Diagnostic methods and compositions for cancer immunotherapy

Abstract

The present invention provides diagnostic and therapeutic methods and compositions for the treatment of cancer. The compositions and methods described herein can be used, for example, to determine a patient's propensity to benefit from treatment with a PD-L1 axis-binding antagonist and to treat such patients accordingly. Using the compositions and methods of the disclosure, it can be determined that a patient, e.g., a human cancer patient, is likely to benefit from treatment with a PD-L1 axis-binding antagonist if the patient exhibits elevated pre-treatment expression levels of one or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2. Exemplary PD-L1 axis binding antagonists that may be used in combination with the compositions and methods of the disclosure are PD-L1 binding antagonists, such as anti-PD-L1 antibodies and antigen-binding fragments thereof, including atezolizumab, and PD-1 binding antagonists, such as anti-PD-1 antibodies and antigen-binding fragments thereof.

Description

[Technical Field]

[0001] Sequence Listing This application has been submitted electronically in ASCII format and contains a Sequence Listing, which is incorporated herein by reference in its entirety. This ASCII copy, created on November 27, 2019, is named 50474-188WO3_Sequence_Listing_11.27.19_ST25 and is 195,457 bytes in size.

[0002] The present invention relates to diagnostic and therapeutic methods for the treatment of cancer using PD-L1 axis-binding antagonists, and related assays and kits are also provided. [Background technology]

[0003] Cancer remains one of the greatest threats to human health. In the United States, cancer affects approximately 1.3 million new cases each year and is the second leading cause of death after heart disease, accounting for approximately one in four deaths. It is also predicted that cancer may surpass cardiovascular disease as the leading cause of death within five years. Solid tumors are responsible for the majority of these deaths.

[0004] Human studies using immune checkpoint inhibitors have shown promise in harnessing the immune system to suppress and eradicate tumor growth. The programmed death-1 (PD-1) receptor and its ligand, programmed death-ligand 1 (PD-L1), are immune checkpoint proteins involved in suppressing immune system responses during chronic infection, pregnancy, tissue allografts, autoimmune diseases, and cancer. PD-L1 regulates immune responses by binding to the inhibitory receptor PD-1, which is expressed on the surface of T cells, B cells, and monocytes. PD-L1 also negatively regulates T cell function through its interaction with another receptor, B7-1. The formation of PD-L1 / PD-1 and PD-L1 / B7-1 complexes negatively regulates T cell receptor signaling, subsequently downregulating T cell activation and suppressing antitumor immune activity.

[0005] Although there have been significant advances in the medical treatment of certain cancers, the overall 5-year survival rate for all cancers has improved by only about 10% over the past 20 years. Malignant solid tumors in particular tend to metastasize and grow rapidly and uncontrolled, making their timely detection and treatment extremely difficult.

[0006] Despite significant advances in cancer treatment, improved diagnostic methods and cancer therapies remain needed. Summary of the Invention

[0007] The present disclosure provides therapeutic and diagnostic methods and compositions for treating individuals with cancer (e.g., lung cancer (e.g., non-small cell lung cancer (NSCLC)), endometrial cancer, colon adenocarcinoma, renal cell carcinoma, bladder cancer (e.g., urothelial carcinoma (UC)), kidney cancer (e.g., renal cell carcinoma (RCC)), and breast cancer (e.g., triple-negative breast cancer (TNBC))). The disclosure is based, at least in part, on: peripheral The present invention is based on the discovery that individuals with cancers that exhibit clonally expanded T cells in the circulation (e.g., lung cancer (e.g., NSCLC), endometrial cancer, colon cancer, renal cell carcinoma, bladder cancer (e.g., UC), kidney cancer (e.g., RCC), and breast cancer (e.g., TNBC)) are particularly likely to benefit from treatments that include a PD-L1 axis-binding antagonist (e.g., a PD-L1-binding antagonist (e.g., an anti-PD-L1 antibody, e.g., atezolizumab (MPDL3280A)) or a PD-1-binding antagonist (e.g., an anti-PD-1 antibody). Clonally expanded T cells can be identified by sequencing the patient's T cell receptor to identify the clonal expansion and / or by identifying the patient's T cell receptor. peripheralIdentification can be achieved by a variety of methods, including detecting elevated immune score expansion levels in the blood of one or more genes associated with clonal expansion, including CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, PATL2, GPR56, IFITM2, IFITM1, TMSB10, CD247, COX2, COX1, CLIC3, S100A4, and CYBA.

[0008] Using the compositions and methods of the present disclosure, patients peripheral Based on the finding that the patient exhibits a clonally expanded T cell population in the blood, the patient may be identified as likely to respond to treatment comprising a PD-L1 axis-binding antagonist (e.g., a PD-L1 binding antagonist (e.g., an anti-PD-L1 antibody, e.g., atezolizumab (MPDL3280A)) or a PD-1 binding antagonist (e.g., an anti-PD-1 antibody)). peripheralThe determination of the presence of clonally expanded T cells in the blood can be made by sequencing the T cell receptor and / or by assessing the patient's gene expression profile in a blood sample obtained from the patient. In this context, a finding that a patient has elevated immune score expression levels of one or more genes selected from, for example, CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, PATL2, GPR56, IFITM2, IFITM1, TMSB10, CD247, COX2, COX1, CLIC3, S100A4, and CYBA, indicates that the patient has a high risk of developing a leukemia. peripheral This may indicate that patients have clonally expanded T cells in the circulation and that they are more likely to respond to treatment with PD-L1 axis-binding antagonists.

[0009] In a first aspect, the disclosure features a method of generating a chimeric antigen receptor (CAR) T cell, the method including: (a) from individuals with cancer peripheral Obtaining a blood sample; (b) determining (i) the amino acid sequence of one or more complementarity determining regions (CDRs) of a T cell receptor (TCR) expressed by one or more T cells in the sample, and / or (ii) the nucleic acid sequence encoding said amino acid sequence; and (c) modifying the progenitor cells to express a CAR comprising one or more CDRs, thereby generating CAR T cells. Includes:

[0010] In another aspect, the disclosure features a method of generating CAR T cells, the method comprising: peripheralThe method includes modifying progenitor cells to express a CAR that comprises one or more CDRs of a TCR expressed by one or more T cells in the blood sample.

[0011] In some embodiments, the T cells in the sample are clonally expanded T cells.

[0012] In some embodiments, the clonally expanded T cells are CD8+ T cells.

[0013] In some embodiments, the CD8+ T cells are CTLs.

[0014] In some embodiments, the TCR expressed by one or more T cells specifically binds to an antigen expressed by a cancer cell in the individual.

[0015] In some embodiments, clonally expanded T cells comprise at least 0.01% of all T cells in the sample.

[0016] In some embodiments, clonally expanded T cells comprise at least 0.1% of all T cells in the sample.

[0017] In some embodiments, clonally expanded T cells comprise at least 1% of all T cells in the sample.

[0018] In some embodiments, clonally expanded T cells comprise at least 2% of all T cells in the sample.

[0019] In some embodiments, clonally expanded T cells comprise at least 3% of all T cells in the sample.

[0020] In some embodiments, clonally expanded T cells comprise at least 4% of all T cells in the sample.

[0021] In some embodiments, clonally expanded T cells comprise at least 5% of all T cells in the sample.

[0022] In some embodiments, clonally expanded T cells comprise at least 10% of all T cells in the sample.

[0023] In some embodiments, clonally expanded T cells comprise at least 15% of all T cells in the sample.

[0024] In some embodiments, clonally expanded T cells comprise at least 20% of all T cells in the sample.

[0025] In some embodiments, clonally expanded T cells comprise at least 25% of all T cells in the sample.

[0026] In some embodiments, the progenitor cells are T cells.

[0027] In some embodiments, the method comprises administering at least about 1×10 3 Generate CAR T cells.

