Compositions and methods for treating cancer by subcutaneous administration of anti-PD1 antibodies

Abstract

The present invention relates to compositions and methods for treating cancer in a patient, comprising administering a PD-1 antagonist, such as an anti-PD-1 antibody (e.g., pembrolizumab) or an antigen-binding fragment thereof, with or without hyaluronidase, subcutaneously to the patient in a specific amount every six weeks. In a specific embodiment, the amount of the anti-PD-1 antibody or antigen-binding fragment thereof is about 600 mg to about 1000 mg. In a specific embodiment, administration is about every three weeks and the amount of the anti-PD-1 antibody or antigen-binding fragment thereof is about 300 mg to about 500 mg. In a specific embodiment, the PD-1 antagonist is pembrolizumab or an antigen-binding fragment thereof. Compositions and kits formulated for subcutaneous administration comprising specific dosage amounts of an anti-PD-1 antibody or antigen-binding fragment thereof, and uses thereof for treating cancer are also provided.

Description

[Technical field]

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS This application is an international patent application that claims priority to and the benefit of U.S. Provisional Application No. 63 / 449,478, filed March 2, 2023, U.S. Provisional Application No. 63 / 415,928, filed October 13, 2022, and U.S. Provisional Application No. 63 / 415,526, filed October 12, 2022, each of which is incorporated by reference in its entirety herein.

[0002] The present invention relates to compositions and therapies useful for the treatment of cancer. In particular, the present invention relates to methods of treating cancer comprising administering an anti-PD-1 antibody or antigen-binding fragment thereof to a patient in need of such treatment using the dosing regimen specified herein. Compositions and kits containing specific doses of an anti-PD-1 antibody or antigen-binding fragment thereof formulated for subcutaneous administration are also provided. [Background technology]

[0003] PD-1 is recognized as a key player in immune regulation and the maintenance of peripheral tolerance. PD-1 is moderately expressed on naive T, B and natural killer T (NKT) cells and is upregulated by T / B cell receptor signaling on lymphocytes, monocytes and myeloid cells (Sharpe et al., The function of programmed cell death 1 and its ligands in regulating autoimmunity and infection. Nature Immunology (2007); 8: 239-245).

[0004] The two known ligands of PD-1, PD-L1 (B7-H1) and PD-L2 (B7-DC), are expressed in human cancers arising in a variety of tissues. In large sample sets of cancers, such as ovarian, renal, colorectal, pancreatic and liver cancers, as well as melanoma, PD-L1 expression has been shown to correlate with poor prognosis and reduced overall survival, regardless of subsequent treatment (Dong et al., Nat Med. 8(8):793-800 (2002); Yang et al. Invest Ophthalmol Vis Sci. 49:2518-2525 (2008); Ghebeh et al. Neoplasia 8:190-198 (2006); Hamanishi et al., Proc. Natl. Acad. Sci. USA 104:3360-3365 (2007); Thompson et al., Cancer 5:206-211 (2006); Nomi et al., Clin. Cancer Research 13:2151-2157 (2007); Ohigashi et al., J. Med. 1999:202-203 (2008); J. Med. 1999:203-204 (2008); J. Med. 1999:204-205 (2008); J. Med. 1999:205-206 (2008); J. Med. 2002 ... al.,Clin.Cancer Research 11:2947-2953(2005);Inman et al.,Cancer 109:1499-1505(2007);Shimauchi et al.Int.J.Cancer 121:2585-2590(2007);Gao et al.Clin.Cancer Research 15:971-979 (2009); Nakanishi J. Cancer Immunol Immunother. 56:1173-1182 (2007); and Hino et al., Cancer 00:1-9 (2010)).

[0005] Similarly, PD-1 expression on tumor-infiltrating lymphocytes has been found to indicate dysfunctional T cells in breast cancer and melanoma (Ghebeh et al., BMC Cancer. 2008 8:5714-15 (2008); and Ahmadzadeh et al., Blood 114:1537-1544 (2009)) and to correlate with poor prognosis in renal cancer (Thompson et al., Clinical Cancer Research 15:1757-1761 (2007)). Thus, it has been proposed that PD-L1-expressing tumor cells interact with PD-1-expressing T cells to attenuate T cell activation and evasion of immune surveillance, thereby contributing to impaired immune responses against tumors.

[0006] Immune checkpoint therapy targeting the PD-1 axis has led to revolutionary improvements in clinical responses in multiple human cancers (Brahmer et al., N Engl J Med 2012,366:2455-65; Garon et al. N Engl J Med 2015,372:2018-28; Hamid et al., N Engl J Med 2013,369:134-44; Robert et al., Lancet 2014,384:1109-17; Robert et al., N Engl J Med 2015,372:2521-32; Robert et al., N Engl J Med 2015,372:320-30; Topalian et al., N Engl J Med 2012,366:2443-54; Topalian et al. al., J Clin Oncol 2014, 32:1020-30; and Wolchok et al., N Engl J Med 2013, 369:122-33). Immunotherapies targeting the PD-1 axis include monoclonal antibodies against the PD-1 receptor (KEYTRUDA™ (pembrolizumab), Merck and Co., Inc., Kenilworth, NJ, USA, and OPDIVO™ (nivolumab), Bristol-Myers Squibb Company, Princeton, NJ, USA), and monoclonal antibodies that bind to PD-L1 ligands (MPDL3280A; TECENTRIQ™ (atezolizumab), Genentech, San Francisco, CA, USA; IMFINZI™ (durvalumab), AstraZeneca Pharmaceuticals LP, Wilmington, DE; and BAVENCIO™ (avelumab), Merck KGaA, Darmstadt, Germany). Both therapeutic approaches have demonstrated antitumor efficacy in multiple cancer types.

[0007] Hyaluronidase is an enzyme that breaks down hyaluronic acid present in the extracellular matrix. There are six types of hyaluronidase known to exist in humans: Hyall, Hyal2, Hyal3, Hyal4, HyalPS1, and PH20 / SPAM1. PH20 / SPAM1 (hereinafter referred to as PH20) is expressed in the sperm plasma membrane and acrosomal membrane.

[0008] Hyaluronidase reduces the viscosity of hyaluronic acid in the extracellular matrix by hydrolyzing hyaluronic acid, increasing its permeability to tissues (skin). The subcutaneous region of the skin has a neutral pH of about 7.0-7.5. Among the various types of hyaluronidase, PH20 is widely used (Bookbinder et al., 2006). In instances where PH20 is used, it is often co-administered with antibody therapeutics that are injected subcutaneously (Bookbinder et al., 2006). rHuPH20, also known as Hylenex® and approved by the FDA, is indicated as an adjuvant to increase the dispersion and absorption of other injected drugs.

[0009] Currently approved anti-PD-1 antibody treatments for use in multiple cancer indications are administered as an IV infusion at doses of either (i) 200 mg or 2 mg / kg Q3W or (ii) 400 mg Q6W. It would be beneficial to develop a dosing schedule that allows for the administration of a safe and effective subcutaneous dose of anti-PD-1 antibody with hyaluronidase that provides equivalent exposure to the approved IV infusion doses. Alternatives to IV infusion, such as subcutaneous administration, would offer convenience and flexibility to patients, reduce patient time in the treatment room, and reduce the time required by providers to administer the treatment. [Prior art documents] [Non-patent literature]

[0010] [Non-Patent Document 1] Sharpe et al.,The function of programmed cell death 1 and its ligands in regulating autoimmunity and infection.Nature Immunology(2007);8:239-245

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[0011] The present invention provides an alternative, convenient, and cost-effective subcutaneous dosing regimen for treating cancer patients with an anti-PD-1 antibody or antigen-binding fragment thereof, where the dosing schedule is expected to provide a safe and effective dose of the anti-PD-1 antibody or antigen-binding fragment thereof. Specifically, the present invention provides a method of treating cancer in a human patient in need of such treatment, comprising subcutaneously administering to the patient every six weeks a dose of about 600 mg to about 1000 mg of an anti-PD-1 antibody or antigen-binding fragment thereof, wherein the anti-PD-1 antibody or antigen-binding fragment thereof comprises: (a) light chain (LC) complementarity determining regions (CDRs) LC-CDR1, LC-CDR2, and LC-CDR3 comprising the sequences of amino acids set forth in SEQ ID NOs: 1, 2, and 3, respectively, and heavy chain (HC) CDRs HC-CDR1, HC-CDR2, and HC-CDR3 comprising the sequences of amino acids set forth in SEQ ID NOs: 6, 7, and 8, respectively. In a specific embodiment, the antibody or antigen-binding fragment thereof is administered in combination with a hyaluronidase, which in a specific embodiment includes human hyaluronidase. In an embodiment of the invention, the antibody or antigen-binding fragment is pembrolizumab or an antigen-binding fragment thereof. In a further embodiment, the anti-PD-1 antibody is pembrolizumab.

[0012] The invention also provides a method of treating cancer in a human patient in need thereof, comprising subcutaneously administering to the patient about every three weeks a dose of about 300 mg to about 500 mg of an anti-PD-1 antibody or antigen-binding fragment thereof, the anti-PD-1 antibody or antigen-binding fragment thereof comprising: (a) light chain (LC) complementarity determining regions (CDRs) LC-CDR1, LC-CDR2, and LC-CDR3 comprising the sequences of amino acids set forth in SEQ ID NOs: 1, 2, and 3, respectively; heavy chain (HC) CDRs HC-CDR1, HC-CDR2, and HC-CDR3 comprising the sequences of amino acids set forth in SEQ ID NOs: 6, 7, and 8, respectively; and hyaluronidase. In an embodiment of the invention, the hyaluronidase is human hyaluronidase. In an embodiment of the invention, the antibody or antigen-binding fragment is pembrolizumab or an antigen-binding fragment thereof. In a further embodiment, the anti-PD-1 antibody is pembrolizumab.

[0013] The present invention also relates to a pharmaceutical composition for subcutaneous injection comprising an anti-PD-1 antibody or antigen-binding fragment thereof at a dose of about 600 mg to about 1000 mg, and about 700 units to about 50000 units of human hyaluronidase.The present invention also relates to a pharmaceutical composition for subcutaneous injection comprising an anti-PD-1 antibody or antigen-binding fragment thereof at a dose of about 300 mg to about 500 mg, and about 700 units to about 50000 units of human hyaluronidase.

[0014] In all of the therapeutic methods, compositions, and uses disclosed herein, the anti-PD-1 antibody or antigen-binding fragment inhibits the binding of PD-L1 to PD-1, and preferably also inhibits the binding of PD-L2 to PD-1. In specific embodiments of the therapeutic methods, compositions, and uses of the invention, the anti-PD-1 antibody or antigen-binding fragment is a monoclonal antibody that specifically binds to PD-1 and blocks the binding of PD-L1 to PD-1. In one particular embodiment, the anti-PD-1 antibody comprises a heavy chain and a light chain, and the light chain and heavy chain comprise the amino acid sequences of SEQ ID NO:5 and SEQ ID NO:10 or 11, respectively.

[0015] In specific embodiments of the disclosed therapeutic methods, compositions and uses, the cancer expresses one or both of PD-L1 and PD-L2, hi specific embodiments, PD-L1 expression is present or elevated in the cancer. [Brief description of the drawings]

[0016] [Figure 1] Figure 13 shows a flat exposure response for Ctrough versus tumor size change at 200mg Q3W IV infusion in KEYNOTE-252.Ctrough at 400mg Q6W IV infusion from KEYNOTE-555 was within this range of Ctrough where a flat exposure response was established. [Figure 2A] FIG. 14. Observed pharmacokinetic data from KEYNOTE-555 compared to model-predicted pharmacokinetic profile for pembrolizumab 400 mg Q6W (Cycle 1). [Figure 2B] FIG. 14. Observed pharmacokinetic data from KEYNOTE-555 compared to model-predicted pharmacokinetic profile for pembrolizumab 400 mg Q6W (steady state). [Figure 2C] FIG. 13. Model-predicted Ctrough for pembrolizumab 200 mg Q3W and 2 mg / kg Q3W (6 weeks of initial treatment) compared to 400 mg Q6W. [Figure 2D] FIG. 13. Model-predicted Ctrough for pembrolizumab 200 mg Q3W and 2 mg / kg Q3W (steady state) compared to 400 mg Q6W. [Figure 2E] FIG. 13. Model predicting Cmax for pembrolizumab 200 mg Q3W, 2 mg / kg Q3W, and 10 mg / kg Q2W compared to 400 mg Q6W (initial 6 weeks of treatment). [Figure 2F] FIG. 13. Model predicting Cmax (steady state) for pembrolizumab 200 mg Q3W, 2 mg / kg Q3W and 10 mg / kg Q2W compared to 400 mg Q6W. [Figure 3A]FIG. 1 shows the distribution of Ctrough (5th, 25th, 50th, 75th, and 95th percentiles; solid lines from bottom to top) in cycle 1 using simulations based on a pharmacokinetic ("PK") model at pembrolizumab doses of 790 mg Q6W SC and 400 mg Q6W IV. Lowest IV refers to the 5th percentile value from the distribution of exposure measurements for the 400 mg Q6W IV dose, median IV refers to the 50th percentile value from the distribution of exposure measurements for the 400 mg Q6W IV dose, and highest IV refers to the 95th percentile value from the distribution of exposure measurements for the 400 mg Q6W IV dose, with lowest IV, median IV, and highest IV being dashed lines from bottom to top. [Figure 3B] FIG. 1 shows the distribution of Ctrough at steady state (Cycle 3) using a simulation based on the PK model for pembrolizumab doses 790 mg Q6W SC and 400 mg Q6W IV (5th, 25th, 50th, 75th, and 95th percentiles; solid lines from bottom to top). Lowest IV refers to the 5th percentile value from the distribution of exposure measurements for the 400 mg Q6W IV dose, median IV refers to the 50th percentile value from the distribution of exposure measurements for the 400 mg Q6W IV dose, and highest IV refers to the 95th percentile value from the distribution of exposure measurements for the 400 mg Q6W IV dose, with lowest, median, and highest IV being dashed lines from bottom to top. [Figure 4A] FIG. 1 shows the distribution of AUC0-6wks in cycle 1 using a simulation based on the PK model for pembrolizumab doses of 790 mg Q6W SC and 400 mg Q6W IV (5th, 25th, 50th, 75th, and 95th percentile solid lines from bottom to top). Lowest IV refers to the 5th percentile value from the distribution of exposure measurements for the 400 mg Q6W IV dose, median IV refers to the 50th percentile value from the distribution of exposure measurements for the 400 mg Q6W IV dose, and highest IV refers to the 95th percentile value from the distribution of exposure measurements for the 400 mg Q6W IV dose, with the lowest, median, and highest IV being dashed lines from bottom to top. [Figure 4B]FIG. 1 shows the distribution of AUC0-6wks at steady state (Cycle 3) using a simulation based on the PK model for pembrolizumab doses of 790 mg Q6W SC and 400 mg Q6W IV (5th, 25th, 50th, 75th, and 95th percentile solid lines from bottom to top). Lowest IV refers to the 5th percentile value from the distribution of exposure measurements for the 400 mg Q6W IV dose, median IV refers to the 50th percentile value from the distribution of exposure measurements for the 400 mg Q6W IV dose, and highest IV refers to the 95th percentile value from the distribution of exposure measurements for the 400 mg Q6W IV dose, with the lowest, median, and highest IV being dashed lines from bottom to top. [Diagram 5] FIG. 1 shows the distribution of Cmax at steady state (Cycle 3) using a simulation based on the PK model at pembrolizumab doses of 790 mg Q6W SC and 400 mg Q6W IV (5th, 25th, 50th, 75th, and 95th percentiles; solid lines from bottom to top). Lowest IV refers to the 5th percentile value from the distribution of exposure measurements for the 400 mg Q6W IV dose, median IV refers to the 50th percentile value from the distribution of exposure measurements for the 400 mg Q6W IV dose, and highest IV refers to the 95th percentile value from the distribution of exposure measurements for the 400 mg Q6W IV dose, with lowest, median, and highest IV being dashed lines from bottom to top. [Figure 6A] FIG. 1 shows the distribution of Ctrough (5th, 25th, 50th, 75th, and 95th percentiles; solid lines from bottom to top) in cycle 1 using a simulation based on the PK model for doses of pembrolizumab-HLN 395 mg Q3W (left panel: Ctrough after 3 weeks; right panel: Ctrough after 6 weeks) and 790 mg Q6W SC. Minimum Ref refers to the 5th percentile value from the distribution of exposure measurements for the 790 mg Q6W SC dose, median Ref refers to the 50th percentile value from the distribution of exposure measurements for the 790 mg Q6W SC dose, and maximum Ref refers to the 95th percentile value from the distribution of exposure measurements for the 790 mg Q6W SC dose, with minimum Ref, median Ref, and maximum Ref being dashed lines from bottom to top. [Figure 6B] FIG. 1 shows the distribution of Ctrough at steady state (5th, 25th, 50th, 75th, and 95th percentiles; solid lines from bottom to top) using a PK model-based simulation for pembrolizumab-HLN doses of 395 mg Q3W and 790 mg Q6W SC. Minimum Ref refers to the 5th percentile value from the distribution of exposure measurements for the 790 mg Q6W SC dose, median Ref refers to the 50th percentile value from the distribution of exposure measurements for the 790 mg Q6W SC dose, and maximum Ref refers to the 95th percentile value from the distribution of exposure measurements for the 790 mg Q6W SC dose; minimum Ref, median Ref, and maximum Ref are dashed lines from bottom to top. [Figure 7A] FIG. 1 shows the distribution of AUC0-6wks in cycle 1 (5th, 25th, 50th, 75th, and 95th percentiles; solid lines from bottom to top) using a PK model-based simulation for pembrolizumab-HLN doses 395 mg Q3W and 790 mg Q6W SC. Minimum Ref refers to the 5th percentile value from the distribution of exposure measurements for the 790 mg Q6W SC dose, median Ref refers to the 50th percentile value from the distribution of exposure measurements for the 790 mg Q6W SC dose, and maximum Ref refers to the 95th percentile value from the distribution of exposure measurements for the 790 mg Q6W SC dose, with minimum Ref, median Ref, and maximum Ref being dashed lines from bottom to top. [Figure 7B] FIG. 1 shows the distribution of AUC0-6wks at steady state using a simulation based on the PK model for doses of pembrolizumab-HLN 395 mg Q3W and 790 mg Q6W SC (5th, 25th, 50th, 75th, and 95th percentiles; solid lines from bottom to top). Minimum Ref refers to the 5th percentile value from the distribution of exposure measurements for the 790 mg Q6W SC dose, median Ref refers to the 50th percentile value from the distribution of exposure measurements for the 790 mg Q6W SC dose, and maximum Ref refers to the 95th percentile value from the distribution of exposure measurements for the 790 mg Q6W SC dose, with minimum Ref, median Ref, and maximum Ref being dashed lines from bottom to top. [Figure 8] Similarity of PK model-based pembrolizumab clearance across indications. Percentiles of the distribution of post-hoc estimated individual baseline clearance values ​​among subjects per indication (number of subjects per indication shown above) are represented by lines (50th), boxes (25th-75th), and whiskers (5th-95th). Above each box plot, sample size (N) per group is shown. NSCLC=non-small cell lung cancer; HN=head and neck squamous cell carcinoma; UC=urothelial carcinoma; MSIH=microsatellite instability high cancer; HCC=hepatocellular carcinoma; cHL=classical Hodgkin lymphoma; PMBCL=primary mediastinal B-cell lymphoma. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0017] The present invention provides methods of treatment of a patient (e.g., a human patient) (e.g., a method of treating cancer) comprising subcutaneous administration of a particular dosage of an anti-PD-1 antibody (e.g., pembrolizumab) or an antigen-binding fragment thereof, and optionally a hyaluronidase (and in a specific embodiment, a human hyaluronidase). Such administration is expected to provide a safe and effective dose of an anti-PD-1 antibody or an antigen-binding fragment thereof. Also provided are compositions and kits formulated for subcutaneous administration comprising a dosage of an anti-PD-1 antibody or an antigen-binding fragment thereof, and optionally a hyaluronidase (e.g., human hyaluronidase), and uses thereof to treat cancer. In certain embodiments of the present invention, the anti-PD-1 antibody is pembrolizumab or an antigen-binding fragment of pembrolizumab.

[0018] I. Abbreviations and Definitions As used throughout the specification and appended claims, the following abbreviations apply:

[0019] AUC area under the concentration-time curve AUCss Area under the concentration-time curve at steady state CDR Complementarity Determining Region Cl Confidence interval CL Clearance Cmax,ss Peak concentration at steady state CPS Combined Positive Score CV Coefficient of variation of the between-subject distribution of the parameter; ECOG Eastern Cooperative Oncology Group eGFR: Estimated glomerular filtration rate ER Exposure (concentration) response F. Bioavailability FFPE Formalin-fixed Paraffin-embedded FR Framework Area GM geometric mean HCC hepatocellular carcinoma HNSCC head and neck squamous cell carcinoma HL Hodgkin's lymphoma IgG Immunoglobulin G IHC immunohistological staining or immunohistochemistry IMAX:Maximum effect of time on CL IV Intravenous ka First-order absorption rate constant LPS Lymphoma Percentage Score mAb Monoclonal antibody MCC Merkel cell carcinoma MEL Melanoma MMR mismatch repair MPS Modified Proportional Score MRI Magnetic Resonance Imaging MSI-H Microsatellite instability high NCI National Cancer Institute NSCLC non-small cell lung cancer OS overall survival PD-1 Programmed Death 1 (also known as Programmed Cell Death-1 and Programmed Death Receptor 1) PD-L1 Programmed Cell Death 1 Ligand 1 PD-L2 Programmed cell death 1 ligand 2 PFS Progression-free survival PK Pharmacokinetics Q Intercompartment Clearance Q2W Once every two weeks Q3W Once every three weeks Q6W Once every 6 weeks RCC renal cell carcinoma RSE Relative Standard Error SC subcutaneous T.I. 50 Time to achieve 50% of maximum clearance effect t lag Absorption lag time TPS Tumor Proportion Score Vc central volume of distribution V H Immunoglobulin heavy chain variable region V L Immunoglobulin light chain variable region Marginal volume of the Vp distribution

[0020] The population parameter estimates presented exclude the effects of covariates; therefore, such estimates apply to a hypothetical typical patient with average characteristics.

[0021] So that the present invention may be more readily understood, certain technical and scientific terms are specifically defined below. Unless specifically defined elsewhere herein, all other technical and scientific terms used herein have the meaning commonly understood by one of ordinary skill in the art to which this invention belongs.

[0022] As used throughout this specification and the appended claims, the singular forms "a," "an," and "the" include plural references unless the context clearly dictates otherwise. References to "or" indicate either or both possibilities, unless the context clearly dictates one of the stated possibilities. In some cases, "and / or" has been used to emphasize either or both possibilities.

[0023] The term "about" or "approximately" refers to variations in numerical quantities that may occur when varying amounts (e.g., mg) of a substance or composition, or values ​​of parameters characterizing a method step, etc., for example, through typical measuring, handling, and sampling procedures involved in preparing, characterizing, and / or using a substance or composition, due to inadvertent errors in these procedures, due to differences in manufacture, source, or purity of ingredients used to make or use a composition or to carry out a procedure, etc. In certain embodiments, "about" or "approximately" may mean a variation of ±0.1%, ±0.5%, ±1%, ±2%, ±3%, ±4%, ±5%, ±6%, ±7%, ±8%, ±9%, ±10%, or ±11%.

[0024] "Administration" and "treatment", when applied to an animal, human, experimental subject, cell, tissue, organ, or biological fluid, refer to contact of an exogenous pharmaceutical, therapeutic, diagnostic agent, or composition with an animal, human, subject, cell, tissue, organ, or biological fluid. As used herein, "treating" or "treating" cancer means administering an anti-PD-1 antibody or antigen-binding fragment to a subject having or diagnosed with cancer to achieve at least one positive therapeutic effect on the cancer, e.g., a reduction in the number of cancer cells, a reduction in tumor size, a reduction in the rate of cancer cell invasion into peripheral organs, or a reduction in the rate of tumor metastasis or tumor growth. "Treatment" may include one or more of the following: inducing / increasing an anti-tumor immune response, reducing the number of one or more tumor markers, halting or slowing the growth of a tumor or blood cancer or the progression of a disease associated with PD-1 binding to its ligands PD-L1 and / or PD-L2 (a "PD-1 associated disease"), stabilizing a PD-1 associated disease, inhibiting the growth or survival of tumor cells, eliminating or reducing the size of one or more cancerous lesions or tumors, reducing the levels of one or more tumor markers, ameliorating or abrogating clinical symptoms of a PD-1 associated disease, reducing the severity or duration of clinical symptoms of a PD-1 associated disease, such as cancer, prolonging survival compared to expected survival in similar untreated patients, and inducing the progression of a cancerous condition or other PD-1 or other PD-1 associated disease.

[0025] Positive therapeutic effects in cancer can be measured in several ways (see WA Weber, J. Nucl. Med. 50:1S-10S (2009)). For example, with regard to tumor growth inhibition, according to NCI standards, T / C≦42% is the minimum level of antitumor activity. T / C<10% is considered a high level of antitumor activity, where T / C(%)=median treated tumor volume / median control tumor volume×100. In specific embodiments, the treatment achieved by the therapeutically effective amount is either progression-free survival (PFS), disease-free survival (DFS) or overall survival (OS). PFS, also known as "time to tumor progression," refers to the length of time during and after treatment that the cancer does not grow, including the length of time the patient experiences a complete or partial response, as well as the length of time the patient experiences stable disease. DFS refers to the length of time during and after treatment that the patient remains disease-free. OS refers to an extension of life expectancy compared to a naive or untreated individual or patient. Although the therapeutic methods, compositions and use embodiments of the present invention may not be effective in achieving a positive therapeutic effect in all patients, they should do so in a statistically significant number of subjects as determined by any statistical test known in the art, such as Student's t-test, chi-square test, Mann-Whitney U test, Kruskal-Wallis test (H test), Jonkheel-Terpstra test, and Wilcoxon test.

[0026] "Antibody" refers to any form of antibody that exhibits the desired biological activity or binding activity. It is therefore used in the broadest sense and specifically encompasses, but is not limited to, monoclonal antibodies (including full-length monoclonal antibodies), polyclonal antibodies, humanized antibodies, fully human antibodies, and chimeric antibodies. A "parent antibody" is an antibody obtained by exposing the immune system to an antigen prior to modification of the antibody for its intended use, such as humanizing the antibody for use as a human therapeutic.

[0027] Generally, a basic antibody structural unit comprises a tetramer. Each tetramer comprises two identical pairs of polypeptide chains, each pair having one "light" chain (about 25 kDa) and one "heavy" chain (about 50-70 kDa). The amino-terminal portion of each chain comprises a variable region of about 100-110 or more amino acids primarily responsible for antigen recognition. The carboxy-terminal portion of the heavy chain may define a constant region primarily responsible for effector function. Typically, human light chains are classified as kappa light chains and lambda light chains. Human heavy chains are further typically classified as mu, delta, gamma, alpha, or epsilon, defining the antibody isotype as IgM, IgD, IgG, IgA, and IgE, respectively. Within the light and heavy chains, the variable and constant regions are linked by a "J" region of about 12 or more amino acids, and the heavy chain also comprises a "D" region of about 10 or more amino acids. See generally, Fundamental Immunology Ch. 7 (Paul, W., ed., 2nd ed. Raven Press, NY (1989)).

[0028] The variable regions of each light / heavy chain pair form the antibody binding site. Thus, an intact antibody generally has two binding sites. Except for bifunctional or bispecific antibodies, the two binding sites are generally the same.

