Anti-PD-1 antibody for use in method of treating tumor

Abstract

To provide a method for treating a subject afflicted with tumor, the method comprising administering to the subject an anti-Programmed Death-1 (PD-1) antibody.SOLUTION: The method comprises administering to the subject an antibody or an antigen-binding portion thereof that specifically binds to a Programmed Death-1 (PD-1) receptor and inhibits PD-1 activity. In some embodiments, the tumor is derived from a non-small cell lung cancer (NSCLC). In some embodiments, the tumor expresses Programmed Death Ligand 1 (PD-L1), Serine / Threonine Kinase 11 (STK11), or both PD-L1 and STK11.SELECTED DRAWING: None

Description

Technical Field

[0001] Field of the Invention The present invention relates to a method of treating a tumor, comprising administering an anti-programmed death-1 (PD-1) antibody to a subject, wherein the tumor expresses PD-L1 and / or wild-type STK11.

Background Art

[0002] Background of the Invention Human cancers harbor numerous genetic and epigenetic alterations and produce neoantigens that may be recognized by the immune system (Sjoblom et al., (2006) Science 314:268-74). The adaptive immune system, consisting of T and B lymphocytes, has a broad capacity and exquisite specificity to respond to diverse tumor antigens and has a potent anti-cancer ability. Furthermore, the immune system exhibits considerable flexibility and a memory component. The success of harnessing all of these properties of the adaptive immune system makes immunotherapy special among all cancer treatment modalities.

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

[0004] Nivolumab (previously named 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 (U.S. Patent 8,008,449; Wang et al., 2014 Cancer Immunol Res. 2(9):846-56).

[0005] NSCLC is the leading cause of cancer death in the United States and worldwide (NCCN GUIDELINES (登録商標) , Version 3.2014 - Non-Small Cell Lung Cancer, available at www.nccn.org / professionals / physician_gls / pdf / nscl.pdf, last accessed May 14, 2014). NSCLC is relatively insensitive to chemotherapeutic agents, but patients with stage IV disease with good performance status (PS) benefit from chemotherapeutic agents including platinum agents (e.g., cisplatin, carboplatin), taxane agents (e.g., paclitaxel, albumin-bound paclitaxel, docetaxel), vinorelbine, vincristine, etoposide, pemetrexed, gemcitabine and various combinations of these drugs. SUMMARY OF THE INVENTION MEANS FOR SOLVING THE PROBLEM

[0006] SUMMARY OF THE INVENTION The present invention provides a method of treating a subject having a tumor, comprising: (i) determining the expression pattern of programmed death ligand 1 (PD-L1), and (ii) if the tumor exhibits a pan-pattern of PD-L1 expression, administering to the subject an antibody or antigen-binding portion thereof that specifically binds to the programmed death-1 (PD-1) receptor and inhibits PD-1 activity (“anti-PD-1 antibody”). In certain embodiments, the present invention provides a method of treating a subject having a tumor, comprising: (i) determining the expression pattern of PD-L1, and (ii) if the tumor exhibits a heterogeneous pattern of PD-L1 expression, administering to the subject an anti-PD-1 antibody. In other embodiments, the present invention provides a method of treating a subject having a tumor, comprising: (i) determining the expression pattern of PD-L1, and (ii) if the tumor exhibits a tumor-stromal interface pattern of PD-L1 expression, administering to the subject an anti-PD-1 antibody. In other embodiments, the present invention provides a method of identifying a subject having a tumor suitable for anti-PD-1 antibody treatment, comprising: (i) determining the expression pattern of PD-L1, and (ii) if the tumor exhibits a pan-pattern of PD-L1 expression, administering to the subject an anti-PD-1 antibody. In yet other embodiments, the present invention provides a method of identifying a subject having a tumor suitable for anti-PD-1 antibody treatment, comprising: (i) determining the expression pattern of PD-L1, and (ii) if the tumor exhibits a heterogeneous pattern of PD-L1 expression, administering to the subject an anti-PD-1 antibody. In certain embodiments, the methods described herein further comprise identifying a patient as having a tumor that expresses STK11 prior to administration.

[0007] In other aspects, the present invention relates to a method of treating a subject having a tumor, comprising (i) identifying a subject having an STK11-positive tumor, and (ii) administering an anti-PD-1 antibody to the subject. In certain aspects, the present invention provides a method of treating a subject having a tumor, comprising administering an anti-PD-1 antibody, wherein the patient has been identified as having an STK11-positive tumor prior to administration. In certain aspects, the present invention relates to a method of identifying a subject having a tumor suitable for anti-PD-1 antibody treatment, comprising (i) measuring STK11 expression in the tumor, and (ii) administering an anti-PD-1 antibody to the subject if the tumor is STK11-positive. In certain embodiments, STK11 is wild-type STK11.

[0008] In certain embodiments, the tumor is derived from lung cancer. In certain embodiments, the tumor is derived from small cell lung cancer (SCLC) or non-small cell lung cancer (NSCLC). In certain embodiments, the tumor is derived from NSCLC.

[0009] In certain embodiments, diffuse pattern of PD-L1 expression is characterized by a PD-L1 H score of about 60 to about 500, about 80 to about 480, about 100 to about 460, about 120 to about 440, about 140 to about 420, about 160 to about 400, about 180 to about 380, about 200 to about 360, about 200 to about 340, about 200 to about 320 or about 200 to about 300. In certain embodiments, diffuse pattern of PD-L1 expression is characterized by a PD-L1 H score of at least about 200.

[0010] In certain embodiments, heterogeneous pattern of PD-L1 expression is characterized by a PD-L1 H score of about 1 to about 50, about 5 to about 45, about 10 to about 40 or about 15 to about 35, wherein the PD-L1 expression is restricted to one or more different sites of the tumor. In certain embodiments, heterogeneous pattern of PD-L1 expression is characterized by a PD-L1 H score of at least about 15.

[0011] In certain embodiments, the anti-PD-1 antibody cross-competes with nivolumab for binding to human PD-1. In certain embodiments, the anti-PD-1 antibody binds to the same epitope as nivolumab. In certain embodiments, the anti-PD-1 antibody is a chimeric, humanized or human monoclonal antibody or a portion thereof. In certain embodiments, the anti-PD-1 antibody is nivolumab.

[0012] In certain embodiments, the anti-PD-1 antibody is administered once every about one, two or three weeks at a dose in the range of at least about 0.1 mg / kg to at least about 10.0 mg / kg body weight. In certain embodiments, the anti-PD-1 antibody is administered once every about two weeks at a dose of at least about 3 mg / kg body weight. In certain embodiments, the anti-PD-1 antibody or an antigen-binding portion thereof is administered at a uniform dose once every one, two, three or four weeks. In certain embodiments, the anti-PD-1 antibody or an antigen-binding portion thereof is administered at a uniform dose or at about 240 mg.

[0013] In other aspects, the invention provides a kit for treating a subject having a tumor, the kit comprising (a) a single dosage amount in the range of about 4 mg to about 500 mg of an anti-PD-1 antibody; and (b) instructions for using the anti-PD-1 antibody in any of the methods described herein. In certain embodiments for treating a human patient, the kit comprises an anti-human PD-1 antibody as disclosed herein, such as nivolumab or pembrolizumab. In certain embodiments, the kit further comprises an anti-PD-L1 antibody and / or an anti-STK11 antibody.

[0014] Embodiments E1. A method of treating a subject having a tumor, comprising (i) determining the expression pattern of programmed death ligand 1 (PD-L1), and (ii) if the tumor exhibits a widespread pattern of PD-L1 expression, administering to the subject an antibody or an antigen-binding portion thereof that specifically binds to the programmed death-1 (PD-1) receptor and inhibits PD-1 activity (“anti-PD-1 antibody”).

[0015] E2. A method of treating a subject having a tumor, comprising: (i) determining the expression pattern of PD-L1, and (ii) administering an anti-PD-1 antibody to the subject if the tumor exhibits a heterogeneous pattern of PD-L1 expression.

[0016] E3. A method of treating a subject having a tumor, comprising: (i) determining the expression pattern of PD-L1, and (ii) administering an anti-PD-1 antibody to the subject if the tumor exhibits a tumor-stromal interface pattern of PD-L1 expression.

[0017] E4. A method of identifying a subject having a tumor suitable for anti-PD-1 antibody treatment, comprising: (i) determining the expression pattern of PD-L1, and (ii) administering an anti-PD-1 antibody to the subject if the tumor exhibits a diffuse pattern of PD-L1 expression.

[0018] E5. A method of identifying a subject having a tumor suitable for anti-PD-1 antibody treatment, comprising: (i) determining the expression pattern of PD-L1, and (ii) administering an anti-PD-1 antibody to the subject if the tumor exhibits a heterogeneous pattern of PD-L1 expression.

[0019] E6. The method according to any one of embodiments E1-E5, further comprising identifying a patient as having a tumor that expresses STK11 before administration.

[0020] E7. A method of treating a subject having a tumor, comprising: (i) identifying a subject having an STK11-positive tumor, and (ii) administering an anti-PD-1 antibody to the subject.

[0021] E8. A method of treating a subject having a tumor, comprising administering an anti-PD-1 antibody, wherein the patient is identified as having an STK11-positive tumor before administration.

[0022] E9. A method of identifying a subject having a tumor suitable for anti-PD-1 antibody treatment, comprising: (i) measuring STK11 expression by the tumor, and (ii) administering an anti-PD-1 antibody to the subject if the tumor is STK11-positive.

[0023] E10. A method according to any of embodiments E6 - E9, wherein STK11 is wild - type STK11.

[0024] E11. A method according to any of embodiments E6 - E10, further comprising identifying a patient as having a tumor that expresses PD - L1 before administration.

[0025] E12. A method according to any of embodiments E1 - E11, wherein the tumor is derived from lung cancer.

[0026] E13. A method according to embodiment E12, wherein the tumor is derived from small - cell lung cancer (SCLC) or non - small - cell lung cancer (NSCLC).

[0027] E14. A method according to embodiment E13, wherein the tumor is derived from NSCLC.

[0028] E15. A method according to any of embodiments E1 and E12 - E14, wherein diffuse - pattern PD - L1 expression is characterized by a PD - L1 H - score of about 60 - about 500, about 80 - about 480, about 100 - about 460, about 120 - about 440, about 140 - about 420, about 160 - about 400, about 180 - about 380, about 200 - about 360, about 200 - about 340, about 200 - about 320, or about 200 - about 300.

[0029] E16. A method according to any of embodiments E1 and E12 - E15, wherein diffuse - pattern PD - L1 expression is characterized by a PD - L1 H - score of at least about 60, at least about 70, at least about 80, at least about 90, at least about 100, at least about 110, at least about 120, at least about 130, at least about 140, at least about 150, at least about 160, at least about 170, at least about 180, at least about 190, at least about 200, at least about 225, at least about 250, at least about 275, or at least about 300.

[0030] The method of embodiment 15, wherein the PD-L1 expression in a diffuse pattern is characterized by at least about 200 PD-L1 H scores.

[0031] The method of any one of embodiments E1 and E12 - E14, wherein the PD-L1 expression in a heterogeneous pattern is characterized by a PD-L1 H score of about 1 to about 50, about 5 to about 45, about 10 to about 40, or about 15 to about 35, wherein the PD-L1 expression is restricted to one or more different sites of the tumor.

[0032] The method of embodiment E18, wherein the PD-L1 expression in a heterogeneous pattern is characterized 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 35, or at least about 40 PD-L1 H scores.

[0033] The method of embodiment E19, wherein the PD-L1 expression in a heterogeneous pattern is characterized by at least about 15 PD-L1 H scores.

[0034] The method of any one of embodiments E1 - E5 and E12 - E20, wherein the PD-L1 expression pattern is determined using an immunohistochemistry (IHC) assay.

[0035] The method of embodiment E21, wherein the IHC assay is an automated IHC assay.

[0036] The method of embodiment E21 or E22, wherein the IHC assay is performed using an anti-PD-L1 monoclonal antibody that specifically binds to PD-L1, wherein the anti-PD-L1 monoclonal antibody is selected from the group consisting of 28 - 8, 28 - 1, 28 - 12, 29 - 8, 5H1, and any combination thereof.

[0037] The method of any one of embodiments E6 - E14, wherein the expression of STK11 is determined by the presence of STK11 mRNA, the presence of STK11 protein, or the detection of both.

[0038] E25. The method of embodiment E24, wherein the presence of STK11 mRNA is determined using reverse transcriptase PCR.