[0028] In some embodiments, the method comprises administering a 2000 mg / kg / day solution of about 1×10 3 CAR T cells to approximately 1 × 10 12 CAR T cells (e.g., approximately 2 x 10 3 CAR T cells to approximately 9 x 10 11 CAR T cells, approximately 3 x 10 3 CAR T cells to approximately 8 x 10 11 CAR T cells, approximately 4 x 10 3 CAR T cells to approximately 7 x 10 11 CAR T cells, approximately 5 x 10 3 CAR T cells to approximately 6 x 10 11 CAR T cells, approximately 6 x 10 3 CAR T cells to approximately 4 × 10 11 CAR T cells, approximately 7 x 10 3CAR T cells to approximately 3 x 10 11 CAR T cells, approximately 8 x 10 3 CAR T cells to approximately 2 × 10 11 CAR T cells, approximately 9 x 10 3 CAR T cells to approximately 1 × 10 11 CAR T cells, approximately 1 x 10 4 CAR T cells to approximately 9 x 10 10 CAR T cells, approximately 2 x 10 4 CAR T cells to approximately 8 x 10 10 CAR T cells, approximately 3 x 10 4 CAR T cells to approximately 7 x 10 10 CAR T cells, approximately 4 x 10 4 CAR T cells to approximately 6 x 10 10 CAR T cells, approximately 5 x 10 4 CAR T cells to approximately 5 × 10 10 CAR T cells, approximately 6 x 10 4 CAR T cells to approximately 4 × 10 10 CAR T cells, approximately 7 x 10 4 CAR T cells to approximately 3 x 10 10 CAR T cells, approximately 8 x 10 4 CAR T cells to approximately 2 × 10 10 CAR T cells, approximately 9 x 10 4 CAR T cells to approximately 1 × 10 10 CAR T cells, or approximately 1 x 10 5 CAR T cells to approximately 9 x 10 9 CAR T cells, e.g., about 1 x 10 3 CAR T cells, 2 x 10 3 CAR T cells, 3 x 10 3 CAR T cells, 4 x 10 3 CAR T cells, 5 x 10 3 CAR T cells, 6 x 10 3 CAR T cells, 7 x 10 3 CAR T cells, 8 x 10 3 CAR T cells, 9 x 10 3 CAR T cells, 1 x 10 4CAR T cells, 2 x 10 4 CAR T cells, 3 x 10 4 CAR T cells, 4 x 10 4 CAR T cells, 5 x 10 4 CAR T cells, 6 x 10 4 CAR T cells, 7 x 10 4 CAR T cells, 8 x 10 4 CAR T cells, 9 x 10 4 CAR T cells, 1 x 10 5 CAR T cells, 2 x 10 5 CAR T cells, 3 x 10 5 CAR T cells, 4 x 10 5 CAR T cells, 5 x 10 5 CAR T cells, 6 x 10 5 CAR T cells, 7 x 10 5 CAR T cells, 8 x 10 5 CAR T cells, 9 x 10 5 CAR T cells, 1 x 10 6 CAR T cells, 2 x 10 6 CAR T cells, 3 x 10 6 CAR T cells, 4 x 10 6 CAR T cells, 5 x 10 6 CAR T cells, 6 x 10 6 CAR T cells, 7 x 10 6 CAR T cells, 8 x 10 6 CAR T cells, 9 x 10 6 CAR T cells, 1 x 10 7 CAR T cells, 2 x 10 7 CAR T cells, 3 x 10 7 CAR T cells, 4 x 10 7 CAR T cells, 5 x 10 7 CAR T cells, 6 x 10 7 CAR T cells, 7 x 10 7 CAR T cells, 8 x 10 7 CAR T cells, 9 x 10 7 CAR T cells, 1 x 10 8 CAR T cells, 2 x 10 8CAR T cells, 3 x 10 8 CAR T cells, 4 x 10 8 CAR T cells, 5 x 10 8 CAR T cells, 6 x 10 8 CAR T cells, 7 x 10 8 CAR T cells, 8 x 10 8 CAR T cells, 9 x 10 8 CAR T cells, 1 x 10 9 CAR T cells, 2 x 10 9 CAR T cells, 3 x 10 9 CAR T cells, 4 x 10 9 CAR T cells, 5 x 10 9 CAR T cells, 6 x 10 9 CAR T cells, 7 x 10 9 CAR T cells, 8 x 10 9 CAR T cells, 9 x 10 9 CAR T cells, 1 x 10 10 CAR T cells, 2 x 10 10 CAR T cells, 3 x 10 10 CAR T cells, 4 x 10 10 CAR T cells, 5 x 10 10 CAR T cells, 6 x 10 10 CAR T cells, 7 x 10 10 CAR T cells, 8 x 10 10 CAR T cells, 9 x 10 10 CAR T cells, 1 x 10 11 CAR T cells, 2 x 10 11 CAR T cells, 3 x 10 11 CAR T cells, 4 x 10 11 CAR T cells, 5 x 10 11 CAR T cells, 6 x 10 11 CAR T cells, 7 x 10 11 CAR T cells, 8 x 10 11 CAR T cells, 9 x 10 11 CAR T cells or x 10 12 CAR T cells (or more).

[0029] In some embodiments, the method comprises administering a 2000 mg / kg / day solution of about 1×10 4 CAR T cells to approximately 1 × 10 11 CAR T cells (e.g., approximately 1 x 10 4 CAR T cells to approximately 9 x 10 10 CAR T cells, approximately 2 x 10 4 CAR T cells to approximately 8 x 10 10 CAR T cells, approximately 3 x 10 4 CAR T cells to approximately 7 x 10 10 CAR T cells, approximately 4 x 10 4 CAR T cells to approximately 6 x 10 10 CAR T cells, approximately 5 x 10 4 CAR T cells to approximately 5 × 10 10 CAR T cells, approximately 6 x 10 4 CAR T cells to approximately 4 × 10 10 CAR T cells, approximately 7 x 10 4 CAR T cells to approximately 3 x 10 10 CAR T cells, approximately 8 x 10 4 CAR T cells to approximately 2 × 10 10 CAR T cells, approximately 9 x 10 4 CAR T cells to approximately 1 × 10 10 CAR T cells, or approximately 1 x 10 5 CAR T cells to approximately 9 x 10 9 CAR T cells, e.g., approximately 1 x 10 4 CAR T cells, 2 x 10 4 CAR T cells, 3 x 10 4 CAR T cells, 4 x 10 4 CAR T cells, 5 x 10 4 CAR T cells, 6 x 10 4 CAR T cells, 7 x 10 4 CAR T cells, 8 x 10 4 CAR T cells, 9 x 10 4 CAR T cells, 1 x 10 5 CAR T cells, 2 x 10 5 CAR T cells, 3 x 10 5 CAR T cells, 4 x 105 CAR T cells, 5 x 10 5 CAR T cells, 6 x 10 5 CAR T cells, 7 x 10 5 CAR T cells, 8 x 10 5 CAR T cells, 9 x 10 5 CAR T cells, 1 x 10 6 CAR T cells, 2 x 10 6 CAR T cells, 3 x 10 6 CAR T cells, 4 x 10 6 CAR T cells, 5 x 10 6 CAR T cells, 6 x 10 6 CAR T cells, 7 x 10 6 CAR T cells, 8 x 10 6 CAR T cells, 9 x 10 6 CAR T cells, 1 x 10 7 CAR T cells, 2 x 10 7 CAR T cells, 3 x 10 7 CAR T cells, 4 x 10 7 CAR T cells, 5 x 10 7 CAR T cells, 6 x 10 7 CAR T cells, 7 x 10 7 CAR T cells, 8 x 10 7 CAR T cells, 9 x 10 7 CAR T cells, 1 x 10 8 CAR T cells, 2 x 10 8 CAR T cells, 3 x 10 8 CAR T cells, 4 x 10 8 CAR T cells, 5 x 10 8 CAR T cells, 6 x 10 8 CAR T cells, 7 x 10 8 CAR T cells, 8 x 10 8 CAR T cells, 9 x 10 8 CAR T cells, 1 x 10 9 CAR T cells, 2 x 10 9 CAR T cells, 3 x 10 9 CAR T cells, 4 x 10 9 CAR T cells, 5 x 109 CAR T cells, 6 x 10 9 CAR T cells, 7 x 10 9 CAR T cells, 8 x 10 9 CAR T cells, 9 x 10 9 CAR T cells, 1 x 10 10 CAR T cells, 2 x 10 10 CAR T cells, 3 x 10 10 CAR T cells, 4 x 10 10 CAR T cells, 5 x 10 10 CAR T cells, 6 x 10 10 CAR T cells, 7 x 10 10 CAR T cells, 8 x 10 10 CAR T cells, 9 x 10 10 CAR T cells or 1 x 10 11 CAR T cells (or more).

[0030] In some embodiments, the method comprises administering a 2000 mg / kg / day solution of about 1×10 5 CAR T cells to approximately 1 × 10 10 CAR T cells (e.g., approximately 1 x 10 5 CAR T cells to approximately 9 x 10 9 CAR T cells, e.g., approximately 1 x 10 5 CAR T cells, 2 x 10 5 CAR T cells, 3 x 10 5 CAR T cells, 4 x 10 5 CAR T cells, 5 x 10 5 CAR T cells, 6 x 10 5 CAR T cells, 7 x 10 5 CAR T cells, 8 x 10 5 CAR T cells, 9 x 10 5 CAR T cells, 1 x 10 6 CAR T cells, 2 x 10 6 CAR T cells, 3 x 10 6 CAR T cells, 4 x 10 6 CAR T cells, 5 x 10 6 CAR T cells, 6 x 10 6CAR T cells, 7 x 10 6 CAR T cells, 8 x 10 6 CAR T cells, 9 x 10 6 CAR T cells, 1 x 10 7 CAR T cells, 2 x 10 7 CAR T cells, 3 x 10 7 CAR T cells, 4 x 10 7 CAR T cells, 5 x 10 7 CAR T cells, 6 x 10 7 CAR T cells, 7 x 10 7 CAR T cells, 8 x 10 7 CAR T cells, 9 x 10 7 CAR T cells, 1 x 10 8 CAR T cells, 2 x 10 8 CAR T cells, 3 x 10 8 CAR T cells, 4 x 10 8 CAR T cells, 5 x 10 8 CAR T cells, 6 x 10 8 CAR T cells, 7 x 10 8 CAR T cells, 8 x 10 8 CAR T cells, 9 x 10 8 CAR T cells, 1 x 10 9 CAR T cells, 2 x 10 9 CAR T cells, 3 x 10 9 CAR T cells, 4 x 10 9 CAR T cells, 5 x 10 9 CAR T cells, 6 x 10 9 CAR T cells, 7 x 10 9 CAR T cells, 8 x 10 9 CAR T cells, 9 x 10 9 CAR T cells or 1 x 10 10 CAR T cells (or more).

[0031] In some embodiments, the method comprises administering a 2000 mg / kg / day solution of about 1×10 6 CAR T cells to approximately 1 × 10 9 CAR T cells (e.g., approximately 1 x 10 6CAR T cells, 2 x 10 6 CAR T cells, 3 x 10 6 CAR T cells, 4 x 10 6 CAR T cells, 5 x 10 6 CAR T cells, 6 x 10 6 CAR T cells, 7 x 10 6 CAR T cells, 8 x 10 6 CAR T cells, 9 x 10 6 CAR T cells, 1 x 10 7 CAR T cells, 2 x 10 7 CAR T cells, 3 x 10 7 CAR T cells, 4 x 10 7 CAR T cells, 5 x 10 7 CAR T cells, 6 x 10 7 CAR T cells, 7 x 10 7 CAR T cells, 8 x 10 7 CAR T cells, 9 x 10 7 CAR T cells, 1 x 10 8 CAR T cells, 2 x 10 8 CAR T cells, 3 x 10 8 CAR T cells, 4 x 10 8 CAR T cells, 5 x 10 8 CAR T cells, 6 x 10 8 CAR T cells, 7 x 10 8 CAR T cells, 8 x 10 8 CAR T cells, 9 x 10 8 CAR T cells, 1 x 10 9 CAR T cells (or more).