[0029] Typically, both the heavy and light chain variable domains contain three hypervariable regions, also called complementarity determining regions (CDRs), located within relatively conserved framework regions (FRs). The CDRs are usually aligned by the framework regions, allowing binding to a specific epitope. Generally, from the N-terminus to the C-terminus, both the light chain variable domain and the heavy chain variable domain contain FR1, CDR1, FR2, CDR2, FR3, CDR3 and FR4. The assignment of amino acids to each domain is generally based on the Sequences of Proteins of Immunological Interest, Kabat, et al.; National Institutes of Health, Bethesda, Md.; thed.;NIH Publ.No.91-3242(1991);Kabat(1978)Adv.Prot.Chem.32:1-75;Kabat, et al.,(1977)J.Biol.Chem.252:6609-6616;Chothia,et al.,(1987)J Mol.Biol.196:901-917 or according to the definition of Chothia, et al., (1989) Nature 342:878-883.

[0030] Unless otherwise indicated, an "antibody fragment" or "antigen-binding fragment" refers to an antigen-binding fragment of an antibody, i.e., an antibody fragment that retains the ability to specifically bind to the antigen bound by the full-length antibody, e.g., a fragment that retains one or more CDR regions, e.g., three heavy chain CDRs and three light chain CDRs. Examples of antigen-binding fragments include, but are not limited to, Fab, Fab', F(ab')2, and Fv fragments.

[0031] An "anti-PD-1 antibody" as used in any of the therapeutic methods, compositions and uses of the invention includes a monoclonal antibody (mAb) or antigen-binding fragment thereof that specifically binds to human PD-1. Alternative names or synonyms for PD-1 and its ligands include PDCD1, PD1, CD279 and SLEB2 for PD-1, PDCD1L1, PDL1, B7H1, B7-4, CD274 and B7-H for PD-L1, and PDCD1L2, PDL2, B7-DC, Btdc and CD273 for PD-L2. In any of the therapeutic methods, compositions and uses of the invention in which a human individual is being treated, the anti-PD-1 antibody or antigen-binding fragment thereof is a PD-1 antagonist that blocks the binding of human PD-L1 to human PD-1, or blocks the binding of both human PD-L1 and PD-L2 to human PD-1. The human PD-1 amino acid sequence can be found at NCBI Locus No.: NP_005009. The human PD-L1 and PD-L2 amino acid sequences can be found at NCBI Locus No.: NP_054862 and NP_079515, respectively. The anti-PD-1 antibody can be a human antibody, a humanized antibody, or a chimeric antibody and can comprise a human constant region. In a specific embodiment, the human constant region is selected from the group consisting of IgG1, IgG2, IgG3, and IgG4 constant regions, and in a specific embodiment, the human constant region is an IgG1 or IgG4 constant region. In a specific embodiment, the antigen-binding fragment is selected from the group consisting of Fab, Fab'-SH, F(ab')2, scFv, and Fv fragments.

[0032] "AUC" and "Cmax" are pharmacokinetic measures of systemic exposure to a drug (e.g., pembrolizumab) in humans following its administration and are typically considered drivers of drug efficacy. "AUC" represents the average exposure over the dosing interval. "Cmax" is the maximum or highest (peak) drug concentration observed immediately following its administration. In the specific case of pembrolizumab administered as a subcutaneous injection, the peak concentration occurs immediately after the end of the infusion. Cmax is a metric that is typically considered a driver of safety.

[0033] "Biotherapeutic agent" means a biological molecule, such as an antibody or a fusion protein, that blocks ligand / receptor signaling in any biological pathway that supports the maintenance and / or growth of a tumor or suppresses an anti-tumor immune response.

[0034] The term "buffering agent" includes agents that maintain the solution pH of the formulation of the invention in an acceptable range or, for the lyophilized formulations of the invention, provide an acceptable solution pH prior to lyophilization. The terms "lyophilization," "freeze-drying," and "lyophilization" refer to a process in which the material to be dried is first frozen and then the ice or freezing solvent is removed by sublimation in a vacuum environment. Excipients may be included in the pre-lyophilized formulation to enhance the stability of the lyophilized product upon storage.

[0035] "C trough " is the trough concentration achieved at the end of the dosing interval. SC:IV C trough The ratio is the C achieved with the SC dose versus the IV dose at the end of the same dosing interval. trough The ratio of these two values ​​(e.g., the geometric mean ratio) is

[0036] As used herein, "co-administration" refers to agents that are administered to a subject at or near the same time. The agents may or may not be physically combined prior to administration. For example, an anti-PD-1 antibody and a hyaluronidase may be contained in separate vials, combined in liquid solution in the same injection device, and administered simultaneously to a patient.

[0037] As used herein, "co-formulated" or "co-formulation" or "co-formulation" or "co-formulated" refers to at least two different proteins or drugs that are formulated together and stored as a combined product in a single vial, container, device or vessel (e.g., an injection device), rather than being formulated and stored separately and then mixed prior to administration or administered separately.

[0038] The terms "cancer", "cancerous" or "malignant" refer to or describe a physiological condition in a mammal that is typically characterized by unregulated cell proliferation. Examples of cancer include, but are not limited to, carcinoma, lymphoma, leukemia, blastoma and sarcoma. More specific examples of such cancer include, but are not limited to, squamous cell carcinoma, myeloma, small cell lung cancer, non-small cell lung cancer, glioma, Hodgkin's lymphoma, non-Hodgkin's lymphoma, acute myeloid leukemia (AML), multiple myeloma, gastrointestinal (ductal) cancer, renal cancer, ovarian cancer, liver cancer, lymphoblastic leukemia, lymphocytic leukemia, colorectal cancer, endometrial cancer, renal cancer, prostate cancer, thyroid cancer, melanoma, chondrosarcoma, neuroblastoma, pancreatic cancer, glioblastoma multiforme, cervical cancer, brain cancer, stomach cancer, bladder cancer, liver cancer, breast cancer, colon cancer, and head and neck cancer. Further cancers that may be treated according to the present invention include those characterized by elevated expression of one or both of PD-L1 and PD-L2 in examined tissue samples.

[0039] "CDR" or "CDRs" means the complementarity determining region(s) in an immunoglobulin variable region, commonly defined using the Kabat numbering system.

[0040] A "chemotherapeutic agent" is a chemical compound useful in the treatment of cancer. Classes of chemotherapeutic agents include, but are not limited to, alkylating agents, antimetabolites, kinase inhibitors, spindle poison plant alkaloids, cytotoxic / antitumor antibiotics, topoisomerase inhibitors, photosensitizers, antiestrogens and selective estrogen receptor modulators (SERMs), antiprogesterone agents, estrogen receptor downregulators (ERDs), estrogen receptor antagonists, leutinizing hormone releasing hormone agonists, antiandrogens, aromatase inhibitors, EGFR inhibitors, VEGF inhibitors, antisense oligonucleotides that inhibit the expression of genes involved in abnormal cell proliferation or tumor growth. Chemotherapeutic agents useful in the therapeutic methods, compositions and uses of the present invention include cytostatic and / or cytotoxic agents.

[0041] A "chimeric antibody" refers to an antibody in which a portion of the heavy and / or light chain is identical to or homologous to corresponding sequences in antibodies from a particular species (e.g., human) or belonging to a particular antibody class or subclass, and the remainder of the chain(s) is identical to or homologous to corresponding sequences in antibodies from another species (e.g., mouse) or belonging to another antibody class or subclass, as well as fragments of such antibodies.

[0042] The words "comprising" or variations such as "comprise," "comprises," or "comprised of" are used throughout this specification and claims in an inclusive sense, i.e., to specify the presence of stated features, but not to preclude the presence or addition of further features that could materially enhance the operation or utility of any of the embodiments of the invention, unless the context requires otherwise by express language or necessary implication.

[0043] "Conservatively modified variants" or "conservative substitutions" refer to the substitution of amino acids in a protein with other amino acids having similar characteristics (e.g., charge, side chain size, hydrophobicity / hydrophilicity, main chain conformation and rigidity, etc.), such that changes can be made frequently without altering the biological activity or other desired properties of the protein, such as antigen affinity and / or specificity. Those skilled in the art will generally recognize that single amino acid substitutions in non-essential regions of a polypeptide do not substantially alter biological activity (see, e.g., Watson et al. (1987) Molecular Biology of the Gene, The Benjamin / Cummings Pub. Co., p. 224 (4th Ed.)). Furthermore, substitution of structurally or functionally similar amino acids is unlikely to destroy biological activity. Exemplary conservative substitutions are shown in Table 1. [Table 1]

[0044] A "diagnostic anti-PD-L monoclonal antibody" refers to a mAb that specifically binds to the mature form of the designated PD-L (PD-L1 or PD-L2) expressed on the surface of certain mammalian cells. Mature PD-L lacks the precursor leader sequence, also called leader peptide. The terms "PD-L" and "mature PD-L" are used interchangeably herein and shall be understood to refer to the same molecule, unless otherwise indicated or readily apparent from the context.

[0045] As used herein, diagnostic anti-human PD-L1 mAb or anti-hPD-L1 mAb refers to a monoclonal antibody that specifically binds to mature human PD-L1. The mature human PD-L1 molecule consists of amino acids 19 to 290 of the following sequence:

[0046] MRIFAVFIFMTYWHLLNAFTVTVPKDLYVVEYGSNMTIECKFPVEKQLDLAALIVYWEMEDKNIIQFVHGEEDLKVQHSSYRQRARLLKDQLSLGNAALQITDVKLQDAGVYRCMISYGGADYKRITVKVNAPYNKINQRILVVDPVTSEHELTCQAEGYPKAEVIWTSSDHQVLSGKTTTTNSKREEKLFNVTSTLRINTTTNEIFYCTFRRLDPEENHTAELVIPELPLAHPPNERTHLVILGAILLCLGVALTFIFRLRKGRMMDVKKCGIQDTNSKKQSDTHLEET (SEQ ID NO: 21).

[0047] Specific examples of diagnostic anti-human PD-L1 mAbs useful as diagnostic mAbs for immunohistochemical (IHC) detection of PD-L1 expression in formalin-fixed paraffin-embedded (FFPE) tumor tissue sections are antibody 20C3 and antibody 22C3, which are described in WO 2014 / 100079. These antibodies comprise the light chain variable region amino acid sequences and heavy chain variable region amino acid sequences shown in Table 2 below: [Table 2]

[0048] Another anti-human PD-L1 mAb that has been reported to be useful for IHC detection of PD-L1 expression in FFPE tissue sections (Chen, BJ et al., Clin Cancer Res 19:3462-3473(2013)) is the publicly available rabbit anti-human PD-L1 mAb from Sino Biological, Inc. (Beijing, PRChina; Catalog No. 10084-R015).

[0049] As used herein, "framework region" or "FR" means an immunoglobulin variable region excluding the CDR regions.

[0050] A "human antibody" refers to an antibody that contains only human immunoglobulin protein sequences. A human antibody may contain mouse carbohydrate chains if produced in a mouse, in a mouse cell, or in a hybridoma derived from a mouse cell. Similarly, a "mouse antibody" or a "rat antibody" refers to an antibody that contains only mouse or rat immunoglobulin sequences, respectively.

[0051] "Humanized antibody" refers to forms of antibodies that contain sequences derived from non-human (e.g., murine) and human antibodies. Such antibodies contain minimal sequences derived from non-human immunoglobulin. Generally, a humanized antibody contains substantially all of at least one, and typically two, variable domains, with all or substantially all of the hypervariable loops corresponding to those of a non-human immunoglobulin, and all or substantially all of the FR regions being those of a human immunoglobulin sequence. A humanized antibody may also contain at least a portion of an immunoglobulin constant region (Fc), typically at least a portion of an immunoglobulin constant region (Fc) of a human immunoglobulin. When it is necessary to distinguish a humanized antibody from a parent rodent antibody, the prefixes "hum", "hu" or "h" are added to the name of the antibody clone. Humanized forms of rodent antibodies generally contain the same CDR sequences of the parent rodent antibody, but may contain certain amino acid substitutions to increase affinity, to increase the stability of the humanized antibody, or for other reasons.

[0052] "Hypervariable region" refers to the amino acid residues of an antibody which are involved in antigen binding. The hypervariable region comprises amino acid residues from the "complementarity determining regions" or "CDRs" (i.e., LC-CDR1, LC-CDR2 and LC-CDR3 in the light chain variable domain and HC-CDR1, HC-CDR2 and HC-CDR3 in the heavy chain variable domain). See Kabat et al. (1991) Sequences of Proteins of Immunological Interest, 5th Ed. Public Health Service, National Institutes of Health, Bethesda, Md (defining antibody CDR regions by sequence); see also Chothia and Lesk (1987) J. Mol. Biol. 196:901-917 (defining antibody CDR regions by structure). The term "framework" or "FR" residues refers to variable domain residues other than the hypervariable region residues defined herein as CDR residues.

[0053] "Immunogenic agent" refers to a composition capable of inducing a humoral and / or cell-mediated immune response. Immunogenic agents can include, for example, attenuated cancerous cells, tumor antigens, tumor-derived antigens or antigen-presenting cells such as dendritic cells pulsed with nucleic acids, immune-stimulating cytokines (e.g., IL-2, IFNα2, GM-CSF), and cells transfected with genes encoding immune-stimulating cytokines such as, but not limited to, GM-CSF.

[0054] "In need of" refers to needing treatment.

[0055] "Kabat" as used herein refers to the immunoglobulin alignment and numbering system pioneered by Elvin A. Kabat ((1991) Sequences of Proteins of Immunological Interest, 5th Ed. Public Health Service, National Institutes of Health, Bethesda, Md.).

[0056] As used herein, "monoclonal antibody" or "mAb" or "Mab" refers to a population of substantially homogeneous antibodies, i.e., the antibody molecules comprising the population are identical in amino acid sequence, except for possible naturally occurring mutations that may be present in minor amounts. In contrast, conventional (polyclonal) antibody preparations typically contain a large number of different antibodies having different amino acid sequences in their variable domains, particularly their CDRs, which are often specific for different epitopes. The modifier "monoclonal" indicates the character of the antibody as being obtained from a substantially homogeneous antibody population and should not be construed as requiring production of the antibody by any particular method. For example, monoclonal antibodies used in accordance with the present invention may be made by the hybridoma method first described by Kohler et al. (1975) Nature 256:495, or may be made by recombinant DNA methods (see, e.g., U.S. Pat. No. 4,816,567). "Monoclonal antibodies" can also be isolated from phage antibody libraries using the techniques described in, for example, Clackson et al. (1991) Nature 352:624-628 and Marks et al. (1991) J. Mol. Biol. 222:581-597. See also Presta (2005) J. Allergy Clin. Immunol. 116:731.

[0057] "Microsatellite instability (MSI)" refers to a form of genomic instability associated with defective DNA mismatch repair in tumors. See Boland et al., Cancer Research 58, 5258-5257, 1998. In one embodiment, MSI analysis can be performed using five National Cancer Institute (NCI) recommended microsatellite markers: BAT25 (GenBank Accession No. 9834508), BAT26 (GenBank Accession No. 9834505), D5S346 (GenBank Accession No. 181171), D2S123 (GenBank Accession No. 187953), D17S250 (GenBank Accession No. 177030). Additional markers can be used, such as BAT40, BAT34C4, TGF-β-RII, and ACTC. Commercially available kits for MSI analysis include, for example, the Promega MSI multiplex PCR assay, FoundationOne® CDx (F1CDx) next-generation sequencing, based on an in vitro diagnostic device that uses DNA isolated from formalin-fixed paraffin-embedded (FFPE) tumor tissue specimens.

[0058] "High frequency microsatellite instability" or "microsatellite instability high (MSI-H)" refers to a tumor in which two or more of the five NCI markers listed above show instability in its DNA, or more than 30-40% of all markers in its DNA show instability (i.e., have insertion / deletion mutations).

[0059] As used herein, "non-MSI-H cancer" refers to microsatellite stable (MSS) and low frequency MSI (MSI-L) cancers.

[0060] "Microsatellite stable (MSS)" refers to tumors that do not exhibit instability in their DNA (i.e., have insertion / deletion mutations) for any of the five NCI markers listed above.

[0061] A "patient" (also referred to herein as a "subject" or "individual") refers to a mammal (e.g., rat, mouse, dog, cat, rabbit), most preferably a human, that can be treated with the methods and compositions of the present invention. In specific embodiments, the patient is an adult patient. In other embodiments, the patient is a pediatric patient.

[0062] "PD-L1" or "PD-L2" expression means any detectable level of expression of the designated PD-L protein on the cell surface or the designated PD-L mRNA in a cell or tissue, unless otherwise defined. PD-L protein expression may be detected using a diagnostic PD-L antibody in an IHC assay of tumor tissue sections or by flow cytometry. Alternatively, PD-L protein expression by tumor cells may be detected by PET imaging using a binding agent (e.g., an antibody fragment, affibody, etc.) that specifically binds to the desired PD-L target, e.g., PD-L1 or PD-L2. Techniques for detecting and measuring PD-L mRNA expression include reverse transcription polymerase chain reaction (RT-PCR) and real-time quantitative RT-PCR.

[0063] Several approaches have been described for quantifying PD-L1 protein expression in IHC assays of tumor tissue sections, see, e.g., Thompson et al., PNAS 101(49):17174-17179 (2004); Thompson et al., Cancer Res. 66:3381-3385 ​​(2006); Gadiot et al., Cancer 117:2192-2201 (2011); Taube et al., Sci Transl Med 4,127ra37 (2012); and Toplian et al., New Eng. J Med. 366(26):2443-2454 (2012).

[0064] One approach uses a simple binary endpoint of positive or negative for PD-L1 expression, with a positive result defined in terms of the percentage of tumor cells that show histological evidence of cell surface membrane staining. A tumor tissue section is counted as positive for PD-L1 expression if at least 1%, preferably 5%, of all tumor cells show histological evidence of cell surface membrane staining.

[0065] In another approach, PD-L1 expression in tumor tissue sections is quantified in tumor cells as well as in infiltrating immune cells, which mainly include lymphocytes. The percentage of tumor cells and infiltrating immune cells showing membrane staining is quantified separately as less than 5%, 5 to 9%, and then in 10% increments up to 100%. For tumor cells, PD-L1 expression is counted as negative if the score is less than 5% and positive if the score is 5% or more. PD-L1 expression in immune infiltrates is reported as a semiquantitative measure called the adjusted inflammation score (AIS), which is determined by multiplying the percentage of membrane-stained cells by the intensity of the infiltrate, which is graded as none (0), mild (score 1, rare lymphocytes), moderate (score 2, focal infiltration of the tumor with lymphohistiocytic aggregates), or severe (score 3, diffuse infiltration). If the AIS is ≧5, the tumor tissue section is counted as positive for PD-L1 expression by immune infiltrates.

[0066] Tissue sections from tumors stained by IHC with diagnostic PD-L1 antibodies can also be scored for PD-L1 protein expression by assessing PD-L1 expression on both tumor cells and infiltrating immune cells in the tissue section using a scoring process. See WO 2014 / 165422. One PD-L1 scoring process involves examining each tumor nest in a tissue section for staining and assigning one or both of a Modified H Score (MHS) and a Modified Proportional Score (MPS) to the tissue section. To assign an MHS, four separate percentages are estimated across all viable tumor cells and stained mononuclear inflammatory cells in all tumor nests examined: (a) cells with no staining (intensity=0), (b) weak staining (intensity=1+), (c) moderate staining (intensity=2+), and (d) strong staining (intensity=3+). Cells must have at least partial membrane staining to be included in the weak, moderate, or strong staining percentages. The estimated percentages, whose sum is 100%, are then entered into the formula 1x (percent weakly staining cells) + 2x (percent moderately staining cells) + 3x (percent strongly staining cells) and the result is assigned to the tissue section as the MHS. The MPS is assigned by estimating the percentage of cells with at least partial membrane staining of any intensity across all viable tumor cells and stained mononuclear inflammatory cells in all of the tumor nests examined, and the resulting percentage is assigned to the tissue section as the MPS. In a specific embodiment, a tumor is indicated as positive for PD-L1 expression if the MHS or MPS is positive.

[0067] Another method for scoring / quantifying PD-L1 expression in tumors is the "combined positive score" or "CPS," which refers to an algorithm for determining a PD-L1 expression score from a patient's tumor sample. The CPS is useful for selecting patients for treatment with a particular therapeutic regimen, including treatment methods involving administration of anti-PD-1 antibodies in which expression of PD-L1 is associated with a higher response rate in a particular patient population compared to the same patient population that does not express PD-L1. The CPS is determined by determining the number of viable PD-L1-positive tumor cells, the number of viable PD-L1-negative tumor cells, and the number of viable PD-L1-positive mononuclear inflammatory cells (MIC) in tumor tissue from a patient with a tumor, and calculating the CPS using the following formula:

[0068] (#PD-L1 positive tumor cells) + (#PD-L1 positive MIC) x 100% (#PD-L1 positive tumor cells) + (#PD-L1 negative tumor cells).

[0069] In a particular embodiment, the PD-L1 expression scoring method used is the "Lymphoma Percentage Score." Lymphomas are characterized by a homogenous population of confluent cells that promote lymph node structure or metastatic site structure. The "LPS" or "Lymphoma Percentage Score" is the percentage of this cell population that expresses PD-L1. When determining LPS, no attempt is made to distinguish truly neoplastic from reactive cells. PD-L1 expression is characterized by partial or complete membrane staining of any intensity.

[0070] Yet another method of scoring PD-L1 expression is the "TPS" or "tumor proportion score," which is the proportion of tumor cells that express PD-L1 on the cell membrane. The TPS typically includes the percentage of neoplastic cells that express PD-L1 at any intensity (weak, medium, strong), which can be determined using immunohistochemistry assays using diagnostic anti-human PD-L1 mAbs, such as antibody 20C3 and antibody 22C3 described above. Cells are considered to express PD-L1 if membrane staining is present, including cells with partial membrane staining.

[0071] The level of PD-L mRNA expression can be compared to the mRNA expression levels of one or more reference genes frequently used in quantitative RT-PCR, such as ubiquitin C.

[0072] In a specific embodiment, the level of PD-L1 expression (protein and / or mRNA) by malignant cells and / or infiltrating immune cells within a tumor is determined to be "overexpressed" or "elevated" based on a comparison to the level of PD-L1 expression (protein and / or mRNA) by a suitable control. For example, the control PD-L1 protein or mRNA expression level can be the level quantified in sections from non-malignant cells of the same type or matched normal tissue. In a specific embodiment, PD-L1 expression in a tumor sample is determined to be elevated if PD-L1 protein (and / or PD-L1 mRNA) in the sample is at least 10%, 20%, or 30% greater than the control.

[0073] "Pembro" (formerly known as MK-3475, SCH900475, and lambrolizumab), also referred to herein as "pembro," is a humanized IgG4 mAb having the structure set forth in WHO Drug Information, Vol. 27, No. 2, pages 161-162 (2013), and contains the heavy and light chain amino acid sequences and CDRs set forth in Table 3. Pembrolizumab has been approved by the US FDA as set forth in Prescribing Information for KEYTRUDA™ (Merck & Co., Inc., Whitehouse Station, NJ USA; initial US approval 2014, updated March 2021).

[0074] As used herein, "pembrolizumab variant" refers to a monoclonal antibody that contains the same heavy and light chain sequences as pembrolizumab, except that it has 3, 2 or 1 conservative amino acid substitutions at positions located outside the light chain CDRs and 6, 5, 4, 3, 2 or 1 conservative amino acid substitutions located outside the heavy chain CDRs, e.g., the variant positions are located in the FR region or constant region, and the C-terminal lysine or glycine residue of the heavy chain may be deleted. In other words, pembrolizumab and pembrolizumab variants contain the same CDR sequences, but differ from each other by having conservative amino acid substitutions at no more than 3 or 6 other positions in their full-length light and heavy chain sequences, respectively. Pembrolizumab variants are substantially the same as pembrolizumab in terms of the following properties: binding affinity to PD-1 and ability to block the binding of PD-L1 and PD-L2 to PD-1, respectively.

[0075] A "pharmaceutical formulation" or "pharmaceutical composition" refers to a preparation that is in a form that allows the active ingredient to be effective and that does not contain additional ingredients that are toxic to the subject to which the formulation is administered.

[0076] "Pharmaceutically acceptable" refers to excipients (vehicles, additives) and compositions that may be reasonably administered to a subject to provide an effective amount of the active ingredient being used, e.g., are physiologically tolerated, and typically do not cause an allergic or similar adverse reaction, e.g., stomach upset, when administered to a human. In another embodiment, the term refers to molecular entities and compositions approved by a federal or state government regulatory agency, or listed in the United States Pharmacopeia or another generally recognized pharmacopoeia, for use in animals, and more specifically, in humans.

[0077] Pharmacokinetic "steady state" is the period during which accumulation of drug concentrations with multiple doses is maximized and systemic drug exposure is considered to be uniform after each subsequent dose administered; in the specific case of pembrolizumab, steady state is achieved after approximately 16 weeks of dosing.

[0078] "Platinum-containing chemotherapy" (also known as platinum) refers to the use of chemotherapeutic agent(s) used to treat cancer that are coordination complexes of platinum. Platinum-containing chemotherapeutic agents are alkylating agents that crosslink DNA, resulting in ineffective DNA mismatch repair, generally resulting in apoptosis. Examples of platinums include cisplatin, carboplatin, and oxaliplatin.

[0079] As used herein, "RECIST 1.1 response criteria" means the definitions set forth in Eisenhauer, EA et al., Eur. J. Cancer 45:228-247 (2009), for target lesions or non-target lesions, as appropriate, based on the context in which response is being measured.

[0080] "Therapeutic agent" refers to an additional agent compared to an anti-PD-1 antibody or antigen-binding fragment thereof. The therapeutic agent can be, for example, a chemotherapeutic agent, a biotherapeutic agent, or an immunogenic agent.

[0081] "Tissue section" refers to a single portion or piece of a tissue sample, for example a thin section of tissue cut from a normal tissue or tumor sample.

[0082] "Tumor" as applied to a subject diagnosed with or suspected of having cancer refers to a malignant or potentially malignant neoplasm or mass of tissue of any size, including primary tumors and secondary neoplasms. A solid tumor is an abnormal growth or mass of tissue that usually does not contain cysts or liquid areas. Various types of solid tumors are named for the type of cells that form them. Examples of solid tumors are sarcomas, carcinomas, and lymphomas. Leukemias (cancers of the blood) do not generally form solid tumors (National Cancer Institute, Dictionary of Cancer Terms).

[0083] As used herein, "tumor mutation burden" or "TMB" refers to the number of somatic mutations in a tumor genome and / or the number of somatic mutations per area of ​​the tumor genome. TMB high (or TMB-H) refers to a tumor with a high mutation burden. In a specific embodiment, a tumor is said to be TMB-H if it contains ≧10 mutations / megabase (Mut / Mb). FDA-approved tests, such as FoundationOne® CDx, are available for solid tumors to determine whether a solid tumor is TMB-H (i.e., has ≧10 mutations / megabase).

[0084] As used herein, "variable region" or "V region" refers to the segment of an IgG chain that is variable in sequence among different antibodies. It spans Kabat residue 109 in the light chain and Kabat residue 113 in the heavy chain.

[0085] "PH20" refers to the wild-type PH20 hyaluronidase of SEQ ID NO:16.

[0086] As used herein, a "PH20 variant" is a variant of PH20 having amino acid residue substitutions including M345T, S347T, M348K, K349E, L352Q, L353A, L354I, D355K, N356E, E359D, and I361T of SEQ ID NO:16.

[0087] A "PH20 variant fragment" or "a PH20 variant fragment thereof" or "a fragment of a PH20 variant" is a fragment of a PH20 variant having an N-terminal deletion of any of amino acid residues 1 to 36, 1 to 37, 1 to 38, 1 to 39, 1 to 40, 1 to 41, or 1 to 42 of SEQ ID NO: 16, and / or an N-terminal deletion of any of amino acid residues 455 to 509, 456 to 509, 457 to 509, 458 to 509, 459~509, 460~509, 461~509, 462~509, 463~509, 464~509, 465~509, 466~509, 467~509, 468~509, 469~509, 470~509, 471~509, 472~509, 473~509, 474~509, 475~509, 476~509, 477~509, 478 ~509, 479~509, 480~509, 481~509, 482~509, 483~509, 484~509, 485~509, 486~509, 487~509, 488~509, 489~509, 490~509, 491~509, 492~509, 493~509, 494~509, 495~509, 496~509, 497~50 9, 498-509, 499-509, 500-509, 501-509, 502-509, 503-509, 504-509, 505-509, 506-509, 507-509, 508-509, or 509 C-terminal deletions, where numbering is with reference to SEQ ID NO:16.

