration of subject survival. In some embodiments, the anti-PD-1 antibody combination therapy of the present invention increases progression-free survival in subjects. In some embodiments, the anti-PD-1 antibody combination therapy of the present invention increases progression-free survival in subjects compared to standard of care. In some embodiments, the anti-PD-1 antibody combination therapy of the present invention increases progression-free survival in subjects compared to anti-PD-1 antibodies alone. In some embodiments, the anti-PD-1 antibody combination therapy of the present invention increases progression-free survival in subjects compared to other combinations of anti-PD-1 antibodies. After administration of anti-PD-1 antibody combination therapy, subjects with lung cancer tumors can demonstrate overall survival of at least approximately 10 months, at least approximately 11 months, at least approximately 12 months, at least approximately 13 months, at least approximately 14 months, at least approximately 15 months, at least approximately 16 months, at least approximately 17 months, at least approximately 18 months, at least approximately 19 months, at least approximately 20 months, at least approximately 21 months, at least approximately 22 months, at least approximately 23 months, at least approximately 2 years, at least approximately 3 years, at least approximately 4 years, or at least approximately 5 years after administration.

[0060] In other embodiments, when compared to another subject treated with standard care therapy alone (e.g., docetaxel), a different dose regimen of anti-PD-1 antibody alone, or a combination therapy, the subject's duration of survival or overall survival increases by at least about 1 month, at least about 2 months, at least about 3 months, at least about 4 months, at least about 6 months, or at least about 1 year. For example, when compared to another subject treated with standard care therapy alone (e.g., docetaxel), a different dose regimen of anti-PD-1 antibody alone, or a combination therapy, the subject treated with the anti-PD-1 antibody combination disclosed herein increases the duration of survival or overall survival by at least about 5%, at least about 10%, at least about 15%, at least about 20%, at least about 25%, at least about 30%, at least about 40%, at least about 50%, or at least about 75%.

[0061] In some embodiments, the therapy of the present invention effectively increases the duration of progression-free survival in subjects. In some embodiments, subjects demonstrate progression-free survival of at least about 1 month, at least about 2 months, at least about 3 months, at least about 4 months, at least about 5 months, at least about 6 months, at least about 7 months, at least about 8 months, at least about 9 months, at least about 10 months, at least about 11 months, at least about 1 year, at least about 18 months, at least about 2 years, at least about 3 years, at least about 4 years, or at least about 5 years.

[0062] In some implementations, subjects had longer progression-free survival when treated with an anti-PD-1 antibody or its antigen-binding fraction at a dose of 3 mg / kg body weight every 2 weeks and an anti-CTLA-4 antibody or its antigen-binding fraction at a dose of 1 mg / kg body weight every 6-12 weeks (e.g., every 6 or 12 weeks), compared to when subjects were treated with an anti-PD-1 antibody or its antigen-binding fraction at a dose of 3 mg / kg body weight every 2 weeks. In some implementations, the progression-free survival of subjects treated with an anti-PD-1 antibody or its antigen-binding fraction at a dose of 3 mg / kg body weight every 2 weeks and an anti-CTLA-4 antibody or its antigen-binding fraction at a dose of 1 mg / kg body weight every 6 or 12 weeks is at least about 1 week, at least about 2 weeks, at least about 3 weeks, at least about 1 month, at least about 2 months, at least about 3 months, at least about 4 months, at least about 5 months, at least about 6 months, at least about 7 months, at least about 8 months, at least about 9 months, at least about 10 months, at least about 11 months, or at least about 1 year. In some embodiments, when an anti-PD-1 antibody or its antigen-binding portion is administered at a dose of 3 mg / kg body weight every 2 weeks and an anti-CTLA-4 antibody or its antigen-binding portion is administered at a dose of 1 mg / kg body weight every 6 or 12 weeks, the progression-free survival of the subject is at least about 3 months longer than when the subject was treated with an anti-PD-1 antibody or its antigen-binding portion administered at a dose of 3 mg / kg body weight every 2 weeks. In some embodiments, the anti-CTLA-4 antibody or its antigen-binding portion is administered every 6 weeks. In other embodiments, the anti-CTLA-4 antibody or its antigen-binding portion is administered every 12 weeks.

[0063] In some implementations, subjects had longer progression-free survival when treated with an anti-PD-1 antibody or its antigen-binding fraction at a dose of 3 mg / kg body weight every 2 weeks and an anti-CTLA-4 antibody or its antigen-binding fraction at a dose of 1 mg / kg body weight every 6 weeks, compared to when subjects were treated with an anti-PD-1 antibody or its antigen-binding fraction at a dose of 3 mg / kg body weight every 2 weeks and an anti-CTLA-4 antibody or its antigen-binding fraction at a dose of 1 mg / kg body weight every 6 or 12 weeks. In some implementations, the progression-free survival of subjects treated with an anti-PD-1 antibody or its antigen-binding fraction at a dose of 3 mg / kg body weight every 2 weeks and an anti-CTLA-4 antibody or its antigen-binding fraction at a dose of 1 mg / kg body weight every 6 or 12 weeks is at least about 1 week, at least about 2 weeks, at least about 3 weeks, at least about 1 month, at least about 2 months, at least about 3 months, at least about 4 months, at least about 5 months, or at least about 6 months. In some embodiments, when an anti-PD-1 antibody or its antigen-binding fraction is administered at a dose of 3 mg / kg body weight every 2 weeks and an anti-CTLA-4 antibody or its antigen-binding fraction is administered at a dose of 1 mg / kg body weight every 6 or 12 weeks, the progression-free survival is at least about 3 months longer than when the subject was treated with anti-PD-1 antibody or its antigen-binding fraction at a dose of 1 mg / kg body weight every 2 weeks and anti-CTLA-4 antibody or its antigen-binding fraction at a dose of 1 mg / kg body weight every 6 weeks, or when the subject was treated with standard care. In some embodiments, the anti-CTLA-4 antibody or its antigen-binding fraction is administered every 6 weeks. In other embodiments, the anti-CTLA-4 antibody or its antigen-binding fraction is administered every 12 weeks.

[0064] The PD-L1 status of the subject's tumor can be measured prior to administration of any composition or using any of the methods disclosed herein. In one embodiment, the PD-L1 expression level of the tumor is at least about 1%, at least about 2%, at least about 3%, at least about 4%, at least about 5%, at least about 6%, at least about 7%, at least about 8%, at least about 9%, at least about 10%, at least about 11%, at least about 12%, at least about 13%, at least about 14%, at least about 15%, at least about 20%, at least about 25%, at least about 30%, at least about 40%, at least about 50%, at least about 60%, at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, or about 100%. In another embodiment, the PD-L1 status of the tumor is at least about 1%. In other embodiments, the subject's PD-L1 status is at least about 5%. In one embodiment, the PD-L1 status of the tumor is at least about 10%. In one embodiment, the PD-L1 status of the tumor is at least about 25%. In the specific implementation plan, the PD-L1 status of the tumor is at least about 50%.

[0065] In some implementations, the median progression-free survival of subjects with tumors expressing ≥ 1% PD-L1 is at least about 1 week, at least about 2 weeks, at least about 3 weeks, at least about 4 weeks, at least about 1 month, at least about 2 months, at least about 3 months, at least about 4 months, at least about 5 months, at least about 6 months, at least about 7 months, at least about 8 months, at least about 9 months, at least about 10 months, at least about 11 months, or at least about 1 year longer than the median progression-free survival of subjects with tumors expressing < 1% PD-L1. In some implementations, the progression-free survival of subjects with tumors expressing ≥ 1% PD-L1 is at least about 1 month, at least about 2 months, at least about 3 months, at least about 4 months, at least about 5 months, at least about 6 months, at least about 7 months, at least about 8 months, at least about 9 months, at least about 10 months, at least about 11 months, at least about 1 year, at least about 18 months, at least about 2 years, at least about 3 years, at least about 4 years, or at least about 5 years.

[0066] To evaluate PD-L1 expression, in one embodiment, the test tissue sample may be obtained from a patient in need of the therapy. In another embodiment, the evaluation of PD-L1 expression may be performed without obtaining a test tissue sample. In some embodiments, selecting a suitable patient includes (i) optionally providing a test tissue sample obtained from a patient with cancer tissue containing tumor cells and / or tumor-infiltrating inflammatory cells; and (ii) evaluating the proportion of cells expressing PD-L1 on the cell surface in the test tissue sample based on an evaluation that the proportion of cells expressing PD-L1 on the cell surface in the test tissue sample is above a predetermined threshold level.

[0067] However, in any method that includes measuring PD-L1 expression in a test tissue sample, it should be understood that the step of providing a test tissue sample obtained from a patient is optional. It should also be understood that in some embodiments, the "measurement" or "evaluation" step of identifying cells expressing PD-L1 on their cell surface in a test tissue sample or determining their number or proportion is performed by a modified method for measuring PD-L1 expression, such as by performing a reverse transcriptase-polymerase chain reaction (RT-PCR) assay or an IHC assay. In some other embodiments, the modified step is not included, and PD-L1 expression is evaluated, for example, by reviewing test result reports from a laboratory. In some embodiments, the steps up to and including the method for evaluating PD-L1 expression provide intermediate results that can be provided to a physician or other healthcare provider for selecting suitable candidates for anti-PD-1 antibody or anti-PD-L1 antibody therapy. In some embodiments, the step of providing intermediate results is performed by a medical practitioner or a person working under the guidance of a medical practitioner. In other embodiments, these steps are performed by a neutral laboratory or by a neutral person such as a laboratory technician.

[0068] In some embodiments of any of the methods of the present invention, the proportion of cells expressing PD-L1 is evaluated by a assay to determine the presence of PD-L1 RNA. In other embodiments, the presence of PD-L1 RNA is determined by RT-PCR, in situ hybridization, or RNase protection. In other embodiments, the proportion of cells expressing PD-L1 is evaluated by a assay to determine the presence of PD-L1 peptide. In other embodiments, the presence of PD-L1 peptide is determined by immunohistochemistry (IHC), enzyme-linked immunosorbent assay (ELISA), in vivo imaging, or flow cytometry. In some embodiments, PD-L1 expression is determined by IHC. In other embodiments of all these methods, cell surface expression of PD-L1 is determined using, for example, IHC or in vivo imaging.