[0039] E26. The method of embodiment E24, wherein the presence of STK11 protein is determined using an IHC assay.

[0040] E27. The method of embodiment E26, wherein the IHC assay is an automated IHC assay.

[0041] E28. The method of embodiment E26 or E27, wherein the IHC assay is performed using an anti-STK11 monoclonal antibody that specifically binds to STK11.

[0042] E29. The method of any one of embodiments E1 - E6 and E11 - E23, characterized by having tumor cells that express at least about 1%, at least about 2%, at least about 3%, at least about 4%, 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 35%, at least about 40%, at least about 45%, at least about 50%, at least about 55%, at least about 60%, at least about 65%, 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 PD-L1.

[0043] The method of any one of embodiments E7-E14, E24-E28, and E30, characterized by having tumor cells expressing STK11 of at least about 1%, at least about 2%, at least about 3%, at least about 4%, 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 35%, at least about 40%, at least about 45%, at least about 50%, at least about 55%, at least about 60%, at least about 65%, 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%.

[0044] E31. The method of any one of embodiments E1-E30, wherein the tumor exhibits high inflammation.

[0045] E32. The method of embodiment E31, wherein the inflammation is measured by the expression of STK11.

[0046] E33. The method of any one of embodiments E1-E32, wherein the anti-PD-1 antibody cross-competes with nivolumab for binding to human PD-1.

[0047] E34. The method of any one of embodiments E1-E33, wherein the anti-PD-1 antibody binds to the same epitope as nivolumab.

[0048] E35. The method of any one of embodiments E1-E34, wherein the anti-PD-1 antibody is a chimeric, humanized or human monoclonal antibody or a part thereof.

[0049] E36. The method of any one of embodiments E1-E35, wherein the anti-PD-1 antibody comprises a heavy chain constant region of a human IgG1 or IgG4 isotype.

[0050] E37. The method of any one of embodiments E1-E36, wherein the anti-PD-1 antibody is nivolumab.

[0051] The method of any one of embodiments E1 - E37, wherein the anti - PD - 1 antibody is pembrolizumab.

[0052] The method of any one of embodiments E1 - E38, wherein the anti - PD - 1 antibody is administered at a dose in the range of at least about 0.1 mg / kg to at least about 10.0 mg / kg body weight, once every about 1 week, 2 weeks or 3 weeks.

[0053] The method of embodiment E39, wherein the anti - PD - 1 antibody is administered at a dose of at least about 3 mg / kg body weight, once every about 2 weeks.

[0054] The method of any one of embodiments E1 - E38, wherein the anti - PD - 1 antibody or its antigen - binding portion is administered at a uniform dose.

[0055] The method of any one of embodiments E1 - E38 and E41, wherein the anti - PD - 1 antibody or its antigen - binding portion is administered at a uniform dose of at least about 200 mg, at least about 220 mg, at least about 240 mg, at least about 260 mg, at least about 280 mg, at least about 300 mg, at least about 320 mg, at least about 340 mg, at least about 360 mg, at least about 380 mg, at least about 400 mg, at least about 420 mg, at least about 440 mg, at least about 460 mg, at least about 480 mg, at least about 500 mg or at least about 550 mg.

[0056] The method of any one of embodiments E1 - E38, E41 and E42, wherein the anti - PD - 1 antibody or its antigen - binding portion is administered at a uniform dose or at about 240 mg.

[0057] The method of any one of embodiments E1 - E38, E41 and E42, wherein the anti - PD - 1 antibody or its antigen - binding portion is administered at a uniform dose once every about 1 week, 2 weeks, 3 weeks or 4 weeks.

[0058] E45. A method of any one of embodiments E1 - E44, wherein the anti - PD - 1 antibody is administered as long as clinical utility is observed or until untreatable toxicity or disease progression occurs.

[0059] E46. A method of any one of embodiments E1 - E45, wherein the anti - PD - 1 antibody is formulated for intravenous administration.

[0060] E47. A method of any one of embodiments E1 - E46, wherein the anti - PD - 1 antibody is administered at a sub - therapeutic dose.

[0061] E48. A method of any one of embodiments E1 - E47, wherein the administration treats a tumor.

[0062] E49. A method of any one of embodiments E1 - E48, wherein the administration reduces the size of a tumor.

[0063] E50. The method of embodiment E49, wherein the size of the tumor is reduced by at least about 10%, about 20%, about 30%, about 40% or about 50% compared to the tumor size before administration.

[0064] E51. A method of any one of embodiments E1 - E50, wherein the subject shows 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 the start of administration.

[0065] E52. A method of any one of embodiments E1 - E51, wherein the subject shows disease stability after administration.

[0066] E53. A method of any one of embodiments E1 - E51, wherein the subject shows a partial response after administration.

[0067] E54. A method of any one of embodiments E1 - E51, wherein the subject shows a complete response after administration.

[0068] Kit for treating a subject having a tumor, comprising: (a) a dose of an anti-PD-1 antibody in the range of about 4 mg to about 500 mg; and (b) instructions for using the anti-PD-1 antibody in any of the methods of embodiments E1-E54 Kit.

[0069] Kit of embodiment E55, further comprising an anti-PD-L1 antibody.

[0070] Kit of embodiment E55 or E56, further comprising an anti-STK11 antibody. BRIEF DESCRIPTION OF THE DRAWINGS

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[0082] Detailed Description of the Invention The present invention relates to a method of treating a subject having a tumor, comprising: (i) determining the expression pattern of programmed death ligand 1 (PD-L1), and (ii) if the tumor exhibits a pan-pattern of PD-L1 expression, administering to the subject an antibody or antigen-binding portion thereof that specifically binds to the programmed death-1 (PD-1) receptor and inhibits PD-1 activity (“anti-PD-1 antibody”). In other aspects, the present invention relates to a method of treating a subject having a tumor, comprising: (i) identifying a subject having an STK11-positive tumor, and (ii) administering an anti-PD-1 antibody to the subject. In certain embodiments, the tumor is derived from NSCLC.

[0083] Terms To facilitate a better understanding of the present invention, several terms are first defined. As used herein, unless otherwise expressly indicated herein, each of the following terms has the following meaning. Further definitions are set forth throughout the specification.

[0084] "Administering" refers to physically introducing a composition comprising a therapeutic agent to a subject using any of a variety of methods and delivery systems known to those of skill in the art. Routes of administration of the anti-PD-1 antibody include, for example, intravenous, intramuscular, subcutaneous, intraperitoneal, spinal or other parenteral routes of administration by injection or infusion. As used herein, the term "parenteral administration" generally means a route of administration other than enteral and topical administration by injection, and includes, but is not limited to, intravenous, intramuscular, intraarterial, intrathecal, intralymphatic, intralesional, intracapsular, intraorbital, intracardiac, intradermal, intraperitoneal, transtracheal, subcutaneous, subepidermal, intraarticular, subcapsular, subarachnoid, intraspinal, epidural and intrasternal injection and infusion, as well as in vivo electroporation. In certain embodiments, the combination is administered by a route that is not parenteral, and in certain embodiments, orally. Other non-parenteral routes include topical, epithelial or mucosal routes of administration, such as intranasal, vaginal, rectal, sublingual or topical. Administration may also be carried out, for example, once, multiple times and / or over an extended period of time more than once.

[0085] As used herein, “adverse event” (AE) is any undesirable, generally unintended or unwanted sign (including the above examination findings), symptom or disease associated with the use of a medical treatment. For example, an adverse event may be related to immune system activation or increased immune system cells (e.g., T cells) in response to a treatment. A medical treatment may have one or more associated AEs, and the severity level of each AE may be the same or different. A description of a method that can “alter an adverse event” means a treatment regimen that reduces the incidence and / or severity of one or more AEs associated with the use of different treatment regimens.

[0086] “Antibody” (Ab) should include, but is not limited to, a glycoprotein immunoglobulin or an antigen-binding portion thereof that specifically binds to an antigen and contains at least two heavy (H) chains and two light (L) chains interconnected by disulfide bonds. Each H chain includes a heavy chain variable region (abbreviated herein as V H and a heavy chain constant region. The heavy chain constant region includes at least three constant domains, C H1 , C H2 and C H3 . Each light chain includes a light chain variable region (abbreviated herein as V L and a light chain constant region. The light chain constant region includes one constant domain, C L . V H and V L regions can be further subdivided into regions of hypervariability interspersed with more conserved regions referred to as framework regions (FR). Each V H and V L includes 3 CDRs and 4 FRs, arranged in the following order from the amino terminus to the carboxy terminus: FR1, CDR1, FR2, CDR2, FR3, CDR3, FR4. The variable regions of the heavy and light chains include a binding domain that interacts with an antigen. The constant region 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 pathway.

[0087] 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 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, by way of example, both 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 immunogenicity in humans. Unless otherwise specified and unless the context indicates otherwise, the term "antibody" also includes any antigen-binding fragment or antigen-binding portion of said immunoglobulins, including monovalent and bivalent fragments or portions and single-chain antibodies.

[0088] "Isolated antibody" refers to an antibody that is substantially free of other antibodies having different antigen specificities (e.g., an isolated antibody that specifically binds to PD-1 is substantially free of antibodies that specifically bind to antigens other than PD-1). However, an isolated antibody that specifically binds to PD-1 may have cross-reactivity with other antigens, such as PD-1 molecules from different species. Furthermore, an isolated antibody may be substantially free of other cellular materials and / or chemical substances.

[0089] The term "monoclonal antibody" (mAb) refers to a preparation of antibody molecules of a single molecular composition that do not occur naturally, i.e., antibody molecules having essentially the same primary sequence and exhibiting a single binding specificity and affinity for a particular epitope. Monoclonal antibodies are examples of isolated antibodies. Monoclonal antibodies can be produced by hybridoma, recombinant, transgenic, or other techniques known to those skilled in the art.

[0090] "Human antibody" (HuMAb) refers to an antibody having a variable region in which both the FR and CDR are derived from human germline immunoglobulin sequences. Further, if the antibody includes a constant region, the constant region is also derived from human germline immunoglobulin sequences. The human antibodies of the present invention may include amino acid residues not encoded by human germline immunoglobulin sequences (for example, mutations are introduced by random or site-directed mutagenesis in vitro or by somatic mutation in vivo). However, the term "human antibody" as used herein is not intended to include antibodies in which CDR sequences from other mammalian species such as mice are grafted onto human framework sequences. The terms "human" antibody and "fully human" antibody are used synonymously.

[0091] "Humanized antibody" refers to an antibody in which some, most, or all of the amino acids outside the CDRs of a non-human antibody have been replaced with the corresponding amino acids derived from human immunoglobulins. In certain embodiments of the humanized form of the antibody, some, most, or all of the amino acids outside the CDRs have been replaced with human immunoglobulins, while some, most, or all of the amino acids within one or more CDRs remain unchanged. Minor additions, deletions, insertions, substitutions, or modifications of amino acids are tolerated as long as they do not abrogate the ability of the antibody to bind a particular antigen. "Humanized" antibodies maintain antigen specificity similar to the original antibody.

[0092] "Chimeric antibody" refers to an antibody in which the variable region is from one species and the constant region is 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.

[0093] "Antigen-binding antibody" refers to an antibody that specifically binds to an antigen. For example, an anti-PD-1 antibody specifically binds to PD-1.

[0094] The "antigen-binding portion" of an antibody (also referred to as an "antigen-binding fragment") refers to one or more fragments of the antibody that retain the ability to specifically bind to the antigen bound by the whole antibody.

[0095] "Cancer" refers to a broad group of various diseases characterized by the uncontrolled growth of abnormal cells in the body. The uncontrolled cell division and growth of cells result in the formation of malignant tumors that can invade neighboring tissues and metastasize to distant sites via the lymphatic system or bloodstream. In certain embodiments, the cancer is any of the cancers disclosed herein. In certain embodiments, the cancer is lung cancer. In certain embodiments, the lung cancer is non-small cell lung cancer (NSCLC). In certain embodiments, the NSCLC has squamous histology (squamous NSCLC). In other embodiments, the NSCLC has non-squamous histology (non-squamous NSCLC). "Cancer" may include tumors. "Tumor" includes all neoplastic cell growth and proliferation and all precancerous and cancerous cells and tissues, regardless of whether they are malignant or benign.