[0032] In another aspect, the disclosure features a CAR T cell produced by the method of the previous aspect of the disclosure, or any embodiment thereof.

[0033] In another aspect, the disclosure features a pharmaceutical composition that includes (i) one or more CAR T cells, or progeny thereof, generated by the method of any of the above aspects or embodiments of the disclosure, and (ii) one or more carriers, diluents, and / or excipients.

[0034] In another aspect, the disclosure features a method of treating cancer in an individual in need thereof, comprising administering to the individual a CAR T cell of the preceding aspect, or a progeny thereof, or a pharmaceutical composition described above.

[0035] In some embodiments, the cancer is lung cancer, kidney cancer, bladder cancer, breast cancer, colorectal cancer, ovarian cancer, pancreatic cancer, gastric cancer, esophageal cancer, mesothelioma, melanoma, head and neck cancer, thyroid cancer, sarcoma, prostate cancer, glioblastoma, cervical cancer, thymic carcinoma, leukemia, lymphoma, myeloma, mycosis fungoides, Merkel cell carcinoma, or a hematological malignancy.

[0036] In some embodiments, the cancer is lung cancer, kidney cancer, bladder cancer, or breast cancer.

[0037] In some embodiments, the lung cancer is non-small cell lung cancer (NSCLC).

[0038] In some embodiments, the kidney cancer is renal cell carcinoma (RCC).

[0039] In some embodiments, the bladder cancer is urothelial carcinoma (UC).

[0040] In some embodiments, the breast cancer is triple negative breast cancer (TNBC).

[0041] In some embodiments, the patient is further administered a PD-L1 axis-binding antagonist.

[0042] In some embodiments, the PD-L1 axis binding antagonist is selected from the group consisting of a PD-L1 binding antagonist, a PD-1 binding antagonist, and a PD-L2 binding antagonist.

[0043] In some embodiments, the PD-L1 axis binding antagonist is a PD-L1 binding antagonist.

[0044] In some embodiments, the PD-L1 binding antagonist inhibits the binding of PD-L1 to one or more of its ligand binding partners.

[0045] In some embodiments, the PD-L1 binding antagonist inhibits the binding of PD-L1 to PD-1.

[0046] In some embodiments, the PD-L1 binding antagonist inhibits the binding of PD-L1 to B7-1.

[0047] In some embodiments, the PD-L1 binding antagonist inhibits the binding of PD-L1 to both PD-1 and B7-1.

[0048] In some embodiments, the PD-L1 binding antagonist is an antibody or antigen-binding fragment thereof.

[0049] In some embodiments, the antibody is selected from the group consisting of atezolizumab (MPDL3280A), YW243.55.S70, MDX-1105, MEDI4736 (durvalumab), and MSB0010718C (avelumab).

[0050] In some embodiments, the antibody comprises a heavy chain comprising the HVR-H1 sequence of SEQ ID NO: 41, the HVR-H2 sequence of SEQ ID NO: 42, and the HVR-H3 sequence of SEQ ID NO: 43, and a light chain comprising the HVR-L1 sequence of SEQ ID NO: 44, the HVR-L2 sequence of SEQ ID NO: 45, and the HVR-L3 sequence of SEQ ID NO: 46.

[0051] In some embodiments, the antibody comprises a heavy chain variable region comprising the amino acid sequence of SEQ ID NO:47 and a light chain variable region comprising the amino acid sequence of SEQ ID NO:48.

[0052] In some embodiments, the PD-L1 axis binding antagonist is a PD-1 binding antagonist.

[0053] In some embodiments, the PD-1 binding antagonist inhibits the binding of PD-1 to one or more of its ligand binding partners.

[0054] In some embodiments, the PD-1 binding antagonist inhibits the binding of PD-1 to PD-L1.

[0055] In some embodiments, the PD-1 binding antagonist inhibits the binding of PD-1 to PD-L2.

[0056] In some embodiments, the PD-1 binding antagonist inhibits the binding of PD-1 to both PD-L1 and PD-L2.

[0057] In some embodiments, the PD-1 binding antagonist is an antibody or antigen-binding fragment thereof.

[0058] In some embodiments, the antibody is selected from the group consisting of MDX-1106 (nivolumab), MK-3475 (pembrolizumab), MEDI-0680 (AMP-514), PDR001, REGN2810, and BGB-108.

[0059] In some embodiments, the PD-1 binding antagonist is an Fc fusion protein.

[0060] In some embodiments, the Fc fusion protein is AMP-224.

[0061] In another aspect, the disclosure features a method for identifying an individual having cancer who may benefit from treatment comprising a PD-L1 axis-binding antagonist, the method comprising: peripheral determining whether the individual has clonally expanded T cells in a blood sample, wherein clonally expanded T cells in the sample identify the individual as one who may benefit from treatment comprising a PD-L1 axis-binding antagonist.

[0062] In another aspect, the disclosure features a method of selecting a treatment for an individual having cancer, the method including ... peripheral determining whether the individual has clonally expanded T cells in a blood sample, wherein clonally expanded T cells in the sample identify the individual as one who may benefit from treatment comprising a PD-L1 axis-binding antagonist.

[0063] In some embodiments, the sample comprises clonally expanded T cells and the method further comprises administering to the individual an effective amount of a PD-L1 axis-binding antagonist.

[0064] In another aspect, the disclosure features a method of treating an individual having cancer, the method including: (a) Individuals derived from individuals peripheral determining that the blood sample has clonally expanded T cells; and (b) administering to the individual an effective amount of a PD-L1 axis-binding antagonist. Includes:

[0065] In another aspect, the present disclosure provides a method for the detection of a leukemia virus from an individual. peripheral The present invention features a method for treating cancer in an individual determined to have clonally expanded T cells in a blood sample, the method comprising administering to the individual an effective amount of a PD-L1 axis-binding antagonist.

[0066] In another aspect, the disclosure features a method of inducing an expansion of a population of T cells in an individual having cancer, the method comprising: (a) Individuals derived from individuals peripheral determining that the blood sample has clonally expanded T cells; and (b) administering to the individual an effective amount of a PD-L1 axis-binding antagonist. Includes:

[0067] In another aspect, the present disclosure provides a method for the detection of a leukemia virus from an individual. peripheralThe invention features a method for inducing an expansion of a population of T cells in an individual with cancer who has been determined to have clonally expanded T cells in a blood sample, the method comprising administering to the individual an effective amount of a PD-L1 axis-binding antagonist.

[0068] In some embodiments, the clonally expanded T cells are CD8+ T cells.

[0069] In some embodiments, the CD8+ T cells are CTLs.

[0070] In some embodiments, the clonally expanded T cells express a TCR comprising an alpha and a beta chain, each comprising CDRs CDR-1, CDR-2, and CDR-3, and the nucleic acid sequences encoding corresponding CDR-1, CDR-2, or CDR-3 of the alpha chain and / or CDR-1, CDR-2, or CDR-3 of the beta chain are at least 85% identical to each other.

[0071] In some embodiments, the nucleic acid sequences encoding the CDR1 of corresponding alpha chains are at least 85% identical to each other.

[0072] In some embodiments, the nucleic acid sequences encoding the CDR1 of corresponding alpha chains are at least 90% identical to each other.

[0073] In some embodiments, the nucleic acid sequences encoding the CDR1 of corresponding alpha chains are at least 95% identical to each other.

[0074] In some embodiments, the nucleic acid sequences encoding the CDR1 of corresponding alpha chains are 100% identical to each other.

[0075] In some embodiments, the nucleic acid sequences encoding the CDR2 of corresponding alpha chains are at least 85% identical to each other.

[0076] In some embodiments, the nucleic acid sequences encoding the CDR2 of corresponding alpha chains are at least 90% identical to each other.

[0077] In some embodiments, the nucleic acid sequences encoding the CDR2 of corresponding alpha chains are at least 95% identical to each other.

[0078] In some embodiments, the nucleic acid sequences encoding the CDR2 of corresponding alpha chains are 100% identical to each other.

[0079] In some embodiments, the nucleic acid sequences encoding the CDR3 of corresponding alpha chains are at least 85% identical to each other.

[0080] In some embodiments, the nucleic acid sequences encoding the CDR3 of corresponding alpha chains are at least 90% identical to each other.

[0081] In some embodiments, the nucleic acid sequences encoding the CDR3 of corresponding alpha chains are at least 95% identical to each other.

[0082] In some embodiments, the nucleic acid sequences encoding the CDR3 of corresponding alpha chains are 100% identical to each other.

[0083] In some embodiments, the nucleic acid sequences encoding the CDR1 of corresponding beta chains are at least 85% identical to each other.

[0084] In some embodiments, the nucleic acid sequences encoding the CDR1 of corresponding beta chains are at least 90% identical to each other.

[0085] In some embodiments, the nucleic acid sequences encoding the CDR1 of corresponding beta chains are at least 95% identical to each other.

[0086] In some embodiments, the nucleic acid sequences encoding the CDR1 of corresponding beta chains are 100% identical to each other.

[0087] In some embodiments, the nucleic acid sequences encoding the CDR2 of corresponding beta chains are at least 85% identical to each other.