[0069] Imaging technologies have provided important tools in cancer research and treatment. Recent developments in molecular imaging systems, including positron emission tomography (PET), single-photon emission computed tomography (SPECT), fluorescence reflectance imaging (FRI), fluorescence-mediated tomography (FMT), bioluminescence imaging (BLI), laser scanning confocal microscopy (LSCM), and multiphoton microscopy (MPM), may foreshadow even greater applications of these technologies in cancer research. Some of these molecular imaging systems allow clinicians not only to observe the location of tumors within the body, but also to visualize the expression and activity of specific molecules, cells, and biological processes that affect the behavior and / or response of tumors to therapeutic drugs (Condeelis and Weissleder, "In vivo imaging in cancer," Cold Spring Harb. Perspect. Biol. 2(12):a003848 (2010)). The specific binding of antibodies to the sensitivity and resolution of PET makes immunoPET imaging particularly attractive for monitoring and analyzing antigen expression in tissue samples (McCabe and Wu, "Positive progress in immunoPET—not just a coincidence," Cancer Biother. Radiopharm. 25(3):253-61 (2010); Olafsen et al., "ImmunoPET imaging of B-cell lymphoma using 124I-anti-CD20 scFv dimers (diabodies)," Protein Eng. Des. Sel. 23(4):243-9 (2010)). In some embodiments of the method of the present invention, PD-L1 expression is determined by immunoPET imaging. In some embodiments of the method of the present invention, the proportion of cells expressing PD-L1 in the test tissue sample is evaluated by a assay to determine the presence of PD-L1 peptide on the cell surface in the test tissue sample. In some embodiments, the test tissue sample is an FFPE tissue sample. In other embodiments, the presence of PD-L1 peptide is determined by IHC assay. In other embodiments, the IHC assay is performed using an automated process. In some embodiments, the IHC assay is performed using an anti-PD-L1 mAb that binds to the PD-L1 peptide.

[0070] In one embodiment of the method of the present invention, an automated IHC method is used to determine the expression of PD-L1 on the cell surface in FFPE tissue samples. This disclosure provides a method for detecting the presence of human PD-L1 antigen in test tissue samples, or for quantifying the level of human PD-L1 antigen or the proportion of cells expressing the antigen in a sample, said method comprising contacting test samples and negative control samples with a mAb specifically binding to human PD-L1 under conditions allowing the formation of a complex between an antibody or a portion thereof and human PD-L1. In some embodiments, the test and control tissue samples are FFPE samples. The formation of the complex is then detected, wherein the difference in complex formation between the test sample and the negative control sample indicates the presence of human PD-L1 antigen in the sample. Various methods are used to quantify PD-L1 expression.

[0071] In a specific implementation, the automated IHC method includes: (a) dewaxing and rehydrating fixed tissue sections on an automated stainer; (b) restoring antigens using a demasking chamber and pH 6 buffer, and heating to 110°C for 10 min; (c) loading reagents onto the automated stainer; and (d) running the automated stainer to include steps such as neutralizing endogenous peroxidases in the tissue sample; blocking nonspecific protein binding sites on the slide; incubating the slide with a first antibody; incubating with a first post-blocking agent; incubating with NovoLink Polymer; adding chromogen substrate and developing the stain; and repeat staining with hematoxylin.

[0072] To evaluate PD-L1 expression in tumor tissue samples, pathologists examined membrane PD-L1 in each field of view under a microscope. + The number of tumor cells and the estimated percentage of positive cells in the brain are then averaged to obtain the final percentage. Different staining intensities are defined as 0 / negative, 1+ / weak, 2+ / moderate, and 3+ / strong. Typically, percentage values ​​are assigned between 0 and 3+, then the intermediate 1+ and 2+ intensities are considered. For highly heterogeneous tissues, the sample is partitioned, and each partition is scored individually, then the percentage values ​​are combined into a single group. The percentage of negative and positive cells for different staining intensities is determined from each region, and a median is given for each region. For each staining intensity category: negative, 1+, 2+, and 3+, a final percentage value for the tissue is given. The sum of all staining intensities needs to be 100%. In one embodiment, the threshold number of cells required to be PD-L1 positive is at least about 100, at least about 125, at least about 150, at least about 175, or at least about 200 cells. In some embodiments, the threshold number of cells required to be PD-L1 positive is at least about 100 cells.

[0073] Staining was also evaluated in tumor-infiltrating inflammatory cells such as macrophages and lymphocytes. In most cases, macrophages were used as an internal positive control because staining was observed in a large proportion of macrophages. Although staining at intensity 3+ is not required, the absence of macrophage staining should be considered to rule out any technical failure. The plasma membrane staining of macrophages and lymphocytes was evaluated, and all samples were recorded simply as positive or negative for each cell class. Staining was also characterized by nomenclature based on lateral / medial tumor immune cells. "Medial" refers to immune cells located within the tumor tissue and / or at the boundary of the tumor area without physical embedding within tumor cells. "Lateral" refers to immune cells that are not physically associated with the tumor and are present in the periphery associated with connective tissue or any relevant adjacent tissue.

[0074] In some embodiments of these scoring methods, samples are scored by two pathologists operating independently, and then the scores are combined. In some other embodiments, the identification of positive and negative cells is performed using appropriate software scoring.

[0075] Histoscore is used as a more quantitative measure of IHC data. The histoscore is calculated as follows: Tissue score = [(% tumor x 1 (low intensity)) + (% tumor x 2 (moderate intensity)) + (% tumor x 3 (high intensity)] To determine a tissue score, pathologists estimate the percentage of stained cells in each intensity category within a sample. Because the expression of most biomarkers is heterogeneous, the tissue score is a more accurate representation of overall expression. The final tissue score ranges from 0 (no expression) to 300 (maximum expression).

[0076] An alternative method for quantifying PD-L1 expression in IHC of tissue samples is to measure the adjusted inflammation score (AIS), which is defined as inflammation density multiplied by the percentage of PD-L1 expression in tumor-infiltrating inflammatory cells (Taube et al., "Colocalization of inflammatory response with B7-h1 expression in human melanocytic lesions supports an adaptive resistance mechanism of immune escape," Sci. Transl. Med. 4(127):127ra37 (2012)).

[0077] The method of this invention can treat non-squamous NSCLC at any stage. NSCLC exists in at least seven stages: occult (latent) stage, stage 0 (carcinoma in situ), stage I, stage II, stage IIIA, stage IIIB, and stage IV. In the occult stage, the cancer is not visible by imaging or bronchoscopy. In stage 0, cancer cells are found in the lining of the airway.

[0078] In one embodiment, the method of the present invention treats stage I non-squamous NSCLC. Stage I NSCLC is divided into stages IA and IB. In stage IA, the tumor is only in the lung and is 3 cm or smaller. In stage IB, the cancer has not spread to the lymph nodes, and one or more of the following are true: 1) the tumor is larger than 3 cm but not larger than 5 cm; 2) the cancer has spread to the main bronchus and is at least 2 cm below the tracheobronchial junction; 3) the cancer has spread to the innermost membrane covering the lung; or 4) a portion of the lung has collapsed or pneumonia (inflammation of the lung) has occurred in the area of ​​the tracheobronchial junction.

[0079] In another embodiment, the method of the present invention treats stage II non-squamous NSCLC. Stage II NSCLC is divided into stages IIA and IIB. In stage IIA, the cancer has spread to the lymph nodes or not. If the cancer has spread to the lymph nodes, it may have spread only to the lymph nodes on the same side of the chest as the tumor, to the lymph nodes with cancer, or to the lymph nodes in the lungs or near the bronchi, and one or more of the following are true: 1) the tumor is no larger than 5 cm; 2) the cancer has spread to the main bronchus, at least 2 cm below the trachea-bronchus junction; 3) the cancer has spread to the innermost membrane covering the lung; or 4) a portion of the lung has collapsed or pneumonia (inflammation of the lungs) has occurred in the area where the trachea-bronchus junction is located. A tumor is considered stage IIA if it has not yet spread to the lymph nodes and one or more of the following are true: 1) The tumor is larger than 5 cm but not larger than 7 cm; 2) The cancer has spread to the main bronchus, at least 2 cm below the trachea-bronchus junction; 3) The cancer has spread to the innermost membrane covering the lung; or 4) A portion of the lung in the trachea-bronchus junction area has collapsed or developed pneumonia (inflammation of the lung). In stage IIB, the cancer may or may not have spread to the lymph nodes. If the cancer has spread to the lymph nodes, it may have spread only to the lymph nodes on the same side of the chest as the tumor, with the cancerous lymph nodes within the lung or near the bronchus, and one or more of the following are true: 1) The tumor is larger than 5 cm but not larger than 7 cm; 2) The cancer has spread to the main bronchus, at least 2 cm below the trachea-bronchus junction; 3) The cancer has spread to the innermost membrane covering the lung; or 4) A portion of the lung in the trachea-bronchus junction area has collapsed or developed pneumonia (inflammation of the lung). A tumor is considered to be stage IIB if it has not spread to the lymph nodes and one or more of the following are true: 1) the tumor is larger than 7 cm; 2) the cancer has spread to the main bronchus (and at least 2 cm below the trachea-bronchus junction), chest wall, diaphragm, or nerves controlling the diaphragm; 3) the cancer has spread to the heart or the membranes surrounding the chest wall lining; 4) the entire lung has collapsed or pneumonia (inflammation of the lungs) has occurred; or 5) one or more separate tumors are present in the same lobe of the lung.

[0080] In other embodiments, any method of the present invention treats stage III non-squamous NSCLC. Stage IIIA is divided into three parts. These three parts are based on 1) tumor size; 2) location where the tumor is found; and 3) lymph nodes with cancer (if any). In stage IIIA NSCLC of type I, the cancer has spread to lymph nodes in the chest on the same side as the tumor, and to lymph nodes with cancer near the sternum or at the location where the bronchus enters the lung. In addition: 1) the tumor can be of any size; 2) a portion of the lung (at the trachea-bronchus junction) or the entire lung may have collapsed or developed pneumonia (inflammation of the lung); 3) one or more separate tumors may be present in the same lobe of the lung; and 4) the cancer may have spread to any of the following: a) the main bronchus, but not the area where the trachea-bronchus junction occurs, b) the chest wall, c) the diaphragm and the nerves that control it, d) the membranes surrounding the lung or chest wall lining, e) the membranes surrounding the heart. In stage IIIA NSCLC of type II, the cancer has spread to lymph nodes in the chest on the same side as the tumor, and to lymph nodes with cancer in the lungs or near the bronchi. Furthermore: 1) the tumor can be of any size; 2) the entire lung may have collapsed or pneumonia (inflammation of the lungs); 3) one or more separate tumors may be present in any lobe of the lung with cancer; and 4) the cancer may have spread to any of the following: a) the main bronchus, but not the area where the trachea connects to the bronchus, b) the chest wall, c) the diaphragm and the nerves controlling it, d) the membranes surrounding the lung or chest wall lining, e) the heart or the membranes surrounding it, f) the main blood vessels leading to or away from the heart, g) the trachea, h) the esophagus, i) the nerves controlling the voice box, j) the sternum or spine, or k) the vertebral column (the location where the trachea connects to the bronchi). In stage IIIA NSCLC (type 3), the cancer has not yet spread to the lymph nodes, the tumor can be of any size, and the cancer has spread to any of the following: a) the heart, b) the main blood vessels leading to or away from the heart, c) the trachea, d) the esophagus, e) the nerves controlling the larynx, f) the sternum or spine, or g) the vertebral column (where the trachea connects to the bronchi). Stage IIIB is divided into two parts based on 1) tumor size, 2) the location where the tumor was found, and 3) the lymph nodes with cancer. In stage IIIB NSCLC (type 1), the cancer has spread to the lymph nodes in the chest opposite the tumor.Additionally, 1) the tumor can be of any size; 2) a portion of the lung (where the trachea connects to the bronchus) or the entire lung may have collapsed or developed pneumonia (inflammation of the lung); 3) one or more separate tumors may be present in any lobe of the lung with cancer; and 4) the cancer may have spread to any of the following: a) the main bronchus, b) the chest wall, c) the diaphragm and the nerves controlling it, d) the membranes surrounding the lung or the lining of the chest wall, e) the heart or the membranes surrounding it, f) the main blood vessels leading to or leaving the heart, g) the trachea, h) the esophagus, i) the nerves controlling the larynx, j) the sternum or spine, or k) the vertebral column (where the trachea connects to the bronchus). In stage IIIB NSCLC of type II, the cancer has spread to the lymph nodes in the chest on the same side as the tumor. The lymph nodes with cancer are located near the sternum or at the site where the bronchus enters the lung. In addition, 1) the tumor can be of any size; 2) a single tumor can be present in different lobes of the same lung; and 3) the cancer has spread to any of the following: a) the heart, b) the main blood vessels leading to or leaving the heart, c) the trachea, d) the esophagus, e) the nerves controlling the larynx, f) the sternum or spine, or g) the vertebral column (where the trachea connects to the bronchus).