[0088] "Unit" or "U" refers to one unit of hyaluronidase activity: the amount of PH20 variant or fragment thereof that causes a change in optical density at 600 nm under conditions suitable for the reaction of hyaluronic acid with the enzyme, calculated according to a calibration curve using an activity standard. An example of the assay is described in Example 4 of US Patent Application Publication No. 2022 / 0089738. Hyaluronic acid (HA) binds to albumin, and the albumin-HA complex produces turbidity. Hydrolysis of HA with hyaluronidase reduces the turbidity of the albumin-HA complex. Thus, the assay measures turbidity to determine the hyaluronidase enzyme activity of the PH20 variant or fragment thereof. Hyaluronidase activity is based on the following reaction: Hyaluronic acid------------>di- and monosaccharides + smaller hyaluronic acid fragments.Those of skill in the art will appreciate that hyaluronidase activity in units per mg of hyaluronidase can vary depending on the purity of the hyaluronidase, the manufacturing process, etc.

[0089] II. PD-1 Antibodies and Antigen-Binding Fragments Useful in the Invention Examples of mAbs that bind human PD-1 useful in the formulations, methods of treatment, compositions and uses of the invention are described in U.S. Patent No. 7,521,051, U.S. Patent No. 8,008,449 and U.S. Patent No. 8,354,509. Particular anti-human PD-1 mAbs useful as PD-1 antagonists or anti-PD-1 antibodies in the methods of treatment, compositions and uses of the invention include: pembrolizumab (formerly known as MK-3475, SCH900475 and lambrolizumab), a humanized IgG4 mAb having a structure described in WHO Drug Information, Vol. 27, No. 2, pages 161-162 (2013).

[0090] In specific embodiments of the therapeutic methods, compositions, kits, and uses of the invention, the anti-PD-1 antibody, or antigen-binding fragment thereof, comprises: (a) light chain CDRs LC-CDR1, LC-CDR2, and LC-CDR3 comprising the sequences of amino acids set forth in SEQ ID NOs: 1, 2, and 3, respectively, and heavy chain CDRs HC-CDR1, HC-CDR2, and HC-CDR3 comprising the sequences of amino acids set forth in SEQ ID NOs: 6, 7, and 8, respectively. In other embodiments, the anti-PD-1 antibody, or antigen-binding fragment thereof, is a humanized antibody. In other embodiments, the anti-PD-1 antibody, or antigen-binding fragment thereof, is a chimeric antibody. In a specific embodiment, the anti-PD-1 antibody, or antigen-binding fragment thereof, is a monoclonal antibody.

[0091] In other embodiments of the therapeutic methods, compositions, kits, and uses of the invention, the anti-PD-1 antibody, or antigen-binding fragment thereof, that specifically binds to human PD-1 comprises (a) a heavy chain variable region comprising the amino acid sequence set forth in SEQ ID NO:9, or a variant thereof, and (b) a light chain variable region comprising an amino acid sequence selected from the group consisting of SEQ ID NO:4, or a variant thereof. In one embodiment, the anti-PD-1 antibody, or antigen-binding fragment thereof, that specifically binds to human PD-1 comprises (a) a heavy chain variable region comprising the amino acid sequence set forth in SEQ ID NO:9, and (b) a light chain variable region comprising the amino acid sequence of SEQ ID NO:4.

[0092] A variant of a heavy chain variable region sequence or a full-length heavy chain sequence is identical to a reference sequence except for up to 17 conservative amino acid substitutions in the framework regions (i.e. outside the CDRs), preferably having fewer than 10, 9, 8, 7, 6 or 5 conservative amino acid substitutions in the framework regions. A variant of a light chain variable region sequence or a full-length light chain sequence is identical to a reference sequence except for up to 5 conservative amino acid substitutions in the framework regions (i.e. outside the CDRs), preferably having fewer than 4, 3 or 2 conservative amino acid substitutions in the framework regions.

[0093] In another embodiment of the therapeutic methods, compositions, kits, and uses of the invention, the anti-PD-1 antibody, or antigen-binding fragment thereof, is a monoclonal antibody that specifically binds to human PD-1 and comprises (a) a heavy chain comprising, or consisting of, an amino acid sequence as set forth in any one of SEQ ID NOs: 10-15, or a variant thereof, and (b) a light chain comprising, or consisting of, an amino acid sequence as set forth in SEQ ID NO: 5, or a variant thereof. In one embodiment, the anti-PD-1 antibody, or antigen-binding fragment thereof, is a monoclonal antibody that specifically binds to human PD-1 and comprises (a) a heavy chain comprising, or consisting of, an amino acid sequence as set forth in any one of SEQ ID NOs: 10-15, and (b) a light chain comprising an amino acid sequence as set forth in SEQ ID NO: 5.

[0094] In yet another embodiment of the therapeutic methods, compositions, kits, and uses of the invention, the anti-PD-1 antibody, or antigen-binding fragment thereof, is a monoclonal antibody that specifically binds to human PD-1 and comprises (a) a heavy chain that comprises, or consists of, the sequence of amino acids set forth in SEQ ID NO:11, and (b) a light chain that comprises, or consists of, the sequence of amino acids set forth in SEQ ID NO:5.

[0095] Table 3 below provides a listing of exemplary amino acid sequences of anti-PD-1 mAbs for use in the therapeutic methods, compositions, kits and uses of the invention. [Table 3] TIFF2025504278000005.tif144153

[0096] As known to those skilled in the art, pyroglutamic acid is the conjugate acid of pyroglutamic acid and is in equilibrium with pyroglutamic acid in solution.

[0097] III. Human Hyaluronidase There are six known hyaluronidases in humans: Hyall, Hyal2, Hyal3, Hyal4, HyalPS1 and PH20 / SPAM1. Recombinant forms of these hyaluronidases with modifications, mutations, additions, truncations can be used in the disclosed methods, uses, compositions and kits. See, for example, U.S. Pat. Nos. 7,767,429, 8,431,380, 7,871,607, WO 2020 / 022791, U.S. Patent Application Publication No. 2006 / 0104968 and EP 1858926, as well as numerous other patents and publications. Examples of such agents are the known agents PEGPH20 or rHuPH20. The methods, uses, compositions and kits of the present invention encompass the use of any human hyaluronidase or fragment thereof, or variants or fragments thereof.

[0098] PH20 variants and fragments thereof In one embodiment, the PH20 variant or fragment thereof further comprises an amino acid residue substitution at one or more positions selected from the group consisting of T341, L342, S343, I344, and N363. In one embodiment, the PH20 variant or fragment thereof further comprises one or more amino acid residue substitutions selected from the group consisting of T341A, T341C, T341D, T341G, T341S, L342W, S343E, I344N, and N363G.

[0099] In one embodiment of the PH20 variant or fragment thereof, the amino acid residue substitutions are selected from the group consisting of the following amino acid residue substitutions:

[0100] (a) T341S, L342W, S343E, I344N, M345T, S347T, M348K, K349E, L352Q, L353A, L354I, D355K, N356E, E359D and I361T; (b) L342W, S343E, I344N, M345T, S347T, M348K, K349E, L352Q, L353A, L354I, D355K, N356E, E359D and I361T; (c) M345T, S347T, M348K, K349E, L352Q, L353A, L354I, D355K, N356E, E359D, I361T and N363G; (d) T341G, L342W, S343E, I344N, M345T, S347T, M348K, K349E, L352Q, L353A, L354I, D355K, N356E, E359D and I361T; (e) T341A, L342W, S343E, I344N, M345T, S347T, M348K, K349E, L352Q, L353A, L354I, D355K, N356E, E359D and I361T; (f) T341C, L342W, S343E, I344N, M345T, S347T, M348K, K349E, L352Q, L353A, L354I, D355K, N356E, E359D and I361T; (g)T341D, L342W, S343E, I344N, M345T, S347T, M348K, K349E, L352Q, L353A, L354I, D355K, N356E, E359D and I361T; (h) I344N, M345T, S347T, M348K, K349E, L352Q, L353A, L354I, D355K, N356E, E359D, and I361T; and (i)S343E, I344N, M345T, S347T, M348K, K349E, L352Q, L353A, L354I, D355K, N356E, E359D and I361T

[0101] In one embodiment of the PH20 variant or fragment thereof, the amino acid residue substitutions consist of T341S, L342W, S343E, I344N, M345T, S347T, M348K, K349E, L352Q, L353A, L354I, D355K, N356E, E359D, and I361T.

[0102] In one aspect of the foregoing embodiments of the PH20 variant fragment, the PH20 variant fragment has an N-terminal deletion of amino acid residues 1-36, 1-37, 1-38, 1-39, 1-40, 1-41, or 1-42 of SEQ ID NO: 16. In another embodiment, the PH20 variant fragment has an N-terminal deletion of amino acid residues 1-36 of SEQ ID NO: 16. In another embodiment, the PH20 variant fragment has an N-terminal deletion of amino acid residues 1-37 of SEQ ID NO: 16. In another embodiment, the PH20 variant fragment has an N-terminal deletion of amino acid residues 1-38 of SEQ ID NO: 16.

[0103] In another aspect of the foregoing embodiment of the PH20 variant fragment, the PH20 variant fragment comprises (one or more) amino acid residues 455-509, 456-509, 457-509, 458-509, 459-509, 460-509, 461-509, 462-509, 463-509, 464-509, 465-509, 466-509, 467-509, 468-509, 469-509, 470-509, 471-509, 472-509, 473-509, 474-509, 475-509, 476-509, 477-509, 478-509, 479-509, 480-509, 481~509, 482~509, 483~509, 484~509, 485~509, 486~509, 487~509, 488~509, 489~509, 490~509, 491~509, 492~509, 493~509, 494~509, 495~509, 496~509, 497 and having a C-terminal deletion of 509, 498-509, 499-509, 500-509, 501-509, 502-509, 503-509, 504-509, 505-509, 506-509, 507-509, 508-509, or 509, where numbering refers to SEQ ID NO:16. In one embodiment, the PH20 variant fragment has a C-terminal deletion of amino acid residues 455-509, 458-509, 461-509, 464-509, 465-509, 466-509, 467-509, 468-509, 470-509, 471-509, 472-509, 473-509, 474-509, 475-509, 476-509, 478-509, 480-509, 482-509, 484-509, 486-509, 488-509, or 490-509, numbering with reference to SEQ ID NO: 16. In one embodiment, the PH20 variant fragment has a C-terminal deletion of amino acid residues 468-509, numbering with reference to SEQ ID NO: 16.

[0104] In one embodiment, the PH20 variant fragment consists of the amino acid sequence set forth in SEQ ID NO: 17 or 18. In other embodiments, the PH20 variant or fragment thereof is any of the sequences disclosed in Table 11 of EP 3636752. [Table 4]

[0105] In one aspect of the methods, compositions, kits, and uses of the invention, the pharmaceutical composition comprises about 165 mg / mL of an anti-human PD-1 antibody or antigen-binding fragment thereof, about 10 mM histidine buffer; about 10 mM L-methionine or a pharma- ceutical acceptable salt thereof, about 7% w / v sucrose, about 0.02% w / v polysorbate 80, and a PH20 variant or fragment. In one embodiment, the pharmaceutical composition comprises about 130 mg / mL of an anti-human PD-1 antibody or antigen-binding fragment thereof, about 10 mM histidine buffer, about 10 mM L-methionine or a pharma- ceutical acceptable salt thereof, about 7% w / v sucrose, about 0.02% w / v polysorbate 80, and a PH20 variant or fragment.

[0106] In a specific embodiment of the pharmaceutical composition, the PH20 variant or fragment thereof is present at a concentration of about 1000 U / ml. In another embodiment, the concentration of the PH20 variant or fragment thereof is about 1500 U / ml. In another embodiment, the concentration of the PH20 variant or fragment thereof is about 2000 U / ml. In another embodiment, the concentration of the PH20 variant or fragment thereof is about 3000 U / ml. In another embodiment, the concentration of the PH20 variant or fragment thereof is about 4000 U / ml. In another embodiment, the concentration of the PH20 variant or fragment thereof is about 5000 U / ml. In another embodiment, the concentration of the PH20 variant or fragment thereof is about 6000 U / ml. In a further embodiment, the concentration of the PH20 variant or fragment thereof is about 1000-6000 U / ml. In a further embodiment, the concentration of the PH20 variant or fragment thereof is about 2000-5000 U / ml.

[0107] In a specific embodiment of the pharmaceutical composition, the PH20 variant or fragment thereof is present at a concentration of about 150 U / ml. In another embodiment, the concentration of the PH20 variant or fragment thereof is about 300 U / ml. In another embodiment, the concentration of the PH20 variant or fragment thereof is about 600 U / ml. In another embodiment, the concentration of the PH20 variant or fragment thereof is about 750 U / ml. In a further embodiment, the concentration of the PH20 variant or fragment thereof is about 150-5000 U / ml. In a further embodiment, the concentration of the PH20 variant or fragment thereof is about 500-8000 U / ml.

[0108] In other aspects of the methods, compositions, kits and uses of the invention, the pharmaceutical composition comprises about 165 mg / mL of an anti-human PD-1 antibody and 2000 U / mL of a PH20 variant or fragment. In one embodiment, the pharmaceutical composition comprises about 130 mg / mL of an anti-human PD-1 antibody and 2000 U / mL of a PH20 variant or fragment.

[0109] rHuPH20 and its fragments rHuPH20, also known as Hylenex®, consists of the amino acid sequence in SEQ ID NO:20, which is amino acid residues 36-482 of wild-type human PH20 in SEQ ID NO:16 (amino acid residues 1-36 are the signal peptide sequence). In one embodiment, rHuPH20 or a variant or fragment comprises amino acid residues 36-464, 36-465, 36-466, 36-467, 36-468, 36-469, 36-470, 36-471, 36-472, 36-473, 36-474, 36-475, 36-476, 36-477, 36-478, 36-479, 36-480, 36-481, 36-482, or 36-483 of SEQ ID NO:16; 464, 37-465, 37-466, 37-467, 37-468, 37-469, 37-470, 37-471, 37-472, 37-473, 37-474, 37-475, 37-476, 37-477, 37-478, 37-479, 37-480, 37-481, 37-482, or 37-483; amino acid residues 38-464, 38-465, 38-466, 38-467, 38-468, 38-469, 38-470, 38-471, 37-472, 37-473, 37-474, 37-475, 37-476, 37-477, 37-478, 37-479, 37-480, 37-481, 37-482, or 37-483 of SEQ ID NO: 16 8-471, 38-472, 38-473, 38-474, 38-475, 38-476, 38-477, 38-478, 38-479, 38-480, 38-481, 38-482, or 38-483; amino acid residues 39-464, 39-465, 39-466, 39-467, 39-468, 39-469, 39-470, 39-471, 39-472, 39-473, 39-474, 39-475, 39-476, 39-477, 39-478, 38-479, 38-480, 38-481, 38-482, or 38-483 of SEQ ID NO: 16 7, 39-478, 39-479, 39-480, 39-481, 39-482, or 39-483; amino acid residues 40-464, 40-465, 40-466, 40-467, 40-468, 40-469, 40-470, 40-471, 40-472, 40-473, 40-474, 40-475, 40-476, 40-477, 40-478, 40-479, 40-480, 40-481, 340-482, or 40-483 of SEQ ID NO:16;amino acid residues 41 to 464, 41 to 465, 41 to 466, 41 to 467, 41 to 468, 41 to 469, 41 to 470, 41 to 471, 41 to 472, 41 to 473, 41 to 474, 41 to 475, 41 to 476, 41 to 477, 41 to 478, 41 to 479, 41 to 480, 41 to 481, 41 to 482, or 41 to 483 of SEQ ID NO: 16; or are amino acid residues 42-464, 42-465, 42-466, 42-467, 42-468, 42-469, 42-470, 42-471, 42-472, 42-473, 42-474, 42-475, 42-476, 42-477, 42-478, 42-479, 42-480, 42-481, 42-482, or 42-483 of SEQ ID NO: 16. In a preferred embodiment, the rHuPH20 variant consists of amino acid residues 36-483 of SEQ ID NO: 16 (which is SEQ ID NO: 19). In a preferred embodiment, the rHuPH20 fragment consists of amino acid residues 36-477 of SEQ ID NO: 16. In a preferred embodiment, the rHuPH20 fragment consists of amino acid residues 36-478 of SEQ ID NO: 16. In a preferred embodiment, the rHuPH20 fragment consists of amino acid residues 36-479 of SEQ ID NO: 16. In a preferred embodiment, the rHuPH20 fragment consists of amino acid residues 36-480 of SEQ ID NO: 16. In a preferred embodiment, the rHuPH20 fragment consists of amino acid residues 36-481 of SEQ ID NO: 16. In a further embodiment, the rHuPH20 variant or fragment is one disclosed in U.S. Patent No. 7,767,429, which is incorporated herein by reference in its entirety;

[0110] [Table 5]

[0111] In one aspect, the methods, compositions, kits, and uses of the invention comprise about 165 mg / mL of an anti-human PD-1 antibody, about 10 mM histidine buffer; about 10 mM L-methionine or a pharma- ceutically acceptable salt thereof, about 7% w / v sucrose, about 0.02% w / v polysorbate 80, and rHuPH20 or a variant or fragment. In one embodiment, the pharmaceutical composition comprises about 130 mg / mL of an anti-human PD-1 antibody, or an antigen-binding fragment thereof, about 10 mM histidine buffer, about 10 mM L-methionine or a pharma- ceutically acceptable salt thereof, about 7% w / v sucrose, about 0.02% w / v polysorbate 80, and a rHuPH20 variant or fragment.

[0112] In a specific embodiment of the pharmaceutical composition, rHuPH20 or a variant or fragment thereof is present at a concentration of about 1000 U / ml. In another embodiment, the concentration of rHuPH20 or a variant or fragment thereof is about 1500 U / ml. In another embodiment, the concentration of rHuPH20 or a variant or fragment thereof is about 2000 U / ml. In another embodiment, the concentration of rHuPH20 or a variant or fragment thereof is about 3000 U / ml. In another embodiment, the concentration of rHuPH20 or a variant or fragment thereof is about 4000 U / ml. In another embodiment, the concentration of rHuPH20 or a variant or fragment thereof is about 5000 U / ml. In another embodiment, the concentration of rHuPH20 or a variant or fragment thereof is about 6000 U / ml. In a further embodiment, the concentration of rHuPH20 or a variant or fragment thereof is about 1000-6000 U / ml. In a further embodiment, the concentration of rHuPH20, or a variant or fragment thereof, is about 2000-5000 U / ml.

[0113] In a specific embodiment of the pharmaceutical composition, rHuPH20 or a variant or fragment thereof is present at a concentration of about 150 U / ml. In another embodiment, the concentration of rHuPH20 or a variant or fragment thereof is about 300 U / ml. In another embodiment, the concentration of rHuPH20 or a variant or fragment thereof is about 600 U / ml. In another embodiment, the concentration of rHuPH20 or a variant or fragment thereof is about 750 U / ml. In a further embodiment, the concentration of rHuPH20 or a variant or fragment thereof is about 150-5000 U / ml. In a further embodiment, the concentration of rHuPH20 or a variant or fragment thereof is about 500-8000 U / ml.

[0114] In another aspect, the methods, compositions, kits, and uses of the invention employ a pharmaceutical composition comprising about 165 mg / mL of an anti-human PD-1 antibody or antigen-binding fragment thereof, and about 2000 U / ml of rHuPH20 or a variant or fragment. In one embodiment, the pharmaceutical composition comprises about 130 mg / mL of an anti-human PD-1 antibody or antigen-binding fragment thereof, and about 2000 U / ml of rHuPH20 or a variant or fragment.

[0115] IV. METHODS AND USES OF THE PRESENT ART The present invention provides a method of treating cancer in a human patient in need of such treatment, comprising administering to the patient subcutaneously about every six weeks an anti-PD-1 antibody or antigen-binding fragment thereof and human hyaluronidase at a dose of about 600 mg to about 1000 mg. In another aspect, the present invention provides a use of an anti-PD-1 antibody or antigen-binding fragment thereof and human hyaluronidase to prepare a medicament for treating cancer in a human patient, comprising administering to the patient subcutaneously about every six weeks an anti-PD-1 antibody or antigen-binding fragment thereof at a dose of about 600 mg to about 1000 mg. In another aspect, the present invention provides a use of an anti-PD-1 antibody or antigen-binding fragment thereof to prepare a medicament for treating cancer in a human patient, comprising administering to the patient subcutaneously about every six weeks an anti-PD-1 antibody or antigen-binding fragment thereof at a dose of about 600 mg to about 1000 mg. In another aspect, the invention provides a pharmaceutical composition comprising an anti-PD-1 antibody or antigen-binding fragment thereof and human hyaluronidase for treating cancer in a human patient, wherein the patient is administered subcutaneously a dose of about 600 mg to about 1000 mg of the anti-PD-1 antibody or antigen-binding fragment thereof and human hyaluronidase approximately every six weeks.In another aspect, the invention provides a pharmaceutical composition comprising an anti-PD-1 antibody or antigen-binding fragment thereof for treating cancer in a human patient, wherein the patient is administered subcutaneously a dose of about 600 mg to about 1000 mg of the anti-PD-1 antibody or antigen-binding fragment thereof and human hyaluronidase approximately every six weeks.

[0116] The invention also provides a method of treating cancer in a human patient in need of such treatment, comprising subcutaneously administering to the patient an anti-PD-1 antibody or antigen-binding fragment thereof at a dose of about 600 mg to about 1000 mg about every six weeks. In another aspect, the invention provides a use of an anti-PD-1 antibody or antigen-binding fragment thereof for the preparation of a medicament for treating cancer in a human patient, comprising subcutaneously administering to the patient an anti-PD-1 antibody or antigen-binding fragment thereof at a dose of about 600 mg to about 1000 mg about every six weeks. In another aspect, the invention provides an anti-PD-1 antibody or antigen-binding fragment thereof for treating cancer in a human patient, wherein the patient is subcutaneously administered an anti-PD-1 antibody or antigen-binding fragment thereof at a dose of about 600 mg to about 1000 mg about every six weeks.

[0117] The present invention further provides a method of treating cancer in a human patient in need of such treatment, comprising subcutaneously administering to the patient about 300 mg to about 500 mg of an anti-PD-1 antibody or antigen-binding fragment thereof and human hyaluronidase approximately every three weeks. In another aspect, the present invention provides the use of an anti-PD-1 antibody or antigen-binding fragment thereof and human hyaluronidase to prepare a medicament for treating cancer in a human patient, comprising subcutaneously administering to the patient a dose of about 300 mg to about 500 mg of the anti-PD-1 antibody or antigen-binding fragment thereof and human hyaluronidase approximately every three weeks. In another aspect, the present invention provides the use of an anti-PD-1 antibody or antigen-binding fragment thereof to prepare a medicament for treating cancer in a human patient, comprising subcutaneously administering to the patient a dose of about 300 mg to about 500 mg of the anti-PD-1 antibody or antigen-binding fragment thereof and human hyaluronidase approximately every three weeks. In another aspect, the invention provides a pharmaceutical composition comprising an anti-PD-1 antibody or antigen-binding fragment thereof and human hyaluronidase for treating cancer in a human patient, wherein the patient is administered a dose of about 300 mg to about 500 mg of the anti-PD-1 antibody or antigen-binding fragment thereof and human hyaluronidase subcutaneously about every three weeks.In another aspect, the invention provides a pharmaceutical composition comprising an anti-PD-1 antibody or antigen-binding fragment thereof for treating cancer in a human patient, wherein the patient is administered a dose of about 300 mg to about 500 mg of the anti-PD-1 antibody or antigen-binding fragment thereof and human hyaluronidase subcutaneously about every three weeks.

[0118] In certain embodiments of the invention, the anti-PD-1 antibody or antigen-binding fragment thereof is pembrolizumab, hi other embodiments, the anti-PD-1 antibody or antigen-binding fragment thereof is a pembrolizumab variant.

[0119] In any embodiment of the methods or uses of the invention, the bioavailability of the anti-PD-1 antibody or antigen-binding fragment thereof in combination with human hyaluronidase is 55-60%. In any embodiment of the methods or uses described herein, the bioavailability of the anti-PD-1 antibody or antigen-binding fragment thereof in combination with human hyaluronidase is 57-59%. In any embodiment of the methods or uses described herein, the bioavailability of the anti-PD-1 antibody or antigen-binding fragment thereof in combination with human hyaluronidase is 57%.

[0120] In any embodiment of the methods or uses of the invention, the subcutaneous administration of the anti-PD-1 antibody or antigen-binding fragment thereof comprises a C of 400 mg dose of the anti-PD-1 antibody or antigen-binding fragment thereof administered by intravenous (IV) route every 6 weeks. trough C is within 20% of trough In another embodiment, subcutaneous administration of the anti-PD-1 antibody or antigen-binding fragment thereof results in a C of 400 mg dose of the anti-PD-1 antibody or antigen-binding fragment thereof administered by an intravenous (IV) route every 6 weeks. trough At least equal to or less than 35% greater than C trough In another embodiment, subcutaneous administration of the anti-PD-1 antibody or antigen-binding fragment thereof results in a C of 400 mg dose of the anti-PD-1 antibody or antigen-binding fragment thereof administered by an intravenous (IV) route every 6 weeks. trough At least equal to or less than 30% greater than C trough In another embodiment, subcutaneous administration of the anti-PD-1 antibody or antigen-binding fragment thereof results in a C of 400 mg dose of the anti-PD-1 antibody or antigen-binding fragment thereof administered by an intravenous (IV) route every 6 weeks. trough C is about 25-30% larger than trough In another embodiment, subcutaneous administration of the anti-PD-1 antibody or antigen-binding fragment thereof results in a C of 400 mg dose of the anti-PD-1 antibody or antigen-binding fragment thereof administered by an intravenous (IV) route every 6 weeks. trough C is about 30% larger than troughIn another embodiment, subcutaneous administration of the anti-PD-1 antibody or antigen-binding fragment thereof results in a C of 400 mg dose of the anti-PD-1 antibody or antigen-binding fragment thereof administered by an intravenous (IV) route every 6 weeks. trough C is about 20-35% larger than trough results.

[0121] In a specific embodiment of the methods or uses of the invention, the subcutaneous administration of a dose of an anti-PD-1 antibody (e.g., pembrolizumab) or an antigen-binding fragment thereof is administered by an IV route of administration of 400 mg Q6W. trough C equal to or greater than trough In one embodiment of any of the methods or uses described herein, the subcutaneous administration of the anti-PD-1 antibody, or antigen-binding fragment thereof, results in IVC trough Subcutaneous C trough In a specific embodiment, the subcutaneous administration has a ratio (e.g., geometric mean ratio) of at least 0.8, at least 1, at least 1.2, at least 1.3, at least 1.4, at least 1.5, or at least 1.6 of SC:IV C of at least 0.8 or greater. trough In a specific embodiment, subcutaneous administration results in a PK profile having an SC:IV C ratio of at least 1.0 or greater. trough In a specific embodiment, subcutaneous administration results in a PK profile having an SC:IV C ratio of at least 1.2 or greater. trough In a specific embodiment, subcutaneous administration results in an SC:IV C ratio of at least 1.3 or greater. trough In a specific embodiment, subcutaneous administration results in an SC:IV C ratio of at least 1.4 or greater. trough In a specific embodiment, subcutaneous administration results in a SC:IV C ratio of at least 1.5 or greater. trough In a specific embodiment, subcutaneous administration results in an SC:IV C ratio of at least 1.6 or greater. trough This results in a ratio.