[0096] "Serine / threonine kinase 11" or "STK11" (also referred to as "polarity-related protein LKB1", "renal carcinoma antigen NY-REN-19", "liver kinase B1", "EC 2.7.11.1", and "HLKB1") refers to a member of the serine / threonine kinase family that controls cell polarity and functions as a tumor suppressor. STK11 controls the activity of members of the AMP-activated protein kinase (AMPK) family, thereby playing a role in various processes such as cell metabolism, cell polarity, apoptosis, and DNA damage response. STK11 is ubiquitously expressed, with the strongest expression in the testis and fetal liver. STK11 is generally inactivated in NSCLC, particularly in tumors with KRAS mutations. As described herein, mutant STK11, e.g., loss of expression of wild-type STK11, is associated with a decrease or abnormality in PD-L1 expression in tumors derived from SCLC. In certain embodiments, mutant STK11, e.g., loss of expression of wild-type STK11, occurs in tumors derived from SCLC, where the tumor expresses or does not express wild-type KRAS (e.g., the tumor has or does not have a KRAS mutation). In certain embodiments, the STK11 variant is, for example, the STK11 variant previously described in Koyama et al., Cancer Res. 76(5):999-1008 (2016) and / or Skoulidis et al., Cancer Discov. 5(8):860-77 (2015), both of which are incorporated herein by reference in their entirety.

[0097] The term "immunotherapy" refers to the treatment of a subject having, at risk of recurrence, or having a recurrence of a disease by a method that includes inducing, enhancing, suppressing, or otherwise modifying an immune response. "Treatment" or "therapy" of a subject refers to any type of intervention, procedure, or administration of an agent carried out on the subject for the purpose of restoring, reducing, alleviating, arresting, delaying, or preventing the onset, progression, occurrence, severity, or recurrence of symptoms, complications, conditions, or biochemical markers associated with the disease.

[0098] As used herein, "PD-L1 positive" can be used interchangeably with "at least about 1% PD-L1 expression". In certain embodiments, PD-L1 expression can be measured by any method known in the art. In other embodiments, PD-L1 expression is measured by automated IHC. A PD-L1 positive tumor, therefore, has, as measured by automated IHC, 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% PD-L1 expressing tumor cells. In certain embodiments, "PD-L1 positive" means that there are at least 100 cells expressing PD-L1 on the cell surface.

[0099] "Programmed death-1" (PD-1) refers to an immunosuppressive receptor belonging to the CD28 family. PD-1 is present primarily on activated T cells in vivo and binds to two ligands, PD-L1 and PD-L2. The term "PD-1" as used herein includes human PD-1 (hPD-1), variants, isoforms and species homologs of hPD-1 and analogs having at least one common epitope with hPD-1. The complete hPD-1 sequence can be found in GenBank Accession No. U64863.

[0100] "Programmed death ligand-1 (PD-L1)" is one of two cell surface glycoprotein ligands for PD-1 (the other being PD-L2) that downregulates T cell activation and cytokine secretion upon binding to PD-1. The term "PD-L1" as used herein includes human PD-L1 (hPD-L1), variants, isoforms and species homologs of hPD-L1 and analogs having at least one common epitope with hPD-L1. The complete hPD-L1 sequence can be found under GenBank Accession No. Q9NZQ7.

[0101] "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 certain embodiments, the subject is human. The terms "subject" and "patient" are used interchangeably herein.

[0102] A "therapeutically effective amount" or "therapeutically effective dose" of a drug or therapeutic agent is any amount of the drug that, when used alone or in combination with other therapeutic agents, protects the subject against the development of a disease or promotes regression of the disease, as evidenced by a reduction in the severity of the symptoms of the disease, an increase in the frequency and duration of disease-free symptoms, or a prevention of dysfunction or disability due to the disease. The ability of a therapeutic agent to promote regression of a disease can be evaluated using a variety of methods known to those of skill in the art, such as in human subjects during clinical trials, in animal model systems that predict efficacy in humans, or by assays of the activity of the agent in in vitro assays.

[0103] As used herein, "sub-therapeutic dose" means a dose of a therapeutic compound (e.g., an antibody) that is lower than the normal or typical dose of the therapeutic compound when used alone in the treatment of a proliferative disorder (e.g., cancer).

[0104] By way of example, an "anticancer agent" promotes regression of cancer in a subject or inhibits further tumor growth. In certain embodiments, a therapeutically effective amount of a drug promotes regression of cancer to the point of eliminating the cancer. "Promoting regression of cancer" means that administration of an effective amount of the drug, alone or in combination with an antineoplastic agent, results in a decrease in tumor growth or size, tumor necrosis, a reduction in the severity of at least one disease symptom, an increase in the frequency and duration of disease-free symptoms, or a prevention of dysfunction or disability due to the disease. Further, the terms "effective" and "efficacy" as related to treatment include both pharmacological efficacy and physiological safety. Pharmacological efficacy refers to the ability of the drug to promote regression of cancer in a patient. Physiological safety refers to the level of toxicity or other adverse physiological effects (adverse effects) at the cellular, organ, and / or organism level resulting from drug administration.

[0105] As an example of tumor treatment, a therapeutically effective amount of an anti-cancer agent can inhibit cell growth or tumor growth by at least about 10%, at least about 20%, at least about 40%, at least about 60%, at least about 70%, at least about 80%, at least about 90%, at least about 95% or at least about 100% compared to an untreated subject. In other embodiments of the present invention, tumor regression can be observed and continue for a period of at least about 20 days, at least about 30 days, at least about 40 days, at least about 50 days or at least about 60 days. Even with these ultimate assessments of therapeutic efficacy, the evaluation of immunotherapeutic agents must also take into account the "immune-related response pattern".

[0106] "Immune-related response pattern" refers to a clinical response pattern often observed in cancer patients treated with immunotherapeutic agents that produce an anti-tumor effect by inducing a cancer-specific immune response or modifying the innate immune process. This response pattern, in the evaluation of conventional chemotherapeutic agents, is classified as disease progression and is synonymous with drug ineffectiveness, characterized by an initial increase in tumor burden or the appearance of new lesions followed by a beneficial therapeutic effect. Therefore, an appropriate evaluation of immunotherapeutic agents may require long-term monitoring of the effects of these agents on the target disease. A therapeutically effective amount of a drug is any amount of a drug that prevents the onset or recurrence of cancer when administered alone or in combination with an anti-neoplastic agent to a subject at risk of developing cancer (e.g., a subject with a pre-malignant condition) or a subject with a recurrence of cancer, including a "preventively effective amount". In certain embodiments, a preventively effective amount completely prevents the onset or recurrence of cancer. "Prevention" of the onset or recurrence of cancer means a reduction in the likelihood of cancer onset or recurrence or a complete prevention of cancer onset or recurrence.

[0107] The use of alternatives (e.g., "or") should be construed to mean one, both, or any combination of these alternatives. As used herein, singular expressions should be construed to mean "one or more" of any recited or enumerated components.

[0108] The terms "about" or "essentially comprising" refer to a value or composition within an acceptable error range of a particular value or composition, as determined by one of ordinary skill in the art, which depends in part on how the value or composition is measured or determined, i.e., on the limitations of the measuring system. For example, "about" or "essentially comprising" means within one or more standard deviations according to the practice in the art. Alternatively, "about" or "essentially comprising" can mean a range 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 any number between 2.4 mg and 3.6 mg (for 20%). Further, especially with respect to biological systems or processes, the term can mean up to one order of magnitude or up to five-fold of the value. When a particular value or composition is provided in the present specification and claims, unless otherwise specified, it should be assumed that the meaning of "about" or "essentially comprising" is within the acceptable error range for that particular value or composition.

[0109] As used herein, the terms "about once a week", "about once every two weeks" or any other similar dosing interval mean approximate numbers. "About once a week" can include every 7 days ±1 day, i.e., every 6 days to every 8 days. "About once every two weeks" can include every 14 days ±3 days, i.e., every 11 days to every 17 days. Similar approximations apply, for example, to about once every three weeks, about once every four weeks, about once every five weeks, about once every six weeks and about once every twelve weeks. In certain embodiments, a dosing interval of about once every six weeks or about once every twelve weeks means that the first dose can be on any day of the first week, and then the next dose can be on any day of the sixth or twelfth week, respectively. In other embodiments, a dosing interval of about once every six weeks or about once every twelve weeks means that the first dose is administered on a specific day of the first week (e.g., Monday), and then the next dose is administered on the specific day of the sixth or twelfth week (i.e., Monday), respectively.

[0110] As used herein, the term "weight-based dosage" means that the dosage administered to a patient is calculated based on the patient's weight. For example, when a patient weighing 60 kg requires an anti-PD-1 antibody at 3 mg / kg, the appropriate amount of the anti-PD-1 antibody for administration (i.e., 180 mg) can be calculated and used.

[0111] The use of the term "fixed dosage" with respect to the methods of the present invention means that two or more different antibodies (e.g., an anti-PD-1 antibody and a second antibody) in a single composition are present in a specific (fixed) ratio to each other in the composition. In certain embodiments, the fixed dosage is based on the weight (e.g., mg) of the antibodies. In certain embodiments, the fixed dosage is based on the concentration (e.g., mg / ml) of the antibodies. In certain 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 1:180, about 1:200, about 200:1, about 180:1, about 160:1, about 140:1, about 120:1, about 100:1, about 90:1, about 80:1, about 70:1, about 60:1, about 50:1, about 40:1, about 30:1, about 20:1, about 15:1, about 10:1, about 9:1, about 8:1, about 7:1, about 6:1, about 5:1, about 4:1, about 3:1 or about 2:1 mg of a first antibody (e.g., an anti-PD-1 antibody) to mg of a second antibody. For example, an anti-PD-1 antibody and a second antibody in a 3:1 ratio can mean that a vial can contain about 240 mg of the anti-PD-1 antibody and 80 mg of the second antibody or about 3 mg / ml of the anti-PD-1 antibody and 1 mg / ml of the second antibody.

[0112] The use of the term "uniform dosage" with respect to the methods and dosages of the present invention means that the dosage administered to a patient is independent of the patient's weight or body surface area (BSA). Thus, the uniform dosage is provided as an absolute amount of the agent (e.g., an anti-PD-1 antibody) rather than as a mg / kg dosage. For example, a 60 kg human and a 100 kg human receive the same dosage of the antibody (e.g., 240 mg of the anti-PD-1 antibody).

[0113] Unless otherwise indicated, all concentration ranges, percentage ranges, ratio ranges or integer ranges recited herein are to be construed as including any integer values within the recited range and, if appropriate, fractions thereof (e.g., one tenth and one hundredth of an integer).

[0114] The various aspects of the invention are further detailed in the following subsections.

[0115] The method of the invention The present invention provides a method of treating a subject having a tumor, comprising (i) determining the expression pattern of programmed death ligand 1 (PD-L1), and (ii) administering to the subject an antibody or antigen-binding portion thereof that specifically binds to the programmed death-1 (PD-1) receptor and inhibits PD-1 activity (“anti-PD-1 antibody”) if the tumor exhibits a pan-pattern of PD-L1 expression. In certain aspects, the present invention provides a method of treating a subject having a tumor, comprising (i) determining the expression pattern of PD-L1, and (ii) administering an anti-PD-1 antibody to the subject if the tumor exhibits a heterogeneous pattern of PD-L1 expression. In other aspects, the present invention provides a method of treating a subject having a tumor, comprising (i) determining the expression pattern of PD-L1, and (ii) administering an anti-PD-1 antibody to the subject if the tumor exhibits a tumor-stromal interface pattern of PD-L1 expression. In other aspects, the present invention provides a method of identifying a subject having a tumor suitable for anti-PD-1 antibody treatment, comprising (i) determining the expression pattern of PD-L1, and (ii) administering an anti-PD-1 antibody to the subject if the tumor exhibits a pan-pattern of PD-L1 expression. In yet other aspects, the present invention provides a method of identifying a subject having a tumor suitable for anti-PD-1 antibody treatment, comprising (i) determining the expression pattern of PD-L1, and (ii) administering an anti-PD-1 antibody to the subject if the tumor exhibits a heterogeneous pattern of PD-L1 expression. In certain embodiments, the methods described herein further comprise identifying a patient as having a tumor that expresses STK11 prior to administration.

[0116] In other aspects, the invention relates to a method of treating a subject having a tumor, comprising (i) identifying a subject having an STK11-positive tumor (e.g., STK11 wild-type), and (ii) administering an anti-PD-1 antibody to the subject. In certain aspects, the invention relates to a method of treating a subject having a tumor, comprising administering an anti-PD-1 antibody, wherein the patient has been identified as having an STK11-positive tumor prior to administration. In certain aspects, the invention relates to a method of identifying a subject having a tumor suitable for anti-PD-1 antibody treatment, comprising (i) measuring STK11 expression in the tumor, and (ii) administering an anti-PD-1 antibody to the subject if the tumor is STK11-positive. In certain embodiments, STK11 is wild-type STK11.