[0081] In some embodiments, the method of the present invention treats stage IV non-squamous NSCLC. In stage IV NSCLC, the tumor can be of any size, and the cancer may have spread to the lymph nodes. In stage IV NSCLC, one or more of the following are true: 1) one or more tumors are present in both lungs; 2) cancer is found in the fluid surrounding the lungs or heart; and 3) the cancer has spread to other parts of the body, such as the brain, liver, adrenal glands, kidneys, or bones.

[0082] This disclosure provides a combination therapy for treating lung cancer, wherein an anti-PD-1 antibody is combined with another anticancer agent, said other anticancer agent being an antibody that specifically binds to CTLA-4 and inhibits CTLA-4 activity, or an antigen-binding portion thereof. This document has demonstrated (see Example 1) that the combination of the anti-PD-1 antibody nirumab and the anti-CTLA-4 antibody ipilimumab produces early and durable antitumor activity in NSCLC patients, particularly with a specific dosing regimen. Therefore, in some embodiments, the anti-CTLA-4 antibody used in combination with the anti-PD-1 antibody is ipilimumab. In one embodiment, the anti-CTLA-4 antibody is trimemumab. In other embodiments, the anti-CTLA-4 antibody or its antigen-binding portion is an antibody or a portion thereof that cross-competitively binds to human CTLA-4 with ipilimumab. In some other embodiments, the anti-CTLA-4 antibody or its antigen-binding portion is chimeric, humanized, or a human mAb or a portion thereof. In still other embodiments, the anti-CTLA-4 antibody or its antigen-binding portion comprises a heavy chain constant region having a human IgG1 or IgG4 isotype. In some implementations, the anti-CTLA-4 antibody contains a heavy chain constant region with a human IgG1 isotype.

[0083] Because of the persistence of clinical effects previously demonstrated with immunotherapies that inhibit immune checkpoints (see, for example, WO 2013 / 173223), combination therapy in alternative embodiments may include limited doses, such as about 1 to 10 doses, or may include administration at long intervals, such as once every 3 to 6 months or once every 1 to 2 years or longer.

[0084] In some embodiments of the method of the present invention, the anti-PD-1 antibody is nilumab. In other embodiments, it is pembrolizumab. In still other embodiments, the anti-CTLA-4 antibody is ipilimumab. In other embodiments, the anti-CTLA-4 antibody is trimemumab. Typically, the anti-PD-1 and anti-CTLA-4 antibodies are formulated for intravenous administration. In some embodiments, when the anti-PD-1 and anti-CTLA-4 antibodies are administered in combination, they are administered within 30 minutes of each other. Either antibody may be administered first; that is, in some embodiments, the anti-PD-1 antibody is administered before the anti-CTLA-4 antibody, while in other embodiments, the anti-CTLA-4 antibody is administered before the anti-PD-1 antibody. Typically, each antibody is administered by intravenous infusion over a 60-minute period. In some embodiments, the anti-PD-1 and anti-CTLA-4 antibodies are administered simultaneously, either as a single composition in a pharmaceutically acceptable formulation for simultaneous administration, or as separate compositions in a pharmaceutically acceptable formulation of each antibody for simultaneous administration.

[0085] In some embodiments, the anti-PD-1 antibody or its antigen-binding portion is administered at a subtherapeutic dose. In some other embodiments, the anti-CTLA-4 antibody or its antigen-binding portion is administered at a subtherapeutic dose. In other embodiments, both the anti-PD-1 antibody or its antigen-binding portion and the anti-CTLA-4 antibody or its antigen-binding portion are administered at subtherapeutic doses.

[0086] The anti-PD-1 antibody or anti-PD-L1 antibody used in this invention HuMAb that binds to PD-1 with high affinity has been disclosed in U.S. Patent No. 8,008,449. Other anti-PD-1 monoclonal antibodies have been described, for example, in U.S. Patent Nos. 6,808,710, 7,488,802, 8,168,757, and 8,354,509 and PCT Publication No. WO 2012 / 145493. Each anti-PD-1 HuMAb disclosed in U.S. Patent No. 8,008,449 has been shown to exhibit one or more of the following characteristics: (a) binding with 1 x 10 -7 M or smaller K D The anti-PD-1 antibody can be: (a) bound to human PD-1, as determined by surface plasmon resonance using a Biacore biosensor system; (b) substantially unbound to human CD28, CTLA-4, or ICOS; (c) increased T-cell proliferation in a mixed lymphocyte response (MLR) assay; (d) increased interferon-γ production in an MLR assay; (e) increased IL-2 secretion in an MLR assay; (f) bound to both human PD-1 and cynomolgus monkey PD-1; (g) inhibited binding of PD-L1 and / or PD-L2 to PD-1; (h) stimulated an antigen-specific memory response; (i) stimulated an antibody response; and (j) inhibited tumor cell growth in vivo. Anti-PD-1 antibodies useful in this invention comprise monoclonal antibodies that specifically bind to human PD-1 and exhibit at least one (in some embodiments, at least five) of the aforementioned characteristics. In some embodiments, the anti-PD-1 antibody is nirumab. In one embodiment, the anti-PD-1 antibody is pembrolizumab.

[0087] In some embodiments, the anti-PD-1 antibody or its antigen-binding portion cross-competes with nilumab for binding to human PD-1. In one embodiment, the anti-PD-1 antibody is nilumab. Nilumab (also known as “OPDIVO®”; formerly known as 5C4, BMS-936558, MDX-1106, or ONO-4538) is a fully human IgG4 (S228P) PD-1 immune checkpoint inhibitor antibody that selectively blocks interaction with PD-1 ligands (PD-L1 and PD-L2), thereby blocking the downregulation of anti-tumor T-cell function (US Patent No. 8,008,449; Wang et al., 2014 Cancer Immunol Res. 2(9):846-56). In other embodiments, the anti-PD-1 antibody or a fragment thereof binds to the same epitopes as nilumab. In some embodiments, the anti-PD-1 antibody has the same CDR as nilumab.

[0088] In another embodiment, the anti-PD-1 antibody or a fragment thereof cross-competes with pembrolizumab. In some embodiments, the anti-PD-1 antibody or a fragment thereof binds to the same epitope as pembrolizumab. In some embodiments, the anti-PD-1 antibody has the same CDR as pembrolizumab. In another embodiment, the anti-PD-1 antibody is pembrolizumab. Pembrolizumab (also known as “KEYTRUDA®”, lambolizumab, and MK-3475) is a humanized monoclonal IgG4 antibody against the human cell surface receptor PD-1 (programmed death-1 or programmed cell death-1). Pembrolizumab is described, for example, in U.S. Patent Nos. 8,354,509 and 8,900,587; see also www.cancer.gov / drugdictionarycdrid=695789 (last accessed: December 14, 2014). Pembrolizumab has been approved by the FDA for the treatment of relapsed or refractory melanoma.

[0089] In other embodiments, the anti-PD-1 antibody or a fragment thereof cross-competes with MEDI0608. In other embodiments, the anti-PD-1 antibody or a fragment thereof binds to the same epitope as MEDI0608. In some embodiments, the anti-PD-1 antibody has the same CDR as MEDI0608. In other embodiments, the anti-PD-1 antibody is MEDI0608 (formerly known as AMP-514), which is a monoclonal antibody. MEDI0608 is described, for example, in U.S. Patent No. 8,609,089B2.

[0090] In some implementations, the immune checkpoint inhibitor is AMP-224, a B7-DC Fc fusion protein. AMP-224 is discussed in U.S. Publication No. 2013 / 0017199 or www.cancer.gov / publications / dictionaries / cancer-drugcdrid=700595 (last accessed: July 8, 2015).

[0091] In other embodiments, the anti-PD-1 antibody or a fragment thereof cross-competes with BGB-A317. In some embodiments, the anti-PD-1 antibody or a fragment thereof binds to the same epitope as BGB-A317. In some embodiments, the anti-PD-1 antibody has the same CDR as BGB-A317. In some embodiments, the anti-PD-1 antibody is BGB-A317, which is a humanized monoclonal antibody. BGB-A317 is described in U.S. Publication No. 2015 / 0079109.

[0092] Useful anti-PD-1 antibodies in the disclosed methods also include isolated antibodies that specifically bind to human PD-1 and cross-compete with nilumab for binding to human PD-1 (see, for example, U.S. Patents 8,008,449 and 8,779,105; WO 2013 / 173223). The ability of antibodies to cross-compete with an antigen indicates that these antibodies bind to the same epitope region of the antigen and spatially prevent other cross-competitive antibodies from binding to that specific epitope region. Due to their binding to the same epitope region of PD-1, these cross-competitive antibodies are expected to have functional properties very similar to nilumab. Cross-competitive antibodies can be readily identified based on their ability to cross-compete with nilumab in standard PD-1 binding assays such as Biacore assays, ELISA assays, or flow cytometry (see, for example, WO 2013 / 173223) (see, for example, WO 2013 / 173223).

[0093] In some embodiments, the antibody that cross-competes with the human PD-1 antibody nilucanumab for binding to human PD-1 or for binding to the same epitope region of the human PD-1 antibody nilucanumab is a monoclonal antibody. For administration to a human subject, these cross-competitive antibodies are chimeric antibodies, humanized antibodies, or human antibodies. Such chimeric, humanized, or human monoclonal antibodies can be prepared and isolated using methods well-known in the art.

[0094] The anti-PD-1 antibodies useful in the disclosed inventive methods also include the antigen-binding portion of the aforementioned antibody. It has been well demonstrated that the antigen-binding function of an antibody can be performed by a fragment of a full-length antibody. Examples of binding fragments included in the term "antigen-binding portion" of an antibody include: (i) Fab fragments, i.e., fragments composed of V... L V H C L and C H1 (ii) A monovalent segment composed of structural domains; (iii) A F(ab')2 segment, i.e., a divalent segment containing two Fab segments connected by disulfide bonds in the hinge region; H and C H1 (iv) Fd fragments composed of structural domains; and V segments composed of a single arm of the antibody. L and V H Fv segments composed of structural domains.