[0122] In a specific embodiment of the methods or uses of the invention, subcutaneous administration of a dose of an anti-PD-1 antibody (e.g., pembrolizumab) or antigen-binding fragment thereof is achieved with a SC:IV C ratio of 0.8 to 1.6, 1.0 to 1.6, 1.1 to 1.6, 1.2 to 1.6, 1.3 to 1.6, 1.4 to 1.6, 1.2 to 1.5, 1.3 to 1.5, 1.4 to 1.5 or 1.3 to 1.4. trough This results in a ratio.

[0123] In a specific embodiment of the methods or uses of the invention, the subcutaneous administration of a dose of an anti-PD-1 antibody (e.g., pembrolizumab) or an antigen-binding fragment thereof is at a SC:IV C ratio of 1.0 to 1.6. trough In a specific embodiment, subcutaneous administration results in a SC:IV C ratio of 1.1 to 1.6. trough In a specific embodiment, subcutaneous administration results in a SC:IV C ratio of 1.2 to 1.6. trough In a specific embodiment, subcutaneous administration results in a SC:IV C ratio of 1.3 to 1.6. trough In a specific embodiment, subcutaneous administration results in a SC:IV C ratio of 1.4 to 1.6. trough In a specific embodiment, subcutaneous administration results in a SC:IV C ratio of 1.2 to 1.5. trough In a specific embodiment, subcutaneous administration results in a SC:IV C ratio of 1.3 to 1.5. trough In a specific embodiment, subcutaneous administration results in a SC:IV C ratio of 1.4 to 1.5. trough In a specific embodiment, subcutaneous administration results in a SC:IV C ratio of 1.3 to 1.4. trough In a specific embodiment, subcutaneous administration results in a SC:IV C ratio of about 1.2:1.3. trough In a specific embodiment, subcutaneous administration results in a SC:IV C ratio of about 1.2:1.4. trough This results in a ratio.

[0124] In a specific embodiment of the methods or uses of the invention, subcutaneous administration of a dose of an anti-PD-1 antibody (e.g., pembrolizumab) or antigen-binding fragment thereof is greater than or equal to the AUC of a 400 mg dose of an anti-PD-1 antibody or antigen-binding fragment thereof administered by IV route Q6W at cycle 1 or steady state(0~6週) AUC ratio of at least 0.8 (0~6週) In a specific embodiment of the methods or uses of the invention, subcutaneous administration of a dose of an anti-PD-1 antibody (e.g., pembrolizumab) or antigen-binding fragment thereof results in an AUC (0~6週) AUC of about 1.0 ratio (0~6週) In a specific embodiment of the methods or uses of the invention, subcutaneous administration of a dose of an anti-PD-1 antibody (e.g., pembrolizumab) or antigen-binding fragment thereof results in an AUC (0~6週) AUC of about 0.8 to 1.2 (0~6週) In a specific embodiment of the methods or uses of the invention, subcutaneous administration of a dose of an anti-PD-1 antibody (e.g., pembrolizumab) or antigen-binding fragment thereof results in an AUC (0~6週) AUC for ratios of approximately 1.0 to 1.1 (0~6週) In a specific embodiment, subcutaneous administration results in an SC:IV AUC of at least 0.8, 1.0 or greater after six cycles of administration. (0~6週) The PK profile having ratios is obtained.

[0125] In any embodiment of the methods or uses of the invention, subcutaneous administration of an anti-PD-1 antibody (e.g., pembrolizumab) co-formulated with human hyaluronidase (e.g., PH20 variant 2) at 760-790 mg every 6 weeks is approximately equal to the C of a 400 mg dose of an anti-PD-1 antibody administered by an intravenous (IV) route every 6 weeks. trough Within 20% of C troughIn a specific embodiment of the methods or uses of the invention, subcutaneous administration of an anti-PD-1 antibody (e.g., pembrolizumab) co-formulated with human hyaluronidase (e.g., PH20 variant 2) at 760-790 mg every 6 weeks results in a subcutaneous C of approximately 1.0 to 1.35 of a dose administered by the IV route of administration of 400 mg Q6W. trough IV C trough In one embodiment of the foregoing embodiments, the subcutaneous administration of an anti-PD-1 antibody (e.g., pembrolizumab) co-formulated with PH20 variant 2 is 790 mg every 6 weeks. In a preferred embodiment of the method or use of the invention, the subcutaneous administration of an anti-PD-1 antibody (e.g., pembrolizumab) co-formulated with PH20 variant 2 at 790 mg every 6 weeks results in a subcutaneous C of about 1.3 of a dose administered by IV route of administration of 400 mg Q6W. trough IV C trough (e.g., geometric mean ratio).

[0126] In a specific embodiment of the methods or uses of the invention, subcutaneous administration of an anti-PD-1 antibody (e.g., pembrolizumab) co-formulated with human hyaluronidase (e.g., PH20 variant 2) at 760-790 mg every 6 weeks reduces the AUC (0~6週) AUC of about 0.8 to 1.2 ratio (0~6週) In a specific embodiment of the methods or uses of the invention, subcutaneous administration of an anti-PD-1 antibody (e.g., pembrolizumab) co-formulated with human hyaluronidase (e.g., PH20 variant 2) at 760-790 mg every 6 weeks results in an AUC (0~6週) AUC of approximately 1.0 to 1.2 ratio (0~6週)In a specific embodiment of the methods or uses of the invention, subcutaneous administration of an anti-PD-1 antibody (e.g., pembrolizumab) co-formulated with human hyaluronidase (e.g., PH20 variant 2) at 760-790 mg every 6 weeks results in an AUC (0~6週) AUC of approximately 1.0 to 1.1 ratio (0~6週) In one embodiment of the foregoing embodiments, the subcutaneous administration of an anti-PD-1 antibody (e.g., pembrolizumab) co-formulated with PH20 variant 2 is 790 mg every 6 weeks. In a preferred embodiment of the method or use of the invention, the subcutaneous administration of an anti-PD-1 antibody (e.g., pembrolizumab) co-formulated with PH20 variant 2 at 790 mg every 6 weeks results in an AUC (0~6週) AUC of about 1.1 ratio (0~6週) results.

[0127] In any embodiment of the methods or uses of the invention, subcutaneous administration of an anti-PD-1 antibody (e.g., pembrolizumab) co-formulated with human hyaluronidase (e.g., PH20 variant 2) at 760-790 mg every 6 weeks is administered at a C of 400 mg dose of an anti-PD-1 antibody administered by intravenous (IV) route every 6 weeks in Cycle 1. max C is about 40% lower than max In any embodiment of the methods or uses of the invention, subcutaneous administration of an anti-PD-1 antibody (e.g., pembrolizumab) co-formulated with human hyaluronidase (e.g., PH20 variant 2) at 760-790 mg every 6 weeks results in a C of 400 mg dose of anti-PD-1 antibody administered by intravenous (IV) route every 6 weeks in cycle 1. max C, which is about 35-38% lower than maxIn any embodiment of the methods or uses of the invention, subcutaneous administration of an anti-PD-1 antibody (e.g., pembrolizumab) co-formulated with human hyaluronidase (e.g., PH20 variant 2) at 760-790 mg every 6 weeks results in a C of a 400 mg dose of an anti-PD-1 antibody or antigen-binding fragment thereof administered by an intravenous (IV) route every 6 weeks at steady state. max C is about 20-30% lower than max In any embodiment of the methods or uses of the invention, subcutaneous administration of an anti-PD-1 antibody (e.g., pembrolizumab) co-formulated with human hyaluronidase (e.g., PH20 variant 2) at 760-790 mg every 6 weeks results in a C of approximately 1.2 mg / kg / day of a 400 mg dose of an anti-PD-1 antibody administered by an intravenous (IV) route every 6 weeks at steady state. max C, which is about 22-25% lower than max In one embodiment of the foregoing embodiments, the subcutaneous administration of an anti-PD-1 antibody (e.g., pembrolizumab) co-formulated with human hyaluronidase (e.g., PH20 variant 2) is 790 mg every 6 weeks. In any embodiment of the methods or uses of the invention, the subcutaneous administration of an anti-PD-1 antibody (e.g., pembrolizumab) co-formulated with PH20 variant 2 at 790 mg every 6 weeks results in a C of a 400 mg dose of anti-PD-1 antibody administered by an intravenous (IV) route of administration every 6 weeks at steady state. max C, which is about 22% lower than max In any embodiment of the methods or uses of the invention, subcutaneous administration of 790 mg of an anti-PD-1 antibody (e.g., pembrolizumab) co-formulated with PH20 variant 2 every 6 weeks results in a C of 400 mg dose of an anti-PD-1 antibody administered by intravenous (IV) route every 6 weeks in Cycle 1. max C, which is about 35% lower than max results.

[0128] In any embodiment of the methods or uses of the invention, subcutaneous administration of an anti-PD-1 antibody (e.g., pembrolizumab) co-formulated with human hyaluronidase (e.g., PH20 variant 2) at 380-395 mg every three weeks is comparable to the C of a 400 mg dose of an anti-PD-1 antibody administered by an intravenous (IV) route every six weeks. trough Within 20% of C trough In a specific embodiment of the methods or uses of the invention, subcutaneous administration of an anti-PD-1 antibody (e.g., pembrolizumab) co-formulated with human hyaluronidase (e.g., PH20 variant 2) at 380-395 mg every 3 weeks results in a subcutaneous C of approximately 1.0 to 2.0 of the dose administered by the 400 mg Q6W IV route of administration. trough IV C trough In one embodiment of the foregoing embodiments, the subcutaneous administration of an anti-PD-1 antibody (e.g., pembrolizumab) co-formulated with PH20 variant 2 is 395 mg every 3 weeks. In a preferred embodiment of the method or use of the invention, the subcutaneous administration of an anti-PD-1 antibody (e.g., pembrolizumab) co-formulated with PH20 variant 2 at 395 mg every 3 weeks results in a subcutaneous C of about 1.5 to 2.0 of a dose administered by IV route of administration of 400 mg Q6W. trough IV C trough (e.g., geometric mean ratio).

[0129] In a specific embodiment of the methods or uses of the invention, subcutaneous administration of an anti-PD-1 antibody (e.g., pembrolizumab) co-formulated with human hyaluronidase (e.g., PH20 variant 2) at 380-395 mg every 3 weeks reduces the AUC (0~6週) AUC of about 0.8 to 1.2 ratio (0~6週) In a specific embodiment of the methods or uses of the invention, subcutaneous administration of a dose of an anti-PD-1 antibody (e.g., pembrolizumab) co-formulated with human hyaluronidase (e.g., PH20 variant 2) at 380-395 mg every 3 weeks results in an AUC(0~6週) AUC of about 0.8 to 1.1 (0~6週) In one embodiment of the foregoing embodiments, the subcutaneous administration of an anti-PD-1 antibody (e.g., pembrolizumab) co-formulated with PH20 variant 2 is 395 mg every 3 weeks. In a preferred embodiment of the method or use of the invention, the subcutaneous administration of an anti-PD-1 antibody (e.g., pembrolizumab) co-formulated with PH20 variant 2 at 395 mg every 3 weeks results in an AUC (0~6週) AUC of about 1.1 ratio (0~6週) results.

[0130] In one embodiment of the above embodiment, the ratio is a geometric mean ratio. In one embodiment of the above embodiment, the ratio is cycle 1 (week 3 or week 6 depending on the subcutaneous administration cycle). In one embodiment of the above embodiment, the ratio is at steady state. In one embodiment of the above embodiment, the ratio is during treatment.

[0131] In specific embodiments of the methods or uses of the present invention, the cancer is selected from the group consisting of melanoma, lung cancer, head and neck cancer, bladder cancer, breast cancer, gastrointestinal cancer, multiple myeloma, hepatocellular carcinoma, Merkel cell carcinoma, cutaneous squamous cell carcinoma, lymphoma, renal cancer, mesothelioma, ovarian cancer, esophageal cancer, anal cancer, biliary tract cancer, colorectal cancer, endometrial cancer, cervical cancer, thyroid cancer, salivary cancer, prostate cancer (e.g. hormone refractory prostate adenocarcinoma), pancreatic cancer, colon cancer, liver cancer, thyroid cancer, glioblastoma, glioma, and other neoplastic malignancies.

[0132] In a specific embodiment, the lung cancer is non-small cell lung cancer.

[0133] In a specific embodiment, the lung cancer is small cell lung cancer.

[0134] In a specific embodiment, the lymphoma is Hodgkin's lymphoma.

[0135] In a specific embodiment, the lymphoma is non-Hodgkin's lymphoma. In a specific embodiment, the lymphoma is primary mediastinal large B-cell lymphoma (PMBCL). In a specific embodiment, the lymphoma is diffuse large B-cell lymphoma (DLBCL). In a specific embodiment, the lymphoma is mantle cell lymphoma.

[0136] In a specific embodiment, the breast cancer is triple-negative breast cancer.

[0137] In a specific embodiment, the breast cancer is ER+ / HER2- breast cancer.

[0138] In a specific embodiment, the breast cancer is HR+ / HER2- breast cancer.

[0139] In a specific embodiment, the breast cancer is HER2+ breast cancer.

[0140] In a specific embodiment, the breast cancer is ER+ breast cancer.

[0141] In a specific embodiment, the breast cancer is germline BRCA mutated HER2- breast cancer.

[0142] In a specific embodiment, the bladder cancer is urothelial carcinoma.

[0143] In a specific embodiment, the head and neck cancer is nasopharyngeal carcinoma. In a specific embodiment, the cancer is thyroid cancer. In another embodiment, the cancer is salivary cancer. In another embodiment, the cancer is squamous cell carcinoma of the head and neck.

[0144] In a specific embodiment, the cancer is metastatic colorectal cancer with high levels of microsatellite instability (MSI-H).

[0145] In a specific embodiment, the cancer is microsatellite stable (MSS) colorectal cancer.

[0146] In a specific embodiment, the cancer is a solid tumor with high levels of microsatellite instability (MSI-H).

[0147] In a specific embodiment, the cancer is a solid tumor with a high mutational burden.

[0148] In specific embodiments of the method or use of the invention, the cancer is selected from the group consisting of melanoma, non-small cell lung cancer, small cell lung cancer, squamous cell carcinoma of the head and neck, classical Hodgkin lymphoma, primary mediastinal large B-cell lymphoma, urothelial carcinoma, microsatellite instability-high or mismatch repair deficient cancer, gastric cancer, esophageal cancer, cervical cancer, hepatocellular carcinoma, Merkel cell carcinoma, renal cell carcinoma, endometrial cancer, cancer characterized by tumors with high mutational burden, cutaneous squamous cell carcinoma, and triple-negative breast cancer.

[0149] In specific embodiments of the method or use of the invention, the cancer is selected from the group consisting of melanoma, non-small cell lung cancer, squamous cell carcinoma of the head and neck, classical Hodgkin's lymphoma, primary mediastinal large B-cell lymphoma, urothelial carcinoma, microsatellite instability-high or mismatch repair deficient cancer, gastric cancer, esophageal cancer, cervical cancer, hepatocellular carcinoma, Merkel cell carcinoma, renal cell carcinoma, endometrial cancer, cancer characterized by tumors with high mutational burden, cutaneous squamous cell carcinoma, and triple-negative breast cancer.

[0150] In specific embodiments of the methods or uses of the invention, the cancer is selected from the group consisting of melanoma, non-small cell lung cancer, squamous cell carcinoma of the head and neck, urothelial carcinoma, classical Hodgkin lymphoma, primary mediastinal large B-cell lymphoma (PMBCL), MSI-H cancer, MSI-H or mismatch repair deficient colorectal cancer, gastric cancer, gastroesophageal junction adenocarcinoma, esophageal cancer, cervical cancer, hepatocellular carcinoma, Merkel cell carcinoma, renal cell carcinoma, endometrial cancer, cutaneous squamous cell carcinoma, high tumor mutation burden (TMB-H) cancer, and triple negative breast cancer.

[0151] In a first embodiment (Embodiment E1), the invention includes a method of treating cancer in a human patient in need of such treatment, comprising subcutaneously administering to the patient a dose of about 600 mg to about 1000 mg of an anti-PD-1 antibody (e.g., pembrolizumab) or antigen-binding fragment thereof approximately once every six weeks. In a specific embodiment, the anti-PD-1 antibody or antigen-binding fragment thereof is administered once every six weeks.

[0152] In a second embodiment (Embodiment E2), the invention includes a method of treating unresectable or metastatic melanoma in a human patient in need thereof, comprising administering subcutaneously to the patient an anti-PD-1 antibody (e.g., pembrolizumab) or antigen-binding fragment thereof at a dose of about 600 mg to about 1000 mg approximately once every six weeks. In a specific embodiment, the anti-PD-1 antibody or antigen-binding fragment thereof is administered once every six weeks. In a third embodiment (Embodiment E3), the invention includes a method of treating metastatic non-small cell lung cancer (NSCLC) in a human patient in need thereof, comprising administering subcutaneously to the patient an anti-PD-1 antibody (e.g., pembrolizumab) or antigen-binding fragment thereof at a dose of about 600 mg to about 1000 mg approximately once every six weeks. In a specific embodiment, the anti-PD-1 antibody or antigen-binding fragment thereof is administered once every six weeks.

[0153] In a subembodiment of embodiment E3 (embodiment E3-A), the patient has a tumor with high PD-L1 expression ((Tumor Proportion Score (TPS)≧50%)) and has not been previously treated with platinum-containing chemotherapy.

[0154] In a further subembodiment of embodiment E3 (embodiment E3-B), the patient has a tumor with PD-L1 expression (TPS > 1%) and has been previously treated with a platinum-containing chemotherapy. In a specific embodiment of embodiment E3-B, the patient has had disease progression during or after receiving a platinum-containing chemotherapy or at least one prior chemotherapy.

[0155] In another subembodiment of embodiment E3 (embodiment E3-C), the patient has a tumor with PD-L1 expression (TPS > 1%) and has not been previously treated with a platinum-containing chemotherapy. In one embodiment, the patient has stage III non-small cell lung cancer and is not a candidate for surgical resection or definitive chemoradiotherapy. In one embodiment, the patient has metastatic non-small cell lung cancer. In yet another subembodiment of embodiment E3 (embodiment E3-D), the patient's tumor is not tested for PD-L1 expression. In this embodiment, the patient is treated with an anti-PD-1 antibody or antigen-binding fragment thereof regardless of PD-L1 expression. In a specific embodiment, the patient has not been previously treated with a platinum-containing chemotherapy.

[0156] In certain embodiments of Embodiment E3 (including Embodiments E3-A, E3-B, and E3-C), PD-L1 TPS is determined by an FDA-approved test.

[0157] In certain embodiments of embodiment E3 (including embodiments E3-A, E3-B, E3-C, and E3-D), the patient's tumor does not have an EGFR or ALK genomic abnormality.

[0158] In certain embodiments of Embodiment E3 (including Embodiments E3-A, E3-B, E3-C, and E3-D), the patient's tumor has an EGFR or ALK genomic abnormality and has had disease progression after or after treatment for the EGFR or ALK abnormality(s) prior to receiving the anti-PD-1 antibody or antigen-binding fragment thereof.

[0159] In a fourth embodiment (Embodiment E4), the invention includes a method of treating metastatic or stage III non-small cell lung cancer (NSCLC) in a human patient in need of such treatment, comprising: (1) subcutaneously administering to the patient an anti-PD-1 antibody (e.g., pembrolizumab) or antigen-binding fragment thereof at a dose of about 600 mg to about 1000 mg about once every six weeks; and (2) administering pemetrexed and platinum chemotherapy (e.g., carboplatin) to the patient. In a subembodiment of embodiment E4, the patient has not been previously treated with an anti-cancer therapeutic agent prior to initiating the combination treatment regimen with the anti-PD-1 antibody or antigen-binding fragment thereof, pemetrexed, and carboplatin.

[0160] In certain embodiments of embodiments E3 and E4 (including subembodiments thereof), the patient has non-squamous non-small cell lung cancer.

[0161] In certain embodiments of embodiments E3 and E4, including subembodiments thereof, the patient is also treated with carboplatin and paclitaxel or nab-paclitaxel. In one embodiment, carboplatin is administered by intravenous infusion at an AUC of 5-6 mg / ml / min and paclitaxel is administered at 200 mg / ml every 21 days. 2 100 mg / m2 administered by intravenous infusion every 7 days, and nab-paclitaxel at 100 mg / m2 2 It is administered by intravenous infusion of

[0162] In a subembodiment of embodiment E4, pemetrexed is administered at 500 mg / m 2 is administered to the patient in an amount of

[0163] In a subembodiment of embodiment E4, pemetrexed is administered at 500 mg / m every 3 weeks. 2 is administered to the patient in an amount of

[0164] In a subembodiment of embodiment E4, pemetrexed is administered to the patient by intravenous infusion every 21 days. In a specific embodiment, the infusion time is about 10 minutes.

[0165] In a subembodiment of embodiment E4 (embodiment E4-A), the invention further comprises administering to the patient about 400 μg to about 1000 μg of folic acid once daily, beginning about 7 days prior to administering pemetrexed to the patient and continuing until about 21 days after the patient has been administered the last dose of pemetrexed. In certain embodiments, the folic acid is administered orally.

[0166] In a subembodiment (Embodiment E4-B) of embodiment E4 and E4-A, the present invention provides a method for treating a patient suffering from atopic dermatitis comprising administering to said patient about 1 mg of vitamin B 12 to the patient about 1 week prior to the first administration of pemetrexed and about every 3 cycles of administration of pemetrexed (i.e., approximately every 9 weeks). 12 is administered intramuscularly.

[0167] In a subembodiment (Embodiment E4-C) of embodiments E4, E4-A, and E4-B, the invention further comprises administering about 4 mg of dexamethasone to the patient twice daily, the day before, the day of, and the day after administration of pemetrexed. In certain embodiments, the dexamethasone is administered orally. In a fifth embodiment (Embodiment E5), the invention comprises a method of treating recurrent or metastatic head and neck squamous cell carcinoma (HNSCC) in a human patient in need thereof, comprising subcutaneously administering to the patient about once every six weeks a dose of about 600 mg to about 1000 mg of an anti-PD-1 antibody (e.g., pembrolizumab), or an antigen-binding fragment thereof.

[0168] In a subembodiment of embodiment E5 (embodiment E5-A), the patient has been previously treated with platinum-containing chemotherapy. In certain embodiments, the patient has had disease progression during or after platinum-containing chemotherapy.

[0169] In a subembodiment of embodiment E5 (embodiment E5-B), the patient has metastatic or unresectable recurrent HNSCC, and the method further comprises administering platinum and 5-FU (fluorouracil) for first line treatment of the HNSCC.

[0170] In a subembodiment of embodiment E5 (embodiment E5-C), an anti-PD-1 antibody (e.g., pembrolizumab) is administered as a single agent for the first-line treatment of patients with metastatic or unresectable recurrent HNSCC, whose tumors express PD-L1 (CPS≧1%).

[0171] In a subembodiment of embodiment E5 (embodiment E5-D), an anti-PD-1 antibody (e.g., pembrolizumab) is administered in combination with platinum and 5-fluorouracil (5-FU) chemotherapy in patients for the first-line treatment of metastatic or unresectable recurrent head and neck squamous cell carcinoma whose tumors express PD-L1 with a CPS of >1.

[0172] In a subembodiment of embodiment E5 (embodiment E5-E), an anti-PD-1 antibody (e.g., pembrolizumab) is administered for the treatment of recurrent or metastatic head and neck squamous cell carcinoma in patients whose tumors express PD-L1 with a TPS of 50% or greater and progress on or after platinum-containing chemotherapy.

[0173] In embodiments of embodiments E5-B and E5-D, the platinum therapy is carboplatin administered by intravenous infusion at an AUC of 5 mg / ml / min every 3 weeks, or 100 mg / ml every 3 weeks. 2 cisplatin administered by intravenous infusion at 1000 mg / m2 every 3 weeks. 5-FU administered at 1000 mg / m2 every 3 weeks. 2 / day for 4 days.

[0174] In a sixth embodiment (Embodiment E6), the invention includes a method of treating refractory or relapsed classical Hodgkin lymphoma (cHL) in a human patient in need thereof, the method comprising subcutaneously administering to the patient a dose of about 600 mg to about 1000 mg of an anti-PD-1 antibody (e.g., pembrolizumab), or an antigen-binding fragment thereof, about once every six weeks.

[0175] In a seventh embodiment (Embodiment E7), the invention includes a method of treating classical Hodgkin lymphoma (cHL) in a human patient in need thereof, comprising subcutaneously administering to the patient about once every six weeks an anti-PD-1 antibody (e.g., pembrolizumab) or antigen-binding fragment thereof at a dose of about 600 mg to about 1000 mg, wherein the patient has relapsed after (a) one or more lines of treatment for cHL, (b) two or more lines of treatment for cHL, or (c) three or more lines of treatment for cHL.

[0176] In subembodiments of embodiments E6 and E7, the patient is an adult patient.

[0177] In alternative subembodiments of embodiments E6 and E7, the patient is a pediatric patient.

[0178] In an eighth embodiment (embodiment E8), the invention includes a method of treating locally advanced or metastatic urothelial carcinoma in a human patient in need of such treatment, comprising administering subcutaneously to the patient about once every six weeks an anti-PD-1 antibody (e.g., pembrolizumab) or antigen-binding fragment thereof at a dose of about 600 mg to about 1000 mg. In one embodiment, the patient is ineligible for platinum-containing chemotherapy or has disease progression during or following platinum-containing chemotherapy or within 12 months of neoadjuvant or adjuvant treatment with platinum-containing chemotherapy.

[0179] In a subembodiment of embodiment E8, the patient is not eligible for cisplatin-containing chemotherapy.

[0180] In a subembodiment of embodiment E8, the patient has had disease progression during or after platinum-containing chemotherapy, or within 12 months of neoadjuvant or adjuvant treatment with platinum-containing chemotherapy.

[0181] In a subembodiment of embodiment E8, the patient's tumor expresses PD-L1.In another subembodiment of embodiment E8, the patient's tumor expresses PD-L1 (CPS>10).

[0182] In a ninth embodiment (embodiment E9), the invention includes a method of treating an unresectable or metastatic microsatellite instability-high (MSI-H) or mismatch repair (MMR) deficient solid tumor in a human patient in need thereof, comprising subcutaneously administering to the patient about once every six weeks a dose of about 600 mg to about 1000 mg of an anti-PD-1 antibody (e.g., pembrolizumab), or an antigen-binding fragment thereof.

[0183] In a subembodiment of embodiment E9, the patient has had disease progression after previous anti-cancer treatment.

[0184] In a subembodiment of embodiment E9, the patient has advanced or recurrent endometrial cancer, hi one embodiment, there has been disease progression on or after prior treatment with a platinum-containing therapy, and the patient is not a candidate for curative surgery or radiation.

[0185] In a subembodiment of embodiment E9, the patient has unresectable or metastatic gastric, small intestine, or cholangiocarcinoma. In one embodiment, the patient has disease progression on or after at least one prior treatment.

[0186] In a tenth embodiment (embodiment E10), the invention includes a method of treating unresectable or metastatic MSI-H or MMR deficient colorectal cancer in a human patient in need of such treatment, comprising subcutaneously administering to the patient about once every six weeks a dose of about 600 mg to about 1000 mg of an anti-PD-1 antibody (e.g., pembrolizumab), or an antigen-binding fragment thereof.

[0187] In a subembodiment of embodiment E10, the patient has had progressive disease following prior treatment with a fluoropyrimidine, oxaliplatin, and irinotecan.