[0088] In some embodiments, the nucleic acid sequences encoding the CDR2 of corresponding beta chains are at least 90% identical to each other.

[0089] In some embodiments, the nucleic acid sequences encoding the CDR2 of corresponding beta chains are at least 95% identical to each other.

[0090] In some embodiments, the nucleic acid sequences encoding the CDR2 of corresponding beta chains are 100% identical to each other.

[0091] In some embodiments, the nucleic acid sequences encoding the CDR3 of corresponding beta chains are at least 85% identical to each other.

[0092] In some embodiments, the nucleic acid sequences encoding the CDR3 of corresponding beta chains are at least 90% identical to each other.

[0093] In some embodiments, the nucleic acid sequences encoding the CDR3 of corresponding beta chains are at least 95% identical to each other.

[0094] In some embodiments, the nucleic acid sequences encoding the CDR3 of corresponding beta chains are 100% identical to each other.

[0095] In some embodiments, the clonally expanded T cells express a TCR comprising an alpha chain and / or a beta chain having nucleic acid sequences that are at least 85% identical to each other.

[0096] In some embodiments, the clonally expanded T cells express TCRs that comprise alpha chains with nucleic acid sequences that are at least 85% identical to each other.

[0097] In some embodiments, the clonally expanded T cells express TCRs that comprise alpha chains with nucleic acid sequences that are at least 90% identical to each other.

[0098] In some embodiments, the clonally expanded T cells express TCRs that comprise alpha chains with nucleic acid sequences that are at least 95% identical to each other.

[0099] In some embodiments, the clonally expanded T cells express TCRs that comprise alpha chains with nucleic acid sequences that are 100% identical to each other.

[0100] In some embodiments, the clonally expanded T cells express TCRs that comprise beta chains with nucleic acid sequences that are at least 85% identical to each other.

[0101] In some embodiments, the clonally expanded T cells express TCRs that comprise beta chains with nucleic acid sequences that are at least 90% identical to each other.

[0102] In some embodiments, the clonally expanded T cells express TCRs that comprise beta chains with nucleic acid sequences that are at least 95% identical to each other.

[0103] In some embodiments, the clonally expanded T cells express TCRs that comprise beta chains with nucleic acid sequences that are 100% identical to each other.

[0104] In some embodiments, clonally expanded T cells comprise at least 0.01% of all T cells in the sample.

[0105] In some embodiments, clonally expanded T cells comprise at least 0.1% of all T cells in the sample.

[0106] In some embodiments, clonally expanded T cells comprise at least 1% of all T cells in the sample.

[0107] In some embodiments, clonally expanded T cells comprise at least 2% of all T cells in the sample.

[0108] In some embodiments, clonally expanded T cells comprise at least 3% of all T cells in the sample.

[0109] In some embodiments, clonally expanded T cells comprise at least 4% of all T cells in the sample.

[0110] In some embodiments, clonally expanded T cells comprise at least 5% of all T cells in the sample.

[0111] In some embodiments, clonally expanded T cells comprise at least 10% of all T cells in the sample.

[0112] In some embodiments, clonally expanded T cells comprise at least 15% of all T cells in the sample.

[0113] In some embodiments, clonally expanded T cells comprise at least 20% of all T cells in the sample.

[0114] In some embodiments, clonally expanded T cells comprise at least 25% of all T cells in the sample.

[0115] In another aspect, the disclosure features a method for identifying an individual having cancer who may benefit from treatment comprising a PD-L1 axis-binding antagonist, the method comprising sequencing a T cell receptor or detecting a PD-L1 axis-binding antagonist from the individual. peripheraldetermining the expression level of one or more of the genes NKG7, GNLY, PRF1, GZMH, GPR56, IFITM2, IFITM1, GZMB, TMSB10, PLAC8, LITAF, CD247, CST7, COX2, KLRD1, CLIC3, S100A4, CYBA, and COX1 in the blood sample, wherein an Immunoscore expression level of the one or more genes that exceeds a reference Immunoscore expression level of the one or more genes identifies the individual as one who may benefit from treatment comprising a PD-L1 axis binding antagonist.

[0116] In another aspect, the disclosure features a method of selecting a treatment for an individual with cancer, the method including sequencing a T cell receptor or detecting a T cell receptor from the individual. peripheral determining the expression level of one or more of the genes NKG7, GNLY, PRF1, GZMH, GPR56, IFITM2, IFITM1, GZMB, TMSB10, PLAC8, LITAF, CD247, CST7, COX2, KLRD1, CLIC3, S100A4, CYBA, and COX1 in a sample of blood, wherein an Immunoscore expression level of one or more genes in the sample that exceeds a reference Immunoscore expression level of the one or more genes identifies the individual as one who may benefit from treatment comprising a PD-L1 axis-binding antagonist.

[0117] In some embodiments, the immunoscore expression level of one or more genes in the sample is greater than a reference immunoscore expression level, and the method further comprises administering to the individual an effective amount of a PD-L1 axis binding antagonist.

[0118] In some embodiments, the reference immunoscore expression level is the immunoscore expression level of one or more genes in a reference population.

[0119] In another aspect, the disclosure features a method of treating an individual having cancer, the method including: (a) Individual peripheraldetermining the expression level of one or more of the genes NKG7, GNLY, PRF1, GZMH, GPR56, IFITM2, IFITM1, GZMB, TMSB10, PLAC8, LITAF, CD247, CST7, COX2, KLRD1, CLIC3, S100A4, CYBA, and COX1 in the sample of blood, wherein the immunoscore expression level of the one or more genes in the sample is determined to be greater than a reference immunoscore expression level of the one or more genes; and (b) administering to the individual an effective amount of a PD-L1 axis-binding antagonist. Includes:

[0120] In another aspect, the present disclosure provides a method for the detection of a leukemia virus from an individual. peripheral The present invention features a method of treating cancer in an individual in which an immunoscore expression level of one or more of the genes NKG7, GNLY, PRF1, GZMH, GPR56, IFITM2, IFITM1, GZMB, TMSB10, PLAC8, LITAF, CD247, CST7, COX2, KLRD1, CLIC3, S100A4, CYBA, and COX1 in a blood sample is determined to be greater than a reference immunoscore expression level of the one or more genes, the method comprising administering to the individual an effective amount of a PD-L1 axis binding antagonist.

[0121] In another aspect, the disclosure features a method of inducing an expansion of a population of T cells in an individual having cancer, the method comprising: (a) Individual peripheral determining the expression level of one or more of the genes NKG7, GNLY, PRF1, GZMH, GPR56, IFITM2, IFITM1, GZMB, TMSB10, PLAC8, LITAF, CD247, CST7, COX2, KLRD1, CLIC3, S100A4, CYBA, and COX1 in the sample of blood, wherein the immunoscore expression level of the one or more genes in the sample is determined to be greater than a reference immunoscore expression level of the one or more genes; and (b) administering to the individual an effective amount of a PD-L1 axis-binding antagonist. Includes:

[0122] In another aspect, the present disclosure provides a method for the detection of a leukemia virus from an individual. peripheral Featured is a method of inducing expansion of a population of T cells in an individual having cancer in which an immunoscore expression level of one or more of the genes NKG7, GNLY, PRF1, GZMH, GPR56, IFITM2, IFITM1, GZMB, TMSB10, PLAC8, LITAF, CD247, CST7, COX2, KLRD1, CLIC3, S100A4, CYBA, and COX1 in a blood sample is determined to be above a reference immunoscore expression level of said one or more genes, the method comprising administering to the individual an effective amount of a PD-L1 axis-binding antagonist.

[0123] In some embodiments, the T cells are CD8+ T cells.

[0124] In some embodiments, the CD8+ cells are cytotoxic T cells (CTL) or effector memory T cells (Tem).

[0125] In some embodiments, the reference immunoscore expression level is the immunoscore expression level of one or more genes in a reference population.

[0126] In some embodiments, the reference population is a population of individuals with cancer.

[0127] In some embodiments, the population of individuals comprises a first subset of individuals treated with a PD-L1 axis-binding antagonist, and a second subset of individuals treated with a therapy that does not include a PD-L1 axis-binding antagonist.

[0128] In some embodiments, the reference immunoscore expression level significantly separates each of the first and second subsets of individuals based on a significant difference between the individual's responsiveness to treatment with a PD-L1 axis-binding antagonist and the individual's responsiveness to treatment with a therapy that does not include a PD-L1 axis-binding antagonist above the reference immunoscore expression level, wherein the individual's responsiveness to treatment with the PD-L1 axis-binding antagonist is significantly improved compared to the individual's responsiveness to treatment with a therapy that does not include a PD-L1 axis-binding antagonist.

[0129] In some embodiments, the reference immunoscore expression level significantly separates each of the first and second subsets of individuals based on a significant difference between the individual's responsiveness to treatment with a PD-L1 axis-binding antagonist and the individual's responsiveness to treatment with a therapy that does not include a PD-L1 axis-binding antagonist that is below the reference immunoscore expression level, wherein the individual's responsiveness to treatment with a therapy that does not include a PD-L1 axis-binding antagonist is significantly improved compared to the individual's responsiveness to treatment with a PD-L1 axis-binding antagonist.

[0130] In some embodiments, the PD-L1 axis-binding antagonist-free therapy comprises an anti-tumor agent, a chemotherapeutic agent, a growth inhibitory agent, an anti-angiogenic agent, radiation therapy, a cytotoxic agent, or a combination thereof.

[0131] In some embodiments, the PD-L1 axis-binding antagonist-free therapy comprises a chemotherapeutic agent.

[0132] In some embodiments, the chemotherapeutic agent is docetaxel.

[0133] In some embodiments, responsiveness to treatment comprises an increase in progression-free survival (PFS).