[0117] In other aspects, the invention relates to a method of treating a subject having a tumor, comprising (i) identifying a subject having an STK11-negative tumor, and (ii) not administering an anti-PD-1 antibody to the subject or enhancing anti-PD-1 antibody therapy. Other aspects of the invention relate to a method of identifying a subject having a tumor unsuitable for anti-PD-1 antibody treatment, comprising (i) measuring STK11 expression in the tumor, and (ii) not administering an anti-PD-1 antibody to the subject or enhancing anti-PD-1 antibody therapy if the tumor is STK11-negative or the tumor expresses an inactive STK11 variant.

[0118] In certain embodiments, the tumor is derived from NSCLC. In certain embodiments, the subject is a human patient. In certain embodiments, the subject is a chemotherapy-naive patient (e.g., a patient who has not previously received any chemotherapy agent). In other embodiments, the subject for the combination therapy has received other cancer treatments (e.g., chemotherapy agents), but is resistant or refractory to such other cancer treatments. In certain specific embodiments, the subject for the treatment has tumor cells that express mutant forms of the EGFR, KRAS, and / or STK11 genes. In certain embodiments, the subject for the treatment has tumor cells that express both wild-type STK11 and mutant STK11. In other embodiments, the subject for the treatment has tumor cells that express only the wild-type form of STK11. In certain embodiments, the tumor expresses one or more genes selected from the group consisting of TP53, KEAP1, KRAS, EGFR, MET, and one or more variant variants thereof. In certain embodiments, the tumor expresses STK11 and one or more genes selected from TP53, KEAP1, KRAS, EGFR, MET, and one or more variant variants thereof.

[0119] In certain embodiments, the subject has tumor cells that are PD-L1 positive (PD-L1+). In certain embodiments, the subject has cancer cells that are PD-L1 negative (PD-L1-). In certain embodiments, the subject has never smoked. In certain embodiments, the subject has previously smoked. In certain embodiments, the subject is currently smoking. In certain embodiments, the subject has cancer cells that are squamous. In certain embodiments, the subject has cancer cells that are non-squamous.

[0120] In certain embodiments, diffuse pattern of PD-L1 expression is characterized by a PD-L1 H score of about 60 to about 500, about 70 to about 490, about 80 to about 480, about 90 to about 470, about 100 to about 460, about 110 to about 450, about 120 to about 440, about 130 to about 430, about 140 to about 420, about 150 to about 410, about 160 to about 400, about 170 to about 390, about 180 to about 380, about 190 to about 370, about 200 to about 360, about 200 to about 350, about 200 to about 340, about 200 to about 330, about 200 to about 320, about 200 to about 310 or about 200 to about 300. In certain embodiments, diffuse pattern of PD-L1 expression is characterized by a PD-L1 H score of at least about 60, at least about 70, at least about 80, at least about 90, at least about 100, at least about 110, at least about 120, at least about 130, at least about 140, at least about 150, at least about 160, at least about 170, at least about 180, at least about 190, at least about 200, at least about 210, at least about 220, at least about 225, at least about 230, at least about 240, at least about 250, at least about 260, at least about 270, at least about 275, at least about 280, at least about 290 or at least about 300. In certain embodiments, diffuse pattern of PD-L1 expression is characterized by a PD-L1 H score of at least about 200. In other embodiments, diffuse pattern of PD-L1 expression is characterized by a PD-L1 H score of at least about 240. In certain embodiments, diffuse pattern of PD-L1 expression is characterized by a PD-L1 H score of at least about 260.

[0121] In certain embodiments, the heterogeneous pattern of PD-L1 expression is characterized by a PD-L1 H score of about 1 to about 50, about 5 to about 45, about 10 to about 40, or about 15 to about 35, where the PD-L1 expression is restricted to one or more different sites of the tumor. In certain embodiments, the heterogeneous pattern of PD-L1 expression is characterized by a PD-L1 H score of 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 35, or at least about 40. In certain embodiments, the heterogeneous pattern of PD-L1 expression is characterized by a PD-L1 H score of at least about 15. In other embodiments, the heterogeneous pattern of PD-L1 expression is characterized by a PD-L1 H score of at least about 20. In certain embodiments, the heterogeneous pattern of PD-L1 expression is characterized by a tumor comprising at least 10, at least 20, at least 30, at least 40, at least 50, at least 60, at least 70, at least 80, at least 90, or 100, at least 120, or at least 150 PD-L1-expressing portions. In certain embodiments, at least about 30%, at least about 40%, at least about 50%, at least about 60%, at least about 70%, at least about 80%, at least about 90%, at least about 95%, or about 100% of the cells within a portion of the tumor express PD-L1.

[0122] In certain embodiments, the tumor-stromal interface PD-L1 expression is characterized by the expression of PD-L1 by tumor cells adjacent to the stroma (e.g., within about 1 cell diameter, about 2 cell diameters, about 3 cell diameters, about 4 cell diameters, about 5 cell diameters, about 6 cell diameters, about 7 cell diameters, about 8 cell diameters, about 9 cell diameters, or about 10 cell diameters). In certain embodiments, the tumor-stromal interface PD-L1 expression is characterized by the PD-L1 expression on the tumor surface.

[0123] In certain embodiments, the treatment of the present invention (e.g., administration of an anti-PD-1 antibody) effectively prolongs the survival period of a subject. In certain embodiments, the treatment of the anti-PD-1 antibody of the present invention prolongs the progression-free survival of a subject. In certain embodiments, the treatment of the anti-PD-1 antibody of the present invention prolongs the progression-free survival of a subject as compared to a standard therapeutic agent. After administration of the anti-PD-1 antibody treatment, a subject having a tumor may exhibit an overall survival period of at least about 10 months, at least about 11 months, at least about 12 months, at least about 13 months, at least about 14 months, at least about 15 months, at least about 16 months, at least about 17 months, at least about 18 months, at least about 19 months, at least about 20 months, at least about 21 months, at least about 22 months, at least about 23 months, at least about 2 years, at least about 3 years, at least about 4 years or at least about 5 years after administration.

[0124] In other embodiments, the survival period or overall survival period of a subject is extended 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 as compared to other subjects treated with only a standard therapeutic agent (e.g., docetaxel) or with a treatment of a different dosing schedule. For example, the survival period or overall survival period of a subject treated with an anti-PD-1 antibody disclosed herein is extended 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% as compared to other subjects treated with only a standard therapeutic agent (e.g., docetaxel) or with a different dosing schedule of combination therapy.

[0125] In certain embodiments, the treatment of the present invention effectively extends the duration of progression-free survival of a subject. In certain embodiments, the subject exhibits 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.

[0126] The PD-L1 or STK11 status of the tumor in a subject can be measured before administration of any of the compositions disclosed herein or before use of any of the methods. In certain embodiments, the PD-L1 or STK11 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 other embodiments, the PD-L1 or STK11 status of the tumor is at least about 1%. In other embodiments, the PD-L1 or STK11 status of the subject is at least about 5%. In certain embodiments, the PD-L1 or STK11 status of the tumor is at least about 10%. In certain embodiments, the PD-L1 or STK11 status of the tumor is at least about 25%. In certain embodiments, the PD-L1 status of the tumor is at least about 50%.

[0127] In certain embodiments, tumors may exhibit high levels of inflammation. Increased inflammation may be an indicator of a pan-PD-L1 expression pattern. Thus, high tumor inflammation may be an indicator of responsiveness to anti-PD-1 antibody therapy. In certain embodiments, inflammation may be measured by the expression of STK11, PD-L1, TP53, KEAP1, KRAS, EGFR, and / or MET.

[0128] In certain embodiments, the median progression-free survival of a subject having a tumor with ≧1% PD-L1 expression 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 a subject having a tumor with <1% PD-L1 expression. In certain embodiments, the progression-free survival of a subject having use of ≧1% PD-L1 expression 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.

[0129] In certain embodiments, administration of the anti-PD-1 antibody treats the tumor. In certain embodiments, the administration reduces the size of the tumor. In certain embodiments, the size of the tumor is reduced by at least about 10%, about 20%, about 30%, about 40% or about 50% compared to the tumor size before administration. In other embodiments, the subject exhibits 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 the start of administration. In certain embodiments, the subject exhibits disease stability after administration. In certain embodiments, the subject exhibits a partial response after administration. In certain embodiments, the subject exhibits a complete response after administration. In certain embodiments, the subject shows an improvement in objective response rate (ORR) after administration compared to a subject treated with standard therapy.

[0130] In certain embodiments, test tissue samples can be obtained from a patient in need of treatment in order to evaluate PD-L1 expression and / or STK11 expression. In other embodiments, evaluation of PD-L1 and / or STK11 expression can be achieved without obtaining test tissue samples. In certain embodiments, appropriate patient selection includes (i) preparing a test tissue sample obtained from the tissue of a patient having cancer, if desired, the test tissue sample comprising tumor cells and / or tumor-infiltrating inflammatory cells; and (ii) evaluating the percentage of cells expressing PD-L1 on the cell surface in the test tissue sample, based on the percentage of cells expressing PD-L1 on the cell surface in the test tissue sample being higher than a predetermined threshold level.

[0131] However, in any of the methods involving the measurement of PD-L1 and / or STK11 expression in a test tissue sample, it should be understood that the step involving the supply of the test tissue sample obtained from the patient is an optional step. In certain embodiments, the "measurement" or "evaluation" step for identifying or determining the number or percentage of cells expressing PD-L1 and / or STK11 (e.g., PD-L1 expression on the cell surface) in the test tissue sample is carried out by, for example, performing a reverse transcriptase-polymerase chain reaction (RT-PCR) assay or an IHC assay, or by a transformative method of assaying PD-L1 and / or STK11 expression. In certain other embodiments, the transformative method step is not included and PD-L1 and / or STK11 expression is evaluated, for example, by reviewing a test result report from a laboratory. In certain embodiments, the steps of the method up to and including the evaluation of PD-L1 and / or STK11 expression provide intermediate results that can be used by a physician or other healthcare provider to select suitable candidates for anti-PD-1 antibody or anti-PD-L1 antibody treatment. In certain embodiments, the step of providing the intermediate report is carried out by a physician or someone working under the physician's direction. In other embodiments, these steps are carried out at an independent laboratory or by an independent person such as a medical technologist.

[0132] In certain embodiments of the method, the percentage of cells expressing PD-L1 and / or STK11 is evaluated by performing an assay to determine the presence of PD-L1 and / or STK11 RNA. In further embodiments, the presence of PD-L1 and / or STK11 RNA is determined by RT-PCR, in situ hybridization or RNase protection. In other embodiments, the percentage of cells expressing PD-L1 and / or STK11 is evaluated by performing an assay to determine the presence of PD-L1 and / or STK11 polypeptide. In further embodiments, the presence of PD-L1 and / or STK11 polypeptide is determined by immunohistochemistry (IHC), enzyme-linked immunosorbent assay (ELISA), in vivo imaging or flow cytometry. In certain embodiments, PD-L1 and / or STK11 expression is assayed by IHC. In other embodiments of all of these methods, cell surface expression of PD-L1 and / or STK11 is assayed using, for example, IHC or in vivo imaging.

[0133] Imaging techniques provide 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 imaging (FRI), fluorescence-mediated tomography (FMT), bioluminescence imaging (BLI), confocal laser scanning microscopy (LSCM), and multiphoton microscopy (MPM), may herald the advent of even wider use of these techniques in cancer research. Some of these molecular imaging systems enable physicians not only to see where tumors are in the body, but also to visualize the expression and activity of specific molecules, cells, and biological processes that affect tumor behavior and / or response to therapeutic agents (Condeelis and Weissleder, "In vivo imaging in cancer," Cold Spring Harb. Perspect. Biol. 2(12):a003848 (2010)). The antibody specificity combined with the sensitivity and resolution of PET results in immunoPET imaging, which is particularly attractive for monitoring and assaying 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 certain embodiments of any of the methods, PD-L1 and / or STK11 expression is assayed by immunoPET imaging. In certain embodiments of any of the methods, the percentage of cells expressing PD-L1 and / or STK11 in a test tissue sample is evaluated by performing an assay to determine the presence of PD-L1 and / or STK11 polypeptide on the cell surface in the test tissue sample. In one embodiment, the test tissue sample is a FFPE tissue sample.In other embodiments, the presence of the PD-L1 and / or STK11 polypeptide is carried out by an IHC assay. In further embodiments, the IHC assay is carried out by an automated process. In certain embodiments, the IHC assay is carried out using an anti-PD-L1 monoclonal antibody that binds to the PD-L1 polypeptide. In other embodiments, the IHC assay is carried out using an anti-STK11 monoclonal antibody for binding to the STK11 polypeptide.