[0188] In an eleventh embodiment (embodiment E11), the invention includes a method of treating recurrent locally advanced unresectable or metastatic gastric or gastroesophageal junction adenocarcinoma in a human patient in need of such treatment, comprising subcutaneously administering to the patient about once every six weeks an anti-PD-1 antibody (e.g., pembrolizumab) or antigen-binding fragment thereof at a dose of about 600 mg to about 1000 mg. In a specific embodiment, the gastric or gastroesophageal junction adenocarcinoma is HER2 positive. In a specific embodiment, the invention further includes treating the patient with trastuzumab, a fluoropyrimidine, and a platinum-containing chemotherapy. In a specific embodiment, the treatment with the anti-PD-1 antibody, trastuzumab, a fluoropyrimidine, and a platinum-containing chemotherapy is the first line of treatment. In one embodiment, trastuzumab is administered at 8 mg / kg for the first infusion and 6 mg / kg for subsequent cycles, followed by chemotherapy: cisplatin 80 mg / m 2 up to 6 cycles, 5-FU 800 mg / m 2 / day for 5 days, or oxaliplatin 130 mg / m 2 6–8 cycles, each given every 3 weeks, with capecitabine 1,000 mg / m 2 In one embodiment, an anti-PD-1 antibody (e.g., pembrolizumab) administered subcutaneously every 6 weeks is administered on day 1 of each cycle prior to trastuzumab and chemotherapy.

[0189] In a twelfth embodiment (embodiment E12), the invention includes a method of treating recurrent locally advanced or metastatic esophageal or gastroesophageal junction adenocarcinoma in a human patient in need thereof, comprising subcutaneously administering to the patient about once every six weeks a dose of about 600 mg to about 1000 mg of an anti-PD-1 antibody (e.g., pembrolizumab), or an antigen-binding fragment thereof.

[0190] In a subembodiment of embodiment E12, the method further comprises administering a fluoropyrimidine-based chemotherapy in combination with a platinum-containing chemotherapy.

[0191] In another twelfth embodiment (Embodiment E12), the invention includes a method of treating recurrent locally advanced or metastatic esophageal or HER-2-negative gastroesophageal junction adenocarcinoma in a human patient in need of treatment, comprising subcutaneously administering to the patient approximately once every six weeks a dose of about 600 mg to about 1000 mg of an anti-PD-1 antibody (e.g., pembrolizumab), or an antigen-binding fragment thereof, in combination with a fluoropyrimidine-based chemotherapy and a platinum-containing chemotherapy, wherein the patient's tumor has a PD-L1 Combined Positive Score (CPS) of ≧10.

[0192] In a subembodiment of embodiment E12, the fluoropyrimidine-based chemotherapy and the platinum-containing chemotherapy are cisplatin 80 mg / m 2 up to 6 cycles, 5-FU 800mg / m 2 / day for 5 days, or oxaliplatin 130 mg / m 2 1,000 mg / m 2 is administered twice daily for 14 days.

[0193] In a subembodiment of embodiments E11 and E12, the patient's tumor expresses PD-L1. In a subembodiment of embodiments E11 and E12, the patient's tumor has a PD-L1 combined positive score (CPS) > 1.

[0194] In a subembodiment of embodiments E11 and E12, the patient has had disease progression during or after one or more prior lines of therapy. In a specific embodiment, the prior lines of therapy include fluoropyrimidine and platinum-containing chemotherapy.

[0195] In subembodiments of embodiments E11 and E12, the patient has had disease progression on or after two or more prior lines of treatment including fluoropyrimidine and platinum-containing chemotherapy.

[0196] In a subembodiment of embodiments E11 and E12, the patient has had disease progression on or after one or more prior lines of therapy that include a HER2 / neu targeted therapy.

[0197] In a subembodiment of embodiments E11 and E12, the patient has had disease progression on or after two or more prior lines of therapy that included a HER2 / neu targeted therapy.

[0198] In a thirteenth embodiment (Embodiment E13), the invention includes a method of treating cancer in a human patient in need of treatment, comprising subcutaneously administering to the patient about once every six weeks a dose of about 600 mg to about 1000 mg of an anti-PD-1 antibody (e.g., pembrolizumab), or an antigen-binding fragment thereof, wherein the patient has a cancer selected from the group consisting of melanoma, lung cancer, head and neck cancer, bladder cancer, breast cancer, gastrointestinal cancer, multiple myeloma, hepatocellular carcinoma, lymphoma, renal cancer, mesothelioma, ovarian cancer, esophageal cancer, anal cancer, biliary tract cancer, colorectal cancer, cervical cancer, hepatocellular carcinoma, Merkel cell carcinoma, renal cell carcinoma, endometrial cancer, cutaneous squamous cell carcinoma, thyroid cancer, and salivary cancer.

[0199] In a fourteenth embodiment (embodiment E14), the invention includes a method of treating cancer in a human patient in need of treatment, comprising subcutaneously administering to the patient about once every six weeks a dose of about 600 mg to about 1000 mg of an anti-PD-1 antibody (e.g., pembrolizumab) or an antigen-binding fragment thereof, wherein the patient has small cell lung cancer. In a subembodiment, the patient has been previously treated with platinum-based chemotherapy and at least one other prior line of treatment.

[0200] In a fifteenth embodiment (Embodiment E15), the invention includes a method of treating non-Hodgkin's lymphoma in a human patient in need of such treatment, comprising subcutaneously administering to the patient a dose of about 600 mg to about 1000 mg of an anti-PD-1 antibody (e.g., pembrolizumab), or an antigen-binding fragment thereof, about once every six weeks.

[0201] In a subembodiment of embodiment E15, the non-Hodgkin's lymphoma is primary mediastinal large B-cell lymphoma (PMBCL). In a specific embodiment where the patient has PMBCL, the patient has refractory PMBCL. In a specific embodiment, the patient has relapsed after one or more prior lines of therapy. In a specific embodiment, the patient has relapsed after two or more prior lines of therapy. In a specific embodiment, the patient has not been previously treated with another line of therapy. In a specific embodiment, the patient is an adult. In a specific embodiment, the patient is a pediatric patient.

[0202] In a sixteenth embodiment (Embodiment E16), the invention includes a method of treating metastatic squamous NSCLC in a human patient in need of treatment for metastatic squamous NSCLC, comprising: (1) subcutaneously administering to the patient an anti-PD-1 antibody (e.g., pembrolizumab) or an antigen-binding fragment thereof at a dose of about 600 mg to about 1000 mg about once every six weeks; and (2) administering to the patient (i) carboplatin and paclitaxel, or (ii) carboplatin and nab-paclitaxel.

[0203] In a seventeenth embodiment (embodiment E17), the invention includes a method of treating Merkel cell carcinoma (MCC) in a human patient in need of treatment, comprising subcutaneously administering to the patient about once every six weeks an anti-PD-1 antibody (e.g., pembrolizumab) or antigen-binding fragment thereof at a dose of about 600 mg to about 1000 mg. In particular, in a subembodiment of embodiment E17, the cancer is recurrent locally advanced MCC. In particular, in a subembodiment of embodiment E17, the cancer is metastatic MCC.

[0204] In a subembodiment of embodiment E17, the patient is an adult patient. In an alternative subembodiment of embodiment E17, the patient is a pediatric patient.

[0205] In an eighteenth embodiment (embodiment E18), the invention includes a method for adjuvant therapy of melanoma in a human patient in need thereof, comprising subcutaneously administering to the patient about once every six weeks a dose of about 600 mg to about 1000 mg of an anti-PD-1 antibody (e.g., pembrolizumab) or an antigen-binding fragment thereof, wherein the patient has previously had one or more melanoma lesions resected. In a subembodiment of embodiment E18, the method includes treating resected high-risk stage III melanoma. In a subembodiment of embodiment E18, the method includes treating resected stage IIB or stage IIC melanoma.

[0206] In a nineteenth embodiment (embodiment E19), the invention includes a method of treating hepatocellular carcinoma (HCC) in a human patient in need of such treatment, comprising subcutaneously administering to the patient about once every six weeks a dose of about 600 mg to about 1000 mg of an anti-PD-1 antibody (e.g., pembrolizumab) or an antigen-binding fragment thereof. In a specific embodiment of embodiment E19, the patient has previously been treated with sorafenib.

[0207] In a twentieth embodiment (Embodiment E20), the invention includes a method of treating renal cell carcinoma (RCC) in a human patient in need of treatment, the method comprising subcutaneously administering to the patient a dose of about 600 mg to about 1000 mg of an anti-PD-1 antibody (e.g., pembrolizumab), or an antigen-binding fragment thereof, about once every six weeks.

[0208] In a subembodiment of embodiment E20, the cancer is advanced clear cell RCC.

[0209] In a subembodiment of embodiment E20, the patient has advanced or metastatic renal cell carcinoma (RCC).

[0210] In a subembodiment of embodiment E20 (embodiment E20C), the anti-PD-1 antibody (e.g., pembrolizumab) is for the adjuvant treatment of patients with intermediate-high or high risk RCC that recurs after nephrectomy, or after nephrectomy and resection of metastatic disease.

[0211] In a twenty-first embodiment (embodiment E21), the invention includes a method of treating breast cancer in a human patient in need of such treatment, comprising subcutaneously administering to the patient a dose of about 600 mg to about 1000 mg of an anti-PD-1 antibody (e.g., pembrolizumab), or an antigen-binding fragment thereof, approximately once every six weeks.

[0212] In a subembodiment of embodiment E21, the breast cancer is triple negative breast cancer. In a further subembodiment, the patient is further treated with chemotherapy. In a further subembodiment, the TNBC is recurrent unresectable or metastatic TNBC, and the patient's tumor expresses PD-L1 (CPS≧10).

[0213] In a subembodiment of embodiment E21, the breast cancer is ER+ / HER2- breast cancer.

[0214] In a further subembodiment of embodiment E21, the patient has high-risk early stage or locally advanced TNBC, and the method comprises treating the patient with an anti-PD-1 antibody (e.g., pembrolizumab) in combination with chemotherapy as a neoadjuvant treatment, and then treating the patient with an anti-PD-1 antibody (e.g., pembrolizumab) as a single agent as an adjuvant treatment following surgery. In one embodiment, the patient is treated with 4 cycles of neoadjuvant anti-PD-1 antibody (e.g., pembrolizumab) on day 1 of cycles 1-4 of the treatment regimen, carboplatin at AUC 5 mg / mL / min every 3 weeks on day 1 of cycles 1-4 of the treatment regimen, or carboplatin at AUC 1.5 mg / mL / min weekly on days 1, 8, and 15 of cycles 1-4 of the treatment regimen, and paclitaxel 80 mg / m weekly on days 1, 8, and 15 of cycles 1-4 of the treatment regimen. 2 in combination with doxorubicin 60 mg / m2 or epirubicin 90 mg / m 2 on days 1 of cycles 5 to 8 of the treatment regimen every 3 weeks, and cyclophosphamide 600 mg / m 2 Administer every 3 weeks on day 1 of cycles 5 to 8 of the treatment regimen, in combination with an additional 4 cycles of a neoadjuvant anti-PD-1 antibody (e.g., pembrolizumab), followed by 9 cycles of an adjuvant anti-PD-1 antibody (e.g., pembrolizumab) following surgery.

[0215] In a twenty-second embodiment (embodiment E22), the invention includes a method of treating nasopharyngeal carcinoma in a human patient in need of such treatment, comprising subcutaneously administering to the patient a dose of about 600 mg to about 1000 mg of an anti-PD-1 antibody (e.g., pembrolizumab) or an antigen-binding fragment thereof approximately once every six weeks.

[0216] In a twenty-third embodiment (embodiment E23), the invention includes a method of treating thyroid cancer in a human patient in need of such treatment, comprising administering to the patient a dose of about 600 mg to about 1000 mg of an anti-PD-1 antibody (e.g., pembrolizumab) or an antigen-binding fragment thereof about once every six weeks.

[0217] In a twenty-fourth embodiment (embodiment E24), the invention includes a method of treating salivary cancer in a human patient in need of such treatment, comprising subcutaneously administering to the patient a dose of about 600 mg to about 1000 mg of an anti-PD-1 antibody (e.g., pembrolizumab), or an antigen-binding fragment thereof, about once every six weeks.

[0218] In a twenty-fifth embodiment (Embodiment E25), the invention includes a method of treating cancer in a human patient in need of treatment, comprising subcutaneously administering to the patient about once every six weeks a dose of about 600 mg to about 1000 mg of an anti-PD-1 antibody (e.g., pembrolizumab), or an antigen-binding fragment thereof, wherein the cancer is selected from the group consisting of melanoma, non-small cell lung cancer (NSCLC), head and neck squamous cell carcinoma (HNSCC), relapsed or refractory classical Hodgkin lymphoma (cHL), primary mediastinal large B-cell lymphoma (PMBCL), urothelial carcinoma, microsatellite instability-high or mismatch repair deficient colorectal cancer, gastric cancer, esophageal cancer, cervical cancer, hepatocellular carcinoma, Merkel cell carcinoma, renal cell carcinoma, endometrial cancer, TMB-H cancer, cutaneous squamous cell carcinoma, and triple-negative breast cancer.

[0219] In a subembodiment of embodiment 25 (embodiment 25B), the invention includes a method of treating cancer in a human patient in need of treatment, comprising subcutaneously administering to the patient about once every six weeks a dose of about 600 mg to about 1000 mg of an anti-PD-1 antibody (e.g., pembrolizumab), or an antigen-binding fragment thereof, wherein the cancer is selected from the group consisting of melanoma, non-small cell lung cancer, relapsed or refractory classical Hodgkin lymphoma, primary mediastinal large B-cell lymphoma, squamous cell carcinoma of the head and neck, urothelial carcinoma, esophageal cancer, gastric cancer, cervical cancer, PMBCL, MSI-H cancer, hepatocellular carcinoma, Merkel cell carcinoma, renal cell carcinoma, endometrial cancer, TMB-H cancer, cutaneous squamous cell carcinoma, and triple-negative breast cancer.

[0220] In a twenty-sixth embodiment (embodiment E26), the invention includes a method of treating cancer in a human patient in need of such treatment, comprising subcutaneously administering to the patient a dose of about 600 mg to about 1000 mg of an anti-PD-1 antibody (e.g., pembrolizumab) or an antigen-binding fragment thereof approximately once every six weeks, wherein the cancer is a heme malignancy.

[0221] In a subembodiment of embodiment E26, the heme malignancy is selected from the group consisting of acute lymphoblastic leukemia (ALL), acute myeloid leukemia (AML), chronic lymphocytic leukemia (CLL), chronic myeloid leukemia (CML), diffuse large B-cell lymphoma (DLBCL), EBV-positive DLBCL, primary mediastinal large B-cell lymphoma, T-cell / histiocyte-rich large B-cell lymphoma, follicular lymphoma, Hodgkin lymphoma (HL), mantle cell lymphoma (MCL), multiple myeloma (MM), myeloid cell leukemia-1 protein (MCL-1), myelodysplastic syndrome (MDS), non-Hodgkin lymphoma (NHL), and small lymphocytic lymphoma (SLL).

[0222] In a twenty-seventh embodiment (embodiment E27), the invention includes a method of treating cancer in a human patient in need of such treatment, comprising subcutaneously administering to the patient about once every six weeks a dose of about 600 mg to about 1000 mg of an anti-PD-1 antibody (e.g., pembrolizumab) or an antigen-binding fragment thereof, wherein the patient has a tumor with a high mutational burden. In a subembodiment of embodiment E27, the tumor is a solid tumor. In a specific embodiment, the patient is an adult patient. In a specific embodiment, the patient is a pediatric patient.

[0223] In a subembodiment of embodiment 27, the high mutation burden is at least about 10 mutations per megabase of the genome interrogated. In other embodiments, the high mutation burden is at least about 11 mutations per megabase of the genome interrogated, at least about 12 mutations per megabase of the genome interrogated, or at least about 13 mutations per megabase of the genome interrogated.

[0224] In a twenty-eighth embodiment (embodiment E28), the invention includes a method of treating esophageal cancer in a human patient in need of such treatment, comprising subcutaneously administering to the patient a dose of about 600 mg to about 1000 mg of an anti-PD-1 antibody (e.g., pembrolizumab), or an antigen-binding fragment thereof, about once every six weeks.

[0225] In a subembodiment of embodiment E28, the patient has progressed on one prior line of standard of care prior to receiving the anti-PD-1 antibody or antigen-binding fragment thereof. In a further embodiment, the patient has progressed on one or more lines of standard of care prior to receiving the anti-PD-1 antibody or antigen-binding fragment thereof. In another embodiment, the patient has progressed on two or more standards of care prior to receiving the anti-PD-1 antibody or antigen-binding fragment thereof. In certain embodiments, the standard of care comprises one or more of paclitaxel, docetaxel, or irinotecan.

[0226] In a subembodiment of embodiment E28, the patient has advanced or metastatic esophageal adenocarcinoma or squamous cell carcinoma.

[0227] In a subembodiment of embodiment E28, the patient has advanced or metastatic Siewert type I adenocarcinoma of the gastroesophageal junction.

[0228] In a subembodiment of embodiment E28, the patient's tumor expresses PD-L1 (combined positive score [CPS] > 10).

[0229] In a twenty-ninth embodiment (embodiment E29), the invention includes a method of treating high-risk non-muscle invasive bladder cancer (NMIBC) in a human patient in need of such treatment, comprising subcutaneously administering to the patient a dose of about 600 mg to about 1000 mg of an anti-PD-1 antibody (e.g., pembrolizumab) or an antigen-binding fragment thereof approximately once every six weeks. In a specific embodiment, the patient has NMIBC with carcinoma in situ (CIS) or CIS+papillary disease.

[0230] In a subembodiment of embodiment E29, the patient has been previously treated with standard therapy prior to being treated with the anti-PD-1 antibody or antigen-binding fragment thereof. In a specific embodiment, the prior therapy is Bacillus Calmette-Guerin (BCG) therapy. In a particular embodiment, the patient did not respond to BCG therapy. In a specific embodiment, the patient was ineligible for radical cystectomy or chose not to undergo radical cystectomy.

[0231] In a thirtieth embodiment (embodiment E30), the invention includes a method of treating cutaneous squamous cell carcinoma (cSCC) in a human patient in need of treatment comprising subcutaneously administering to the patient about once every six weeks a dose of about 600 mg to about 1000 mg of an anti-PD-1 antibody (e.g., pembrolizumab) or antigen-binding fragment thereof. In a specific embodiment, the cutaneous squamous cell carcinoma is not curable by surgery or radiation.

[0232] In a thirty-first embodiment (embodiment E31), the invention includes a method of treating endometrial cancer in a human patient in need of such treatment, the method comprising subcutaneously administering to the patient a dose of about 600 mg to about 1000 mg of an anti-PD-1 antibody (e.g., pembrolizumab), or an antigen-binding fragment thereof, about once every six weeks.

[0233] In some subembodiments of embodiment E31, the endometrial cancer is an aggressive endometrial cancer that is not MSI-H or mismatch repair deficient (dMMR). In a specific embodiment, the patient has had disease progression after prior systemic therapy.

[0234] In some subembodiments of embodiment E31, the endometrial cancer is advanced endometrial cancer that is MSI-H or dMMR as determined by an FDA approved test, and the patient has had disease progression after prior systemic therapy in any setting. In specific embodiments, the patient is not a candidate for curative surgery or radiation.

[0235] In a thirty-second embodiment (embodiment E32), the invention includes a method of treating cervical cancer in a human patient in need of treatment comprising subcutaneously administering to the patient about once every six weeks an anti-PD-1 antibody (e.g., pembrolizumab) or antigen-binding fragment thereof at a dose of about 600 mg to about 1000 mg. In one embodiment, the cervical cancer is recurrent or metastatic cervical cancer and the patient has had disease progression during or after chemotherapy.

[0236] In a subembodiment of embodiment E32, the method further comprises treating the patient with chemotherapy with or without bevacizumab. In a specific embodiment, the cervical cancer is persistent, recurrent, or metastatic cervical cancer and the patient's tumor expresses PD-L1 (CPS>1). In one embodiment, the chemotherapy comprises paclitaxel 175 mg / m 2 and cisplatin 50 mg / m 2 or paclitaxel 175 mg / m 2 and carboplatin AUC 5 mg / mL / min administered on day 1 every 3 weeks. In one embodiment, the bevacizumab chemotherapy is 2 and cisplatin 50 mg / m 2 or paclitaxel 175 mg / m 2 and carboplatin AUC 5 mg / mL / min and bevacizumab 15 mg / kg administered on day 1 every 3 weeks.

[0237] In a subembodiment of embodiment E32, the cervical cancer is recurrent or metastatic cervical cancer with disease progression on or after chemotherapy, and the patient's tumor expresses PD-L1 (CPS>1).

[0238] In a thirty-third embodiment (embodiment E33), the invention comprises a method of treating stage IB, II, or IIIA non-small cell lung cancer (NSCLC) in a human patient in need of such treatment, comprising administering subcutaneously to the patient a dose of about 600 mg to about 1000 mg of an anti-PD-1 antibody (e.g., pembrolizumab), or an antigen-binding fragment thereof, about once every six weeks for adjuvant treatment following resection.

[0239] In a subembodiment of embodiment E33 (E33-A), the patient has previously received platinum-based chemotherapy.

[0240] In any of the methods or uses of the invention described herein (including embodiments E1-E33), the anti-PD-1 antibody or antigen-binding fragment thereof is any of the antibodies or antigen-binding fragments described in Section II "PD-1 Antibodies and Antigen-Binding Fragments Useful in the Invention" of the Detailed Description of the Invention herein. In a specific embodiment, the anti-PD-1 antibody is pembrolizumab or an antigen-binding fragment thereof, or an antibody that cross-competes with pembrolizumab for binding to human PD-1. In a specific embodiment, the anti-PD-1 antibody is a pembrolizumab variant.

[0241] In any of the methods or uses of the invention described herein (including embodiments E1-E33), the anti-PD-1 antibody or antigen-binding fragment thereof may be co-administered or co-formulated with a hyaluronidase as described in Section III. In one embodiment, the anti-PD-1 antibody or antigen-binding fragment thereof is co-administered with human hyaluronidase. In another embodiment, the anti-PD-1 antibody or antigen-binding fragment thereof is co-formulated with human hyaluronidase. In a specific embodiment, the human hyaluronidase is rHuPH20. In a specific embodiment, the human hyaluronidase is PH20 variant 2.

[0242] In any one of the methods or uses of the invention described herein (including embodiments E1 to E33), the anti-PD-1 antibody or antigen-binding fragment thereof is administered to the patient at a dose of about 600 mg to about 1000 mg every 6 weeks. In a further embodiment, the anti-PD-1 antibody or antigen-binding fragment thereof is administered to the patient at a dose of about 600 mg to about 950 mg every 6 weeks. In a further embodiment, the anti-PD-1 antibody or antigen-binding fragment thereof is administered to the patient at a dose of about 600 mg to about 900 mg every 6 weeks. In a further embodiment, the anti-PD-1 antibody or antigen-binding fragment thereof is administered to the patient at a dose of about 650 mg to about 800 mg every 6 weeks. In a further embodiment, the anti-PD-1 antibody or antigen-binding fragment thereof is administered to the patient at a dose of about 660 mg to about 800 mg every 6 weeks. In a further embodiment, the anti-PD-1 antibody or antigen-binding fragment thereof is administered to the patient at a dose of about 700 mg to about 800 mg every 6 weeks. In a further embodiment, the anti-PD-1 antibody or antigen-binding fragment is administered at a dose of about 760 mg to about 800 mg every six weeks.In a further embodiment, the anti-PD-1 antibody or antigen-binding fragment is administered at a dose of about 760 mg to about 790 mg every six weeks.

[0243] In any one of the methods or uses of the invention described herein (including embodiments E1-E33), the anti-PD-1 antibody or antigen-binding fragment is administered at a dose of 600 mg to 1000 mg every 6 weeks. In a further embodiment, the anti-PD-1 antibody or antigen-binding fragment is administered to the patient at a dose of 600 mg to 950 mg every 6 weeks. In a further embodiment, the anti-PD-1 antibody or antigen-binding fragment is administered at a dose of 600 mg to 900 mg every 6 weeks. In a further embodiment, the anti-PD-1 antibody or antigen-binding fragment is administered to the patient at a dose of 650 mg to 800 mg every 6 weeks. In a further embodiment, the anti-PD-1 antibody or antigen-binding fragment is administered to the patient at a dose of 660 mg to 800 mg every 6 weeks. In a further embodiment, the anti-PD-1 antibody or antigen-binding fragment is administered to the patient at a dose of 700 mg to 800 mg every 6 weeks. In a further embodiment, the anti-PD-1 antibody or antigen-binding fragment is administered at a dose of 760 mg to 800 mg every six weeks.In a further embodiment, the anti-PD-1 antibody or antigen-binding fragment is administered at a dose of 760 mg to 790 mg every six weeks.

[0244] In any one of the methods or uses of the invention described herein (including embodiments E1-E33), the anti-PD-1 antibody or antigen-binding fragment thereof is administered at a dose of 760 mg every 6 weeks. In one embodiment, the anti-PD-1 antibody or antigen-binding fragment is administered at a dose of 770 mg every 6 weeks. In another embodiment, the anti-PD-1 antibody or antigen-binding fragment is administered at a dose of 780 mg every 6 weeks. In another embodiment, the anti-PD-1 antibody or antigen-binding fragment is administered at a dose of 790 mg every 6 weeks. In another embodiment, the anti-PD-1 antibody or antigen-binding fragment is administered at a dose of 800 mg every 6 weeks. In another embodiment, the anti-PD-1 antibody or antigen-binding fragment is administered at a dose of 810 mg every 6 weeks. In another embodiment, the anti-PD-1 antibody or antigen-binding fragment is administered at a dose of 650 mg every 6 weeks. In another embodiment, the anti-PD-1 antibody or antigen-binding fragment is administered at a dose of 750 mg every 6 weeks.

[0245] In any of the methods or uses of the invention described herein (including embodiments E1-E33), the anti-PD-1 antibody or antigen-binding fragment thereof is administered subcutaneously to the patient at a dose of about 300 mg to about 500 mg together with human hyaluronidase (including as described in Section III) approximately every three weeks. In any of the methods or uses of the invention described herein (including embodiments E1-E33), the anti-PD-1 antibody or antigen-binding fragment thereof is administered subcutaneously to the patient at a dose of about 300 mg to about 500 mg together with PH20 variant 2 approximately every three weeks. In any of the methods or uses of the invention described herein (including embodiments E1-E33), the anti-PD-1 antibody or antigen-binding fragment thereof is administered subcutaneously to the patient at a dose of about 300 mg to about 500 mg together with rHuPH20 approximately every three weeks. In certain embodiments, the anti-PD-1 antibody or antigen-binding fragment thereof is administered subcutaneously to a patient every 3 weeks, every 3 weeks ± every 5 days, ± every 4 days, ± every 3 days, ± every 2 days, or ± every 1 day.

[0246] In a further embodiment, the anti-PD-1 antibody or antigen-binding fragment is administered with human hyaluronidase at a dose of about 300 mg to about 500 mg every three weeks. In a further embodiment, the anti-PD-1 antibody or antigen-binding fragment is administered with human hyaluronidase at a dose of about 350 mg to about 450 mg every three weeks. In a further embodiment, the anti-PD-1 antibody or antigen-binding fragment is administered with human hyaluronidase at a dose of about 360 mg to about 420 mg every three weeks. In a further embodiment, the anti-PD-1 antibody or antigen-binding fragment is administered with human hyaluronidase at a dose of about 380 mg to about 410 mg every three weeks. In a further embodiment, the anti-PD-1 antibody or antigen-binding fragment is administered with human hyaluronidase at a dose of 380 mg every three weeks. In a further embodiment, the anti-PD-1 antibody or antigen-binding fragment is administered with human hyaluronidase at a dose of 385 mg every three weeks. In another embodiment, the anti-PD-1 antibody or antigen-binding fragment is administered with human hyaluronidase at a dose of 390 mg every three weeks. In another embodiment, the anti-PD-1 antibody or antigen-binding fragment is administered with human hyaluronidase at a dose of 395 mg every three weeks. In another embodiment, the anti-PD-1 antibody or antigen-binding fragment is administered with human hyaluronidase at a dose of 325 mg every three weeks. In another embodiment, the anti-PD-1 antibody or antigen-binding fragment is administered with human hyaluronidase at a dose of 400 mg every three weeks.