[0134] In some embodiments, responsiveness to treatment comprises an increase in overall survival (OS).

[0135] In some embodiments, the reference immune score expression level is the median expression level of each of the one or more genes in the reference population.

[0136] In some embodiments, the immunoscore expression level of the one or more genes in the reference population is the median of the normalized expression levels of each of the one or more genes in the reference population.

[0137] In some embodiments, the reference immune score expression level is the average of the expression levels of each of the one or more genes in the reference population.

[0138] In some embodiments, the mean expression level of each of the one or more genes in the reference population is the mean normalized expression level of each of the one or more genes in the reference population.

[0139] In some embodiments, the normalized expression level of each of the one or more genes is the expression level of each of the one or more genes normalized to a reference gene.

[0140] In some embodiments, the reference gene is CD8A.

[0141] In some embodiments, the reference immune score expression level is a pre-assigned expression level of one or more genes.

[0142] In some embodiments, the benefit of the treatment comprising the PD-L1 axis-binding antagonist comprises an increase in OS.

[0143] In some embodiments, the benefit of treatment comprising a PD-L1 axis-binding antagonist comprises prolongation of PFS.

[0144] In some embodiments, the benefit of treatment comprising a PD-L1 axis-binding antagonist comprises prolongation of OS and PFS.

[0145] In some embodiments, the genes include two or more of NKG7, GNLY, PRF1, GZMH, GPR56, IFITM2, IFITM1, GZMB, TMSB10, PLAC8, LITAF, CD247, CST7, COX2, KLRD1, CLIC3, S100A4, CYBA, and COX1.

[0146] In some embodiments, the genes include three or more of NKG7, GNLY, PRF1, GZMH, GPR56, IFITM2, IFITM1, GZMB, TMSB10, PLAC8, LITAF, CD247, CST7, COX2, KLRD1, CLIC3, S100A4, CYBA, and COX1.

[0147] In some embodiments, the genes include four or more of NKG7, GNLY, PRF1, GZMH, GPR56, IFITM2, IFITM1, GZMB, TMSB10, PLAC8, LITAF, CD247, CST7, COX2, KLRD1, CLIC3, S100A4, CYBA, and COX1.

[0148] In some embodiments, the expression level is a nucleic acid expression level.

[0149] In some embodiments, the nucleic acid expression level is an mRNA expression level.

[0150] In some embodiments, mRNA expression levels are determined by RNA-seq, RT-qPCR, qPCR, multiplex qPCR or RT-qPCR, microarray analysis, SAGE, MassARRAY techniques, FISH, or a combination thereof.

[0151] In some embodiments, mRNA expression levels are determined using RNA-seq.

[0152] In some embodiments, mRNA expression levels are determined using RT-qPCR.

[0153] In some embodiments, the cancer is lung cancer, kidney cancer, bladder cancer, breast cancer, colorectal cancer, ovarian cancer, pancreatic cancer, gastric cancer, esophageal cancer, mesothelioma, melanoma, head and neck cancer, thyroid cancer, sarcoma, prostate cancer, glioblastoma, cervical cancer, thymic carcinoma, leukemia, lymphoma, myeloma, mycosis fungoides, Merkel cell carcinoma, or a hematological malignancy.

[0154] In some embodiments, the cancer is lung cancer, kidney cancer, bladder cancer, or breast cancer.

[0155] In some embodiments, the lung cancer is non-small cell lung cancer (NSCLC).

[0156] In some embodiments, the kidney cancer is renal cell carcinoma (RCC).

[0157] In some embodiments, the bladder cancer is urothelial carcinoma (UC).

[0158] In some embodiments, the breast cancer is triple negative breast cancer (TNBC).

[0159] In some embodiments, the PD-L1 axis binding antagonist is selected from the group consisting of a PD-L1 binding antagonist, a PD-1 binding antagonist, and a PD-L2 binding antagonist.

[0160] In some embodiments, the PD-L1 axis binding antagonist is a PD-L1 binding antagonist.

[0161] In some embodiments, the PD-L1 binding antagonist inhibits the binding of PD-L1 to one or more of its ligand binding partners.

[0162] In some embodiments, the PD-L1 binding antagonist inhibits the binding of PD-L1 to PD-1.

[0163] In some embodiments, the PD-L1 binding antagonist inhibits the binding of PD-L1 to B7-1.

[0164] In some embodiments, the PD-L1 binding antagonist inhibits the binding of PD-L1 to both PD-1 and B7-1.

[0165] In some embodiments, the PD-L1 binding antagonist is an antibody or antigen-binding fragment thereof.

[0166] In some embodiments, the antibody is selected from the group consisting of atezolizumab (MPDL3280A), YW243.55.S70, MDX-1105, MEDI4736 (durvalumab), and MSB0010718C (avelumab).

[0167] In some embodiments, the antibody comprises a heavy chain comprising the HVR-H1 sequence of SEQ ID NO: 41, the HVR-H2 sequence of SEQ ID NO: 42, and the HVR-H3 sequence of SEQ ID NO: 43, and a light chain comprising the HVR-L1 sequence of SEQ ID NO: 44, the HVR-L2 sequence of SEQ ID NO: 45, and the HVR-L3 sequence of SEQ ID NO: 46.

[0168] In some embodiments, the antibody comprises a heavy chain variable region comprising the amino acid sequence of SEQ ID NO:47 and a light chain variable region comprising the amino acid sequence of SEQ ID NO:48.

[0169] In some embodiments, the PD-L1 axis binding antagonist is a PD-1 binding antagonist.

[0170] In some embodiments, the PD-1 binding antagonist inhibits the binding of PD-1 to one or more of its ligand binding partners.

[0171] In some embodiments, the PD-1 binding antagonist inhibits the binding of PD-1 to PD-L1.

[0172] In some embodiments, the PD-1 binding antagonist inhibits the binding of PD-1 to PD-L2.

[0173] In some embodiments, the PD-1 binding antagonist inhibits the binding of PD-1 to both PD-L1 and PD-L2.

[0174] In some embodiments, the PD-1 binding antagonist is an antibody or antigen-binding fragment thereof.

[0175] In some embodiments, the antibody is selected from the group consisting of MDX-1106 (nivolumab), MK-3475 (pembrolizumab), MEDI-0680 (AMP-514), PDR001, REGN2810, and BGB-108.

[0176] In some embodiments, the PD-1 binding antagonist is an Fc fusion protein.

[0177] In some embodiments, the Fc fusion protein is AMP-224.

[0178] In some embodiments, the individual has not been previously treated for cancer.

[0179] In some embodiments, the individual has not previously been administered a PD-L1 axis-binding antagonist.

[0180] In some embodiments, the individual has previously been treated for cancer by administering to the individual a platinum-containing chemotherapeutic agent, and the individual did not respond to the chemotherapeutic agent.

[0181] In some embodiments, the treatment comprising the PD-L1 axis-binding antagonist is a monotherapy.

[0182] In some embodiments, the methods further comprise administering to the individual an effective amount of one or more additional therapeutic agents.

[0183] In some embodiments, the one or more additional therapeutic agents comprise an anti-tumor agent, a chemotherapeutic agent, a growth inhibitory agent, an anti-angiogenic agent, radiation therapy, a cytotoxic agent, an immunomodulatory agent, or a combination thereof.

[0184] In some embodiments, the one or more additional therapeutic agents comprises an immunomodulatory agent.

[0185] In some embodiments, the immunomodulatory agent is an antibody or antigen-binding fragment thereof.

[0186] In some embodiments, the individual is a human.

[0187] In another aspect, the disclosure features a method of identifying an individual (e.g., a human) with cancer who may benefit from a treatment comprising a PD-L1 axis-binding antagonist. The method can include sequencing T cell receptors to measure the extent of clonal expansion, or determining the expression levels of one or more of the genes CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2 in a sample from the individual (e.g., determining the expression levels of one or more of the genes CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2). , 31, 32, or all 33 of: 4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2), where an Immunoscore expression level of the one or more genes that exceeds a reference Immunoscore expression level of the one or more genes identifies the individual as one that may benefit from treatment comprising a PD-L1 axis-binding antagonist.

[0188] In another aspect, the disclosure features a method of selecting a treatment for an individual (e.g., a human) with cancer. The method includes determining, in a sample from the individual, the expression level of one or more of the genes CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2 (e.g., determining the expression level of one or more of the genes CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2). ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2), where an Immunoscore expression level of the one or more genes in the sample that is greater than a reference Immunoscore expression level of the one or more genes identifies the individual as one who may benefit from treatment comprising a PD-L1 axis-binding antagonist.

[0189] In some embodiments of any of the above aspects, the immunoscore expression level of the one or more genes in the sample is greater than a reference immunoscore expression level. In some embodiments, the method further comprises administering to the individual an effective amount of a PD-L1 axis-binding antagonist.

[0190] In some embodiments, the reference immunoscore expression level is the immunoscore expression level of one or more genes in a reference population.

[0191] In an additional aspect, the disclosure features a method of treating an individual (e.g., a human) with cancer. The method includes: (A) determining, in a sample from the individual, the expression level of one or more of the genes CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2 (e.g., determining the expression level of one or more of the genes CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2). - determining the expression levels of one or more genes in the sample that are greater than a reference immunoscore expression level; and (b) administering to the individual an effective amount of a PD-L1 axis-binding antagonist.