[0095] Suitable anti-PD-1 antibodies for use in the disclosed methods or compositions are antibodies that bind to PD-1 with high specificity and affinity, block the binding of PD-L1 and / or PD-L2, and inhibit the immunosuppressive effect of the PD-1 signaling pathway. In any of the compositions or methods disclosed herein, an anti-PD-1 “antibody” comprises an antigen-binding portion or fragment that binds to the PD-1 receptor and exhibits functional properties similar to those of a full antibody in inhibiting ligand binding and upregulating the immune system. In some embodiments, the anti-PD-1 antibody or its antigen-binding portion cross-competes with nirushumab for binding to human PD-1. In other embodiments, the anti-PD-1 antibody or its antigen-binding portion is a chimeric, humanized, or human monoclonal antibody or a portion thereof. In some embodiments, the antibody is a humanized antibody. In other embodiments, the antibody is a human antibody. Antibodies of IgG1, IgG2, IgG3, or IgG4 isotypes may be used.

[0096] In some embodiments, the anti-PD-1 antibody or its antigen-binding portion comprises a heavy chain constant region having a human IgG1 or IgG4 isotype. In some other embodiments, the sequence of the IgG4 heavy chain constant region of the anti-PD-1 antibody or its antigen-binding portion contains an S228P mutation, which replaces a serine residue in the hinge region with a proline residue typically found at the corresponding position in IgG1 isotype antibodies. This mutation present in nilucanab prevents Fab arm exchange with endogenous IgG4 antibodies while retaining low affinity for activating the Fc receptor associated with wild-type IgG4 antibodies (Wang et al., 2014 Cancer Immunol Res. 2(9):846-56). In other embodiments, the antibody comprises a light chain constant region, which is a human κ or λ constant region. In other embodiments, the anti-PD-1 antibody or its antigen-binding portion is a mAb or its antigen-binding portion. In some embodiments of any treatment method described herein that includes administration of an anti-PD-1 antibody, the anti-PD-1 antibody is nilucanab. In other embodiments, the anti-PD-1 antibody is pembrolizumab. In other embodiments, the anti-PD-1 antibody is selected from human antibodies 17D8, 2D3, 4H1, 4A11, 7D3, and 5F4 described in U.S. Patent No. 8,008,449. In other embodiments, the anti-PD-1 antibody is MEDI0608 (formerly known as AMP-514), AMP-224, or BGB-A317.

[0097] The anti-PD-1 antibodies useful in the methods of the disclosed invention also include the antigen-binding portion of the above-mentioned antibodies. It has been well demonstrated that the antigen-binding function of an antibody can be performed by fragments of a full-length antibody. Examples of binding fragments included in the term "antigen-binding portion" of an antibody include: (i) Fab fragments, i.e., fragments composed of V... L V H C L and C H1 (ii) A monovalent segment composed of structural domains; (iii) A F(ab'')2 segment, i.e., a divalent segment containing two Fab segments connected by disulfide bonds in the hinge region; H and C H1 (iv) Fd fragments composed of structural domains; and V segments composed of a single arm of the antibody. L and V H Fv segments composed of structural domains.

[0098] In some embodiments, the anti-PD-1 antibody used in the method can be replaced with another PD-1 or anti-PD-L1 antagonist. For example, because the anti-PD-L1 antibody blocks the interaction between PD-1 and PD-L1, thereby playing a similar role in the PD-1 signaling pathway, the anti-PD-L1 antibody can replace the use of the anti-PD-1 antibody in the methods disclosed herein. Therefore, in one embodiment, the present invention relates to a method for treating a subject with NSCLC, the method comprising administering to the subject a therapeutically effective amount of an anti-PD-L1 antibody and an anti-CTLA-4 antibody.

[0099] In some embodiments, the anti-PD-L1 antibody used in the method is BMS-936559 (formerly known as 12A4 or MDX-1105) (see, for example, U.S. Patent No. 7,943,743; WO 2013 / 173223).

[0100] In other embodiments, the anti-PD-L1 antibody is MPDL3280A (also known as RG7446) (see, for example, Herbst et al. (2013) J Clin Oncol 31(Supplement):3000. Abstract; U.S. Patent No. 8,217,149).

[0101] In other embodiments, the anti-PD-L1 antibody is MEDI4736 (also known as Durvalumab; Khleif (2013) see: Proceedings from the European Cancer Congress 2013; September 27–October 1, 2013; Amsterdam, The Netherlands. Abstract 802, see U.S. Patent No. 8,779,108 or US 2014 / 0356353, filed May 6, 2014).

[0102] In other embodiments, the anti-PD-L1 antibody is MSB0010718C (also known as Avelumab; see US2014 / 0341917).

[0103] Because anti-PD-1 and anti-PD-L1 antibodies target the same signaling pathway and have been shown in clinical trials to exhibit similar potency against a variety of cancers, including RCC (see Brahmer et al. (2012) N Engl J Med 366:2455-65; Topalian et al. (2012a) N Engl J Med 366:2443-54; WO 2013 / 173223), anti-PD-L1 antibodies can replace anti-PD-1 antibodies in any treatment method disclosed herein. In some embodiments, the anti-PD-L1 antibody is BMS-936559 (formerly known as 12A4 or MDX-1105) (see, for example, U.S. Patent No. 7,943,743; WO 2013 / 173223). In other embodiments, the anti-PD-L1 antibody is MPDL3280A (also known as RG7446) (see, for example, Herbst et al. (2013) J Clin Oncol 31(Supplement):3000. Abstract; U.S. Patent No. 8,217,149) or MEDI4736 (Khleif (2013) see: Proceedings from the European Cancer Congress 2013; September 27–October 1, 2013; Amsterdam, The Netherlands. Abstract 802). In some embodiments, the antibody that cross-competes with the above-mentioned reference PD-L1 antibody for binding to human PD-L1, or that binds to the same epitope region of human PD-L1 with the above-mentioned reference PD-L1 antibody, is a monoclonal antibody. For administration to a human subject, these cross-competitive antibodies may be chimeric antibodies, or may be humanized or human antibodies. Such chimeric, humanized, or human monoclonal antibodies can be prepared and isolated using methods well known in the art.

[0104] In some embodiments, the immune checkpoint inhibitors (e.g., anti-PD-1 antagonists) used in this invention are PD-1 Fc fusion proteins.

[0105] The anti-PD-1 antibody or its antigen-binding portion thereof, or the anti-PD-L1 antibody or its antigen-binding portion thereof, of the present invention may be administered to a subject at a dose selected from the following (first dose or second dose): about 0.1 mg / kg, about 0.2 mg / kg, about 0.3 mg / kg, about 0.4 mg / kg, about 0.5 mg / kg, about 0.6 mg / kg, about 0.7 mg / kg, about 0.8 mg / kg, about 0.9 mg / kg, about 1.0 mg / kg, about 1.1 mg / kg, about 1.2 mg / kg, about 1.3 mg / kg, about 1.4 mg / kg, about 1.5 mg / kg, about 2.0 mg / kg, about 2.5 mg / kg, about 3.0 mg / kg, about 3.5 mg / kg, about 4.0 mg / kg, about 4.5 mg / kg, about 5.5 mg / kg, about 6.5 mg / kg, about 7.5 mg / kg, about 8.0 mg / kg, about 8.5 mg / kg. mg / kg, about 9.0 mg / kg, about 9.5 mg / kg, about 10 mg / kg, about 11 mg / kg, about 12 mg / kg, about 13 mg / kg, about 14 mg / kg, about 15 mg / kg, about 20 mg / kg or greater than about 20 mg / kg. In other embodiments, the anti-PD-1 antibody or its antigen-binding portion, or the anti-PD-L1 antibody or its antigen-binding portion, may be administered at doses selected from the following: about 0.1 to about 20.0 mg / kg, about 0.1 to about 15.0 mg / kg, about 0.1 to about 10.0 mg / kg, about 0.1 to about 9.5 mg / kg, about 0.1 to about 9.0 mg / kg, about 0.1 to about 8.5 mg / kg, about 0.1 to about 8.0 mg / kg, about 0.1 to about 7.5 mg / kg, about 0.1 to about 7.0 mg / kg, about 0.1 to about 6.5 mg / kg, about 0.1 to about 6.0 mg / kg, about 0.1 to about 5.5 mg / kg, about 0.1 to about 5.0 mg / kg, about 0.1 to about 4.5 mg / kg, about 0.1 to about 4.0 mg / kg, about 0.1 to about 3.5 mg / kg, about 0.1 to about 3.0 mg / kg. The dosage is approximately 0.3 to 10.0 mg / kg, approximately 0.3 to 9.0 mg / kg, approximately 0.3 to 6.0 mg / kg, approximately 0.3 to 3.0 mg / kg, approximately 3.0 to 10.0 mg / kg, approximately 3.0 to 9.0 mg / kg, or approximately 3.0 to 6.0 mg / kg. In some embodiments, the subject is given 0.3 mg / kg of an anti-PD-1 antibody, such as nilumab, or an anti-PD-L1 antibody. In other embodiments, the subject is given 2.0 mg / kg of an anti-PD-1 antibody, such as nilumab, or an anti-PD-L1 antibody.In other embodiments, the subject is given 10 mg / kg of an anti-PD-1 antibody, such as nilumab, or an anti-PD-L1 antibody.

[0106] The anti-CTLA-4 antibody used in this invention The anti-CTLA-4 antibody of the present invention binds to human CTLA-4 to disrupt the interaction between CTLA-4 and the human B7 receptor. Because the interaction between CTLA-4 and B7 transduces signals that lead to the inactivation of T cells carrying the CTLA-4 receptor, disruption of this interaction effectively induces, enhances, or prolongs the activation of such T cells, thereby inducing, enhancing, or prolonging the immune response.

[0107] HuMAb that specifically binds to CTLA-4 with high affinity has been disclosed in U.S. Patent Nos. 6,984,720 and 7,605,238. Other anti-CTLA-4 monoclonal antibodies have been described, for example, in U.S. Patent Nos. 5,977,318, 6,051,227, 6,682,736, and 7,034,121. The anti-CTLA-4 HuMAb disclosed in U.S. Patent Nos. 6,984,720 and 7,605,238 has been shown to exhibit one or more of the following characteristics: (a) binding with at least about 10 7 M -1 or about 10 9 M -1 or about 10 10 M -1 Up to 10 11 M -1 Or a higher equilibrium association constant (K) a (a) Reflects binding affinity specifically binding to human CTLA-4, as determined by Biacore analysis; (b) At least about 10 3 Approximately 10 4 Or about 10 5 m -1 s -1 The dynamic association constant (k) a (c) at least about 10 3 Approximately 10 4 Or about 10 5 m -1 s -1 The kinetic dissociation constant (k) d(d) Inhibits the binding of CTLA-4 to B7-1 (CD80) and B7-2 (CD86). Anti-CTLA-4 antibodies useful in this invention comprise monoclonal antibodies that specifically bind to human CTLA-4 and exhibit at least one, at least two, or in one embodiment at least three of the aforementioned characteristics. An exemplary clinical anti-CTLA-4 antibody is human mAb 10D1 (now known as ipilimumab and marketed as YERVOY®), disclosed in U.S. Patent No. 6,984,720. Ipilimumab is an anti-CTLA-4 antibody used in the methods disclosed herein. Another anti-CTLA-4 antibody useful in the methods of this invention is trimemumab.