[0134] In certain embodiments of the method, an automated IHC method is used to assay for PD-L1 and / or STK11 expression on the cell surface of FFPE tissue specimens. The present invention provides a method for detecting the presence of human PD-L1 and / or STK11 antigen in a test tissue sample or quantifying the level of human PD-L1 and / or STK11 antigen or the percentage of cells expressing the antigen in the sample, the method comprising contacting the test sample and a negative control sample with a monoclonal antibody that specifically binds to human PD-L1 and / or STK11 under conditions that allow the formation of a complex of the antibody or a portion thereof and human PD-L1 and / or STK11. In certain embodiments, the test and control tissue samples are FFPE samples. Complex formation is then detected, where a difference in complex formation between the test sample and the negative control sample is an indicator of the presence of human PD-L1 and / or STK11 antigen in the sample. Various methods are used for quantifying PD-L1 and / or STK11 expression.

[0135] In certain embodiments, the automated IHC method comprises (a) deparaffinizing and rehydrating a tissue section mounted on an automated stainer, (b) heating at 110° C. for 10 minutes using a decloaking chamber and a pH 6 buffer to retrieve antigens, (c) setting up reagents in the automated stainer, and (d) activating the automated stainer to include the steps of neutralizing endogenous peroxidase in the tissue specimen; blocking non-specific protein binding sites on the slide; incubating the slide with a primary antibody; incubating with a post-primary blocker; incubating with a Novolink polymer; adding and developing a chromogenic substrate; and counterstaining with hematoxylin.

[0136] For the evaluation of PD-L1 and / or STK11 expression in tumor tissue samples, a pathologist examines the membranous PD-L1 + and / or STK11 + in tumor cells in each microscopic field, subjectively estimates the percentage of cells that are positive, and then normalizes to obtain a final percentage. Various staining intensities are defined as 0 / negative, 1+ / weak, 2+ / moderate, and 3+ / strong. Generally, percentage values are first assigned to the 0 and 3+ buckets, and then the intermediate 1+ and 2+ intensities are considered. For highly heterogeneous tissue, the specimen is divided into zones, each zone is scored separately, and then adjusted to one set of percentage values. The percentage of negative and positive cells of various staining intensities is determined from each region, and the median value is given to each zone. A final percentage value is assigned to the tissue for each staining intensity category of negative, 1+, 2+, and 3+. The sum of all staining intensities should be 100%. In certain embodiments, the threshold number of cells required to be PD-L1 and / or STK11 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 certain embodiments, the threshold number of cells required to be PD-L1 and / or STK11 positive is at least about 100 cells.

[0137] The staining is also evaluated in tumor-infiltrating inflammatory cells such as macrophages and lymphocytes. In most cases, macrophages serve as an internal positive control since staining is observed in most macrophages. It is not necessary to be stained at an intensity of 3+, but the absence of macrophage staining should be considered to rule out any technical failure. Macrophages and lymphocytes are evaluated for plasma membrane staining and are recorded for each sample only as positive or negative for each cell category. The staining is also characterized by outer / inner tumor immune cell designation. "Inner" means that immune cells are on the tumor area boundary without being physically intervened within the tumor tissue and / or between tumor cells. "Outer" means that there is no physical connection to the tumor and immune cells are seen in the periphery associated with connective tissue or some related adjacent tissue.

[0138] In some embodiments of these scoring methods, the samples are scored by two pathologists operating independently and the scores are then fixed. In some other embodiments, the identification of positive and negative cells is scored using appropriate software.

[0139] The histoscore (also referred to as the H-score) is used as a more quantitative means of IHC data. The histoscore is calculated as follows. Histoscore = [( % tumor × 1 (low intensity)) + ( % tumor × 2 (medium intensity)) + ( % tumor × 3 (high intensity)

[0140] For histoscore determination, the pathologist estimates the percentage of stained cells in each intensity category in the specimen. Since the expression of most biomarkers is heterogeneous, the histoscore is a true representation of the overall expression. The final histoscore range is 0 (no expression) to 300 (maximum expression).

[0141] Another means of quantifying PD-L1 and / or STK11 expression in test tissue samples by IHC is the determination of an Adaptive Inflammatory Score (AIS), which is defined as the density of inflammation multiplied by the percentage of PD-L1 and / or STK11 expression by 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)).

[0142] This method can treat tumors at all stages. In certain embodiments, the tumor is derived from NSCLC at all stages. There are at least 7 stages of NSCLC for which it can be used: potential (occult) stage, stage 0 (carcinoma in situ), stage I, stage II, stage IIIA, stage IIIB, and stage IV. In the potential stage, the cancer cannot be seen by imaging or bronchoscopy. In stage 0, cancer cells are seen in the lining of the airway.

[0143] In certain embodiments, the method treats stage I non-squamous NSCLC. Stage I NSCLC is divided into stage IA and IB. In stage IA, the tumor is only in the lung and is 3 cm or less. In stage IB, the cancer has not spread to the lymph nodes and one or more of the following apply: 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 junction of the trachea and bronchus; 3) it has spread to the innermost layer of the membrane covering the lung; or 4) part of the lung is collapsed or interstitial pneumonia (inflammation of the lung) has developed in the area where the trachea and bronchus connect.

[0144] In other embodiments, the method of the invention treats stage II non-small cell lung cancer (NSCLC). Stage II NSCLC is divided into stage IIA and IIB. In stage IIA, the cancer has or has not spread to the lymph nodes. If the cancer has spread to the lymph nodes, the cancer can only spread to the lymph nodes on the same side as the chest tumor, the lymph nodes with cancer, or inside the lung or near the bronchus. And one or more of the following apply: 1) The tumor is not larger than 5 cm; 2) The cancer has spread to the main bronchus, at least 2 cm below the junction of the trachea and the bronchus; 3) It has spread to the innermost layer of the membrane covering the lung; or 4) A part of the lung is collapsed or interstitial pneumonia (inflammation of the lung) has developed in the region where the trachea and the bronchus are connected. If the tumor has not spread to the lymph nodes and one or more of the following apply, it is considered stage IIA: 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 junction of the trachea and the bronchus; 3) It has spread to the innermost layer of the membrane covering the lung; or 4) A part of the lung is collapsed or interstitial pneumonia (inflammation of the lung) has developed in the region where the trachea and the bronchus are connected. In stage IIB, the cancer has or has not spread to the lymph nodes. If the cancer has spread to the lymph nodes, the cancer can only spread to the lymph nodes on the same side as the chest tumor, the lymph nodes with cancer are only inside the lung or near the bronchus, and one or more of the following apply: 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 junction of the trachea and the bronchus; 3) It has spread to the innermost layer of the membrane covering the lung; or 4) A part of the lung is collapsed or interstitial pneumonia (inflammation of the lung) has developed in the region where the trachea and the bronchus are connected. If the tumor has not spread to the lymph nodes and one or more of the following apply, it is considered stage IIB: 1) The tumor is larger than 7 cm; 2) The cancer has spread to the main bronchus (and at least 2 cm below the junction of the trachea and the bronchus), the chest wall, the diaphragm or the nerve controlling the diaphragm; 3) The cancer has spread around the heart or to the membrane on the inner layer of the chest wall; 4) The whole lung is collapsed or has developed interstitial pneumonia (inflammation of the lung); or 5) There are one or more metastatic tumors in the same lobe of the lung.

[0145] In other embodiments, any of the methods of the invention treat stage III non-squamous NSCLC. Stage IIIA is divided into three sections. These three sections are based on 1) the size of the tumor; 2) where the tumor is found; and 3) which (if any) lymph nodes have cancer. In the first type of stage IIIA NSCLC, the cancer has spread to lymph nodes on the same side of the chest as the tumor in the lung, and the lymph nodes with cancer are near the sternum or where the bronchus enters the lung. Further: 1) the tumor can be any size; 2) part of the lung (where the trachea connects to the bronchus) or the whole lung may be collapsed or have developed interstitial pneumonia (inflammation of the lung); 3) there may be one or more metastatic tumors in the same lobe of the lung; and 4) the cancer may have spread to any of the following: a) the main bronchus outside the area where the trachea connects to the bronchus, b) the chest wall, c) the diaphragm and the nerve that controls it, d) the membrane around the lung or the inner layer of the chest wall, e) the membrane around the heart. In the second type of stage IIIA NSCLC, the cancer has spread to lymph nodes on the same side of the chest as the tumor in the lung, and the lymph nodes with cancer are in the lung or near the bronchus. Further: 1) the tumor can be any size; 2) the whole lung may be collapsed or have developed interstitial pneumonia (inflammation of the lung); 3) there may be one or more other tumors in any of the lobes of the lung with cancer; and 4) the cancer may have spread to any of the following: a) the main bronchus, not in the area where the trachea connects to the bronchus, b) the chest wall, c) the diaphragm and the nerve that controls it, d) the membrane around the lung or the inner layer of the chest wall, e) the heart or the membrane around it, f) the major blood vessels to or from the heart, g) the trachea, h) the esophagus, i) the nerve that controls the larynx (voice box), j) the sternum (bone in the chest) or the backbone or k) the tracheal bifurcation (where the trachea connects to the bronchi). In the third type of stage IIIA NSCLC, the cancer has not spread to the lymph nodes, the tumor can be any size, and the cancer has spread to any one of the following: a) the heart, b) the major blood vessels to or from the heart, c) the trachea, d) the esophagus, e) the nerve that controls the larynx (voice box), f) the sternum (bone in the chest) or the backbone or g) the tracheal bifurcation (where the trachea connects to the bronchi). Stage IIIB is divided into two sections based on 1) the size of the tumor, 2) where the tumor is found, and 3) which lymph nodes have cancer.In the first type of stage IIIB NSCLC, the cancer has spread to lymph nodes on the side opposite the chest tumor. Additionally, 1) the tumor can be of any size; 2) part of the lung (where the trachea connects to the bronchi) or the entire lung may be collapsed or have developed interstitial pneumonia (inflammation of the lung); 3) one or more additional tumors may be present in any of the lobes of the lung with the 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 nerve controlling it, d) the membrane around the lung or the inner layer of the chest wall, e) the heart or the membrane around it, f) the major blood vessels to or from the heart, g) the trachea, h) the esophagus, i) the nerve controlling the larynx (voice box), j) the breastbone (bone of the chest) or the backbone or k) the tracheal bifurcation (where the trachea connects to the bronchi). In the second type of stage IIIB NSCLC, the cancer has spread to lymph nodes on the same side as the chest tumor. The lymph nodes with cancer are near the breastbone (bone of the chest) or where the bronchi enter the lungs. Additionally, 1) the tumor can be of any size; 2) additional tumors may be present in different lobes of the same lung; and 3) the cancer may have spread to any of the following: a) the heart, b) the major blood vessels to or from the heart, c) the trachea, d) the esophagus, e) the nerve controlling the larynx (voice box), f) the breastbone (bone of the chest) or the backbone or g) the tracheal bifurcation (where the trachea connects to the bronchi).

[0146] In certain embodiments, the methods of the invention treat stage IV non-squamous NSCLC. In stage IV NSCLC, the tumor can be of any size and the cancer may have spread to lymph nodes. In stage IV NSCLC, one or more of the following apply: 1) there is one or more tumors in both lungs; 2) the cancer is found in the body fluids around the lung or the heart; and 3) the cancer has spread to other parts of the body such as the brain, liver, adrenal glands, kidneys or bones.

[0147] In one embodiment of the method, the anti-PD-1 antibody is nivolumab. In another embodiment, it is pembrolizumab. In general, the anti-PD-1 antibody is formulated for intravenous administration. In one embodiment, the anti-PD-1 antibody is administered by intravenous infusion over a period of 60 minutes. In one embodiment, the anti-PD-1 antibody is administered as a pharma- ceutically acceptable formulation. In one embodiment, the anti-PD-1 antibody or antigen-binding portion thereof is administered in a subtherapeutic dose.