[0247] In a further embodiment, the anti-PD-1 antibody or antigen-binding fragment is administered at a dose of 300 mg-500 mg every three weeks with human hyaluronidase. In a further embodiment, the anti-PD-1 antibody or antigen-binding fragment is administered at a dose of 350 mg-450 mg every three weeks with human hyaluronidase. In a further embodiment, the anti-PD-1 antibody or antigen-binding fragment is administered at a dose of 360 mg-420 mg every three weeks with human hyaluronidase. In a further embodiment, the anti-PD-1 antibody or antigen-binding fragment is administered at a dose of 380 mg-410 mg every three weeks with human hyaluronidase.

[0248] In any one of the above methods or uses of the invention (including embodiments E1-E33), the anti-PD-1 antibody or antigen-binding fragment thereof is co-administered with a human hyaluronidase PH20 variant or fragment as defined herein. In another embodiment, the anti-PD-1 antibody or antigen-binding fragment thereof is co-formulated with PH20 variant 2 in the composition. For example, U.S. Patent Application Publication No. 20220089739, the contents of which are incorporated herein by reference, describes the preparation of a liquid composition comprising pembrolizumab and a PH20 variant.

[0249] In one embodiment, the composition comprises 130 mg / ml of the anti-PD-1 antibody or antigen-binding fragment thereof. In another embodiment, the composition comprises 165 mg / ml of the anti-PD-1 antibody or antigen-binding fragment thereof.

[0250] In a further embodiment, the composition further comprises L-methionine, hi a particular embodiment, L-methionine is present at a concentration of about 10 mM.

[0251] In a further embodiment, the composition further comprises a histidine buffer at about pH 5.0 to pH 6.0. In a particular embodiment, the histidine is present at a concentration of about 10 mM.

[0252] In further embodiments, the composition further comprises sucrose. In certain embodiments, the sucrose is present at a concentration of about 70 mg / mL. In certain embodiments, the sucrose is present at a concentration of 7% (w / v).

[0253] In a further implementation of the invention, the composition further comprises polysorbate 80. In certain embodiments, polysorbate 80 is present at a concentration of about 0.2 mg / mL. In certain embodiments, polysorbate 80 is present at a concentration of 0.02% (w / v).

[0254] In a specific embodiment, the composition comprises 10 mM L-methionine, 10 mM histidine, pH 5.5, 7% sucrose, 0.02% polysorbate 80, and 130 mg / mL of an anti-PD-1 antibody or antigen-binding fragment thereof.

[0255] In a specific embodiment, the composition comprises 10 mM L-methionine, 10 mM histidine, pH 5.5, 7% sucrose, 0.02% polysorbate 80, and 165 mg / mL of an anti-PD-1 antibody or antigen-binding fragment thereof.

[0256] In a specific embodiment of the methods or uses described herein, the anti-PD-1 antibody or antigen-binding fragment thereof is administered subcutaneously in one or more injections. In a specific embodiment, the anti-PD-1 antibody or antigen-binding fragment thereof is administered in two injections.

[0257] In one embodiment, 790 mg of the anti-PD-1 antibody or antigen-binding fragment thereof is administered subcutaneously in one injection in a composition containing 130 mg / mL, hi one embodiment, 790 mg of the anti-PD-1 antibody or antigen-binding fragment thereof is administered subcutaneously in two injections.

[0258] In one embodiment, 790 mg of the anti-PD-1 antibody or antigen-binding fragment thereof is administered subcutaneously in a single injection of a composition containing 165 mg / mL. In one embodiment, 790 mg of the anti-PD-1 antibody or antigen-binding fragment thereof is administered subcutaneously in a composition containing 165 mg / mL in two injections. In a further embodiment, 4.8 mL of a composition containing 165 mg / mL of the anti-PD-1 antibody or antigen-binding fragment thereof is administered subcutaneously in a single injection.

[0259] In one embodiment, 395 mg of the anti-PD-1 antibody or antigen-binding fragment thereof is administered subcutaneously in one injection in a composition containing 130 mg / mL, hi one embodiment, 395 mg of the anti-PD-1 antibody or antigen-binding fragment thereof is administered subcutaneously in two injections.

[0260] In one embodiment, 395 mg of the anti-PD-1 antibody or antigen-binding fragment thereof is administered subcutaneously in one injection in a composition comprising 165 mg / mL. In one embodiment, 395 mg of the anti-PD-1 antibody or antigen-binding fragment thereof is administered subcutaneously in two injections in a composition comprising 165 mg / mL. In a further embodiment, 2.4 mL of a composition comprising 165 mg / mL of the anti-PD-1 antibody or antigen-binding fragment thereof is administered subcutaneously in one injection. In a further embodiment, 1.2 mL of a composition comprising 165 mg / mL of the anti-PD-1 antibody or antigen-binding fragment thereof is administered subcutaneously in each of two injections.

[0261] In any of the methods and uses of the invention, a dose of an anti-PD-1 antibody or antigen-binding fragment thereof described herein is co-formulated or co-administered with a dose of about 700 Units to about 50,000 Units of human hyaluronidase. In one embodiment, the dose is about 2,000 Units to about 20,000 Units of human hyaluronidase. In one embodiment, the dose is about 5,000 Units to about 15,000 Units of human hyaluronidase. In one embodiment, about 10,000 Units to about 30,000 Units of human hyaluronidase are co-formulated or co-administered with a dose of an anti-PD-1 antibody or antigen-binding fragment thereof. In one embodiment, the dose is about 7,000 Units to about 13,000 Units of human hyaluronidase. In one embodiment, the dose is about 8,000 Units to about 10,000 Units of human hyaluronidase. In one embodiment, the dose is about 4000 Units to about 6000 Units of human hyaluronidase. In one embodiment, the dose is 4800 Units of human hyaluronidase. In one embodiment, the dose is 9600 Units of human hyaluronidase.

[0262] In one embodiment, the human hyaluronidase is co-formulated or co-administered with the anti-PD-1 antibody or antigen-binding fragment thereof in a ratio of about 12.15 units:1 mg. In one embodiment, the human hyaluronidase is co-formulated or co-administered with the anti-PD-1 antibody or antigen-binding fragment thereof in a ratio of about 12 units:1 mg. In one embodiment, the human hyaluronidase is co-formulated or co-administered with the anti-PD-1 antibody or antigen-binding fragment thereof in a ratio of about 10-14 units:1 mg. In one embodiment, the human hyaluronidase is co-formulated or co-administered with the anti-PD-1 antibody or antigen-binding fragment thereof in a ratio of about 8-18 units:1 mg. In one embodiment, the human hyaluronidase is co-formulated or co-administered with the anti-PD-1 antibody or antigen-binding fragment thereof in a ratio of about 6-25 units:1 mg. In one embodiment, the human hyaluronidase is co-formulated or co-administered with the anti-PD-1 antibody, or antigen-binding fragment thereof, in a ratio of about 3-36 units:1 mg.

[0263] In any of the methods or uses described herein, including embodiments E1-E33 and subembodiments thereof, the method may further comprise administering one or more "additional therapeutic agents" (as used herein, "additional therapeutic agent" refers to an additional agent to an anti-PD-1 antibody or antigen-binding fragment thereof). The additional therapeutic agent may be, for example, a chemotherapeutic agent, a biotherapeutic agent (including but not limited to antibodies to CTLA4, TIGIT, VEGF, EGFR, Her2 / neu, VEGF receptors, other growth factor receptors, CD20, CD40, CD-40L, OX-40, 4-1BB, and ICOS), an immunogenic agent (e.g., attenuated cancerous cells, tumor antigens, tumor-derived antigens, or antigen-presenting cells such as dendritic cells pulsed with nucleic acid, immune stimulating cytokines (e.g., IL-2, IFNα2, GM-CSF), and cells transfected with a gene encoding an immune stimulating cytokine such as, but not limited to, GM-CSF).

[0264] As described above, in specific embodiments of the method or use of the invention, the method further comprises administering an additional therapeutic agent. In certain embodiments, the additional therapeutic agent is an anti-CTLA4 antibody or an antigen-binding fragment thereof, an anti-LAG3 antibody or an antigen-binding fragment thereof, an anti-GITR antibody or an antigen-binding fragment thereof, an anti-TIGIT antibody or an antigen-binding fragment thereof, an anti-CD27 antibody or an antigen-binding fragment thereof, an anti-ILT3 antibody or an antigen-binding fragment thereof, or an anti-ILT4 antibody or an antigen-binding fragment thereof. In one embodiment, the additional therapeutic agent is a Newcastle Disease Virus vector expressing IL-12. In a further embodiment, the additional therapeutic agent is dinaciclib. In another embodiment, the additional therapeutic agent is navarixin. In a further embodiment, the additional therapeutic agent is vicriviroc.

[0265] In a further embodiment, the additional therapeutic agent is an oncolytic virus. In one embodiment, the additional therapeutic agent is a Coxsackievirus or CVA21. In one embodiment, the additional therapeutic agent is CAVATAK™.

[0266] In yet another embodiment, the additional therapeutic agent is a STING agonist. In a further embodiment, the additional therapeutic agent is an IL-27 antagonist. In one embodiment, the additional therapeutic agent is a PARP inhibitor. In one embodiment, the additional therapeutic agent is a multikinase inhibitor. In one embodiment, the additional therapeutic agent is a MEK inhibitor. In one embodiment, the additional therapeutic agent is a 4-1BB agonist. In one embodiment, the additional therapeutic agent is nemtabrutinib. In another embodiment, the additional therapeutic agent is velzutifan.

[0267] Examples of chemotherapeutic agents include alkylating agents such as thiotepa and cyclophosphamide; alkylsulfonates, e.g., busulfan, improsulfan, and piposulfan; aziridines, such as benzodopa, carboquone, metoledopa, and uredopa; ethylenimines and methylamelanamines, including altretamine, triethylenemelamine, triethylenephosphoramide, triethylenethiophosphoramide, and trimethylomelanamine; acetogenins (especially bullatacin and bullatacinone); camptothecins (including the synthetic analog topotecan); bryostatin; kallistatin; CC-1065 (including its adozelesin, carzelesin, and bizelesin synthetic analogs); cryptophycins (especially cryptophycin 1 and cryptophycin 8); dolastatins; duocarmycins (including the synthetic analogs, KW-2189 and CBI-TMI); eleu terobin; pancratistatin; sarcodictin; spongiostatin; nitrogen mustards such as chlorambucil, chlornaphazine, chlorophosphamide, estramustine, ifosfamide, mechlorethamine, mechlorethamine oxide hydrochloride, melphalan, nobembitine, phenesterine, prednimustine, trophosfamide, uracil mustard, etc.; nitrosoureas such as carmustine, chlorozotocin, fotemustine, lomustine, nimustine, ranimustine, etc.; enediyne antibiotics (e.g., calicheamicin, particularly calicheamicin gamma 1I and calicheamicin phiI1 (e.g., Agnew, Chem. Intl. Ed. Engl., 33:183-186 (1994)); dynemicins (including dynemicin A); bisphosphonates such as clodronate; esperamicin;Also included are neocarzinostatin chromophore and related chromoprotein enediyne antibiotic chromophores), aclacinomycin, actinomycin, ausramycin, azaserine, bleomycin, cactinomycin, carabicin, caminomycin, carzinophilin, chromomycin, dactinomycin, daunorubicin, detrevicin, 6-diazo-5-oxo-L-norleucine, doxorubicin (including morpholino-doxorubicin, cyanomorpholino-doxorubicin, 2-pyrrolidine-doxorubicin and deoxydoxorubicin), epirubicin, mitomycins such as esorubicin, idarubicin, marcelomycin, and mitomycin C, mycophenolic acid, nogalamycin, olivomycin, peplomycin, puromycin, queramycin, rodorubicin, streptonigrin, streptozocin, tubercidin, ubenimex, zinostatin, and zorubicin; metabolic antagonists such as methotrexate and 5-fluorouracil (5-FU); folic acid analogs such as denopterin, methotrexate, pteropterin, and trimetrexate; fludarabine, 6-mercaptopurine, thiamine Purines, purine analogues such as thioguanine; pyrimidine analogues such as ancitabine, azacitidine, 6-azauridine, carmoful, cytarabine, dideoxyuridine, doxifluridine, enocitabine, floxuridine; androgens such as calsterone, dromostanolone propionate, epithiostanol, mepitiostane, testolactone; antiadrenergics such as aminoglutethimide, mitotane, trilostane; folic acid supplements such as floric acid; aceglatone; aldophosphamide glycosides; aminolevulinic acid; enyluracil; amsacrine; bessacrine; Trabucil;Bisantrene;Edatraxate;Defofamine;Demecolcine;Diazicon;Elformitin;Elliptinium acetate;Epothilone;Etoglucide;Gallium nitrate;Hydroxyurea;Lentinan;Lonidamine;Maytansinoids such as maytansine and ansamitocin;Mitoguazone;Mitoxantrone;Mopidamol;Nitracrine;Pentostatin;Fenamet;Pirarubicin;Rosoxantrone;Podophyllic acid;2-Ethylhydrazide;Procarbazine;Razoxane;Rhizoxin;Schizofuran;Spirogermanium;Tenuazonic acid;Triazicon;2,2',2"-Trichlorotriethylamine;Tricothecenes (especially T-2 toxin, veracrine A, roridin A and anguidin);Urethane;Vindesine;Dacarbazine;Mannomustine;Mitobronitol;Mitolactol;Pipobroman;Gacytosine;Arabinoside ("Ara-C");Cyclophosphamide;Thiotepa;Taxoids, such as paclitaxel and doxetaxel;Chlorambucil;Gemcitabine;6-Thioguanine;Mercaptopurine;Methotrexate;Cisplatin platinum analogs such as rifapril and carboplatin; vinblastine; platinum; etoposide (VP-16); ifosfamide; mitoxantrone; vincristine; vinorelbine; novantrone; teniposide; edatrexate; daunomycin; aminopterin; xeloda; ibandronate; CPT-11; the topoisomerase inhibitor RFS2000; difluoromethylornithine (DMFO); retinoids such as retinoic acid; capecitabine; and pharmaceutically acceptable salts, acids, or derivatives of any of the above. Also included are anti-hormonal agents that act to regulate or inhibit hormone action on tumors, such as anti-estrogens and selective estrogen receptor modulators (SERMs), including tamoxifen, raloxifene, droloxifene, 4-hydroxytamoxifen, trioxyphene, keoxifene, LY117018, onapristone, and toremifene (Fareston); aromatase inhibitors that inhibit the enzyme aromatase, which regulates estrogen production in the adrenal glands, such as 4(5)-imidazole, aminoglutethimide, megestrol acetate, exemestane, formestane, fadrozole, vorozole, letrozole, and anastrozole; and anti-androgens such as flutamide, nilutamide, bicalutamide, leuprolide, and goserelin; and pharmaceutically acceptable salts, acids, or derivatives of any of the above;

[0268] In specific embodiments involving administering an additional therapeutic agent (i.e., in addition to an anti-PD-1 antibody (e.g., pembrolizumab) or antigen-binding fragment thereof), the additional therapeutic agent in the combination therapy may be administered using the same dosing regimen (dosage, frequency and duration of treatment) that is typically used when the agent is used as a monotherapy to treat the same cancer. In other embodiments, the patient receives a lower total amount of the additional therapeutic agent in the combination therapy, e.g., lower doses, less frequent doses, and / or a shorter duration of treatment, than when the agent is used as a monotherapy.

[0269] The determination of the appropriate dose is made by the clinician, for example, using parameters or factors known or suspected in the art to affect or be expected to affect treatment. The appropriate dosage of the drug ("therapeutically effective amount") depends, for example, on the condition being treated, the severity and course of the condition, whether the drug is administered for prophylactic or therapeutic purposes, previous treatments, the patient's medical history and response to the drug, the type of drug used, and the discretion of the attending physician. Generally, the dose is started at a somewhat lower amount than the optimal dose, and then increased by small increments until the desired or optimal effect is achieved against any negative side effects.

[0270] The additional therapeutic agents in the combination therapy may be administered orally, intratumorally, or parenterally, including intravenous, intramuscular, intraperitoneal, subcutaneous, rectal, topical, and transdermal routes of administration. For example, the combination therapy may include an anti-PD-1 antibody or antigen-binding fragment thereof, and an anti-CTLA antibody or antigen-binding fragment thereof, both of which may be administered intravenously or subcutaneously, and a chemotherapeutic agent may be administered orally.

[0271] In a specific embodiment, the combination therapy of the invention may be used before or after surgery to remove the tumor, and may be used before, during, or after radiation therapy. The combination therapy of the invention may also be used when a patient's tumor is unresectable.

[0272] In specific embodiments, the combination therapy of the invention is administered to patients who have not been previously treated with a biotherapeutic or chemotherapeutic agent, i.e., treatment naive patients. In other embodiments, the combination therapy is administered to patients who have failed to achieve a durable response after previous therapy with a biotherapeutic or chemotherapeutic agent, i.e., treatment experienced patients.

[0273] The combination therapy of the present invention may be used to treat tumors large enough to be detected by palpation or by imaging techniques well known in the art, such as MRI, ultrasound, or CAT scan. In a specific embodiment, the combination therapy of the present invention is used to treat tumors large enough to be detected by palpation or by imaging techniques well known in the art, such as MRI, ultrasound, or CAT scan. 3 , 300mm 3 , 400mm 3 , 500mm 3 , 750mm 3 , or up to 1000mm 3 It is used to treat advanced stage tumors having dimensions of

[0274] In a specific embodiment, the combination therapy of the invention is administered to a human patient having a cancer that expresses PD-L1. In a specific embodiment, PD-L1 expression is detected using a diagnostic anti-human PD-L1 antibody, or antigen-binding fragment thereof, in an IHC assay on FFPE or frozen tissue sections of a tumor sample removed from the patient. The patient's physician may order a diagnostic test to determine PD-L1 expression in a tumor tissue sample removed from the patient prior to initiation of treatment with an anti-PD-1 antibody, or antigen-binding fragment thereof, although it is envisioned that the physician may order a first or subsequent diagnostic test at any time after initiation of treatment, such as after completion of a treatment cycle.

[0275] V. Kits and Compositions The present invention also relates to a pharmaceutical composition for subcutaneous injection comprising an anti-PD-1 antibody or antigen-binding fragment thereof at a dose of about 600 mg to about 1000 mg, and about 700 units to about 50000 units of human hyaluronidase. The present invention also relates to a pharmaceutical composition for subcutaneous injection comprising an anti-PD-1 antibody or antigen-binding fragment thereof at a dose of about 600 mg to about 1000 mg, and about 7290 units to about 12150 units of human hyaluronidase. In one embodiment, the pharmaceutical composition for subcutaneous injection comprises an anti-PD-1 antibody or antigen-binding fragment thereof at a dose of about 700 mg to about 800 mg, and about 700 units to about 50000 units of human hyaluronidase. In one embodiment, the pharmaceutical composition for subcutaneous injection comprises an anti-PD-1 antibody or antigen-binding fragment thereof at a dose of about 700 mg to about 800 mg, and about 8505 units to about 9720 units of human hyaluronidase. In one embodiment, a pharmaceutical composition for subcutaneous injection comprises a dose of 790 mg of an anti-PD-1 antibody or antigen-binding fragment thereof and 9600 units of human hyaluronidase.

[0276] The present invention also relates to a pharmaceutical composition for subcutaneous injection comprising an anti-PD-1 antibody or antigen-binding fragment thereof at a dose of about 600 mg to about 1000 mg. In one embodiment, the pharmaceutical composition for subcutaneous injection comprises an anti-PD-1 antibody or antigen-binding fragment thereof at a dose of about 700 mg to about 800 mg. In one embodiment, the pharmaceutical composition for subcutaneous injection comprises an anti-PD-1 antibody or antigen-binding fragment thereof at a dose of about 760 mg to about 790 mg. In one embodiment, the pharmaceutical composition for subcutaneous injection comprises an anti-PD-1 antibody or antigen-binding fragment thereof at a dose of 790 mg.

[0277] The present invention also relates to a pharmaceutical composition for subcutaneous injection comprising an anti-PD-1 antibody or antigen-binding fragment thereof at a dose of about 300 mg to about 500 mg, and about 700 units to about 50,000 units of human hyaluronidase. The present invention also relates to a pharmaceutical composition for subcutaneous injection comprising an anti-PD-1 antibody or antigen-binding fragment thereof at a dose of about 300 mg to about 500 mg, and about 3,645 units to about 6,075 units of human hyaluronidase. In one embodiment, the pharmaceutical composition for subcutaneous injection comprises an anti-PD-1 antibody or antigen-binding fragment thereof at a dose of about 350 mg to about 400 mg, and about 700 units to about 50,000 units of human hyaluronidase. In one embodiment, the pharmaceutical composition for subcutaneous injection comprises an anti-PD-1 antibody or antigen-binding fragment thereof at a dose of about 350 mg to about 400 mg, and about 4,252 units to about 4,860 units of human hyaluronidase. In one embodiment, the pharmaceutical composition for subcutaneous injection comprises a 395 mg dose of an anti-PD-1 antibody or antigen-binding fragment thereof and 4800 units of human hyaluronidase. In one embodiment, the pharmaceutical composition for subcutaneous injection comprises a 395 mg dose of pembrolizumab and about 33 μg of human hyaluronidase of SEQ ID NO: 18. In another embodiment, the pharmaceutical composition for subcutaneous injection comprises a 790 mg dose of pembrolizumab and about 66 μg of human hyaluronidase of SEQ ID NO: 18. In one embodiment, the pharmaceutical composition for subcutaneous injection comprises a 395 mg dose of an anti-PD-1 antibody and about 33 μg of human hyaluronidase of SEQ ID NO: 18. In another embodiment, the pharmaceutical composition for subcutaneous injection comprises a 790 mg dose of an anti-PD-1 antibody and about 66 μg of human hyaluronidase of SEQ ID NO: 18. In one embodiment of the foregoing embodiments, the anti-PD-1 antibody is a monoclonal antibody comprising a heavy chain consisting of the sequence of amino acids set forth in SEQ ID NO:11 and a light chain consisting of the sequence of amino acids set forth in SEQ ID NO:5.

[0278] In any of the methods, uses, and compositions described herein, about 700 Units to about 50,000 Units of human hyaluronidase are co-formulated with the dose of anti-PD-1 antibody or antigen-binding fragment thereof. In one embodiment, about 3,000 Units to about 15,000 Units of human hyaluronidase are co-formulated with the dose of anti-PD-1 antibody or antigen-binding fragment thereof. In one embodiment, about 3,000 Units to about 7,000 Units of human hyaluronidase are co-formulated with the dose of anti-PD-1 antibody or antigen-binding fragment thereof. In one embodiment, about 7,000 Units to about 14,000 Units of human hyaluronidase are co-formulated with the dose of anti-PD-1 antibody or antigen-binding fragment thereof. In one embodiment, about 10,000 Units to about 30,000 Units of human hyaluronidase are co-formulated with the dose of anti-PD-1 antibody or antigen-binding fragment thereof. In one embodiment, about 8,000 Units to about 10,000 Units of human hyaluronidase are co-formulated with the dose of anti-PD-1 antibody or antigen-binding fragment thereof. In one embodiment, about 4,000 Units to about 5,000 Units of human hyaluronidase are co-formulated with the dose of anti-PD-1 antibody or antigen-binding fragment thereof. In one embodiment, about 9,600 Units of human hyaluronidase are co-formulated with the dose of anti-PD-1 antibody or antigen-binding fragment thereof. In one embodiment, about 4,800 Units of human hyaluronidase are co-formulated with the dose of anti-PD-1 antibody or antigen-binding fragment thereof.

[0279] In one embodiment, the human hyaluronidase is co-formulated with the anti-PD-1 antibody or antigen-binding fragment thereof in a ratio of about 12.15 units:1 mg. In one embodiment, the human hyaluronidase is co-formulated or co-administered with the anti-PD-1 antibody or antigen-binding fragment thereof in a ratio of about 12 units:1 mg. In one embodiment, the human hyaluronidase is co-formulated or co-administered with the anti-PD-1 antibody or antigen-binding fragment thereof in a ratio of about 10-14 units:1 mg. In one embodiment, the human hyaluronidase is co-formulated or co-administered with the anti-PD-1 antibody or antigen-binding fragment thereof in a ratio of about 8-18 units:1 mg. In one embodiment, the human hyaluronidase is co-formulated or co-administered with the anti-PD-1 antibody or antigen-binding fragment thereof in a ratio of about 6-25 units:1 mg. In one embodiment, the human hyaluronidase is co-formulated or co-administered with the anti-PD-1 antibody, or antigen-binding fragment thereof, in a ratio of about 3-36 units:1 mg.

[0280] In the foregoing embodiments, the pharmaceutical composition can be contained in a vial, an on-body device, or one or more pre-filled syringes. Any pharmaceutical composition described herein can be used in the methods or uses described in Section IV.

[0281] The invention also relates to kits for treating a patient having cancer, comprising: (a) a subcutaneous composition comprising an anti-PD-1 antibody or antigen-binding fragment thereof at a dose of about 600 mg to about 1000 mg, or about 300 mg to about 500 mg (or a dose or composition described in Sections IV or V), and optionally human hyaluronidase (in any of the amounts described in Sections III, IV, or V), or any of the pharmaceutical compositions in the preceding embodiments under this section; and (b) instructions for using the composition or pharmaceutical composition of (a) in any of the methods or uses for treating cancer described herein.

[0282] The kits of the invention may provide an anti-PD-1 antibody or antigen-binding fragment thereof and optionally human hyaluronidase in a container and include a package insert. The container contains a composition comprising an anti-PD-1 antibody or antigen-binding fragment thereof at a dose of at least about 600 mg to about 1000 mg, or about 300 mg to about 500 mg, and optionally about 700 units to about 50,000 units of human hyaluronidase, and the package insert or label includes instructions for treating a patient having cancer with the composition. The container may be constructed of any shape and / or material (e.g., plastic or glass). For example, the container may be a vial, syringe, or bottle. The kit may further include other materials that may be useful for administration of the medicament, such as an on-body device, needles, and syringes. In a specific embodiment of the kit, the instructions state that the medicament is intended for use in treating a patient, as described in any of embodiments E1 to E33 above under the heading "Methods and Uses of the Invention" in IV.

[0283] In one embodiment, the composition comprises 130 mg / ml of the anti-PD-1 antibody or antigen-binding fragment thereof. In another embodiment, the composition comprises 165 mg / ml of the anti-PD-1 antibody or antigen-binding fragment thereof.

[0284] In a further embodiment, the composition further comprises L-methionine, hi a particular embodiment, L-methionine is present at a concentration of about 10 mM.

[0285] In a further embodiment, the composition further comprises a histidine buffer at about pH 5.0 to pH 6.0. In a particular embodiment, the histidine is present at a concentration of about 10 mM.

[0286] In further embodiments, the composition further comprises sucrose. In certain embodiments, the sucrose is present at a concentration of about 70 mg / mL. In certain embodiments, the sucrose is present at a concentration of 7% (w / v).

[0287] In a further implementation of the invention, the composition further comprises polysorbate 80. In certain embodiments, polysorbate 80 is present at a concentration of about 0.2 mg / mL. In certain embodiments, polysorbate 80 is present at a concentration of 0.02% (w / v).

[0288] In a specific embodiment, the composition comprises 10 mM L-methionine, 10 mM histidine, pH 5.5, 7% sucrose, 0.02% polysorbate 80, and 130 mg / mL of an anti-PD-1 antibody or antigen-binding fragment thereof.

[0289] In a specific embodiment, the composition comprises 10 mM L-methionine, 10 mM histidine, pH 5.5, 7% sucrose, 0.02% polysorbate 80, and 165 mg / mL of an anti-PD-1 antibody or antigen-binding fragment thereof.