[0108] An exemplary clinical anti-CTLA-4 antibody is human mAb 10D1 disclosed in U.S. Patent No. 6,984,720 (now known as ipilimumab and marketed as YERVOY®). Ipilimumab is an anti-CTLA-4 antibody used in the methods disclosed herein. Ipilimumab is a fully human IgG1 monoclonal antibody that blocks the binding of CTLA-4 to its B7 ligand, thereby stimulating T cell activation and improving overall survival (OS) in patients with advanced melanoma.

[0109] Another anti-CTLA-4 antibody that can be used in the methods of this invention is trimemumab (also known as CP-675,206). Trimemumab is a human IgG2 monoclonal anti-CTLA-4 antibody. Trimemumab is described in WO / 2012 / 122444, US Publication No. 2012 / 263677, or WO Publication No. 2007 / 113648 A2.

[0110] Useful anti-CTLA-4 antibodies in the disclosed methods also include isolated antibodies that specifically bind to human PD-1 and cross-competitively bind to human CTLA-4 with ipilimumab or trimemumab, or bind to the same epitope region of human CTLA-4 with ipilimumab or trimemumab. In some embodiments, the antibody that cross-competitively binds to human CTLA-4 with ipilimumab or trimemumab, or binds to the same epitope region of human PD-1 with ipilimumab or trimemumab, is an antibody containing a heavy chain of the human IgG1 isotype. For administration to human subjects, these cross-competitive antibodies are chimeric antibodies, or humanized or human antibodies. Useful anti-CTLA-4 antibodies also include antigen-binding portions of the above antibodies, such as Fab, F(ab'')2, Fd, or Fv fragments.

[0111] Ipilimumab (YERVOY®) is a fully human IgG1 monoclonal antibody that blocks the binding of CTLA-4 to its B7 ligand, thereby stimulating T-cell activation and improving overall survival (OS) in patients with advanced melanoma (Hodi et al. (2010) N Engl J Med 363:711-23). ​​In a phase 1 clinical trial, concomitant therapy with nirushumab and ipilimumab produced rapid and substantial tumor regression in a high proportion of patients with advanced melanoma, significantly more effective than either antibody alone (Wolchok et al. (2013) N Engl J Med 369(2):122-33; WO 2013 / 173223). However, it remains unknown whether this combination of immunomodulatory antibodies is similarly effective in other tumor types.

[0112] Standard nursing care for lung cancer The standard care treatments for different types of cancer are well known to those skilled in the art. For example, the National Comprehensive Cancer Network (NCCN), a consortium of 21 major cancer centers in the United States, publishes the NCCN Clinical Practice Guidelines in Oncology (NCCN GUIDELINES®), which provides detailed and recently updated information on standard care treatments for various cancers (see NCCN GUIDELINES® (2014), available at: www.nccn.org / professionals / physician_gls / f_guidelines.asp, last accessed: May 14, 2014).

[0113] Non-sclerotic lung cancer (NSCLC) is the leading cause of cancer death in the United States and worldwide, surpassing the combined causes of breast, colon, and prostate cancer. An estimated 228,190 new cases of lung and bronchial cancer will be diagnosed in the US, and approximately 159,480 deaths will occur due to this disease (Siegel et al. (2014) CA Cancer J Clin 64(1):9-29). The majority of patients (approximately 78%) are diagnosed with advanced / recurrent or metastatic disease. Metastasis from lung cancer to the adrenal glands is a common event, occurring in approximately 33% of patients. NSCLC therapy has shown progressively improving overall survival (OS), but the benefit has plateaued (median OS in advanced patients is only 1 year). Progression after 1L therapy occurs in almost all of these subjects, and the 5-year survival rate in refractory cases is only 3.6%. From 2005 to 2009, the overall 5-year relative survival rate for lung cancer in the United States was 15.9% (NCCN GUIDELINES®, version 3.2014 – Non-small cell lung cancer, available at www.nccn.org / professionals / physician_gls / pdf / nscl.pdf, last accessed: May 14, 2014).

[0114] Surgery, radiation therapy (RT), and chemotherapy are the three commonly used treatments for NSCLC patients. As a type, NSCLC is relatively insensitive to chemotherapy and RT compared to small cell carcinoma. Typically, for patients with stage I or II disease, surgical resection offers the best chance of cure, followed by chemotherapy administered gradually before and after surgery. RT can also be used as adjuvant therapy for patients with resectable NSCLC, as initial local treatment, or as palliative care for patients with refractory NSCLC.

[0115] Patients with stage IV disease who have good performance status (PS) benefit from chemotherapy. Many drugs, including platinum-based agents (e.g., cisplatin, carboplatin), taxanes (e.g., paclitaxel, albumin-bound paclitaxel, docetaxel), vinorelbine, vinblastine, etoposide, pemetrexed, and gemcitabine, are available for stage IV NSCLC. Combinations of many of these drugs produce 30-40% 1-year survival rates, superior to single-agent therapy. Specific targeted therapies have also been developed for the treatment of advanced lung cancer. For example, bevacizumab (AVASTIN®) is a mAb that blocks vascular endothelial growth factor A (VEGF-A). Erlotinib (TARCEVA®) is a small molecule TKI targeting the epidermal growth factor receptor (EGFR). Crizotinib (XALKORI®) is a small molecule TKI targeting ALK and MET, and is used to treat NSCLC patients with mutated ALK fusion genes. Cetuximab (ERBITUX®) is a mAb targeting EGFR.

[0116] There is a particularly unmet need in patients with squamous cell NSCLC (representing up to 25% of all NSCLC) due to limited treatment options after first-line (1L) therapy. Single-agent chemotherapy is the standard of care after progression with platinum-based doublet chemotherapy (Pt-doublet), resulting in a median overall survival (OS) of approximately 7 months. Docetaxel remains the baseline treatment in this line of therapy, although erlotinib can also be used less frequently. In second-line (2L) treatment of patients with advanced NSCLC, pemetrexed has also shown clinically equivalent results compared to docetaxel, but with significantly fewer side effects (Hanna et al. (2004) J Clin Oncol 22:1589-97). No therapy is currently approved for lung cancer in a context beyond third-line (3L). Pemetrexed and bevacizumab are not approved in squamous NSCLC, and molecularly targeted therapies have limited use. The recent failures of Oncothyreon and Merck KgaA's STIMUVAX® to improve overall survival in a phase 3 trial, ArQule and Daiichi Sankyo's c-Met kinase inhibitor tivantinib to fail to meet the survival endpoint, Eli Lilly's ALIMTA® combined with Roche's AVASTIN® to improve overall survival in a late-stage study, and Amgen and Takeda Pharmaceutical's failure to meet the clinical endpoint with the small molecule VEGF-R antagonist motesanib in a late-stage trial, have increased the unmet need in advanced lung cancer.

[0117] Immunotherapy for lung cancer There is a clear need for effective agents for patients who have progressed on multiple lines of targeted therapy, and for therapies that extend survival beyond current standard treatments. Newer approaches, including immunotherapy, particularly those blocking immune checkpoints including CTLA-4, PD-1, and PD-L1 inhibitory pathways, have recently shown promise (Creelan et al. (2014) CancerControl 21(1):80-89). Thus, the combination of ipilimumab with chemotherapy has shown encouraging results in small cell and non-small cell lung cancers. Furthermore, dual checkpoint blockade strategies, such as those combining anti-PD-1 and anti-CTLA-4, have proven highly effective in treating melanoma (Wolchok et al. (2013) N Engl J Med 369(2):122-33; WO2013 / 173223), and other combinations including anti-PD-L1, anti-LAG-3, or anti-KIR have been tested to increase the proportion and durability of tumor responses. Similar to melanoma, NSCLC patients can benefit from combinations of different immunotherapies or such drugs combined with targeted agents or other treatments, including surgery, radiation, standard cancer chemotherapy, or vaccines. Although the combination of nirushumab and ipilimumab has been shown to be very effective in treating melanoma with manageable toxicity (Wolchok et al. (2013) N Engl J Med 369(2):122-33), it is unknown to date whether this combination will be significantly more effective in human subjects compared to treating NSCLC and other cancers with the drugs alone.

[0118] Drug composition and dosage The therapeutic agents of the present invention can be incorporated into compositions, for example, pharmaceutical compositions containing one or more antibodies and pharmaceutically acceptable carriers. As used herein, "pharmaceutically acceptable carrier" includes any and all physiologically compatible solvents, dispersion media, coating agents, antibacterial and antifungal agents, isotonic agents, and absorption delay agents. In one embodiment, the carrier used in the antibody-containing composition is suitable for intravenous, intramuscular, subcutaneous, parenteral, spinal, or epidermal administration (e.g., by injection or infusion). The pharmaceutical compositions of the present invention may include one or more pharmaceutically acceptable salts, antioxidants, aqueous and non-aqueous carriers, and / or adjuvants such as preservatives, wetting agents, emulsifiers, and dispersants.

[0119] This disclosure provides dosing regimens that can provide the required response, such as maximum therapeutic response and / or minimal side effects. For administration of anti-PD-1 antibodies, particularly in combination with anti-CTLA-4 antibodies, the dose range can be about 0.01 to about 20 mg / kg, about 0.1 to about 10 mg / kg, about 0.01 to about 5 mg / kg, about 1 to about 5 mg / kg, about 2 to about 5 mg / kg, about 1 to about 3 mg / kg, about 7.5 to about 12.5 mg / kg, or about 0.1 to about 30 mg / kg of the subject's body weight. For example, the dose can be about 0.1, about 0.3, about 1, about 2, about 3, about 4, about 5, or about 10 mg / kg of body weight. Based on the typical pharmacokinetic properties of the antibody, the dosing regimen is generally designed to achieve exposure leading to sustained receptor occupancy (RO). Exemplary treatment regimens must be administered about once a week, about once every 2 weeks, about once every 3 weeks, about once every 4 weeks, about once a month, about once every 3-6 months, or longer. In some embodiments, an anti-PD-1 antibody, such as nilumumab, is administered to the subject approximately every two weeks. The anti-PD-1 antibody may be administered in at least two doses, each dose being an amount of about 0.01 mg / kg to about 5 mg / kg, for example, 3 mg / kg, with an interval of every two weeks between the two doses. In some embodiments, the anti-PD-1 antibody is administered in at least 3, 4, 5, 6, or 7 doses (i.e., multiple doses), each dose being an amount of about 0.01 mg / kg to about 5 mg / kg, for example, 3 mg / kg, with an interval of every two weeks between two adjacent doses. The dosage and timing may vary during treatment. In one embodiment, the dosage regimen of the anti-PD-1 antibody of the present invention comprises intravenous administration of about 0.3-1 mg / kg body weight, about 5 mg / kg body weight, 1-5 mg / kg body weight, or about 1-3 mg / kg body weight, wherein the antibody is administered every about 14-21 days over a period of up to about 6 weeks or about 12 weeks, until a complete response or confirmed disease progression. In some implementations, the antibody treatment or any combination treatment disclosed herein lasts for at least about 1 month, at least about 3 months, at least about 6 months, at least about 9 months, at least about 1 year, at least about 18 months, at least about 24 months, at least about 3 years, at least about 5 years, or at least about 10 years.