[0148] Anti-PD-1 or anti-PD-L1 antibodies useful in the present invention Human monoclonal antibodies that specifically bind to PD-1 with high affinity are disclosed in U.S. Patent 8,008,449. Other anti-PD-1 monoclonal antibodies are disclosed, for example, in U.S. Patents 6,808,710, 7,488,802, 8,168,757, and 8,354,509 and PCT Publication WO 2012 / 145493. Each of the anti-PD-1 human monoclonal antibodies disclosed in U.S. Patent 8,008,449 has been shown to exhibit one or more of the following characteristics: (a) a specific binding affinity of 1×10 to human PD-1 as determined by surface plasmon resonance using a Biacore biosensor system; -7 K below M D (b) does not substantially bind to human CD28, CTLA-4 or ICOS; (c) increases T cell proliferation in a mixed lymphocyte reaction (MLR) assay; (d) increases interferon-γ production in an MLR assay; (e) increases IL-2 secretion in an MLR assay; (f) binds to human PD-1 and cynomolgus PD-1; (g) inhibits the binding of PD-L1 and / or PD-L2 to PD-1; (h) stimulates an antigen-specific memory response; (i) stimulates an antibody response; and (j) inhibits tumor cell proliferation in vivo. Anti-PD-1 antibodies that can be used in the present invention include monoclonal antibodies that specifically bind to human PD-1 and exhibit at least one, and in some embodiments, at least five, of the above properties. In some embodiments, the anti-PD-1 antibody is nivolumab. In some embodiments, the anti-PD-1 antibody is pembrolizumab.

[0149] In one embodiment, the anti-PD-1 antibody is nivolumab. Nivolumab (also known as “Opdivo (登録商標) ”; previously named 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 the PD-1 ligands (PD-L1 and PD-L2), thereby blocking the downregulation of anti-tumor T cell function (U.S. Patent 8,008,449; Wang et al., In vitro characterization of the anti-PD-1 antibody nivolumab, BMS-936558, and in vivo toxicology in non-human primates, Cancer Imm Res, 2(9):846-56 (2014)). In other embodiments, the anti-PD-1 antibody or fragment thereof cross-competes with nivolumab. In other embodiments, the anti-PD-1 antibody or fragment thereof binds to the same epitope as nivolumab. In one embodiment, the anti-PD-1 antibody has the same CDRs as nivolumab.

[0150] In other embodiments, the anti-PD-1 antibody (or antigen-binding portion thereof) cross-competes with pembrolizumab. In one embodiment, the anti-PD-1 antibody binds to the same epitope as pembrolizumab. In one embodiment, the anti-PD-1 antibody has the same CDRs as pembrolizumab. In other embodiments, the anti-PD-1 antibody is pembrolizumab. Pembrolizumab (also known as “Keytruda (登録商標)Pembrolizumab (also known as lambrolizumab 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. Patents 8,354,509 and 8,900,587; see also www.cancer.gov / drugdictionary?cdrid=695789 (last accessed December 14, 2014). Pembrolizumab is approved by the FDA for the treatment of recurrent or refractory melanoma.

[0151] In other embodiments, the anti-PD-1 antibody (or antigen-binding portion thereof) cross-competes with MEDI0680. In still other embodiments, the anti-PD-1 antibody or fragment thereof binds to the same epitope as MEDI0680. In certain embodiments, the anti-PD-1 antibody has the same CDRs as MEDI0680. In other embodiments, the anti-PD-1 antibody is MEDI0680, a monoclonal antibody (previously AMP-514). MEDI0680 is described, for example, in U.S. Patent 8,609,089 B2.

[0152] In certain embodiments, the immune checkpoint inhibitor is AMP-224, a B7-DC Fc fusion protein. AMP-224 is described in U.S. Publication 2013 / 0017199 or www.cancer.gov / publications / dictionaries / cancer-drug?cdrid=700595 (last accessed July 8, 2015).

[0153] In other embodiments, the anti-PD-1 antibody (or antigen-binding portion thereof) cross-competes with BGB-A317. In certain embodiments, the anti-PD-1 antibody binds to the same epitope as BGB-A317. In certain embodiments, the anti-PD-1 antibody has the same CDRs as BGB-A317. In certain embodiments, the anti-PD-1 antibody is BGB-A317, a humanized monoclonal antibody. BGB-A317 is described in U.S. Publication 2015 / 0079109.

[0154] In other embodiments, the anti-PD-1 antibody (or antigen-binding portion thereof) cross-competes with INCSHR1210 (SHR-1210). In certain embodiments, the anti-PD-1 antibody binds to the same epitope as INCSHR1210 (SHR-1210). In certain embodiments, the anti-PD-1 antibody has the same CDRs as INCSHR1210 (SHR-1210). In certain embodiments, the anti-PD-1 antibody is INCSHR1210 (SHR-1210), a human monoclonal antibody. INCSHR1210 (SHR-1210) is described in WO2015 / 085847.

[0155] In other embodiments, the anti-PD-1 antibody (or antigen-binding portion thereof) cross-competes with REGN-2810. In certain embodiments, the anti-PD-1 antibody binds to the same epitope as REGN-2810. In certain embodiments, the anti-PD-1 antibody has the same CDRs as REGN-2810. In certain embodiments, the anti-PD-1 antibody is REGN-2810, a human monoclonal antibody. REGN-2810 is described in WO2015 / 112800.

[0156] In other embodiments, the anti-PD-1 antibody (or antigen-binding portion thereof) cross-competes with PDR001. In certain embodiments, the anti-PD-1 antibody binds to the same epitope as PDR001. In certain embodiments, the anti-PD-1 antibody has the same CDRs as PDR001. In certain embodiments, the anti-PD-1 antibody is PDR001, a humanized monoclonal antibody. PDR001 is described in WO2015 / 112900.

[0157] In other embodiments, the anti-PD-1 antibody (or antigen-binding portion thereof) cross-competes with TSR-042 (ANB011). In certain embodiments, the anti-PD-1 antibody binds to the same epitope as TSR-042 (ANB011). In certain embodiments, the anti-PD-1 antibody has the same CDRs as TSR-042 (ANB011). In certain embodiments, the anti-PD-1 antibody is TSR-042 (ANB011), a humanized monoclonal antibody. TSR-042 (ANB011) is described in WO2014 / 179664.

[0158] In other embodiments, the anti-PD-1 antibody (or antigen-binding portion thereof) cross-competes with STI-1110. In certain embodiments, the anti-PD-1 antibody binds to the same epitope as STI-1110. In certain embodiments, the anti-PD-1 antibody has the same CDRs as STI-1110. In certain embodiments, the anti-PD-1 antibody is STI-1110, a human monoclonal antibody. STI-1110 is described in WO2014 / 194302.

[0159] Anti-PD-1 antibodies that can be used in the methods of the invention also include isolated antibodies that specifically bind to human PD-1 and cross-compete with nivolumab for binding to human PD-1 (see, e.g., U.S. Patents 8,008,449 and 8,779,105; WO2013 / 173223). The ability of antibodies to cross-compete for binding to an antigen indicates that these antibodies bind to the same epitope region of the antigen and sterically interfere with the binding of the other cross-competing antibody to that particular epitope region. These cross-competing antibodies are predicted to have functional properties very similar to nivolumab by virtue of binding to the same epitope region of PD-1. Cross-competing antibodies can be readily identified based on their ability to cross-compete with nivolumab in standard PD-1 binding assays such as Biacore analysis, ELISA assays or flow cytometry (see, e.g., WO2013 / 173223).

[0160] In certain embodiments, antibodies that cross-compete with the human PD-1 antibody nivolumab for binding to human PD-1 or that bind to the same epitope region of the human PD-1 antibody are monoclonal antibodies. For administration to a human subject, these cross-competing antibodies are chimeric antibodies, humanized antibodies, or human antibodies. Such chimeric, humanized, or human monoclonal antibodies can be produced and isolated by methods well known in the art.

[0161] Anti-PD-1 antibodies that can be used in the methods of the invention also include the antigen-binding portions of the above antibodies. It has been well demonstrated that the antigen-binding function of an antibody can be exerted by fragments of the full-length antibody. Examples of binding fragments included within the term "antigen-binding portion" of an antibody are: (i) a Fab fragment, which is a monovalent fragment consisting of the V L -, V H -, C L - and C H1 -domains; (ii) an F(ab') 2 fragment, which is a bivalent fragment comprising two Fab fragments linked by a disulfide bridge in the hinge region; (iii) an Fd fragment consisting of the V H - and C H1 -domains; (iv) an Fv fragment consisting of the V L - and V H -domains of a single arm of the antibody; or any combination thereof.

[0162] An antibody that binds to PD-1 with high specificity and affinity suitable for use in the methods or compositions of the present invention, blocks the binding of PD-L1 and / or PD-L2, and inhibits the immunosuppressive effect of the PD-1 signaling pathway. In any of the compositions or methods described herein, an anti-PD-1 "antibody" includes an antigen-binding portion or fragment that binds to the PD-1 receptor and exhibits functional properties similar to those of an intact antibody in ligand binding inhibition and immune system upregulation. In certain embodiments, the anti-PD-1 antibody or its antigen-binding portion cross-competes with nivolumab 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 certain embodiments, the antibody is a humanized antibody. In other embodiments, the antibody is a human antibody. Antibodies of the IgG1, IgG2, IgG3 or IgG4 isotype can be used.

[0163] In certain embodiments, the anti-PD-1 antibody or antigen-binding portion thereof comprises a heavy chain constant region that is of the human IgG1 or IgG4 isotype. In certain other embodiments, the sequence of the IgG4 heavy chain constant region of the anti-PD-1 antibody or antigen-binding portion thereof comprises the S228P mutation, which replaces the serine residue in the hinge region with a proline residue commonly found at the corresponding position in IgG1 isotype antibodies. This mutation present in nivolumab prevents Fab arm exchange with endogenous IgG4 antibodies while maintaining low affinity for Fc receptor activation associated with wild-type IgG4 antibodies (Wang et al., 2014 Cancer Immunol Res. 2(9):846-56). In yet other embodiments, the antibody comprises a light chain constant region that is a human kappa or lambda constant region. In other embodiments, the anti-PD-1 antibody or antigen-binding portion thereof is a monoclonal antibody or antigen-binding portion thereof. In certain embodiments of any of the treatment methods described herein, including administration of an anti-PD-1 antibody, the anti-PD-1 antibody is nivolumab. In other embodiments, the anti-PD-1 antibody is pembrolizumab. In other embodiments, the anti-PD-1 antibody is a further antibody selected from human antibodies 17D8, 2D3, 4H1, 4A11, 7D3, and 5F4 described in U.S. Patent 8,008,449. In yet other embodiments, the anti-PD-1 antibody is MEDI0680 (formerly AMP-514), AMP-224, or BGB-A317.

[0164] In certain embodiments, the anti-PD-1 antibody used in the methods herein can be replaced with other anti-PD-1 or anti-PD-L1 antagonists. For example, because anti-PD-L1 antibodies block the interaction of PD-1 and PD-L1 and thereby exert an effect similar to that of the PD-1 signaling pathway, anti-PD-L1 antibodies can be substituted for the use of anti-PD-1 antibodies in the methods disclosed herein. Accordingly, in certain embodiments, the present invention relates to a method of treating a subject having a tumor, comprising administering to the subject a therapeutically effective amount of an anti-PD-L1 antibody. In certain embodiments, an anti-PD-L1 antibody useful in the methods herein is BMS-936559 (formerly 12A4 or MDX-1105) (see, e.g., U.S. Patent No. 7,943,743; WO2013 / 173223). In other embodiments, the anti-PD-L1 antibody is MPDL3280A (also known as RG7446 or atezolizumab) (see, e.g., Herbst et al., (2013) J Clin Oncol 31(suppl):3000. Abstract; U.S. Patent No. 8,217,149), MEDI4736 (also known as durvalumab; In: 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 US2014 / 0356353, filed May 6, 2014), or MSB0010718C (also known as avelumab; see US2014 / 0341917). In other embodiments, the anti-PD-L1 antibody is CX-072 (also known as CytomX; see WO2016 / 149201). In other embodiments, the anti-PD-L1 monoclonal antibody is selected from the group consisting of 28-8, 28-1, 28-12, 29-8, 5H1, and any combination thereof. In certain embodiments, antibodies that cross-compete for binding to the above PD-L1 antibody or bind to the same epitope of human PD-L1 are monoclonal antibodies. For administration to human subjects, these cross-competing antibodies can be chimeric, humanized, or human antibodies.Such chimeric, humanized or human monoclonal antibodies can be produced and isolated by methods well known in the art.