[0192] In another aspect, the disclosure provides a method for detecting an immunoscore expression level of one or more of the genes CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2 in a sample from the individual, the immunoscore expression level being greater than a reference immunoscore expression level of the one or more genes. Featured are methods of treating cancer in an individual (e.g., a human) determined to have immunoscore expression levels of one, two, three, four, five, six, seven, eight, nine, ten, eleven, twelve, thirteen, thirteen, fourteen, fifteen, six, seven, eight, nine, ten, eleven, twelve, thirteen, thirteen, twenty, twenty-one, twenty-two, twenty-three, twenty-four, twenty-five, twenty-six, twenty-seven, twenty-eight, twenty-nine, twenty-nine, twenty-one, twenty-two, twenty-three, twenty-five, twenty-six, twenty-seven, twenty-eight, twenty-nine, twenty-nine, twenty-one ... The method may include administering to the individual an effective amount of a PD-L1 axis binding antagonist.

[0193] In an additional aspect, the disclosure features a method of inducing an expansion of a population of T cells in an individual (e.g., a human) with cancer. The method includes: (A) determining, in a sample from the individual, the expression level of one or more of the genes CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2 (e.g., determining the expression level of one or more of the genes CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2). - determining the expression levels of one or more genes in the sample that are greater than a reference immunoscore expression level; and (b) administering to the individual an effective amount of a PD-L1 axis-binding antagonist.

[0194] In a further aspect, the disclosure features a method of inducing an expansion of a population of T cells in an individual (e.g., a human) with cancer. The method may comprise administering to the individual an effective amount of a PD-L1 axis binding antagonist, wherein prior to treatment, the expression level of one or more of the genes CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2 has been determined in a sample from the individual, and the immunoscore expression level of the one or more genes in the sample has been determined to be greater than a reference immunoscore expression level of the one or more genes. For example, prior to treatment, a sample from an individual may be enriched for the genes CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRS The expression levels of one, two, three, four, five, six, seven, eight, nine, ten, eleven, twelve, thirty, thirty-one, thirty-two, or all thirty-three of S23, GNLY, PRF1, and PATL2 may be determined, and the immunoscore expression level of one or more genes in the sample may be determined to be greater than a reference immunoscore expression level of one or more genes. In some embodiments, the T cell is a CD8+ T cell, e.g., a cytotoxic T cell (CTL, also referred to herein as "Tcyt") or an effector memory T cell (Tem).

[0195] In some embodiments of the above aspects, the reference immunoscore expression level is the immunoscore expression level of one or more genes in a reference population. In some embodiments, the reference population is a population of individuals with cancer. In some embodiments, the population of individuals includes a first subset of individuals treated with a PD-L1 axis-binding antagonist and a second subset of individuals treated with a therapy that does not include a PD-L1 axis-binding antagonist. The reference immunoscore expression level significantly separates each of the first and second subsets of individuals, for example, based on a significant difference between the individual's responsiveness to treatment with the PD-L1 axis-binding antagonist and the individual's responsiveness to treatment with the therapy that does not include a PD-L1 axis-binding antagonist that is greater than the reference immunoscore expression level. In such cases, the individual's responsiveness to treatment with the PD-L1 axis-binding antagonist may be significantly improved compared to the individual's responsiveness to treatment with the therapy that does not include a PD-L1 axis-binding antagonist.

[0196] In some embodiments, the reference immunoscore expression level significantly separates each of the first and second subsets of individuals based on a significant difference between the individual's responsiveness to treatment with the PD-L1 axis-binding antagonist and the individual's responsiveness to treatment with a therapy that does not include a PD-L1 axis-binding antagonist that is below the reference immunoscore expression level. In such cases, the individual's responsiveness to treatment with the therapy that does not include a PD-L1 axis-binding antagonist is significantly improved compared to the individual's responsiveness to treatment with the PD-L1 axis-binding antagonist.

[0197] In some embodiments, the PD-L1 axis binding antagonist-free therapy comprises an anti-tumor agent, a chemotherapeutic agent, a growth inhibitory agent, an anti-angiogenic agent, radiation therapy, a cytotoxic agent, or a combination thereof. For example, the PD-L1 axis binding antagonist-free therapy comprises a chemotherapeutic agent, such as docetaxel.

[0198] In some embodiments, responsiveness to treatment comprises an increase in progression-free survival (PFS). In some embodiments, responsiveness to treatment comprises an increase in overall survival (OS).

[0199] In some embodiments, the reference immunoscore expression level is the median of the expression level of one or more genes in the reference population.For example, the immunoscore expression level of one or more genes in the reference population can be the median of the normalized expression level of one or more genes in the reference population.

[0200] In some embodiments, reference immune score expression level is the average of each expression level of one or more genes in reference population.For example, the immune score expression level of one or more genes in reference population can be the average of each normalized expression level of one or more genes in reference population.In some embodiments, each normalized expression level of one or more genes is the expression level of one or more genes normalized to reference gene, for example, CD8A.

[0201] In some embodiments, the reference immune score expression level is a pre-assigned expression level of one or more genes.

[0202] In some embodiments, the benefit of treatment comprising a PD-L1 axis binding antagonist comprises an increase in OS. In some embodiments, the benefit of treatment comprising a PD-L1 axis binding antagonist comprises an increase in PFS. In some embodiments, the benefit of treatment comprising a PD-L1 axis binding antagonist comprises an increase in OS and PFS.

[0203] In some embodiments, the one or more genes include more than one of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2. For example, in some embodiments, the one or more genes include two or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2. In some embodiments, the one or more genes include three or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2. In some embodiments, the one or more genes include four or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2.In some embodiments, the one or more genes include five or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2. In some embodiments, the one or more genes include six or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2. In some embodiments, the one or more genes include seven or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2. In some embodiments, the one or more genes include eight or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2.In some embodiments, the one or more genes include nine or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2. In some embodiments, the one or more genes include ten or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2. In some embodiments, the one or more genes include 11 or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2. In some embodiments, the one or more genes include 12 or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2.In some embodiments, the one or more genes include 13 or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2. In some embodiments, the one or more genes include 14 or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2. In some embodiments, the one or more genes include 15 or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2. In some embodiments, the one or more genes include 16 or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2.In some embodiments, the one or more genes include 17 or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2. In some embodiments, the one or more genes include 18 or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2. In some embodiments, the one or more genes include 19 or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2. In some embodiments, the one or more genes include 20 or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2.In some embodiments, the one or more genes include 21 or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2. In some embodiments, the one or more genes comprise 22 or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2. In some embodiments, the one or more genes comprise 22 or more of CST7, NKG7, GZMH, MT-ND4, These include 23 or more of HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2. In some embodiments, the one or more genes include 24 or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2. In some embodiments, the one or more genes include 25 or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2. In some embodiments, the one or more genes include 26 or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2.In some embodiments, the one or more genes include 27 or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2. In some embodiments, the one or more genes include 28 or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2. In some embodiments, the one or more genes include 29 or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2. In some embodiments, the one or more genes include 30 or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2.In some embodiments, the one or more genes include 31 or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2. In some embodiments, the one or more genes include 32 or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2. In some embodiments, the one or more genes include all 33 of: CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2.

[0204] In some embodiments, the one or more genes include one or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, and LITAF. For example, the one or more genes can include one, two, three, four, five, six, seven, eight, nine, ten, eleven, twelve, thirteen, fourteen, fifteen, sixteen, seventeen, eighteen, nineteen, or all twenty of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, and LITAF.

[0205] In some embodiments, the one or more genes comprise two or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, and LITAF. In some embodiments, the one or more genes comprise three or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, and LITAF. In some embodiments, the one or more genes include four or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, and LITAF. In some embodiments, the one or more genes include five or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, and LITAF. In some embodiments, the one or more genes include six or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, and LITAF. In some embodiments, the one or more genes include seven or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, and LITAF.In some embodiments, the one or more genes include eight or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, and LITAF. In some embodiments, the one or more genes include nine or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, and LITAF. In some embodiments, the one or more genes include 10 or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, and LITAF. In some embodiments, the one or more genes include 11 or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, and LITAF. In some embodiments, the one or more genes include 12 or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, and LITAF. In some embodiments, the one or more genes include 13 or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, and LITAF.In some embodiments, the one or more genes include 14 or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, and LITAF. In some embodiments, the one or more genes include 15 or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, and LITAF. In some embodiments, the one or more genes include 16 or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, and LITAF. In some embodiments, the one or more genes include 17 or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, and LITAF. In some embodiments, the one or more genes include 18 or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, and LITAF. In some embodiments, the one or more genes include 19 or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, and LITAF.In some embodiments, the one or more genes include all 20 of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, and LITAF. In some embodiments, the one or more genes consist of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, and LITAF.

[0206] In some embodiments, the one or more genes comprise a two-gene panel shown in Table 1 below.

[0207] Table 1: Exemplary two-gene panels for use in combination with the compositions and methods described herein TIFF0007671248000001.tif38170

[0208] In some embodiments, the one or more genes comprise a three-gene panel, as shown in Table 2 below.

[0209] Table 2: Exemplary 3-gene panels for use in combination with the compositions and methods described herein TIFF0007671248000002.tif72170

[0210] In some embodiments, the one or more genes comprise a four-gene panel, as shown in Table 3 below.

[0211] Table 3: Exemplary 4-gene panel for use in combination with the compositions and methods described herein TIFF0007671248000003.tif38170

[0212] In some embodiments, the one or more genes include a panel of CST7, NKG7, GZMH, MT-ND4, and HLA-H.

[0213] In some embodiments, the one or more genes include one or more of GZMB, KLRD1, FGFBP2, NKG7, CST7, KLRC4-KLRK1, KLRK1, GZMH, B2M, LITAF, CTSW, GZMA, ID2, CX3CR1, PRSS23, CCL5, GNLY, PRF1, PATL2, and CD8A. For example, the one or more genes can include one, two, three, four, five, six, seven, eight, nine, ten, eleven, twelve, thirteen, fourteen, fifteen, sixteen, seventeen, eighteen, nineteen, or all twenty of GZMB, KLRD1, FGFBP2, NKG7, CST7, KLRC4-KLRK1, KLRK1, GZMH, B2M, LITAF, CTSW, GZMA, ID2, CX3CR1, PRSS23, CCL5, GNLY, PRF1, PATL2, and CD8A.