[0120] When used in combination with an anti-CTLA-4 antibody, the dose of the anti-PD-1 antibody can be reduced relative to the monotherapy dose. A dose of nirumab below the typical 3 mg / kg, but not less than 0.001 mg / kg, is a subtherapeutic dose. The subtherapeutic dose of the anti-PD-1 antibody used in the methods described herein is greater than 0.001 mg / kg and less than 3 mg / kg. In some embodiments, the subtherapeutic dose is about 0.001 mg / kg to about 1 mg / kg, about 0.01 mg / kg to about 1 mg / kg, about 0.1 mg / kg to about 1 mg / kg, or about 0.001 mg / kg to about 0.1 mg / kg body weight. In some embodiments, the subtherapeutic dose is at least about 0.001 mg / kg, at least about 0.005 mg / kg, at least about 0.01 mg / kg, at least about 0.05 mg / kg, at least about 0.1 mg / kg, at least about 0.5 mg / kg, or at least about 1.0 mg / kg body weight. Receptor occupancy data from 15 subjects who received nirumab at doses ranging from 0.3 mg / kg to 10 mg / kg indicate that PD-1 occupancy appears to be dose-independent within this dose range. The mean occupancy rate was 85% (range, 70% to 97%) across all doses, and the mean plateau occupancy rate was 72% (range, 59% to 81%) (Brahmer et al. (2010) J Clin Oncol 28:3167-75). Therefore, a dose of 0.3 mg / kg may allow sufficient exposure to result in maximal biological activity.

[0121] In some embodiments of the invention, the anti-PD-1 antibody is administered at a dose of 3 mg / kg. In other embodiments of the invention, the anti-PD-1 antibody is administered at a dose of 1 mg / kg.

[0122] In some embodiments, the dose of the anti-PD-1 antibody (or anti-PD-L1 antibody) is a fixed dose in the pharmaceutical composition. In other embodiments, the method of the present invention can be used at a uniform dose (the dose administered to the patient regardless of the patient's weight). In embodiments, the uniform dose of the anti-PD-1 antibody or its antigen-binding portion is at least about 100 mg, 120 mg, 140 mg, 160 mg, 180 mg, 200 mg, 220 mg, 240 mg, 260 mg, 280 mg, 300 mg, 360 mg, 400 mg, 420 mg, 440 mg, 460 mg, 480 mg, 500 mg, 520 mg, 540 mg, 560 mg, or 600 mg. For example, the uniform dose of nirumab can be about 240 mg. For example, the uniform dose of pembrolizumab can be about 200 mg. In embodiments, the anti-PD-1 antibody or its antigen-binding portion is administered at a dose of about 240 mg. In one embodiment, the anti-PD-1 antibody or its antigen-binding portion is administered at a dose of approximately 360 mg. In another embodiment, the anti-PD-1 antibody or its antigen-binding portion is administered at a dose of approximately 480 mg. In yet another embodiment, a uniform dose of the anti-PD-1 antibody or its antigen-binding portion is administered approximately weekly, every 2 weeks, every 3 weeks, every 4 weeks, every 5 weeks, or every 6 weeks. In one embodiment, 360 mg of the anti-PD-1 antibody or antigen-binding fragment is administered every 3 weeks. In another embodiment, 480 mg of the anti-PD-1 antibody or antigen-binding fragment is administered every 4 weeks.

[0123] Ipilimumab (YERVOY) ®It is approved for use in the treatment of melanoma by intravenous administration of 3 mg / kg every 3 weeks for 4 consecutive doses. Therefore, in some embodiments, approximately 3 mg / kg is the highest dose of ipilimumab used in combination with an anti-PD-1 antibody, but in other embodiments, when combined with an anti-PD-1 antibody, the anti-CTLA-4 antibody, such as ipilimumab, can be administered approximately every 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, or 20 weeks in the range of approximately 0.3 to approximately 10 mg / kg, approximately 0.5 to approximately 10 mg / kg, approximately 0.5 to approximately 5 mg / kg, approximately 1 to approximately 5 mg / kg, approximately 1 to approximately 4 mg / kg, or approximately 1 to approximately 3 mg / kg body weight. In other embodiments, the anti-CTLA-4 antibody is administered at a different dosing regimen than the anti-PD-1 antibody. In some embodiments, ipilimumab is administered approximately weekly, approximately every 2 weeks, approximately every 3 weeks, approximately every 4 weeks, approximately every 5 weeks, approximately every 6 weeks, approximately every 7 weeks, approximately every 8 weeks, approximately every 9 weeks, approximately every 10 weeks, approximately every 11 weeks, approximately every 12 weeks, approximately every 13 weeks, approximately every 14 weeks, approximately every 15 weeks, or approximately every 20 weeks. A dose of ipilimumab below the typical 3 mg / kg, but not less than 0.001 mg / kg, is a subtherapeutic dose. The subtherapeutic dose of the anti-CTLA-4 antibody used in the methods described herein is greater than 0.001 mg / kg and less than 3 mg / kg. In some embodiments, the subtherapeutic dose is approximately 0.001 mg / kg to approximately 1 mg / kg, approximately 0.01 mg / kg to approximately 1 mg / kg, approximately 0.1 mg / kg to approximately 1 mg / kg, or approximately 0.001 mg / kg to approximately 0.1 mg / kg body weight. In some embodiments, the subtherapy dose is at least about 0.001 mg / kg, at least about 0.005 mg / kg, at least about 0.01 mg / kg, at least about 0.05 mg / kg, at least about 0.1 mg / kg, at least about 0.5 mg / kg, or at least about 1.0 mg / kg body weight. It has been shown that the combination of 3 mg / kg nilumab and 3 mg / kg ipilimumab exceeds the MTD in the melanoma population, while the combination of 1 mg / kg nilumab + 3 mg / kg ipilimumab, or 3 mg / kg nilumab + 1 mg / kg ipilimumab, has been found to be tolerable in melanoma patients (Wolchok et al. (2013) N Engl J Med 369(2):122-33). Therefore, although niluzumab is tolerated up to an intravenous dose of 10 mg / kg every 2 weeks, in some embodiments, when combined with an anti-CTLA-4 antibody, the dose of the anti-PD-1 antibody does not exceed about 3 mg / kg. In some embodiments, the dose of the anti-CTLA-4 antibody is about 1 mg / kg.

[0124] In some embodiments, based on risk-benefit and PK-PD assessments, the dosage used comprises a combination of approximately 1 mg / kg nilumab plus approximately 1 mg / kg ipilimumab, or approximately 3 mg / kg nilumab plus approximately 1 mg / kg ipilimumab. In some embodiments, nilumab is administered approximately every 2 weeks. In some embodiments, ipilimumab is administered approximately every 6 weeks or 12 weeks. In some other embodiments, in combination with ipilimumab administered at a dose of approximately 1 mg / kg, nilumab is administered at a dose of approximately 1 or approximately 3 mg / kg, wherein nilumab is administered every 2 weeks and ipilimumab is administered every 6 weeks or every 12 weeks. In some other embodiments, in combination with ipilimumab administered at a dose of approximately 1 mg / kg every 6 weeks, nilumab is administered at a dose of 1 mg / kg every 2 weeks. In some embodiments, niruzumab is administered in combination with ipilimumab at a dose of about 1 mg / kg every 12 weeks, and niruzumab is administered at a dose of 1 mg / kg every 2 weeks. In some embodiments, niruzumab is administered in combination with ipilimumab at a dose of about 1 mg / kg every 6 weeks, and niruzumab is administered at a dose of 3 mg / kg every 2 weeks. In other embodiments, niruzumab is administered in combination with ipilimumab at a dose of about 1 mg / kg every 12 weeks, and niruzumab is administered at a dose of 3 mg / kg every 2 weeks.

[0125] In some embodiments, the anti-PD-1 antibody and the anti-CTLA-4 antibody are formulated as a single composition, wherein the doses of the anti-PD-1 antibody and the anti-CTLA-4 antibody are combined in a fixed ratio of 1:50, 1:40, 1:30, 1:20, 1:10, 1:5, 1:3, 1:1, 3:1, 5:1, 10:1, 20:1, 30:1, 40:1, or 50:1. In some embodiments, the dose of the anti-CTLA-4 antibody is a uniform dose administered to the patient regardless of weight. In some embodiments, the uniform dose of the anti-CTLA-4 antibody is at least about 40 mg, 50 mg, 60 mg, 70 mg, 80 mg, 90 mg, 100 mg, 120 mg, 140 mg, 160 mg, 180 mg, or 200 mg. In a particular embodiment, the uniform dose of the anti-CTLA-4 antibody is about 80 mg.

[0126] The dosage and frequency vary with the half-life of the antibody in the subject. Generally, human antibodies exhibit the longest half-life, followed by humanized antibodies, chimeric antibodies, and non-human antibodies. The dosage and frequency of administration can vary depending on whether the treatment is prophylactic or therapeutic. In prophylactic use, relatively low doses are typically administered at relatively infrequent intervals over a long period. Some patients continue treatment for the rest of their lives. In therapeutic use, relatively higher doses are sometimes required at relatively short intervals until the disease progression slows or ceases, or until the patient shows partial or complete improvement in disease symptoms. Afterward, a prophylactic regimen can be administered to the patient.

[0127] The actual dose level of the active ingredient in the pharmaceutical composition of the present invention can be varied to obtain an amount of active ingredient that effectively achieves the desired therapeutic response for a particular patient, composition, and administration mode, without undue toxicity to the patient. The selected dose level will depend on a variety of pharmacokinetic factors, including the activity of the particular composition of the present invention used, the route of administration, the time of administration, the excretion rate of the particular compound being used, the duration of treatment, other drugs, compounds, and / or materials used in combination with the particular composition used, the age, sex, weight, condition, general health, and prior medical history of the patient being treated, and similar factors well known in the medical field. The compositions of the present invention can be administered via one or more routes of administration using one or more methods well known in the art. Those skilled in the art will understand that the route and / or mode of administration will vary depending on the desired outcome.