[0165] Standard therapeutic agents for lung cancer Standard therapeutic agents for various types of cancer are well known to those skilled in the art. For example, the National Comprehensive Cancer Network (NCCN), an alliance of 21 major cancer centers in the United States, issues the NCCN Clinical Practice Guidelines in Oncology (NCCN GUIDELINES (登録商標) ) which provides detailed up-to-date information on standard treatment regimens for a variety of cancers (see NCCN GUIDELINES (登録商標) (2014), available at www.nccn.org / professionals / physician_gls / f_guidelines.asp, last accessed May 14, 2014).

[0166] NSCLC is the leading cause of cancer death in the United States and worldwide, excluding breast, colon, and prostate cancers combined. In the United States, an estimated 228,190 new cases of lung and bronchus are diagnosed, and approximately 159,480 people die from the 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 gland is a common occurrence, and approximately 33% of patients have such metastases. NSCLC treatment has gradually improved OS, but the benefit has reached a plateau (the median OS for advanced patients is only 1 year). Progression after 1L treatment occurs in almost all of these subjects, and the 5-year survival rate is only 3.6% in refractory situations. From 2005 to 2009, the overall 5-year relative survival rate for lung cancer in the United States was 15.9% (NCCN GUIDELINES (登録商標) , Version 3.2014 - Non-Small Cell Lung Cancer, available at www.nccn.org / professionals / physician_gls / pdf / nscl.pdf, last accessed May 14, 2014).

[0167] Surgery, radiotherapy (RT), and chemotherapeutic agents are the three modalities commonly used in NSCLC patients. Collectively, NSCLC is relatively insensitive to chemotherapeutic agents and RT compared to small cell carcinoma. Generally, in patients with stage I or II disease, surgical resection offers the greatest chance of cure, and the use of chemotherapeutic agents before and after surgery is increasing. RT is also an adjuvant treatment for patients with resectable NSCLC and a primary local or palliative treatment for patients with incurable NSCLC.

[0168] Patients with stage IV disease with a good performance status (PS) benefit from chemotherapeutic agents. Many drugs, including platinum agents (e.g., cisplatin, carboplatin), taxane agents (e.g., paclitaxel, albumin-bound paclitaxel, docetaxel), vinorelbine, vincristine, etoposide, pemetrexed, and gemcitabine, are useful in stage IV NSCLC. Combinations using multiple of these drugs result in a 1-year survival rate of 30% - 40%, which is superior to single-agent therapy. Specific targeted therapies have also been developed for the treatment of advanced lung cancer. For example, bevacizumab (Avastin (登録商標) ) is a monoclonal antibody that blocks vascular endothelial growth factor A (VEGF-A). Erlotinib (Tarceva (登録商標) ) is a small molecule TKI of the epidermal growth factor receptor (EGFR). Crizotinib (Xalkori (登録商標) ) is a small molecule TKI that targets ALK and MET and is used in the treatment of NSCLC in patients with the mutant ALK fusion gene. Cetuximab (Erbitux (登録商標) ) is a monoclonal antibody that targets the EGFR.

[0169] Among patients with squamous NSCLC (which accounts for up to 25% of all NSCLC), there are certain unmet needs due to few treatment options after first-line (1L) therapy. Monotherapy agents are the standard of care after progression with platinum-based doublet chemotherapy agents (Pt doublets), which provide a median OS of approximately 7 months. Docetaxel remains the standard of care for this line of treatment, although erlotinib may also be used infrequently. Pemetrexed has been shown to produce clinically equivalent efficacy outcomes compared to docetaxel in second-line (2L) treatment of patients with advanced NSCLC, but with significantly fewer side effects (Hanna et al. (2004) J Clin Oncol 22:1589-97). There are currently no treatments approved for use in lung cancer beyond third-line (3L) settings. Pemetrexed and bevacizumab are not approved for squamous NSCLC, and molecularly targeted therapies have limited applicability. Unmet needs in advanced lung cancer are complicated by the failure of Oncothyreon and Merck KGaA's STIMUVAX (登録商標) to improve OS in a phase 3 trial, the recent failure of ArQule and Daiichi Sankyo's c-Met kinase inhibitor, tivantinib, to meet survival endpoints, and the failure of the combination of Eli Lilly's Alimta (登録商標) and Roche's Avastin (登録商標) to improve OS in late-stage trials and the failure of Amgen and Takeda's motesanib, a small molecule VEGF-R antagonist, to meet clinical endpoints in late-stage trials.

[0170] Immunotherapy for Lung Cancer There is a clear need for effective agents for patients undergoing multi-line targeted therapy and for therapies that extend survival beyond current standard of care. New attempts, including immunotherapy, particularly blockade of immune checkpoint including CTLA-4, PD-1 and PD-L1 inhibitory pathways, have recently shown promise (Creelan et al. (2014) Cancer Control 21(1):80-89). However, there remains a need to identify patients who will respond to immunotherapy, particularly those who are likely to respond to anti-PD-1 or anti-PD-L1 antibody therapy.

[0171] Pharmaceutical Compositions and Dosages The therapeutic agent of the present invention can be constituted as a composition comprising one or more antibodies and a pharmaceutically acceptable carrier, for example, a pharmaceutical composition. As used herein, "pharmaceutically acceptable carrier" includes any and all solvents, dispersion media, coatings, antibacterial and antifungal agents, isotonic and absorption delaying agents, and the like that are physiologically compatible. In certain embodiments, the carrier for the antibody-containing composition is suitable for intravenous, intramuscular, subcutaneous, parenteral, spinal or epithelial administration (e.g., by injection or infusion). The pharmaceutical compositions of the present invention may contain one or more pharmaceutically acceptable salts, antioxidants, aqueous and non-aqueous carriers and / or adjuvants such as preservatives, wetting agents, emulsifying agents and dispersing agents.

[0172] The present invention provides a dosing regimen that can provide a desired response, such as a maximum therapeutic response and / or a minimum adverse effect. For anti-PD-1 antibody administration, the dose can range from about 0.01 to about 10 mg / kg, about 1 to about 9 mg / kg, about 2 to about 8 mg / kg, about 3 to about 7 mg / kg, about 3 to about 6 mg / kg, 0.01 to about 5 mg / kg or about 1 to about 3 mg / kg of patient body weight. For example, the dose can be about 0.1 mg / kg, about 0.3 mg / kg, about 1 mg / kg, about 2 mg / kg, about 3 mg / kg, about 4 mg / kg, about 5 mg / kg, about 6 mg / kg, about 7 mg / kg, about 8 mg / kg, about 9 mg / kg or about 10 mg / kg body weight. The dosing schedule is generally designed to achieve an exposure that results in sustained receptor occupancy (RO) based on the typical pharmacokinetic properties of the antibody. Examples of treatment regimens are once about every week, once about every two weeks, once about every three weeks, once about every four weeks, once a month, once about every 3 to 6 months or longer. In certain embodiments, an anti-PD-1 antibody, such as, is administered to a subject once about every two weeks. The anti-PD-1 antibody can be administered at least twice, each administration being about 0.01 mg / kg to about 5 mg / kg, for example, 3 mg / kg, with an administration interval of every two weeks between the two administrations. In certain embodiments, the anti-PD-1 antibody is administered at least 3, 4, 5, 6 or 7 times (i.e., multiple times), each of these administrations having an administration interval of every two weeks between two successive administrations and being in an amount of about 0.01 mg / kg to about 10 mg / kg, for example, 1 mg / kg, 3 mg / kg or 6 mg / kg. The dose and schedule can be changed during the course of treatment. In certain embodiments, the dosing regimen of the anti-PD-1 antibody of the present invention includes about 0.1 to about 5 mg / kg body weight, about 1 to about 5 mg / kg body weight or about 1 to about 3 mg / kg body weight by intravenous administration, and the antibody is administered about every 14 to 21 days until complete response or disease progression is confirmed, up to about 6 weeks or about 12 week cycles. In certain embodiments, the antibody treatment or any combination treatment disclosed herein is continued 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.

[0173] When used in combination with other treatments (e.g., other immunotherapies), the dosage of the anti-PD-1 antibody can be reduced compared to the monotherapy dosage. A dosage of nivolumab that is typically lower than 3 mg / kg but does not fall below 0.001 mg / kg is a sub-therapeutic dosage. The sub-therapeutic dosages of the anti-PD-1 antibody used in the methods herein exceed 0.001 mg / kg and are lower than 3 mg / kg. In certain embodiments, the sub-therapeutic dosages are from 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 of body weight. In certain embodiments, the sub-therapeutic dosages are 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 of body weight. Receptor occupancy data from 15 subjects receiving nivolumab at 0.3 mg / kg to 10 mg / kg indicate that the PD-1 occupancy is thought to be dose-independent within this dosing range. Across all doses, the mean occupancy was 85% (range, 70% - 97%) and the mean plateau occupancy was 72% (range, 59% - 81%) (Brahmer et al. (2010) J Clin Oncol 28:3167 - 75). Thus, a 0.3 mg / kg dose may allow sufficient exposure to reach maximal biological activity.

[0174] In certain embodiments of the invention, the anti-PD-1 antibody is administered at a dosage of 3 mg / kg. In other embodiments of the invention, the anti-PD-1 antibody is administered at a dosage of 1 mg / kg.

[0175] In certain embodiments, the dosage of the anti-PD-1 antibody (or anti-PD-L1 antibody) is a fixed dosage in the pharmaceutical composition. In other embodiments, the methods of the invention can be used with a uniform dosage (a dosage given to a patient regardless of the patient's weight). In certain embodiments, the uniform dosage of the anti-PD-1 antibody or an antigenic portion thereof 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, 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 dosage of nivolumab can be about 240 mg. For example, the uniform dosage of pembrolizumab can be about 200 mg. In certain embodiments, the anti-PD-1 antibody or an antigen-binding portion thereof is administered at a dosage of about 240 mg. In certain embodiments, the anti-PD-1 antibody or an antigen-binding portion thereof is administered at a dosage of about 360 mg. In certain embodiments, the anti-PD-1 antibody or an antigen-binding portion thereof is administered at a dosage of about 480 mg. In certain embodiments, the uniform dosage of the anti-PD-1 antibody or an antigenic portion thereof is administered once every about 1 week, 2 weeks, 3 weeks, 4 weeks, 5 weeks or 6 weeks. In certain embodiments, 360 mg of the anti-PD-1 antibody or antigen-binding fragment is administered to a subject once every 3 weeks. In other embodiments, 480 mg of the anti-PD-1 antibody or antigen-binding fragment is administered to a subject once every 4 weeks.

[0176] The dosage and frequency vary depending on 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 dosing frequency can vary depending on whether the treatment is prophylactic or therapeutic. In prophylactic applications, relatively low dosages are generally administered at relatively infrequent intervals over a long period of time. Some patients receive treatment throughout their lives. In therapeutic applications, relatively high dosages at relatively short intervals may be required until disease progression is reduced or halted and the patient shows partial or complete improvement of disease symptoms. Thereafter, the patient may be administered in a prophylactic regimen.

[0177] The actual dosage level of the active ingredient in the pharmaceutical composition of the present invention can vary in order to obtain an amount of the active ingredient that is effective to achieve the desired therapeutic effect for a particular patient, composition, and mode of administration without undue toxicity to the patient. The selected dosage is determined by a variety of pharmacokinetic factors including the activity of the particular composition of the present invention employed, the route of administration, the time of administration, the rate of excretion of the particular compound used, the duration of treatment, other drugs, compounds and / or substances used in combination with the particular composition employed, the age, sex, weight, condition, general health, and prior medical history of the patient to be treated, and similar factors well known in the medical arts. The compositions of the present invention can be administered by one or more routes of administration using one or more of the various methods well known in the art. As is well known to those skilled in the art, the route of administration and / or mode will vary depending on the desired results.

[0178] Kit Also within the scope of the present invention is a kit comprising an anti-PD-1 antibody or an anti-PD-L1 antibody. The kit generally includes a label indicating the intended use and instructions for use of the contents of the kit. The term label includes any written or recorded medium on, with, or otherwise associated with the kit. Accordingly, the present invention provides a kit for treating a subject having a tumor, the kit comprising (a) a single dosage in the range of about 4 mg to about 500 mg of an anti-PD-1 antibody; and (b) instructions for using the anti-PD-1 antibody in any of the methods described herein. In certain embodiments for treating human patients, the kit comprises an anti-human PD-1 antibody disclosed herein, such as nivolumab or pembrolizumab. In certain embodiments, the kit further comprises an anti-PD-L1 antibody and / or an anti-STK11 antibody. In other embodiments, the kit further comprises instructions for detecting PD-L1 and / or STK11 expression in a tumor sample.