[0290] In a further embodiment, the composition further comprises 2000 U / ml of PH20 variant 2. In a further embodiment, the composition further comprises 2000 U / ml of rHuPH20.

[0291] In one embodiment, the composition is contained in a vial. In another embodiment, the composition is contained in one or more pre-filled syringes. In one embodiment, the composition is contained in two pre-filled syringes.

[0292] In any of the kits or compositions of the invention, the anti-PD-1 antibody or antigen-binding fragment can be any of the antibodies or antigen-binding fragments described in Section II of the Detailed Description of the Invention, "PD-1 Antibodies and Antigen-Binding Fragments Useful in the Invention." In one embodiment, the anti-PD-1 antibody or antigen-binding fragment thereof is pembrolizumab. In another embodiment, the anti-PD-1 antibody or antigen-binding fragment thereof is a pembrolizumab variant.

[0293] In any of the kits or compositions of the invention, the anti-PD-1 antibody, or antigen-binding fragment thereof, may be co-formulated with a human hyaluronidase, as described in Section III.

[0294] These and other aspects of the present invention will become apparent from the teachings contained herein, including the illustrative specific embodiments listed below.

[0295] Common methods The standard method in molecular biology is the method described by Sambrook, Fritsch and Maniatis (1982 & 1989 2 nd Edition, 2001 3 rd Edition)Molecular Cloning,A Laboratory Manual,Cold Spring Harbor Laboratory Press,Cold Spring Harbor,NY;Sambrook and Russell(2001)Molecular Cloning,3 rd ed., Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY; Wu (1993) Recombinant DNA, Vol. 217, Academic Press, San Diego, CA). Standard methods are described in Ausbel, et al. (2001) Current Protocols in Molecular Biology, Vols. 1-4, John Wiley and Sons, Inc. New York, NY, which describe cloning in bacterial cells and DNA mutagenesis (Vol. 1), cloning in mammalian cells and yeast (Vol. 2), glycoconjugates and protein expression (Vol. 3), and bioinformatics (Vol. 4).

[0296] All publications mentioned herein are incorporated by reference for the purpose of describing and disclosing methodology and materials that might be used in connection with the present invention.

[0297] Although different embodiments of the present invention have been described herein with reference to the accompanying drawings, it should be understood that the invention is not limited to those precise embodiments, and various changes and modifications may be made by those skilled in the art without departing from the scope or spirit of the present invention as defined in the appended claims.

[0298] [Example 1] Population PK modeling Pembrolizumab is currently approved for use in multiple cancer indications at doses of either 200 mg or 2 mg / kg Q3W or 400 mg Q6W administered as an IV infusion. An alternative subcutaneous ("SC") formulation containing hyaluronidase would provide convenience and flexibility to patients and prescribers. A Phase I clinical trial designed to estimate the bioavailability of an SC formulation of pembrolizumab in combination with hyaluronidase PH20 variant 2 ("SC pembrolizumab-HLN" or "pembrolizumab-HLN SC") at two different solution strengths of pembrolizumab, at concentrations of 165 mg / ml (arm 1) and 130 mg / ml (arm 2), was conducted. The bioavailability of two different subcutaneous solution strengths of pembrolizumab (two independent study arms) was estimated in this study using a subcutaneous dose (650 mg with a 6-week dosing interval (cycle) given at two different concentrations / volumes of pembrolizumab (shown in Table 6) compared with the IV dose of pembrolizumab (400 mg with a 6-week dosing interval). [Table 6]

[0299] Patients in each study arm with advanced melanoma or non-small cell lung cancer (NSCLC) or renal cell carcinoma (RCC) received both SC and IV pembrolizumab, respectively, in the following crossover design: Cycle 1: SC 650 mg; Cycle 2: IV 400 mg; Cycle 3: SC 650 mg; Cycles 4-18: IV 400 mg.

[0300] Eligible patients were aged 18 years or older, had unresectable stage III or IV melanoma not amenable to local therapy, measurable disease by RECIST v1.1, Eastern Cooperative Oncology Group performance status of 0 or 1, or had not received prior therapy for progressive disease (except for BRAF V600 Patients were required to have received a BRAF / MEK inhibitor for mutant disease and prior adjuvant or neoadjuvant therapy ≥ 4 weeks from randomization), a histologically or cytologically confirmed diagnosis of NSCLC that was either metastatic (stage IV [M1a, M1b, or M1c by current AJCC criteria]) non-squamous NSCLC without EGFR, ALK, or ROS1 genomic tumor aberrations, or metastatic (stage IV [M1a, M1b, or M1c by current AJCC criteria]) squamous NSCLC with or without clear cell components, unresectable locally advanced / metastatic (i.e., stage IV RCC by AJCC), and had not received prior systemic therapy for advanced ccRCC.

[0301] For participants with NSCLC or RCC, treatment with pembrolizumab-HLN SC and pembrolizumab IV included combination with standard of care as follows: NSCLC: with pemetrexed and platinum (carboplatin or cisplatin) chemotherapy for the 1L treatment of patients with metastatic non-squamous NSCLC without EGFR or ALK genomic tumor aberrations; or with carboplatin and either paclitaxel or nab-paclitaxel for the 1L treatment of patients with metastatic squamous NSCLC.

[0302] RCC: With axitinib for 1L treatment of patients with advanced RCC. [Table 7]

[0303] Using serum concentration data from 81 subjects collected throughout cycle 1 (i.e., weeks 0–6) and cycle 2 (i.e., weeks 7–12) of a Phase I clinical trial, the PK of SC pembrolizumab given as a co-formulation with hyaluronidase was characterized along with extensive historical pembrolizumab IV PK data using population PK analysis. Nonlinear mixed-effects modeling was applied to the Phase I data with prior information from a previously established pembrolizumab reference PK model. The reference pembrolizumab PK model was based on pembrolizumab PK data collected from 2993 patients with various cancers who received pembrolizumab doses of 1–10 mg / kg Q2W, 2–10 mg / kg Q3W, or 200 mg Q3W in Phase I or Phase III clinical trials. Pembrolizumab absorption phase PK parameters were estimated for SC administration from the Phase I data, and differences between the two solution strengths were also assessed. The distribution and elimination parameters (clearance (CL), central volume of distribution (Vc), intercompartmental clearance (Q), and peripheral volume of distribution (Vp)) were fixed from the reference IV model since these phases are the same for IV and SC administration. Given the small sample size and short duration of SC administration in the study, i.e., two treatment cycles, parameters describing the time dependence and effect of patient baseline characteristics on pembrolizumab PK were also fixed from the previously established reference IV PK model.

[0304] The new population PK model developed was able to simultaneously describe pembrolizumab PK after IV or SC (with hyaluronidase) administration. The final parameter estimates of the combined SC and IV population PK model are shown in Table 8. The absorption phase for SC administration was characterized by first-order absorption rate (ka) and bioavailability (F) parameters. The distribution and elimination phases were described by a two-compartment model with time-dependent clearance and fixed effects of body weight, as historically established in the reference pembrolizumab PK model. The inclusion of covariate effects for the strength of the SC solution was not statistically significant, indicating no significant differences in bioavailability and absorption rate between the two pembrolizumab-HLN SC solution strengths (i.e., arm 1 165 mg / mL and arm 2 130 mg / ml). Goodness-of-fit assessment demonstrated the absence of structural bias as a function of drug concentration or time. Analysis showed that pembrolizumab-HLN had an estimated bioavailability of 57% when administered SC (range: 38%-75%). The median time to achieve maximum pembrolizumab serum concentrations in the subcutaneous formulation with hyaluronidase was estimated to be 4 days (range 2-35 days). Additionally, anti-drug antibodies (ADAs) were observed in 1 of 83 subjects (<2%) in the Phase I study. [Table 8]

[0305] Ongoing results from the Phase 1 study indicate no new systemic safety signals with SC pembrolizumab-HLN. All injection site reactions were non-serious, mostly mild (grade 1), and effectively managed. The overall safety profile during both SC pembrolizumab-HLN and pembrolizumab IV cycles was consistent with the known safety profile of pembrolizumab monotherapy and standard of care administered to participants with NSCLC and RCC.

[0306] The two tested SC solution strengths of pembrolizumab-HLN (130 mg / ml and 165 mg / ml) had similar absorption PK. SC administration of pembrolizumab-HLN was well tolerated without significant ADAs. Based on analysis of a simulation based on a PK model using estimated bioavailability and intersubject variability from this study, a subcutaneous dose of pembrolizumab-HLN of 760 to 790 mg Q6W is expected to provide similar exposure as the approved dose of pembrolizumab IV of 400 mg Q6W.

[0307] [Example 2] A 6-week (Q6W) dosing schedule of SC pembrolizumab co-formulated with hyaluronidase across multiple tumor types, based on evaluation using modeling and simulation with 400 mg Q6W IV as the reference regimen.

[0308] Pembrolizumab is an anti-PD-1 checkpoint inhibitor currently approved for use in multiple cancer indications and has demonstrated safety and efficacy when administered by intravenous infusion (IV) at doses of 200mg Q3W, 2mg / kg Q3W or 400mg Q6W. Robust characterization of pembrolizumab's pharmacokinetic (PK) and exposure (concentration) response (ER) relationships for both efficacy and safety enables the use of model-based approaches to support alternative routes of administration of pembrolizumab. Previously, KN-252 efficacy data in 294 advanced melanoma patients demonstrated a flat exposure-response relationship over a Ctrough range of 3.5-32.5ug / ml with a 200mg Q3W IV dose of pembrolizumab. KN-555 PK data in 89 advanced melanoma patients indicates that the observed Ctrough of 400mg Q6W IV is well within the range of the previously established flat exposure-response relationship. Therefore, a Ctrough of 400 mg Q6W IV is sufficient to maximize pembrolizumab efficacy [Figure 1].

[0309] KEYNOTE-555 (NCT03665597) was an open-label, multicenter, phase 1 clinical trial investigating pembrolizumab in patients with advanced melanoma. Cohort B investigated the pharmacokinetics, efficacy, and safety of intravenous pembrolizumab at a dose of 400 mg every 6 weeks. 101 patients were screened and treated. Clinically meaningful objective response rates and durable progression-free survival within the expected range for first-line pembrolizumab were observed. The objective response rate was 50.5% (95% CI, 40.4-60.6), with 19 patients (18.8%) having a complete response and 32 patients (31.7%) having a partial response. Median duration of response was not reached (NR; range 2.4+-21.0+ months). Median progression-free survival was 13.8 months (95% CI, 4.1-NR). Observed pharmacokinetic exposures were consistent with model predictions for pembrolizumab 400 mg Q6W IV and within exposures with other approved and tested regimens (2 mg / kg or 200 mg Q3W IV and 10 mg / kg Q2W IV) (see Figure 2). Safety in cohort B was comparable to the known profile of pembrolizumab in melanoma with the 200 mg Q3W IV regimen. Grade 3-5 treatment-related adverse events occurred in 13 patients (12.9%). Three patients (3.0%) discontinued due to treatment-related adverse events.

[0310] The phase 2 KEYNOTE-B68 trial investigated the efficacy and safety of pembrolizumab administered 400 mg IV every 6 weeks in relapsed / refractory (R / R) classical Hodgkin lymphoma (cHL) or primary mediastinal B-cell lymphoma (PMBCL). At data cutoff, 66 patients (60 R / R cHL, 6 R / R PMBCL) were enrolled. Median (range) follow-up was 8.9 months (1-15.9) for patients with R / R cHL and 10.6 months (5.1-15.4) for patients with R / R PMBCL. The ORR was 65% (95% CI, 51.6 to 76.9 [33.3% CR; 31.7% PR]) for patients with R / R cHL and 50% (95% CI, 11.8 to 88.2 [33.3% CR; 16.7% PR]) for R / R PMBCL. Drug-related adverse events (AEs) occurred in 24 patients (40%) with R / R cHL and in 2 patients (33.3%) with R / R PMBCL. Grade ≥3 drug-related AEs occurred in 3 patients (5%) with R / R cHL and in 1 patient (16.7%) with R / R PMBCL. Immune-mediated AEs occurred in 13 patients (21.7%) with R / R cHL and in 1 patient (16.7%) with R / R PMBCL. Grade ≥3 immune-mediated AEs occurred in 2 patients (3.3%) with R / R cHL. With approximately 9 months of follow-up, pembrolizumab 400 mg Q6W IV demonstrated robust antitumor activity in patients with R / R cHL and R / R PMBCL without new safety concerns. The observed ORR was similar to pembrolizumab 200 mg Q3W IV in R / R cHL and R / R PMBCL.

[0311] Simulations based on the PK model were performed to select the SC dose, targeting consistency of the SC PK exposure profile with the approved 400 mg Q6W IV dose and an overall exposure profile based on clinical experience with IV pembrolizumab. Simulations were performed on a pooled dataset (total N=3105 subjects) from Phase 1 studies evaluating pembrolizumab-HLN SC, as well as a reference pembrolizumab IV PK dataset that included 2993 subjects with melanoma or non-small cell lung cancer (NSCLC) from previous Phase I and Phase III studies of IV pembrolizumab.

[0312] A combined SC and IV PK model (described in Table 8), including estimates of population mean PK parameters as well as between-subject variability and residual error for each parameter, was used to simulate pembrolizumab serum concentrations for a range of doses (18 weeks, steady state achieved) of 600 mg to 1200 mg Q6W pembrolizumab-HLN SC and 400 mg Q6W IV from Cycle 1 to Cycle 3. For each subject in the dataset, simulated trough concentrations at the end of the dosing interval (C trough ) and area under the curve (AUC) exposure were determined in both cycle 1 (first dose) and cycle 3 (representing steady state). trough and AUC 0-6wks represents the PK exposure measurement and is considered a driver of pembrolizumab efficacy. Cycle 1 represents the PK exposure achieved after the first dose was administered. Cycle 3 represents the PK exposure achieved at steady state, which is the exposure subsequently maintained throughout the treatment period. For each SC dose and the 400 mg IV dose of pembrolizumab, C trough and AUC 0-6wks The geometric mean ratios (GMRs) of SC to IV pembrolizumab (as the ratio of the GMs for each formulation group) for both of these PK exposure measures were then calculated for treatment cycles 1 and 3.

[0313] Simulations based on the PK model show that pembrolizumab-HLN SC (containing hyaluronidase PH20 variant 2) doses ranging from 600 to 1000 mg Q6W provide equivalent exposure to the approved dose of 400 mg Q6W of pembrolizumab administered via IV infusion and are safe considering the highest clinically evaluated dose at 10 mg / kg Q2W IV with established safety. In principle, similar PK exposure will provide similar efficacy and safety of pembrolizumab, given that the exposure-response relationship for both efficacy and safety has already been well established for pembrolizumab. Thus, SC doses in this range are expected to have equivalent efficacy and safety to 400 mg Q6W IV.

[0314] Lower limit of exposure (AUC or C trough ), it would be appropriate to establish non-inferiority using a commonly accepted margin of the lower limit of the 90% CI around a GMR > 0.8. This would confirm that the efficacy of a dose of pembrolizumab co-formulated with hyaluronidase and administered subcutaneously is no worse than that of a dose of pembrolizumab administered by the IV route of administration ("IV pembrolizumab") (400 mg Q6W).

[0315] The simulations developed showed that all SC pembrolizumab-HLN doses ≥ 600 mg Q6W had a mean SC:IV C value > 1. trough The Cycle 1 C:IV AUC ratios showed that SC pembrolizumab-HLN doses of 600 mg Q6W and above all had mean SC:IV AUC ratios greater than 0.8. See Tables 9A and 9B. Tables 9A-B show the C:IV AUC ratios for Cycle 1 across the different evaluated Q6W SC doses compared to pembrolizumab 400 mg Q6W IV. trough and steady-state C trough Tables 9C-D show the SC (subcutaneous):IV (intravenous) geometric mean (GM) ratios (GMR) of the 100 mg / kg / day group (Ctroughss). Tables 9C-D show the AUC.C1 and 100 mg / kg / day group (Ctroughss) in cycle 1 (AUC.C1) and steady state (AUC.ss). 0-6wks The SC:IV GMR is shown.

[0316] [Table 9] [Table 10] [Table 11] [Table 12]

[0317] SC pembrolizumab-HLN doses ranging from 760-790 mg Q6W have similar PK exposure to the approved dose of pembrolizumab of 400 mg Q6W administered via IV infusion. Based on the analyses performed, the efficacy of SC pembrolizumab-HLN at doses of 760-790 mg Q6W is expected to be similar to that of 400 mg Q6W IV pembrolizumab, based on the following:

[0318] C at a dose of 760–790 mg Q6W SC trough is expected to be approximately 25-30% higher than 400 mg Q6W IV throughout the treatment period. trough The distributions of AUC100 and AUC101 overlap significantly between SC and IV at both cycle 1 and steady state. See Figures 3A and 3B. AUC100 and AUC101 at 760-790 mg Q6W SC doses 0-6wks Exposure is expected to be approximately 2-10% higher than 400 mg Q6W IV throughout the treatment period. 0-6wks The distributions of α, β, and β largely overlap between SC and IV at both cycle 1 and steady state (see Figures 4A and 4B).

[0319] When comparing SC pembrolizumab to IV pembrolizumab, it is important to note the significant differences inherent in the PK profiles between SC and IV administration. Typically, at comparable doses (adjusted for bioavailability), concentrations with SC administration develop gradually over approximately 4 days, with peak concentrations (C max ) is the C achieved at the end of the IV infusion. max Specifically, at a Q6W SC dose of 760–790 mg, C max was achieved with 400 mg Q6W IV max (approximately 38-35% decrease in cycle 1 and approximately 25-22% decrease at steady state) (see Figure 5). Thus, with the pembrolizumab-HLN SC dose range, C is expected to be significantly lower throughout treatment compared to the approved IV dose of 400 mg Q6W. max No increase in PK is expected. Furthermore, all PK exposures of 760-790 mg Q3W SC pembrolizumab-HLN remain reliably well below the 5-fold higher dose / exposure at 10 mg / kg Q2W IV, the highest clinically evaluated dose with established safety. Thus, the safety profile following SC administration of 760-790 mg Q6W SC pembrolizumab-HLN is unlikely to differ from the previously established safety profile for the identified 400 mg Q6W IV pembrolizumab dose, and therefore, a quantitative assessment of the upper limit of exposure was not further evaluated.

[0320] The selected dose for the Phase III clinical trial of pembrolizumab SC with hyaluronidase PH20 variant 2 is 790 mg Q6W. Figures 3A and 3B show the C-values ​​for the 790 mg Q6W SC and 400 mg Q6W IV doses of pembrolizumab. trough The authors summarize the results of a population simulation that included variation in C. The simulations developed showed that the 790 mg Q6W SC dose was associated with a C. troughFigures 4A and 4B show the AUC 0-6wks The results of a population simulation including variability in AUC 100 and 150 mg Q6W SC across a range of patients that generally overlap with the 400 mg Q6W IV dose are summarized in Table 1. 0-6wks Figure 5 shows the C for pembrolizumab doses of 790 mg Q6W SC and 400 mg Q6W IV. max The results of a population simulation including variability in C. The simulations show that the 790 mg Q6W SC dose generally results in a lower C than the 400 mg Q6W IV dose. max 3A and 3B, and 4A and 4B, respectively, show that the C trough and AUC 0-6wks Figure 5 shows the distribution (5th, 25th, 50th, 75th, and 95th percentiles) of C at cycle 1 and steady state, and Figure 6 shows the C at cycle 1 and steady state distribution (5th, 25th, 50th, 75th, and 95th percentiles) at cycle 1 and steady state, and Figure 7 shows the C at cycle 1 and steady state distribution (5th, 25th, 50th, 75th, and 95th percentiles) at cycle 1 and steady state, and Figure 8 shows the C at cycle 1 and steady state distribution (5th, 25th, 50th, 75th, and 95th percentiles) at cycle 1 and steady state, and Figure 9 shows the C at cycle 1 and steady state distribution (5th, 25th, 50th, 75th, and 95th percentiles) at cycle 1 and steady state, and Figure 10 shows the max The steady-state distribution (5th, 25th, 50th, 75th, and 95th percentiles) of PK is shown. Simulated PK exposure measurements in 3105 subjects are shown.

[0321] Overall, the model-based simulations supported by Tables 9A-D, Figures 3A, 3B, 4A, 4B, and 5 indicate that a dose of 790 mg Q6W of pembrolizumab-HLN administered SC results in an optimal PK exposure profile similar to the approved dose of 400 mg Q6W of pembrolizumab IV and should therefore maintain efficacy while remaining within clinical safety margins.

[0322] [Example 3] A Phase 3 Randomized, Open-Label Clinical Trial to Evaluate the Pharmacokinetics and Safety of Subcutaneous Pembrolizumab Coformulated with Hyaluronidase (Pembrolizumab-HLN) Versus Intravenous Pembrolizumab Administered Concomitantly with Chemotherapy in the First-Line Treatment of Participants with Metastatic Non-Small Cell Lung Cancer This is a phase 3, randomized, validated, controlled, parallel-arm, multicenter, open-label study of pembrolizumab-HLN and platinum doublet therapy versus pembrolizumab IV and platinum doublet therapy in participants with treatment-naïve metastatic NSCLC.

[0323] After a screening period of up to 28 days, participants will be randomly assigned in a 2:1 ratio to Arm 1 or Arm 2. Each cycle will last 6 weeks. Arm 1 will be pembrolizumab-HLN Q6W in combination with platinum doublet chemotherapy for up to 18 cycles. Arm 2 will be pembrolizumab IV Q6W in combination with platinum doublet chemotherapy for up to 18 cycles.

[0324] Platinum doublet chemotherapy is as follows:For non-squamous NSCLC: pemetrexed Q3W with platinum chemotherapy (cisplatin Q3W or carboplatin Q3W) for up to four infusions, followed by maintenance pemetrexed until one of the discontinuation criteria for the study intervention is met.For squamous NSCLC: carboplatin Q3W with taxane (paclitaxel Q3W or nab-paclitaxel [days 1, 8, 15, 22, 29, and 36 of cycles 1 and 2]) for up to four infusions.

[0325] Randomization will be stratified by ECOG performance status, histology, PD-L1 TPS, and region. Each participant will receive the study intervention until one of the conditions for discontinuation of the study intervention is met.

[0326] The dual primary objective of this study is to evaluate the cycle 1 AUC 0-6wks and steady state (cycle 3) C troughThe objective of this study is to compare the efficacy and safety of 100 mg / kg / day of chemotherapy with 100 mg / kg / day of chemotherapy alone. Non-inferiority will be assessed with a non-inferiority margin of 0.8. Tumor response will be assessed according to an adjustment to RECIST 1.1. Safety evaluations will also be performed.

[0327] Pembrolizumab IV in combination with platinum doublet therapy is indicated for the first-line treatment of patients with metastatic NSCLC. This study will primarily compare pembrolizumab exposure between pembrolizumab-HLN (arm 1) and pembrolizumab IV (arm 2) over a Q6W dosing regimen, both in combination with chemotherapy, in participants with treatment-naïve metastatic NSCLC. Pembrolizumab C between pembrolizumab-HLN and pembrolizumab 200 mg IV Q3W trough Further descriptive comparisons of are also performed using model-based exposures.

[0328] Study objectives and endpoints: In participants with treatment-naïve metastatic NSCLC: [Table 13] [Table 14]

[0329] Rationale for Pharmacokinetic Endpoints Cycle 1 AUC 0-6wks and steady state (cycle 3) C trough To compare pembrolizumab exposure between pembrolizumab-HLN and pembrolizumab IV Q6W using the dual primary endpoint of AUC in cycle 1 0-6wksis the most conservative PK value to ensure non-inferiority of SC exposure compared to IV. The difference in exposure between SC and IV administration of pembrolizumab should decrease after multiple dosing due to accumulation. Therefore, demonstrating non-inferiority of SC AUC exposure in cycle 1 also implies non-inferiority at steady state. Based on our analysis, the higher AUC exposure at steady state after SC administration is not expected to exceed the clinical safety margin established for pembrolizumab (i.e., exposure at 10 mg / kg Q2W IV).

[0330] Steady state (cycle 3) C trough is the steady-state concentration for pembrolizumab at the end of the 6-week dosing interval after the third dose. Because the pharmacological activity of mAbs is mediated through direct interaction with specific targets, target saturation can be used as a surrogate for maximum pharmacological and therapeutic activity. Pembrolizumab exposure at the approved IV dose is expected to maintain PD-1 target saturation throughout the dosing interval, thereby maintaining efficacy. Thus, the C of the approved IV dose of 400 mg Q6W trough can be considered a threshold above which the target saturation and efficacy are maintained. Pembrolizumab-HLN Pembrolizumab Steady State (Cycle 3) C trough Demonstration of non-inferiority of IV pembrolizumab would allow the inference that efficacy similar to that of IV pembrolizumab is maintained.

[0331] Secondary PK endpoints allow further characterization of pembrolizumab exposure: Comparison with pembrolizumab IV Q6W: Cycle 1:C max , C trough ; Steady state (Cycle 3): AUC 0-6wks and C max For comparison with pembrolizumab IV Q3W: Model-based C trough Simulations based on PK modeling indicate that a pembrolizumab SC dose of 790 mg Q6W provides similar exposure to the approved pembrolizumab IV dose of 400 mg Q6W. Efficacy is expected to be maintained with SC pembrolizumab at a dose of 790 mg Q6W based on the following:

[0332] C at 790mg Q6W SC trough is expected to be approximately 30% higher than 400 mg Q6W IV throughout the treatment period. Furthermore, the C between SC and IV at both cycle 1 and steady state trough Distribution of duplications (see Figures 3A and 3B).

[0333] AUC at 790 mg Q6W SC 0-6wks Exposure is expected to be approximately 10% higher than 400 mg Q6W IV throughout the treatment period. 0-6wks The distributions of overlap between SC and IV in both cycle 1 and steady state (see Figures 4A and 4B).

[0334] Safety is expected to be maintained with SC pembrolizumab at a dose of 790 mg Q6W based on the following:

[0335] C max was achieved with 400 mg Q6W IV max The expected C over the treatment period is therefore lower (approximately 35% in cycle 1 and approximately 22% at steady state) compared to the approved dose of 400 mg Q3W IV. max There is no increase.

[0336] - 6-week dosing interval and all SC exposures during the treatment period (C max , C avg , C trough ) at 400 mg Q6W IV max and initial concentrations, and are expected to remain well below the maximum dose and exposure established for clinical safety (i.e., 10 mg / kg Q2W).

[0337] Overall, as discussed above, the developed model-based simulations indicate that a dose of 790 mg Q6W of pembrolizumab-HLN administered subcutaneously should result in an optimal PK exposure profile similar to the approved dose of 400 mg Q6W of pembrolizumab IV, thereby remaining within the clinical safety margin while maintaining efficacy.

[0338] [Example 4] A three-weekly (Q3W) dosing schedule for SC pembrolizumab-HLN across multiple tumor types, assessed using modeling and simulation with Q6W SC pembrolizumab-HLN as the reference regimen Simulations based on the developed PK model show that the dosing regimen of SC pembrolizumab with hyaluronidase PH20 variant 2 at 790 mg Q6W results in similar exposure to the approved dose of pembrolizumab at 400 mg Q6W administered via IV infusion. In principle, similar PK exposure results in similar efficacy and safety of pembrolizumab, given that the exposure-response relationship for both efficacy and safety has already been well established for pembrolizumab. Using such an exposure-matching approach, the Q3W dose of SC pembrolizumab-HLN was selected using PK model-based simulations, with the Q6W SC pembrolizumab-HLN dose as the reference.

[0339] Simulations based on the PK model were performed to select the Q3W SC dose to target the consistency of the SC PK exposure profile with the Q6W SC dose and the overall exposure profile based on clinical experience with IV pembrolizumab. Simulations were performed on a pooled dataset (total N=3105 subjects) from Phase 1 studies evaluating pembrolizumab-HLN SC, as well as a reference pembrolizumab IV PK dataset that included 2993 subjects with melanoma or non-small cell lung cancer (NSCLC) from previous Phase I and Phase III studies of IV pembrolizumab.