[0214] In some embodiments, the one or more genes comprise two or more of GZMB, KLRD1, FGFBP2, NKG7, CST7, KLRC4-KLRK1, KLRK1, GZMH, B2M, LITAF, CTSW, GZMA, ID2, CX3CR1, PRSS23, CCL5, GNLY, PRF1, PATL2, and CD8A. In some embodiments, the one or more genes comprise three or more of GZMB, KLRD1, FGFBP2, NKG7, CST7, KLRC4-KLRK1, KLRK1, GZMH, B2M, LITAF, CTSW, GZMA, ID2, CX3CR1, PRSS23, CCL5, GNLY, PRF1, PATL2, and CD8A. In some embodiments, the one or more genes include four or more of GZMB, KLRD1, FGFBP2, NKG7, CST7, KLRC4-KLRK1, KLRK1, GZMH, B2M, LITAF, CTSW, GZMA, ID2, CX3CR1, PRSS23, CCL5, GNLY, PRF1, PATL2, and CD8A. In some embodiments, the one or more genes include five or more of GZMB, KLRD1, FGFBP2, NKG7, CST7, KLRC4-KLRK1, KLRK1, GZMH, B2M, LITAF, CTSW, GZMA, ID2, CX3CR1, PRSS23, CCL5, GNLY, PRF1, PATL2, and CD8A. In some embodiments, the one or more genes include six or more of GZMB, KLRD1, FGFBP2, NKG7, CST7, KLRC4-KLRK1, KLRK1, GZMH, B2M, LITAF, CTSW, GZMA, ID2, CX3CR1, PRSS23, CCL5, GNLY, PRF1, PATL2, and CD8A. In some embodiments, the one or more genes include seven or more of GZMB, KLRD1, FGFBP2, NKG7, CST7, KLRC4-KLRK1, KLRK1, GZMH, B2M, LITAF, CTSW, GZMA, ID2, CX3CR1, PRSS23, CCL5, GNLY, PRF1, PATL2, and CD8A.In some embodiments, the one or more genes include eight or more of GZMB, KLRD1, FGFBP2, NKG7, CST7, KLRC4-KLRK1, KLRK1, GZMH, B2M, LITAF, CTSW, GZMA, ID2, CX3CR1, PRSS23, CCL5, GNLY, PRF1, PATL2, and CD8A. In some embodiments, the one or more genes include nine or more of GZMB, KLRD1, FGFBP2, NKG7, CST7, KLRC4-KLRK1, KLRK1, GZMH, B2M, LITAF, CTSW, GZMA, ID2, CX3CR1, PRSS23, CCL5, GNLY, PRF1, PATL2, and CD8A. In some embodiments, the one or more genes include 10 or more of GZMB, KLRD1, FGFBP2, NKG7, CST7, KLRC4-KLRK1, KLRK1, GZMH, B2M, LITAF, CTSW, GZMA, ID2, CX3CR1, PRSS23, CCL5, GNLY, PRF1, PATL2, and CD8A. In some embodiments, the one or more genes include 11 or more of GZMB, KLRD1, FGFBP2, NKG7, CST7, KLRC4-KLRK1, KLRK1, GZMH, B2M, LITAF, CTSW, GZMA, ID2, CX3CR1, PRSS23, CCL5, GNLY, PRF1, PATL2, and CD8A. In some embodiments, the one or more genes include 12 or more of GZMB, KLRD1, FGFBP2, NKG7, CST7, KLRC4-KLRK1, KLRK1, GZMH, B2M, LITAF, CTSW, GZMA, ID2, CX3CR1, PRSS23, CCL5, GNLY, PRF1, PATL2, and CD8A. In some embodiments, the one or more genes include 13 or more of GZMB, KLRD1, FGFBP2, NKG7, CST7, KLRC4-KLRK1, KLRK1, GZMH, B2M, LITAF, CTSW, GZMA, ID2, CX3CR1, PRSS23, CCL5, GNLY, PRF1, PATL2, and CD8A.In some embodiments, the one or more genes include 14 or more of GZMB, KLRD1, FGFBP2, NKG7, CST7, KLRC4-KLRK1, KLRK1, GZMH, B2M, LITAF, CTSW, GZMA, ID2, CX3CR1, PRSS23, CCL5, GNLY, PRF1, PATL2, and CD8A. In some embodiments, the one or more genes include 15 or more of GZMB, KLRD1, FGFBP2, NKG7, CST7, KLRC4-KLRK1, KLRK1, GZMH, B2M, LITAF, CTSW, GZMA, ID2, CX3CR1, PRSS23, CCL5, GNLY, PRF1, PATL2, and CD8A. In some embodiments, the one or more genes include 16 or more of GZMB, KLRD1, FGFBP2, NKG7, CST7, KLRC4-KLRK1, KLRK1, GZMH, B2M, LITAF, CTSW, GZMA, ID2, CX3CR1, PRSS23, CCL5, GNLY, PRF1, PATL2, and CD8A. In some embodiments, the one or more genes include 17 or more of GZMB, KLRD1, FGFBP2, NKG7, CST7, KLRC4-KLRK1, KLRK1, GZMH, B2M, LITAF, CTSW, GZMA, ID2, CX3CR1, PRSS23, CCL5, GNLY, PRF1, PATL2, and CD8A. In some embodiments, the one or more genes include 18 or more of GZMB, KLRD1, FGFBP2, NKG7, CST7, KLRC4-KLRK1, KLRK1, GZMH, B2M, LITAF, CTSW, GZMA, ID2, CX3CR1, PRSS23, CCL5, GNLY, PRF1, PATL2, and CD8A. In some embodiments, the one or more genes include 19 or more of GZMB, KLRD1, FGFBP2, NKG7, CST7, KLRC4-KLRK1, KLRK1, GZMH, B2M, LITAF, CTSW, GZMA, ID2, CX3CR1, PRSS23, CCL5, GNLY, PRF1, PATL2, and CD8A.In some embodiments, the one or more genes include all 20 of GZMB, KLRD1, FGFBP2, NKG7, CST7, KLRC4-KLRK1, KLRK1, GZMH, B2M, LITAF, CTSW, GZMA, ID2, CX3CR1, PRSS23, CCL5, GNLY, PRF1, PATL2, and CD8A. In some embodiments, the one or more genes consist of GZMB, KLRD1, FGFBP2, NKG7, CST7, KLRC4-KLRK1, KLRK1, GZMH, B2M, LITAF, CTSW, GZMA, ID2, CX3CR1, PRSS23, CCL5, GNLY, PRF1, PATL2, and CD8A.

[0215] In some embodiments, the one or more genes comprise one or more cytotoxic genes, such as one or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C. In some embodiments, the one or more genes comprise two or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C. In some embodiments, the one or more genes comprise three or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C. In some embodiments, the one or more genes comprise four or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C. In some embodiments, the one or more genes include five or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C. In some embodiments, the one or more genes include six or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C. In some embodiments, the one or more genes include seven or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C. In some embodiments, the one or more genes include eight or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C. In some embodiments, the one or more genes include nine or more of CST7, PLEK, HCST, CTsSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C. In some embodiments, the one or more genes include ten or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C.In some embodiments, the one or more genes include all 11 of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C. In some embodiments, the one or more genes consist of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C.

[0216] In some embodiments, the one or more genes include one or more mitochondrial genes, such as one or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6. In some embodiments, the one or more genes include two or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6. In some embodiments, the one or more genes include three or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6. In some embodiments, the one or more genes include four or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6. In some embodiments, the one or more genes include all five of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6. In some embodiments, the one or more genes consist of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6.

[0217] In some embodiments, the one or more genes include one or more cytotoxic genes (e.g., one or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C) and one or more mitochondrial genes (e.g., one or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6). For example, in some embodiments, the one or more genes include a cytotoxic gene and one or more mitochondrial genes. In some embodiments, the one or more genes include CST7 and one or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6 (e.g., any one of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; two or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; three or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; four or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; or all five of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6). In some embodiments, the one or more genes include PLEK and one or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6 (e.g., any one of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; two or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; three or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; four or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; or all five of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6).In some embodiments, the one or more genes include HCST and one or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6 (e.g., any one of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; two or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; three or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; four or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; or all five of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6). In some embodiments, the one or more genes include CTSW and one or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6 (e.g., any one of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; two or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; three or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; four or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; or all five of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6). In some embodiments, the one or more genes include KLRG1 and one or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6 (e.g., any one of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; two or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; three or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; four or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; or all five of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6).In some embodiments, the one or more genes include CD8A and one or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6 (e.g., any one of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; two or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; three or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; four or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; or all five of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6). In some embodiments, the one or more genes include CCL5 and one or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6 (e.g., any one of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; two or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; three or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; four or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; or all five of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6). In some embodiments, the one or more genes include CD8B and one or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6 (e.g., any one of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; two or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; three or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; four or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; or all five of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6).In some embodiments, the one or more genes include NKG7 and one or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6 (e.g., any one of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; two or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; three or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; four or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; or all five of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6). In some embodiments, the one or more genes include GZMH and one or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6 (e.g., any one of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; two or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; three or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; four or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; or all five of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6). In some embodiments, the one or more genes include HLA-C and one or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6 (e.g., any one of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; two or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; three or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; four or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; or all five of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6).