[0179] The present invention will be further illustrated by the following examples, which should not be construed as further limiting. The contents of all cited references throughout this specification are hereby expressly incorporated herein by reference.

[0180] Example 1 STK11 Mutation as a Biomarker for Nivolumab Response PD-L1 is expressed in various expression patterns in NSCLC tumors, such as commercially available NSCLC tumors (FIGS. 1A-1D). These patterns were designated as diffuse, heterogeneous, tumor-stromal interface, and negative, respectively. The PD-L1 expression pattern can be associated with a mechanistic hypothesis. For example, in tumors with a diffuse pattern, the expression of PD-L1 is driven not by mutation but by 9p24 amplification in the oncogenic signaling pathway. In tumors with a tumor-stromal interface pattern, there is adaptive resistance instead of epithelial-mesenchymal transition (EMT).

[0181] The PD-L1 expression pattern in commercially available NSCLC tumors correlates with the PD-L1 H score, as shown in FIGS. 2A-2B. There are significant differences in PD-L1 expression levels among the patterns. For example, extremely high H scores were observed in diffuse pattern samples, suggesting that these tumors may be dependent on PD-L1 inhibition.

[0182] The PD-L1 expression pattern observed in commercially available NSCLC tumors was also observed in biopsy samples. FIGS. 3A-3C show the PD-L1 expression pattern in trial biopsy samples corresponding to patients treated with nivolumab monotherapy, which corresponds to the same pattern observed in commercially available NSCLC tumors.

[0183] The possibility of false-negative PD-L1 is affected by the pattern category, pre-analytical variables, and biopsy size. For example, the tumor-stromal interface pattern is heterogeneous and particularly susceptible to false-negative results. Therefore, there is a need for a biomarker that can facilitate the classification of NSCLC tumors and thus be used to predict tumor response to a certain therapeutic agent.

[0184] A correlation was observed between the PD-L1 expression pattern and nivolumab efficacy (Figure 4). The majority of complete responders in grade 3 tumors had a diffuse PD-L1 expression pattern and a high PD-L1 H score. Therefore, identification of biomarkers specific to the diffuse PD-L1 expression pattern can be used to identify patients suitable for treatment with nivolumab based on the presence / absence of the biomarker.

[0185] Immune infiltration was associated with more immune infiltration (higher PD-L1 H score) with a diffuse or heterogeneous PD-L1 pattern in commercially available NSCLC tumors used for generation of the data shown in Figure 5. Therefore, it may be used as a biomarker specific to NSCLC tumors with a diffuse expression pattern (Figure 5A - 5B).

[0186] The multiplex IHC experiment showed a defined specific correlation between tumor cells and immune cell subsets. PD-L1 labeling showed diffuse PD-L1 expression in the tumor. CD68 detection showed that the layer of macrophages at the tumor-stromal interface contributed to the formation of "barrier" activated T cells, while CD3 detection showed that T cells were moderately abundant but mostly limited to the stroma (Figure 6).

[0187] The PD-L1 expression pattern correlates with genomic data (Figure 7A - 7B). Figure 7A shows that the PD-L1 expression level correlates with RNA sequencing data, but RNA sequencing data alone does not provide a geographical context for the PD-L1 expression pattern observed by IHC. Figure 7B, which provides exome sequencing data, shows that the diffuse PD-L1 expression pattern correlates with a high mutation burden.

[0188] A high mutation burden is also associated with inflammatory tumors (Figure 8A - 8B). Figure 8A shows the overall inflammation measured using the "CI score", which is the intensity score of chronic inflammatory infiltration. Figure 8B shows the PD-L1+ inflammation measured using the "PD-L1+CI score", which is the intensity score of the relative ratio of PD-L1+ immune infiltration. There is a correlation between the number of missense mutations and overall inflammation in NSCLC tumors.

[0189] The frequencies of mutations in various biomarkers (TP53, STK11, KEAP1, KRAS, EGFR, and MET) with respect to the observed PD-L1 expression patterns were evaluated (Figs. 9A-9B), and the results showed that negative PD-L1 tumor cell expression and low PD-L1 mRNA expression were associated with STK11 mutations. The presence of the mutant STK11 correlated with the presence of the "N" (PD-L1 negative) expression pattern (Fig. 10A) and inflammation (Figs. 10B-10C). The presence of the mutant STK11 did not correlate with the presence of the "D" (diffuse) pattern, which was the pattern observed in the majority of responders to nivolumab treatment. Thus, the presence of the mutant form of STK11 can be used as a negative biomarker for NSCLC tumor treatment with nivolumab (i.e., its presence predicts lack of response or poor response to nivolumab). Conversely, the presence of the wild-type form of STK11 (or absence of the mutant form) can be used as a positive selection biomarker for nivolumab treatment.

[0190] Loss of STK11 due to mutation is predictive of increased mTOR signaling. Lung adenocarcinomas (both mouse models and human tumors) with KRAS and STK11 mutations show decreased expression of PD-L1 and decreased T cell infiltration. The proposed mechanisms of immunosuppression mediated by mutations in STK11 include alterations to glycolytic metabolism with increased lactate production and frequent co-mutations in KEAP1 leading to an anti-inflammatory transcriptional program.

[0191] Immunoprint analysis of 24 NSCLC tumor samples in which the levels of FOLR2, VSIG4, CD163, CLEC4D, CSF1R, CD86, MS4A1, CD79B, CD19, KIR2DS4, CD3E, CCR4, CCR8 and CD8A were analyzed was used to classify the samples by inflammatory pattern (sigClass). See Figure 11. Samples were classified as low (“sigClass low”), medium (“sigClass med”) and high (“sigClass hi”) inflammation. Samples were also classified by the presence (“STK11 mut”) or absence (“STK11 wt”) of STK11 mutations. Additionally, samples were classified by PD-L1 expression pattern as negative (“PDL1_pattern 2 Negative”), diffuse (“PDL1_pattern 2 Diffuse”), heterogeneous (“PDL1_pattern 2 Heterogeneous”) and tumor-stromal interface (“PDL1_pattern 2 TS”). Tumors with a diffuse PD-L1 expression pattern showed high inflammation and the wild-type form of STK11. PD-L1 negative tumors clustered into two groups: moderate inflammation and low inflammation. There was no clear distinction in the level of inflammation of PD-L1 negative tumors by STK11 mutation status. All tumors with mutant STK11 were also negative for PD-L1.

[0192] This data indicates that PD-L1 expression patterns are associated with distinct phenotypes and genetic backgrounds. Diffuse PD-L1 expression correlates with an inflammatory TME and high mutational burden. Additionally, the presence of STK11 mutations identifies a subset of PD-L1 negative tumors. These findings establish the suitability of STK11 as a biomarker for identifying a subset of PD-L1 negative tumors and the potential for integration of histopathological and genomic data to define the properties that govern the likelihood of response to immunotherapy in various NSCLC subsets.

Claims

1. 1. An antibody or antigen-binding portion thereof that specifically binds to the programmed death-1 (PD-1) receptor and inhibits PD-1 activity (an "anti-PD-1 antibody") for use in a method of treating a subject having a tumor, the method comprising: (i) determining an expression pattern of PD-L1 in the tumor; and (ii) administering the anti-PD-1 antibody if the tumor exhibits a generalized PD-L1 expression pattern, if the tumor exhibits a tumor-stroma interface PD-L1 expression pattern, and / or if the tumor exhibits a heterogeneous PD-L1 expression pattern.

2. 1. An anti-PD-1 antibody for use in a method of treating a subject having a tumor, the method comprising administering the anti-PD-1 antibody to a subject if the tumor exhibits a tumor-stroma interface PD-L1 expression pattern and / or if the tumor exhibits a heterogeneous PD-L1 expression pattern.

3. 3. The anti-PD-1 antibody for use according to claim 1 or 2, wherein the method further comprises identifying the patient as having a tumor that expresses STK11 (an "STK11 positive tumor") prior to administration of the anti-PD-1 antibody.

4. An anti-PD-1 antibody for use in a method of treating a subject having a tumor, the method comprising administering to the subject the anti-PD-1 antibody, wherein the tumor is an STK11-positive tumor.

5. The anti-PD-1 antibody for use according to claim 3 or 4, wherein STK11 is wild-type STK11.

6. The anti-PD-1 antibody for use according to any of claims 1 to 5, wherein the tumor originates from a lung cancer, optionally a small cell lung cancer (SCLC) or a non-small cell lung cancer (NSCLC).

7. (i) a PD-L1 of about 60 to about 500, about 80 to about 480, about 100 to about 460, about 120 to about 440, about 140 to about 420, about 160 to about 400, about 180 to about 380, about 200 to about 360, about 200 to about 340, about 200 to about 320, or about 200 to about 300; (ii) a pan-PD-L1 expression pattern is characterized by a PD-L1 H score of at least about 60, at least about 70, at least about 80, at least about 90, at least about 100, at least about 110, at least about 120, at least about 130, at least about 140, at least about 150, at least about 160, at least about 170, at least about 180, at least about 190, at least about 200, at least about 225, at least about 250, at least about 275, or at least about 300; (iii) a heterogeneous PD-L1 expression pattern is characterized by a PD-L1 H score of about 1 to about 50, about 5 to about 45, about 10 to about 40, or about 15 to about 35.

7. The anti-PD-1 antibody for use according to any of claims 1 to 6, wherein the PD-L1 expression pattern is characterized by a PD-L1 H score of 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 35, or at least about 40.

8. 8. The anti-PD-1 antibody for use according to any of claims 1 to 7, wherein the tumor is characterized by having at least about 1%, at least about 2%, at least about 3%, at least about 4%, 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 35%, at least about 40%, at least about 45%, at least about 50%, at least about 55%, at least about 60%, at least about 65%, 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.

9. 9. The anti-PD-1 antibody for use according to any of claims 1 to 8, wherein the STK11 positive tumor is characterized by having at least about 1%, at least about 2%, at least about 3%, at least about 4%, 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 35%, at least about 40%, at least about 45%, at least about 50%, at least about 55%, at least about 60%, at least about 65%, 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 the tumor cells expressing STK11.

10. The anti-PD-1 antibody for use according to any one of claims 1 to 9, wherein the tumor exhibits severe inflammation.

11. 11. The anti-PD-1 antibody for use according to any of claims 1 to 10, wherein (i) the anti-PD-1 antibody cross-competes with nivolumab for binding to human PD-1, (ii) the anti-PD-1 antibody binds to the same epitope as nivolumab, (iii) the anti-PD-1 antibody is nivolumab, or (iv) the anti-PD-1 antibody is nivolumab.

12. 12. The anti-PD-1 antibody for use according to any of claims 1 to 11, wherein (i) the anti-PD-1 antibody is administered about once every week, every two weeks, or every three weeks at a dose ranging from at least about 0.1 mg / kg to at least about 10.0 mg / kg of body weight, or (ii) the anti-PD-1 antibody is administered about once every two weeks at a dose of at least about 3 mg / kg of body weight.

13. 12. The anti-PD-1 antibody for use according to any of claims 1 to 11, wherein the anti-PD-1 antibody is administered in a flat dose of at least about 200 mg, at least about 220 mg, at least about 240 mg, at least about 260 mg, at least about 280 mg, at least about 300 mg, at least about 320 mg, at least about 340 mg, at least about 360 mg, at least about 380 mg, at least about 400 mg, at least about 420 mg, at least about 440 mg, at least about 460 mg, at least about 480 mg, at least about 500 mg, or at least about 550 mg, optionally about once every 1, 2, 3, or 4 weeks.

14. 14. The anti-PD-1 antibody for use according to any of claims 1 to 13, wherein (i) the administration reduces the size of the tumor by, optionally, at least about 10%, about 20%, about 30%, about 40%, or about 50% compared to the size of the tumor before the administration; (ii) the administration provides 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 the initial administration; (iii) the administration results in stable disease after the administration; (iv) the administration results in a partial response after the administration; or (v) the administration results in a complete response after the administration.

15. (a) a dose of an antibody or antigen-binding portion thereof that specifically binds to the Programmed Death-1 (PD-1) receptor and inhibits PD-1 activity (an "anti-PD-1 antibody"), in the range of about 4 mg to about 500 mg; and (b) instructions for using an anti-PD-1 antibody according to any one of claims 1 to 14. Including the kit.

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