[0128] medicine box The scope of this invention also includes a kit containing an anti-PD-1 antibody and another anticancer agent for therapeutic use. Kits typically include a label indicating the intended use of the contents and instructions for use. Terminology labels include any written or recorded material on or with the kit, or otherwise accompanying the kit. Therefore, this disclosure provides a kit for treating a subject with lung cancer, the kit containing: (a) a PD-1 antibody or its antigen-binding portion in an amount ranging from about 4 mg to about 500 mg; (b) a CTLA-4 antibody or its antigen-binding portion in an amount ranging from about 40 mg to about 500 mg; and (c) instructions for using the PD-1 antibody or its antigen-binding portion and the CTLA-4 antibody or its antigen-binding portion in any of the methods disclosed herein. In some embodiments, the kit contains the PD-1 antibody or its antigen-binding portion and the CTLA-4 antibody or its antigen-binding portion as a separate composition. In some embodiments, the kit contains the PD-1 antibody or its antigen-binding portion and the CTLA-4 antibody or its antigen-binding portion as a single composition. In some embodiments, the anti-PD-1 and anti-CTLA-4 antibodies may be co-packaged in a unit dosage form. In some embodiments for treating human patients, the kit contains an anti-human PD-1 antibody disclosed herein, such as nilumumab or pembrolizumab. In other embodiments, the kit contains an anti-human CTLA-4 antibody disclosed herein, such as ipilimumab or trimemumab.

[0129] The present invention is further illustrated by the following embodiments, which should not be construed as further limitations. All references cited in this application are expressly incorporated herein by reference.

[0130] Example 1 NSCLC treated with nilumumab and ipilimumab Research Design Four distinct groups were developed to test the dosage and dosing regimens of nivo and ipilimumab ("ipi"). These groups were designed to test dosing regimens different from the initial test of nivo + ipi combination therapy. A summary of results from previous combination therapies is shown in Table 1. NR = Not reached.

[0131] Table 1. Groups with advanced NSCLC previously provided. See Gettinger, S. et al. J. Clin Oncol33(suppl): 8025 (2015) and Antonia, SJ et al. Int. J. Radial Oncol Bio Phys 90(suppl5): S32-S33 (2014).

[0132] Data for Nivo 3 mg / kg Q3W were based on the March 2015 database lockout. Data for Nivo 1 mg / kg + Ipi 3 mg / kg Q3W and Nivo 3 mg / kg + Ipi 1 mg / kg Q3W were based on the September 2015 database lockout. The estimated mDOR is the time from first response to confirmed progression, death within 100 days of the last nirumab dose, or the last tumor assessment (for examination + data).

[0133] Four additional groups were tested: 1 mg / kg ipi and 1 mg / kg nivolumab q3w; 1 mg / kg nivolumab q2w and 1 mg / kg ipi q6w; 3 mg / kg nivolumab q2w and 1 mg / kg ipi q12w; and 3 mg / kg nivolumab q2w and 1 mg / kg gibberellic acid pi q6w. Patient demographics, mutations, and baseline disease characteristics are shown in Table 2.

[0134] Table 2. Baseline characteristics of patients Due to rounding, some percentages may total less than 100%. PD-L1 expression percentages are based on patients with known PD-L1 status (nivo 1 + ipi 1 Q3W, n=25; nivo 1 Q2W + ipi 1 Q6W, n=28; nivo 3 Q2W + ipi 1 Q12W, n=31; nivo 3 Q2W + ipi Q6W, n=30). The most common treatment-related adverse events (AEs) are shown in Table 3. At the time of analysis, 47%–81% of patients (across groups) discontinued study treatment, most commonly due to disease progression.

[0135] Table 3. Safety Overview, Exposure, and Patient Arrangements NC = Not calculated. The new dosing regimen resulted in a lower frequency and severity of treatment-related adverse events compared to the earlier nirumab plus ipilimumab group (see Table 1). The new dosing regimen was also associated with a lower frequency of treatment-related adverse events leading to discontinuation (Table 3) compared to the earlier nirumab plus ipilimumab group. The discontinuation rate was comparable to that of nirumab monotherapy (see Table 1). There were no treatment-related deaths.

[0136] The selected treatment-related adverse events are shown in Table 4. These were those with underlying immunological causes requiring frequent monitoring / intervention. Across all groups, the most common types of selected treatment-related adverse events (any grade ≥ 20%) were: skin, endocrine, gastrointestinal, and hepatic.

[0137] Table 4. Reported treatment-related adverse events (AEs) in patients treated with nirumab plus ipilimumab Overall response, progression-free survival, and total survival are summarized in Table 5. The confirmed overall response rate (ORR) ranged from 13% to 39% (across all groups), with an additional 21%–42% of patients achieving disease stabilization. Two patients in the nirumab 3 mg / kg Q2W + ipilimumab 1 mg / kg Q12W group and one patient in the nirumab 3 mg / kg Q2W + ipilimumab 1 mg / kg Q6W group had unconventional immune responses, with maximum reductions in target lesions of 42%, 47%, and 44% respectively following disease progression or the concurrent occurrence of new lesions. The median disease-free survival (DOR) was not reached in any group. Tumor burden reduction was observed in all groups. Figures 2A-2D Surprisingly, compared with nilumab 1 mg / kg Q2W plus ipilimumab 1 mg / kg Q6W, nilumab 3 mg / kg Q2W plus ipilimumab 1 mg / kg Q12W and nilumab 3 mg / kg Q2W plus ipilimumab 1 mg / kg Q6W showed patients with complete responses and a significant increase in the length of progression-free survival.

[0138] Table 5. Response Overview (Database locked in August 2015, unless otherwise noted).

[0139] NC = Not calculated (when >25% of patients are examined); NR = Not achieved due to a high percentage of progressive responses or inadequate number of events and / or follow-up. The + sign indicates the examined value. DCR = Disease Control Rate; mOS = Median OS; mPFS = Median PFS. DCR includes patients with confirmed CR, PR, and SD.

[0140] 7 Database locked in February 2016 The duration of the response of Nivo plus IPI in first-line NSCLC can be observed in Figure 4In the Q6W group, 12 out of 15 respondents (80%) and in the Q12W group, 14 out of 18 respondents (77%) had responses confirmed by the initial scan at week 11 + / - 5 days. At database lockout, 12 out of 15 respondents (80%) in the Q6W group and 12 out of 18 respondents (67%) in the Q12W group had progressive responses.

[0141] Therapeutic efficacy based on baseline tumor PD-L1 expression is shown in Table 6. Tumor PD-L1 expression was assessed in pre-treatment (archived or fresh) tumor samples using an automated Bristol-Myers Squibb / Dako immunohistochemical assay, and ORR and PFS were evaluated. See Phillips, T. et al. Appl. Immunohistochem Mol Morphol 23: 541-549 (2015). All patients had available pre-treatment tumor samples, and 76% (113 / 148) had samples evaluable for PD-L1 expression. Furthermore, clinical activity was observed regardless of tumor PD-L1 expression. Figure 3 While preliminary evidence suggests greater activity in tumors with ≥ 1% PD-L1 expression, 85% (35 / 41) of confirmed responses were in progress at the time of analysis. Median DOR was not reached in any group regardless of tumor PD-L1 expression. Further evaluation is shown in Table 7, and the efficacy of nivolumab plus ipi at PD-L1 expression levels is shown in [Table 7]. Figure 5 .

[0142] Table 6. Efficacy of baseline tumor PD-L1 expression NR is caused by a high percentage of progressive responses or insufficient number of events and / or follow-up.

[0143] Table 7. Efficacy of baseline tumor PD-L1 expression Median PFS and overall response rate (ORR) were higher in current and former smokers. See Table 8 and Figure 6A In patients with non-squamous NSCLC, a response was observed regardless of EGFR mutation status. See Table 9 and... Figure 6B These data demonstrate that clinical activity was observed regardless of smoking status or EGFR mutation status.

[0144] Table 8. Efficacy of Nirutumab plus Ipilimumab in patients treated with smoking status Table 9. Efficacy of EGFR mutation status in non-squamous patients treated with nirucumab plus ipilimumab NR is due to a high percentage of progressive responses or an inadequate number of events and / or follow-up. The + sign indicates a test value. These values ​​include patients with non-squamous histology treated with nivolumab 1 mg / kg Q2W + ipi 1 mg / kg Q6W, nivolumab 3 mg / kg Q2W + ipi 1 mg / kg Q12W, or nivolumab 3 mg / kg Q2W + ipi 1 mg / kg Q6W. DCR includes patients with confirmed complete response, partial response, and stable disease. The + sign indicates a test value.

[0145] in conclusion Although earlier studies showed clinical activity with nirumab at 1 mg / kg and ipilimumab every three weeks, or nirumab at 3 mg / kg and ipilimumab at 1 mg / kg, the dosing regimen was associated with toxicity. In this study, the new dosing regimen unexpectedly demonstrated synergistic activity and increased clinical activity, along with acceptable safety. First-line therapy with nirumab plus ipilimumab demonstrated high levels of clinical activity, characterized by deep and durable responses in patients with advanced NSCLC. Furthermore, combination therapy with nirumab plus ipilimumab was associated with a favorable safety profile. There were a low frequency of treatment-related grade 3–4 adverse events leading to discontinuation, and no treatment-related deaths.

[0146] Example 2 A case of pathological complete remission in a patient who received Nivo 3 Q2W + Ipi 1 Q6W A 54-year-old male (former smoker, 52 pack-years) with metastatic large cell lung cancer (PD-L1 <1%; included because of partial response at database lockout and PD-L1 expression unknown at analysis) was treated with Nivo3 Q2W + Ipi 1 Q6W. The patient achieved a 53% reduction in total tumor size by RECIST and radiographic residual damage in the lungs and mediastinal lymph nodes, and had no distant disease. See Figures 7A-7D .

[0147] Implementation Plan E1. A method for treating a subject with lung cancer, comprising administering to the subject a combination of the following: (a) An antibody or its antigen-binding moiety that specifically binds to the programmed death-1 (PD-1) receptor and inhibits PD-1 activity, wherein the PD-1 antibody or its antigen-binding moiety is administered approximately every 2 weeks at a dose ranging from about 0.1 to about 5.0 mg / kg body weight; and (b) An antibody or antigen-binding moiety thereof that specifically binds to cytotoxic T lymphocyte antigen-4 (CTLA-4) and inhibits CTLA-4 activity, wherein the CTLA-4 antibody or antigen-binding moiety thereof is administered at a dose ranging from about 1 to about 5.0 mg / kg body weight approximately every 6 or 12 weeks.

[0148] The E2.E1 method, where the lung cancer is non-small cell lung cancer (NSCLC).

[0149] The E3.E2 method, in which NSCLC has squamous histology.

[0150] The E4.E2 method, in which NSCLC has non-squamous histology.

[0151] The method of any one of E5.E1-4, wherein the anti-PD-1 antibody or its antigen-binding portion cross-competes with nirushumab for binding to human PD-1.

[0152] The method of any one of E6.E1-5, wherein the anti-PD-1 antibody or its antigen-binding portion is chimeric, humanized or a human monoclonal antibody or a portion thereof.

[0153] The method of any one of E7, E1-E6, wherein the anti-PD-1 antibody or its antigen-binding portion comprises a heavy chain constant region having a human IgG1 or IgG4 isotype.

[0154] The method of any one of E8.E1-E7, wherein the anti-PD-1 antibody is nirumab.

[0155] The method is any one of E9.E1-E7, wherein the anti-PD-1 antibody is pembrolizumab.

[0156] E10. The method of any one of E1-E9, wherein the anti-CTLA-4 antibody or its antigen-binding portion is a chimeric, humanized, or human monoclonal antibody or a portion thereof.