[0340] Pembrolizumab serum concentrations were simulated for a range of pembrolizumab-HLN SC doses from 380 mg to 410 mg Q3W, as well as a range of Q6W SC doses equivalent to twice the Q3W dose from cycle 1 through cycle 6 or cycle 3 (18 weeks, steady state achieved) using a combined SC and IV PK model (described in Table 8) that includes estimates of population mean PK parameters, as well as between-subject variability and residual error in each parameter. For each subject in the dataset, simulated trough concentrations at the end of the dosing interval (C trough ) and area under the curve (AUC) exposure were determined both at cycle 1 (first dose) and at steady state (cycle 6 for Q3W or cycle 3 for Q6W). trough and AUC represent PK exposure measurements and are considered drivers of pembrolizumab efficacy. Cycle 1 represents the PK exposure achieved after the first dose is administered. The PK exposure achieved at steady state is the exposure subsequently maintained throughout the treatment period. For each SC dose of pembrolizumab, C trough Geometric means (GM) of AUC and PK were then calculated. Geometric mean ratios (GMR) of SC to IV pembrolizumab (as the ratio of GM for each formulation group) for both of these PK exposure measures were then calculated for treatment cycle 1 and steady state.

[0341] The developed simulations demonstrated that a Q3W SC pembrolizumab-HLN dose that is half the corresponding Q6W SC pembrolizumab-HLN dose significantly reduces the Q6W SC dose from cycle 1 to steady state (mean SC-Q3W:SC-Q6W C trough The study showed that the PEMBRO-HLN-HLN-treated patients had a Ctrough greater than 1 (mean SC-Q3W:SC-Q6W AUC ratio >0.8), comparable AUC exposure in Cycle 1 (mean SC-Q3W:SC-Q6W AUC ratio >0.8), and the same AUC exposure at steady state (mean SC-Q3W:SC-Q6W AUC ratio 1). See Tables 12A-12D. Tables 12A-B show the Ctrough in Cycle 1 across the different evaluated Q3W and Q6W SC doses of pembrolizumab-HLN. trough and steady-state C trough (C troughss) and Q3W:Q6W geometric mean ratio (GMR). In Table 12A, the sampling period for the SC-Q3W regimen was 3 weeks and the sampling period for the SC-Q6W reference was 6 weeks. In Table 12B, the sampling period for the SC-Q3W regimen was 6 weeks and the sampling period for the SC-Q6W reference was 6 weeks. Tables 12C-D show the AUC at cycle 1 (AUC.C1), cycle 2, and steady state (AUC.ss) across the different evaluated Q3W and Q6W SC doses of pembrolizumab-HLN. 0-6wks The Q3W:Q6W GMR for each treatment group is shown in Tables 12C-D. In Tables 12C-D, the sampling period for the SC-Q3W regimen was 6 weeks, and the sampling period for the SC-Q6W reference was 6 weeks.

[0342] [Table 15] [Table 16] [Table 17] [Table 18]

[0343] The selected dose of pembrolizumab SC with hyaluronidase PH20 variant 2 is 790 mg Q6W. The corresponding selected dose of pembrolizumab SC with hyaluronidase PH20 variant 2 is 395 mg Q3W. Based on our analysis, efficacy is expected to be maintained with SC pembrolizumab-HLN at a dose of 395 mg Q3W based on the following:

[0344] C at a dose of 395 mg Q3W SC trough is expected to be approximately 15-60% higher than 790 mg Q6W SC throughout the treatment period. trough The distributions of σ largely overlap between SC-Q3W and SC-Q6W at both cycle 1 and steady state (see Figures 6A and 6B).

[0345] AUC at 395mg Q3W SC dose 0-6wks Exposure is expected to be similar to that of 790 mg Q6W SC throughout the treatment period. 0-6wks The distributions of σ largely overlap between SC-Q3W and SC-Q6W at both cycle 1 and steady state (see Figures 7A and 7B).

[0346] Safety is expected to be maintained with SC pembrolizumab-HLN at a dose of 395 mg Q3W, assuming Cmax remains lower than the 790 mg Q6W dose throughout cycle 1 through steady state treatment.

[0347] 6A-6B show C-values ​​for doses of pembrolizumab-HLN 395 mg Q3W SC and 790 mg Q6W SC. trough The authors summarize the results of a population simulation that included variation in C. The simulations developed showed that the 395 mg Q3W SC dose was associated with a C. trough Figures 7A and 7B show that the AUC 0-6wks The results of a population simulation including variability in AUC 395 mg Q3W SC across a range of patients generally overlapping with the 790 mg Q6W SC dose. 0-6wks Figures 6A and 6B, and 7A and 7B show the C range using simulations based on the PK model for pembrolizumab-HLN at doses of 395 mg Q3W and 790 mg Q6W SC, respectively. trough and AUC 0-6wks Figure 1 shows the distributions (5th, 25th, 50th, 75th, and 95th percentiles) of PK at cycle 1 and steady state. Simulated PK exposure measurements in 3105 subjects are shown.

[0348] Overall, the model-based simulations, as supported by Tables 12A-D, Figures 6A, 6B, 7A, and 7B, indicate that a 395 mg Q3W dose of pembrolizumab-HLN administered SC provides an optimal PK exposure profile consistent with a 790 mg Q6W pembrolizumab-HLN SC dose, and should therefore maintain efficacy while remaining within clinical safety margins. Furthermore, based on our analysis, a 300-500 mg Q3W SC pembrolizumab-HLN dose is also expected to be safe and efficacious, with PK exposures comparable to a 600-1000 mg Q6W SC pembrolizumab-HLN dose.

[0349] Approximately 50 participants with unresectable advanced melanoma will be assigned to arm 4 of the Phase I clinical trial in Table 7 (165 mg / mL pembrolizumab-HLN SC, 2.39 ml). Treatment cycles will be 21 days (Q3W) with 395 mg pembrolizumab doses containing pembrolizumab-HLN SC. PK exposure measurements at cycle 1 and steady state (cycle 6), including Ctrough, Cmax, and AUC, will be monitored. Overall survival, safety, and anti-pembrolizumab antibodies will be monitored in arm 4. Pembrolizumab trough concentrations (Ctrough) following pembrolizumab-HLN SC treatment will be measured by collecting patient PK samples pre-dose (0-3 hours) at any time on day 1 of cycles 1 and 6 and days 2, 4, 6, 10, and 15 of cycles 1 and 6. Pembrolizumab maximum plasma concentration (Cmax) following pembrolizumab-HLN SC treatment will be measured by collecting patient PK samples pre-dose (0-3 hours) on Day 1 of Cycles 1 and 6 and any time on Days 2, 4, 6, 10, and 15 of Cycles 1 and 6. Pembrolizumab area under the curve (AUC) following pembrolizumab-HLN SC treatment will be measured by collecting patient PK samples pre-dose (0-3 hours) on Day 1 of Cycles 1 and 6 and any time on Days 2, 4, 6, 10, and 15 of Cycles 1 and 6.

[0350] [Example 5] A 6-week (Q6W) dosing schedule of SC pembrolizumab (without hyaluronidase) across multiple tumor types, assessed using PK modeling and simulation with 400 mg Q6W IV as the reference regimen The PK data from the Phase 1 study of SC pembrolizumab-HLN described above were generally consistent with the PK data from the Phase 1 study of SC pembrolizumab without hyaluronidase (KEYNOTE-555 Cohort A, the same formulation in Table 6 without hyaluronidase). A comparison of both studies is shown in Table 13. Thus, the bioavailability ranges largely overlap for these formulations, and the data confirm that the addition of hyaluronidase does not significantly affect the PK of SC pembrolizumab. Thus, the PK exposure (C trough ,AUC,C max ) are expected to be similar. Thus, by inference, the selected 790 mg Q6W dose and the 600-1000 mg dose range identified for SC pembrolizumab with hyaluronidase are also expected to be safe and effective when applied to SC pembrolizumab without hyaluronidase. [Table 19]

[0351] [Example 6] Pharmacokinetics of Pembrolizumab Across Tumor Types and Combination Therapies Over the course of pembrolizumab's clinical development, extensive historical and emerging data across multiple indications and treatment settings have shown that both PK and immunogenicity are generally consistent across tumor types and between monotherapy and combination treatments. Furthermore, the exposure-response relationship of pembrolizumab for both efficacy and safety has been well established based on eight randomized dose comparisons in melanoma and non-small lung cancer (NSCLC) and has been shown to be flat over a dose / exposure range of more than five-fold the clinically tested 2 mg / kg Q3W to 10 mg / kg Q2W IV. Consistent flat exposure-response relationships have also been observed for other indications (e.g., head and neck squamous cell carcinoma (HNSCC), classical Hodgkin lymphoma (cHL), UC (urothelial carcinoma), GC (gastric cancer), PMBCL (primary mediastinal large B-cell lymphoma), and MSI-H cancers) based on pooled analysis of data from treatment arms across studies (Figure 8).

[0352] PK consistency across tumor types The reference PK model of pembrolizumab, which serves as the basis for pembrolizumab labeling, is based on a definitive analysis characterizing pembrolizumab PK based on a robust dataset of 2993 participants with melanoma or NSCLC from KEYNOTE-001, KEYNOTE-002, KEYNOTE-006, KEYNOTE-010, and KEYNOTE-024 (including doses of 2 mg / kg Q3W, 10 mg / kg Q3W, 10 mg / kg Q2W, and 200 mg Q3W). Tumor type did not significantly affect PK in the reference analysis. This model was further evaluated for consistency of PK parameters across other approved indications (HNSCC, UC, GC, MSI-H cancer, cHL, PMBCL, HCC, and cervical cancer). Data from participants with these cancer types were added to the reference dataset (based on melanoma and NSCLC), and the parameters of the reference model were re-estimated. This updated PK analysis, including several approved tumor types, provided model parameter estimates that were consistent with the reference analyses based on melanoma and NSCLC.

[0353] Additionally, PK consistency between individual tumor types and the reference PK model (melanoma and NSCLC) was assessed by overlaying observed concentrations against the 90% prediction intervals of the models. These analyses revealed that observed concentrations fell within the range of predicted concentrations, regardless of weight-based or fixed dosing, with minor differences seen in hematological tumors such as cHL and PMBCL that were not considered clinically meaningful. This indicated that the PK model was able to adequately describe pembrolizumab PK across tumor types and confirmed the similarity of pembrolizumab PK across indications following IV administration.

[0354] PK consistency between monotherapy and combination therapy Chemotherapy is typically metabolized in the liver and typically does not affect the disposition of monoclonal antibodies in the body. As is well documented in the literature, monoclonal antibodies are primarily catabolized by the human reticuloendothelial system. In general, metabolic or transporter pathways are not involved in the disposition of pembrolizumab, so PK interactions between pembrolizumab and small molecules are not pharmacologically predicted.

[0355] Clinical trials administering pembrolizumab with chemotherapy (KEYNOTE-021, KEYNOTE-189: with carboplatin / cisplatin and pemetrexed; KEYNOTE-407: with carboplatin and paclitaxel / nab-paclitaxel; KEYNOTE-048: with carboplatin / cisplatin and 5-FU; and KEYNOTE-426: with axitinib) showed no effect on total pembrolizumab exposure compared with pembrolizumab monotherapy.

[0356] Pembrolizumab - Applicability to HLN Administration In the context of SC administration, bioavailability and absorption are not expected to be affected by tumor type or concomitant treatment (Anselmo AC et al., Nat Rev Drug Discov. 2019;18:19-40), and given that the distribution and elimination phases are the same as IV, PK consistency across tumor types and treatment settings should be maintained. Thus, comparisons of pembrolizumab PK parameters made between SC and IV formulations based on studies conducted in NSCLC in combination with chemotherapy should be equally applicable to draw inferences regarding matched exposure and therefore bridged efficacy and safety between SC and IV for pembrolizumab indications.

Claims

1. 1. A pharmaceutical composition for treating cancer in a human patient comprising an anti-PD-1 antibody, wherein the treatment comprises subcutaneously administering to the patient every six weeks an anti-PD-1 antibody at a dose of about 760 mg to about 790 mg and human hyaluronidase; wherein the anti-PD-1 antibody is (a) a heavy chain comprising a sequence of amino acids set forth in any one of SEQ ID NOs: 10 to 15; and (b) a light chain comprising the sequence of amino acids set forth in SEQ ID NO:5; And wherein the human hyaluronidase consists of the amino acid sequence set forth in SEQ ID NO:

18. Pharmaceutical compositions.

2. 2. The pharmaceutical composition of claim 1, wherein the treatment comprises subcutaneous administration of an anti-PD-1 antibody at a dose of 790 mg and human hyaluronidase every 6 weeks.

3. The pharmaceutical composition of claim 2, wherein the anti-PD-1 antibody and the human hyaluronidase are co-formulated.

4. 2. The pharmaceutical composition of claim 1, wherein the treatment comprises subcutaneous administration of a 790 mg dose of an anti-PD-1 antibody and 9600 units of human hyaluronidase.

5. C of subcutaneous administration of the anti-PD-1 antibody ｔｒｏｕｇｈ C-terminal cytotoxicity profile of a 400 mg dose of an anti-PD-1 antibody administered by intravenous (IV) route every 6 weeks. ｔｒｏｕｇｈ 0.8 to 1.6 SC:IV C ｔｒｏｕｇｈ The pharmaceutical composition of claim 4 having the ratio:

6. C of subcutaneous administration of the anti-PD-1 antibody ｔｒｏｕｇｈ C-terminal cytotoxicity profile of a 400 mg dose of an anti-PD-1 antibody administered by intravenous (IV) route every 6 weeks. ｔｒｏｕｇｈ 1.4 to 1.6 SC:IV C ｔｒｏｕｇｈ The pharmaceutical composition of claim 4 having the ratio:

7. C of subcutaneous administration of the anti-PD-1 antibody ｔｒｏｕｇｈ C-terminal cytotoxicity profile of a 400 mg dose of an anti-PD-1 antibody administered by intravenous (IV) route every 6 weeks. ｔｒｏｕｇｈ At least 1.5 SC:IV C ｔｒｏｕｇｈ The pharmaceutical composition of claim 4 having the ratio:

8. C of subcutaneous administration of the anti-PD-1 antibody ｔｒｏｕｇｈ C-terminal cytotoxicity profile of a 400 mg dose of an anti-PD-1 antibody administered by intravenous (IV) route every 6 weeks. ｔｒｏｕｇｈ 1.2 to 1.6 SC:IV C ｔｒｏｕｇｈ The pharmaceutical composition of claim 4 having the ratio:

9. Said SC:IV C ｔｒｏｕｇｈ 6. The pharmaceutical composition of claim 5, wherein the ratio is at cycle 1 or steady state.

10. Said SC:IV C ｔｒｏｕｇｈ 8. The pharmaceutical composition of claim 7, wherein the ratio is a geometric mean ratio.

11. AUC of subcutaneous administration of the anti-PD-1 antibody （０～６週） is the AUC of a 400 mg dose of an anti-PD-1 antibody administered by intravenous (IV) route every 6 weeks. （０～６週） Cycle 1, SC:IV AUC of about 0.8 to about 1.2 （０～６週） The pharmaceutical composition of claim 4 having the ratio:

12. AUC of subcutaneous administration of the anti-PD-1 antibody （０～６週） is the AUC of a 400 mg dose of an anti-PD-1 antibody administered by intravenous (IV) route every 6 weeks. （０～６週） Cycle 1, SC:IV AUC of about 1.0 to about 1.2 （０～６週） The pharmaceutical composition of claim 4 having the ratio:

13. AUC of subcutaneous administration of the anti-PD-1 antibody （０～６週） is the AUC of a 400 mg dose of an anti-PD-1 antibody administered by intravenous (IV) route every 6 weeks. （０～６週） Cycle 1 had an SC:IV AUC of approximately 1.1 （０～６週） The pharmaceutical composition of claim 4 having the ratio:

14. 14. The pharmaceutical composition of claim 13, wherein the ratio is a geometric mean ratio.

15. C of subcutaneous administration of the anti-PD-1 antibody ｍａｘ C-terminal cytotoxicity profile of a 400 mg dose of an anti-PD-1 antibody administered by intravenous (IV) route every 6 weeks. ｍａｘ The pharmaceutical composition of claim 4, wherein the IL-10 concentration is about 40% lower in cycle 1 than in cycle 2.

16. C of subcutaneous administration of the anti-PD-1 antibody ｍａｘ C-terminal cytotoxicity profile of a 400 mg dose of an anti-PD-1 antibody administered by intravenous (IV) route every 6 weeks. ｍａｘ The pharmaceutical composition of claim 12, wherein the IL-10 level is about 40% lower in cycle 1 than in cycle 2.

17. C of subcutaneous administration of the anti-PD-1 antibody ｍａｘ C-terminal cytotoxicity profile of a 400 mg dose of an anti-PD-1 antibody administered by intravenous (IV) route every 6 weeks. ｍａｘ The pharmaceutical composition according to claim 4, wherein the concentration of α-aminobutyric acid in the pharmaceutical composition is about 20 to 30% lower at steady state than that in the pharmaceutical composition according to claim 4.

18. 1. A pharmaceutical composition for treating cancer in a human patient comprising an anti-PD-1 antibody, wherein the treatment comprises subcutaneously administering to the patient every three weeks an anti-PD-1 antibody at a dose of about 380 mg to about 395 mg and human hyaluronidase; wherein the anti-PD-1 antibody is (a) a heavy chain comprising a sequence of amino acids set forth in any one of SEQ ID NOs: 10 to 15; and (b) a light chain comprising the sequence of amino acids set forth in SEQ ID NO:5; And wherein the human hyaluronidase consists of the amino acid sequence set forth in SEQ ID NO:

18. Pharmaceutical compositions.

19. 19. The pharmaceutical composition of claim 18, wherein the treatment comprises subcutaneous administration of an anti-PD-1 antibody at a dose of 395 mg and human hyaluronidase every three weeks.

20. 20. The pharmaceutical composition of claim 19, wherein the anti-PD-1 antibody and the human hyaluronidase are co-formulated.

21. 21. The pharmaceutical composition of claim 20, wherein the treatment comprises subcutaneous administration of an anti-PD-1 antibody at a dose of 395 mg and 4800 units of human hyaluronidase.

22. The pharmaceutical composition according to any one of claims 1 to 21, wherein the cancer is selected from the group consisting of melanoma, non-small cell lung cancer, head and neck cancer, urothelial carcinoma, breast cancer, gastric cancer, gastroesophageal junction adenocarcinoma, hepatocellular carcinoma, Merkel cell carcinoma, renal cell carcinoma, endometrial cancer, cutaneous squamous cell carcinoma, primary mediastinal large B-cell lymphoma (PMBCL), Hodgkin's lymphoma, mesothelioma, ovarian cancer, small cell lung cancer, esophageal cancer, biliary tract cancer, and cervical cancer.

23. 22. The pharmaceutical composition of any one of claims 1 to 21, wherein the cancer is selected from the group consisting of melanoma, non-small cell lung cancer, squamous cell carcinoma of the head and neck, urothelial carcinoma, classical Hodgkin lymphoma, primary mediastinal large B-cell lymphoma (PMBCL), microsatellite instability high (MSI-H) cancer, MSI-H or mismatch repair deficient colorectal cancer, gastric cancer, gastroesophageal junction adenocarcinoma, esophageal cancer, cervical cancer, hepatocellular carcinoma, Merkel cell carcinoma, renal cell carcinoma, endometrial cancer, cutaneous squamous cell carcinoma, tumor mutation burden high (TMB-H) cancer, and triple negative breast cancer.

24. The pharmaceutical composition according to any one of claims 1 to 21, wherein the cancer is locally advanced or metastatic urothelial carcinoma, locally advanced or metastatic gastric cancer or gastroesophageal junction adenocarcinoma, refractory or recurrent primary mediastinal large B-cell lymphoma (PMBCL), hepatocellular carcinoma, renal cell carcinoma (RCC), recurrent or locally advanced or metastatic Merkel cell carcinoma (MCC).

25. The pharmaceutical composition according to any one of claims 1 to 21, wherein the cancer is microsatellite instability high (MSI-H) colorectal cancer or a solid tumor.

26. The pharmaceutical composition of any one of claims 1 to 21, wherein the patient has a tumor with a high mutational burden comprising ≧10 mutations per megabase (Mut / Mb).

27. The pharmaceutical composition of any one of claims 1 to 21, wherein the cancer is unresectable or metastatic melanoma, or is resected stage IIB, IIC or III melanoma.

28. The pharmaceutical composition according to any one of claims 1 to 21, wherein the cancer is advanced or metastatic non-small cell lung cancer (NSCLC).

29. The pharmaceutical composition of any one of claims 1 to 21, wherein the cancer is resected stage IB, II or IIIA non-small cell lung cancer (NSCLC).

30. 29. The pharmaceutical composition of claim 28, wherein the patient has a tumor with PD-L1 expression as measured by a Tumor Proportion Score (TPS) of 1% or greater or a Tumor Proportion Score (TPS) of 50% or greater.

31. 29. The pharmaceutical composition of claim 28, wherein the patient's tumor does not have an EGFR or ALK genomic abnormality.

32. The pharmaceutical composition of claim 30, wherein the patient's tumor does not have an EGFR or ALK genomic abnormality.

33. 32. The pharmaceutical composition of claim 31, wherein the patient has non-squamous non-small cell lung cancer and further wherein the treatment comprises administering to the patient a therapeutically effective amount of pemetrexed and platinum chemotherapy.

34. The patient has non-squamous non-small cell lung cancer and the pemetrexed is administered at a dose of 500 mg / m every 21 days. ２ and the platinum chemotherapy is administered to the patient by intravenous infusion at a dose of 75 mg / m every 21 days. ２ 34. The pharmaceutical composition of claim 33, wherein the cisplatin is administered to the patient in an amount of

35. 29. The pharmaceutical composition of claim 28, wherein the NSCLC is squamous and the treatment further comprises administration of therapeutically effective amounts of carboplatin and paclitaxel or nab-paclitaxel.

36. Carboplatin was administered by intravenous infusion at an AUC of 5-6 mg / ml / min, and paclitaxel was administered at 200 mg / ml every 21 days. ２ was administered by intravenous infusion at 100 mg / m every 7 days, and nab-paclitaxel was administered at 100 mg / m ２ 36. The pharmaceutical composition of claim 35, wherein the composition is administered by intravenous infusion of

37. 22. The pharmaceutical composition of any one of claims 1 to 21, wherein the cancer is metastatic, unresectable or recurrent head and neck squamous cell carcinoma (HNSCC) and the treatment may further comprise administration of platinum and 5-fluorouracil (5-FU), or the patient's tumor may express PD-L1 with a combined positive score (CPS) >= 1.

38. The pharmaceutical composition according to any one of claims 1 to 21, wherein the cancer is refractory or relapsed classical Hodgkin's lymphoma (cHL).

39. The pharmaceutical composition according to any one of claims 1 to 21, wherein the cancer is refractory or recurrent primary mediastinal large B-cell lymphoma (PMBCL).

40. The pharmaceutical composition according to any one of claims 1 to 21, wherein the cancer is recurrent locally advanced unresectable or metastatic gastric cancer, or HER2-positive gastroesophageal junction adenocarcinoma.

41. 41. The pharmaceutical composition of claim 40, wherein the treatment further comprises administration of trastuzumab, fluoropyrimidine and platinum-containing chemotherapy.

42. The pharmaceutical composition of claim 41, wherein the patient's tumor has a PD-L1 combined positive score (CPS) > 1.

43. The pharmaceutical composition according to any one of claims 1 to 21, wherein the cancer is recurrent locally advanced unresectable or metastatic esophageal cancer, or gastroesophageal junction adenocarcinoma.

44. 44. The pharmaceutical composition of claim 43, wherein the treatment further comprises administration of a fluoropyrimidine-based chemotherapy and a platinum-containing chemotherapy.

45. The pharmaceutical composition of claim 43, wherein the patient's tumor has a PD-L1 combined positive score (CPS) of ≧10.

46. The pharmaceutical composition according to any one of claims 1 to 21, wherein the cancer is recurrent or metastatic cervical cancer.

47. 47. The pharmaceutical composition of claim 46, wherein the treatment further comprises administration of paclitaxel and cisplatin or paclitaxel and carboplatin, with or without bevacizumab.

48. The pharmaceutical composition of claim 46, wherein the patient's tumor expresses a PD-L1 combined positive score (CPS) > 1.

49. The pharmaceutical composition of claim 47, wherein the patient's tumor expresses a PD-L1 combined positive score (CPS) > 1.

50. The pharmaceutical composition according to any one of claims 1 to 21, wherein the cancer is cutaneous squamous cell carcinoma.

51. The pharmaceutical composition according to any one of claims 1 to 21, wherein the cancer is advanced or metastatic renal cell carcinoma (RCC).

52. The pharmaceutical composition of any one of claims 1 to 21, wherein the cancer is renal cell carcinoma (RCC) at moderately high or high risk of recurrence after nephrectomy, or after nephrectomy and resection of metastatic lesions, and the administration of the anti-PD-1 antibody is for adjuvant treatment.

53. The pharmaceutical composition according to any one of claims 1 to 21, wherein the cancer is advanced endometrial cancer.

54. The pharmaceutical composition of any one of claims 1 to 21, wherein the cancer is an advanced endometrial cancer that is not MSI-H or mismatch repair deficient (dMMR).

55. The pharmaceutical composition according to any one of claims 1 to 21, wherein the cancer is recurrent unresectable or metastatic triple-negative breast cancer (TNBC).

56. 56. The pharmaceutical composition of claim 55, wherein the treatment further comprises administration of carboplatin and paclitaxel.

57. 57. The pharmaceutical composition of claim 56, wherein the patient's tumor expresses PD-L1 as measured by a combined positive score (CPS) > 10.

58. 22. The pharmaceutical composition of any one of claims 1 to 21, wherein the patient has high-risk early stage or locally advanced TNBC and the treatment comprises neoadjuvant administration of the anti-PD-1 antibody, carboplatin and paclitaxel, followed by administration of the anti-PD-1 antibody, doxorubicin and cyclophosphamide.

59. 59. The pharmaceutical composition of any one of claims 1-3 and 5-58, wherein about 700 Units to about 50,000 Units of human hyaluronidase are co-formulated with the anti-PD-1 antibody, or antigen-binding fragment thereof.

60. 60. The pharmaceutical composition of any one of claims 1-3 and 5-58, wherein the human hyaluronidase is co-formulated with the anti-PD-1 antibody in a ratio of about 3-36 units:1 mg.

61. 59. The pharmaceutical composition of any one of claims 1-3 and 5-58, wherein the human hyaluronidase is co-formulated with the anti-PD-1 antibody in a ratio of about 6-25 units:1 mg.

62. 59. The pharmaceutical composition of any one of claims 1-3 and 5-58, wherein the human hyaluronidase is co-formulated with the anti-PD-1 antibody in a ratio of about 10-14 units:1 mg.

63. 59. The pharmaceutical composition of any one of claims 1-3 and 5-58, wherein the human hyaluronidase is co-formulated with the anti-PD-1 antibody in a ratio of about 12.15 units:1 mg.

64. The pharmaceutical composition according to any one of claims 1 to 4 and 18 to 21, wherein the anti-PD-1 antibody is a monoclonal antibody comprising a heavy chain consisting of the amino acid sequence shown in SEQ ID NO: 10 and a light chain consisting of the amino acid sequence shown in SEQ ID NO:

5.

65. The pharmaceutical composition according to any one of claims 1 to 4 and 18 to 21, wherein the anti-PD-1 antibody is a monoclonal antibody comprising a heavy chain consisting of the amino acid sequence shown in SEQ ID NO: 11 and a light chain consisting of the amino acid sequence shown in SEQ ID NO: 5.

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