[0157] E11. The method of any one of E1-E10, wherein the anti-CTLA-4 antibody or its antigen-binding portion comprises a heavy chain constant region having a human IgG1 isotype.

[0158] E12. Any of the methods in E1-E11, wherein the anti-CTLA-4 antibody is ipilimumab.

[0159] E13. The method of any one of E1-E11, wherein the anti-CTLA-4 antibody is trimesumab.

[0160] E14. The method of any one of E1-E13, wherein the anti-CTLA-4 antibody or its antigen-binding portion cross-competes with ipilimumab for binding to human CTLA-4.

[0161] E15. The method of any of E1-14, wherein the anti-PD-1 antibody or its antigen-binding portion is administered approximately every 2 weeks at a dose of approximately 1 mg / kg or approximately 3 mg / kg body weight.

[0162] E16. The method of any one of E1-E15, wherein the anti-CTLA-4 antibody or its antigen-binding portion is administered at a dose of about 1 mg / kg body weight.

[0163] E17. The method of any one of E1-E16, wherein an anti-PD-1 antibody or its antigen-binding portion is administered at a dose of about 3 mg / kg body weight about every 2 weeks and an anti-CTLA-4 antibody or its antigen-binding portion is administered at a dose of about 1 mg / kg body weight about every 12 weeks.

[0164] E18. The method of any one of E1-E16, wherein an anti-PD-1 antibody or its antigen-binding portion is administered at a dose of about 1 mg / kg body weight approximately every 2 weeks and an anti-CTLA-4 antibody or its antigen-binding portion is administered at a dose of about 1 mg / kg body weight approximately every 6 weeks.

[0165] E19. The method of any one of E1-E18, wherein the subject demonstrates progression-free survival for at least about 1 month, at least about 2 months, at least about 3 months, at least about 4 months, at least about 5 months, at least about 6 months, at least about 7 months, at least about 8 months, at least about 9 months, at least about 10 months, at least about 11 months, at least about 1 year, at least about 18 months, at least about 2 years, at least about 3 years, at least about 4 years, or at least about 5 years after initial administration.

[0166] The methods of E20 and E19, in which subjects showed at least approximately 8 months of progression-free survival after initial administration.

[0167] E21. The method of any one of E1-E20, wherein, compared with treatment with an anti-PD-1 antibody or its antigen-binding portion administered at a dose of 1 mg / kg body weight every 2 weeks and an anti-CTLA-4 antibody or its antigen-binding portion administered at a dose of 1 mg / kg body weight every 6 weeks ("Regimen B"), the subject has longer progression-free survival when treated with an anti-PD-1 antibody or its antigen-binding portion administered at a dose of about 3 mg / kg body weight every 2 weeks and an anti-CTLA-4 antibody or its antigen-binding portion administered at a dose of about 1 mg / kg body weight every 12 weeks ("Regimen A").

[0168] E22. E21 method, wherein the progression-free survival of the subject receiving regimen A is at least about 1 week, at least about 2 weeks, at least about 3 weeks, at least about 1 month, at least about 2 months, at least about 3 months, at least about 4 months, at least about 5 months, or at least about 6 months longer than that of the subject receiving regimen B.

[0169] Methods E23, E21, or E22, wherein subjects receiving regimen A have a progression-free survival at least approximately 3 months longer than subjects receiving regimen B.

[0170] E24. The method of any one of E1-E22, wherein when the subject is treated with an anti-PD-1 antibody or its antigen-binding portion administered at a dose of 3 mg / kg body weight every 2 weeks ("Regimen C"), the subject has longer progression-free survival when treated with an anti-PD-1 antibody or its antigen-binding portion administered at a dose of 3 mg / kg body weight every 2 weeks and an anti-CTLA-4 antibody or its antigen-binding portion administered at a dose of 1 mg / kg body weight every 12 weeks ("Regimen A").

[0171] E25. E23's method, wherein the progression-free survival of subjects receiving regimen A is at least about 1 week, at least about 2 weeks, at least about 3 weeks, at least about 4 weeks, at least about 5 weeks, at least about 6 weeks, at least about 7 weeks, at least about 8 weeks, at least about 9 weeks, at least about 10 weeks, at least about 11 weeks, at least about 12 weeks, at least about 13 weeks, at least about 14 weeks, or at least about 15 weeks longer than that of subjects receiving regimen C.

[0172] Methods E26, E21, or E22, wherein subjects receiving regimen A have a progression-free survival at least approximately 3 months longer than subjects receiving regimen C.

[0173] E27. The method of any one of E1-E26, wherein the subject has a lung tumor that expresses ≥ 1% PD-L1, ≥ 5% PD-L1, ≥ 10% PD-L1, ≥ 25% PD-L1, or ≥ 50% PD-L1.

[0174] E28. The method of any one of E1-E27, wherein the combination is administered as soon as clinical benefit is observed or until disease progression or uncontrollable toxicity occurs.

[0175] E29. The method of any one of E1-E28, wherein the anti-PD-1 and anti-CTLA-4 antibodies are formulated for intravenous administration.

[0176] The method of any of E30 and E1-29, wherein an anti-PD-1 antibody or its antigen-binding portion and an anti-CTLA-4 antibody or its antigen-binding portion are sequentially administered to a subject.

[0177] E31. The method of any of E1-30, wherein the anti-PD-1 and anti-CTLA-4 antibodies are administered to each other within 30 minutes.

[0178] E32. The method of any one of E1-E31, wherein the anti-PD-1 antibody or its antigen-binding portion is administered prior to the anti-CTLA-4 antibody or its antigen-binding portion.

[0179] E33. The method of any one of E1-E31, wherein the anti-CTLA-4 antibody or its antigen-binding portion is administered prior to the anti-PD-1 antibody or its antigen-binding portion.

[0180] E34. The method of any one of E1-E29, wherein the anti-PD-1 antibody or its antigen-binding portion and the anti-CTLA-4 antibody or its antigen-binding portion are administered simultaneously in separate compositions.

[0181] E35. The method of any one of E1-E29, wherein the anti-PD-1 antibody or its antigen-binding portion and the anti-CTLA-4 antibody or its antigen-binding portion are administered simultaneously as a single composition.

[0182] E36. The method of any one of E1-E35, wherein the anti-PD-1 antibody or its antigen-binding portion is administered at a subtherapeutic dose.

[0183] E37. The method of any of E1-36, wherein the anti-CTLA-4 antibody or its antigen-binding portion is administered at a subtherapeutic dose.

[0184] E38. The method of any one of E1-E37, wherein the anti-PD-1 antibody or its antigen-binding portion and the anti-CTLA-4 antibody or its antigen-binding portion are each administered at a subtherapeutic dose.

[0185] E39. A medicine kit for treating a subject with lung cancer, the medicine kit comprising: (a) Anti-PD-1 antibody or its antigen-binding portion in amounts ranging from about 4 mg to about 500 mg; (b) A CTLA-4 antibody or its antigen-binding moiety in amounts ranging from about 40 mg to about 500 mg; and (c) Instructions for use of the PD-1 antibody or its antigen-binding portion and the CTLA-4 antibody or its antigen-binding portion in any of the methods in E1-E38.

Claims

1. A composition comprising an antibody or antigen-binding portion thereof (anti-PD-1 antibody or antigen-binding portion thereof) that specifically binds to the programmed death-1 (PD-1) receptor and inhibits PD-1 activity, for use in combination with an antibody or antigen-binding portion thereof (anti-CTLA-4 antibody or antigen-binding portion thereof) that specifically binds to cytotoxic T lymphocyte antigen-4 (CTLA-4) and inhibits CTLA-4 activity to treat a subject with lung cancer; The anti-PD-1 antibody or its antigen-binding moiety is administered approximately every 2 weeks at a dose ranging from approximately 0.1 to approximately 5.0 mg / kg body weight; and The anti-CTLA-4 antibody or its antigen-binding fraction is administered at a dose ranging from about 1 to about 5.0 mg / kg body weight, approximately every 6 to 12 weeks.

2. The composition used in claim 1, wherein the lung cancer is non-small cell lung cancer (NSCLC).

3. The composition used in claim 1 or 2, wherein the anti-PD-1 antibody or its antigen-binding portion cross-competes with nilucanab for binding to human PD-1.

4. The composition used in any one of claims 1-3, wherein the anti-CTLA-4 antibody or its antigen-binding portion cross-competes with ipilimumab for binding to human CTLA-4.

5. The composition used in any one of claims 1-4, wherein: (i) The anti-PD-1 antibody is nirumab; (ii) The anti-CTLA-4 antibody is ipilimumab; or (iii) The anti-PD-1 antibody is nilumumab, and the anti-CTLA-4 antibody is ipilimumab.

6. The composition used in any one of claims 1-5, wherein the anti-PD-1 antibody or its antigen-binding portion is administered at a dose of about 1 mg / kg or about 3 mg / kg body weight once every 2 weeks.

7. The composition used in any one of claims 1-6, wherein the anti-CTLA-4 antibody or its antigen-binding portion is administered at a dose of about 1 mg / kg body weight.

8. The composition used in any one of claims 1-7, wherein the anti-PD-1 antibody or its antigen-binding portion is administered at a dose of about 3 mg / kg body weight once every 2 weeks, and the anti-CTLA-4 antibody or its antigen-binding portion is administered at a dose of about 1 mg / kg body weight once every 12 weeks.

9. The composition used in any one of claims 1-7, wherein the anti-PD-1 antibody or its antigen-binding portion is administered at a dose of about 1 mg / kg body weight once every 2 weeks, and the anti-CTLA-4 antibody or its antigen-binding portion is administered at a dose of about 1 mg / kg body weight once every 6 weeks.

10. The composition used in any one of claims 1-9, wherein the subject has a lung tumor, the lung tumor having ≥ 1% PD-L1, ≥ 5% PD-L1, ≥ 10% PD-L1, ≥ 25% PD-L1, or ≥ 50% PD-L1 expression.

11. The composition used in any one of claims 1-10, wherein the composition is administered as soon as clinical benefit is observed or until disease progression or uncontrollable toxicity occurs.

12. The composition used in any one of claims 1-11, wherein the anti-PD-1 and anti-CTLA-4 antibodies are formulated for intravenous administration.

13. The composition used in any one of claims 1-12, wherein the anti-PD-1 antibody or its antigen-binding portion and the anti-CTLA-4 antibody or its antigen-binding portion are sequentially administered to the subject.

14. The composition used in any one of claims 1-12, wherein the anti-PD-1 antibody or its antigen-binding portion and the anti-CTLA-4 antibody or its antigen-binding portion are administered simultaneously in separate compositions or as a single composition.

15. The composition used in any one of claims 1-14, wherein: (i) Anti-PD-1 antibody or its antigen-binding moiety is administered at a subtherapeutic dose; (ii) Anti-CTLA-4 antibody or its antigen-binding moiety is administered at a subtherapeutic dose; or (iii) The anti-PD-1 antibody or its antigen-binding portion and the anti-CTLA-4 antibody or its antigen-binding portion are each administered at a subtherapeutic dose.

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