[0218] In some embodiments, the one or more genes include a mitochondrial gene and one or more cytotoxic genes. For example, in some embodiments, the one or more genes include MT-ND4 and one or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C (e.g., any one of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C, ... G1, two or more of CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C, CST7, PLEK, HCST, CTSW, KLRG1, three or more of CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C, CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C, CST7, PLEK, HCST, CTSW, Five or more of KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C, CST7, PLEK, HCST, CTSW, Six or more of KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C, CST7, PLEK, HCST, CTSW, Seven or more of KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C, CST7, PLEK, HCST, C In some embodiments, the one or more genes are selected from the group consisting of eight or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C, nine or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C, ten or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C, or all eleven of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C. In some embodiments, the one or more genes are selected from the group consisting of MT-CYB and CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C.and one or more of HLA-C (e.g., any one of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C, two or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C, CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, Three or more of CD8B, NKG7, GZMH, and HLA-C; Four or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C; Five or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C; CST7, PLEK, HCST, CTSW, KLRG1, C Six or more of D8A, CCL5, CD8B, NKG7, GZMH, and HLA-C, Seven or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C, Eight or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C, CST7, PLEK, HCST, CTS nine or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C, ten or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C, or all eleven of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C). In some embodiments, the one or more genes are MT-CO2 and one or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C (e.g., any one of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C, CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH,and two or more of HLA-C, CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and three or more of HLA-C, CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and four or more of HLA-C, CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and five or more of HLA-C, CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and six or more of HLA-C, CST7, PLEK, HCST, CTSW, KLRG1, CD8A , seven or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C; eight or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C; nine or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C; ten or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C; or all eleven of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C). In some embodiments, the one or more genes are MT-ND4L and one or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C (e.g., any one of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C, CST7, PLEK, HCST, two or more of CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C; three or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C; four or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C;Five or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C; Six or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C; Seven or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C; CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD eight or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C; nine or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C; ten or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C; or all eleven of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C). In some embodiments, the one or more genes are MT-ATP6 and one or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C (e.g., any one of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C, two or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C, ... Three or more of RG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C; Four or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C; Five or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C; Six or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C; CST7, PLEK, HCST,seven or more of CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C, eight or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C, nine or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C, ten or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C, or all eleven of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C).

[0219] In some embodiments, the one or more genes comprise a combination of a cytotoxic gene and a mitochondrial gene as shown in Table 4.

[0220] Table 4: Exemplary panels of cytotoxic and mitochondrial genes for use in combination with the compositions and methods described herein TIFF0007671248000004.tif234170TIFF0007671248000005.tif159170

[0221] In some embodiments, the one or more genes include one or more of NKG7, CST7, GZMH, LITAF, CTSW, CCL5, and CD8A. For example, the one or more genes can include one, two, three, four, five, six, or all seven of NKG7, CST7, GZMH, LITAF, CTSW, CCL5, and CD8A.

[0222] In some embodiments, the one or more genes include two or more of NKG7, CST7, GZMH, LITAF, CTSW, CCL5, and CD8A. In some embodiments, the one or more genes include three or more of NKG7, CST7, GZMH, LITAF, CTSW, CCL5, and CD8A. In some embodiments, the one or more genes include four or more of NKG7, CST7, GZMH, LITAF, CTSW, CCL5, and CD8A. In some embodiments, the one or more genes include five or more of NKG7, CST7, GZMH, LITAF, CTSW, CCL5, and CD8A. In some embodiments, the one or more genes include six or more of NKG7, CST7, GZMH, LITAF, CTSW, CCL5, and CD8A. In some embodiments, the one or more genes include all seven of NKG7, CST7, GZMH, LITAF, CTSW, CCL5, and CD8A. In some embodiments, the one or more genes consist of NKG7, CST7, GZMH, LITAF, CTSW, CCL5, and CD8A.

[0223] In some embodiments, the expression level is a nucleic acid expression level, such as an mRNA expression level. The mRNA expression level can be determined, for example, by RNA-seq, RT-qPCR, qPCR, multiplex qPCR or RT-qPCR, microarray analysis, SAGE, MassARRAY techniques, FISH, or a combination thereof.

[0224] In some embodiments, the expression level is detected in tumor cells, tumor-infiltrating immune cells, stromal cells, normal adjacent tissue (NAT) cells, or a combination thereof. In some embodiments, the sample is a tissue sample, a cell sample, a whole blood sample, a plasma sample, a serum sample, or a combination thereof. The tissue sample can be, for example, a tumor tissue sample, such as a tumor tissue sample containing tumor cells, tumor-infiltrating immune cells, stromal cells, NAT cells, or a combination thereof. In some embodiments, the tumor tissue sample is a formalin-fixed and paraffin-embedded (FFPE) sample, an archival sample, a fresh sample, or a frozen sample. In some embodiments, the tumor tissue sample is an FFPE sample.

[0225] In some embodiments, the cancer is lung cancer, kidney cancer, bladder cancer, breast cancer, colorectal cancer, ovarian cancer, pancreatic cancer, gastric cancer, esophageal cancer, mesothelioma, melanoma, head and neck cancer, thyroid cancer, sarcoma, prostate cancer, glioblastoma, cervical cancer, thymic carcinoma, leukemia, lymphoma, myeloma, mycosis fungoides, Merkel cell carcinoma, or a hematological malignancy. For example, the cancer can be lung cancer, kidney cancer, bladder cancer, or breast cancer. In some embodiments, the lung cancer is non-small cell lung cancer (NSCLC). In some embodiments, the kidney cancer is renal cell carcinoma (RCC). In some embodiments, the bladder cancer is urothelial carcinoma (UC). In some embodiments, the breast cancer is triple-negative breast cancer (TNBC).

[0226] In some embodiments, the PD-L1 axis binding antagonist is a PD-L1 binding antagonist, a PD-1 binding antagonist, or a PD-L2 binding antagonist. In some embodiments, the PD-L1 axis binding antagonist is a PD-L1 binding antagonist. A PD-L1 binding antagonist may inhibit the binding of PD-L1 to one or more of its ligand binding partners. In some embodiments, the PD-L1 binding antagonist inhibits the binding of PD-L1 to PD-1. In some embodiments, the PD-L1 binding antagonist inhibits the binding of PD-L1 to B7-1. In some embodiments, the PD-L1 binding antagonist inhibits the binding of PD-L1 to both PD-1 and B7-1.

[0227] In some embodiments, the PD-L1 binding antagonist is an antibody or antigen-binding fragment thereof. The antibody can be, for example, atezolizumab (MPDL3280A), YW243.55.S70, MDX-1105, MEDI4736 (durvalumab), and MSB0010718C (avelumab). In some embodiments, the PD-L1 binding antagonist is atezolizumab (MPDL3280A).

[0228] In some embodiments, the antibody comprises a heavy chain comprising the HVR-H1 sequence of SEQ ID NO: 41, the HVR-H2 sequence of SEQ ID NO: 42, and the HVR-H3 sequence of SEQ ID NO: 43, and a light chain comprising the HVR-L1 sequence of SEQ ID NO: 44, the HVR-L2 sequence of SEQ ID NO: 45, and the HVR-L3 sequence of SEQ ID NO: 46.

[0229] In some embodiments, the antibody comprises a heavy chain variable region comprising the amino acid sequence of SEQ ID NO:47 and a light chain variable region comprising the amino acid sequence of SEQ ID NO:48.

[0230] In some embodiments, the PD-L1 axis binding antagonist is a PD-1 binding antagonist. The PD-1 binding antagonist may, for example, inhibit the binding of PD-1 to one or more of its ligand binding partners. In some embodiments, the PD-1 binding antagonist inhibits the binding of PD-1 to PD-L1. In some embodiments, the PD-1 binding antagonist inhibits the binding of PD-1 to PD-L2. In some embodiments, the PD-1 binding antagonist inhibits the binding of PD-1 to both PD-L1 and PD-L2.

[0231] In some embodiments, the PD-1 binding antagonist is an antibody or antigen-binding fragment thereof. The antibody can be, for example, MDX-1106 (nivolumab), MK-3475 (pembrolizumab), MEDI-0680 (AMP-514), PDR001, REGN2810, and BGB-108.

[0232] In some embodiments, the PD-1 binding antagonist is an Fc fusion protein, such as AMP-224.

[0233] In some embodiments, the individual has not been previously treated for cancer, hi some embodiments, the individual has not been previously administered a PD-L1 axis-binding antagonist.

[0234] In some embodiments, the individual has previously been treated for cancer by administering a platinum-containing chemotherapeutic agent to the individual. In some embodiments, the individual did not respond to the chemotherapeutic agent.

[0235] In some embodiments, the treatment comprising the PD-L1 axis-binding antagonist is a monotherapy.

[0236] In some embodiments, the method further comprises administering to the individual an effective amount of one or more additional therapeutic agents. The one or more additional therapeutic agents may include, for example, an anti-tumor agent, a chemotherapeutic agent, a growth inhibitory agent, an anti-angiogenic agent, radiation therapy, a cytotoxic agent, an immunomodulatory agent, or a combination thereof. In some embodiments, the one or more additional therapeutic agents comprises an immunomodulatory agent. The immunomodulatory agent may be, for example, an antibody or an antigen-binding fragment thereof. Exemplary immunomodulatory agents that may be used in combination with the compositions and methods of the present disclosure include anti-TIGIT antibodies and antigen-binding fragments thereof, anti-CTLA-4 antibodies or antigen-binding fragments thereof, anti-CD27 antibodies or antigen-binding fragments thereof, anti-CD30 antibodies or antigen-binding fragments thereof, anti-CD40 antibodies or antigen-binding fragments thereof, anti-4-1BB antibodies or antigen-binding fragments thereof, anti-GITR antibodies or antigen-binding fragments thereof, anti-OX40 antibodies or antigen-binding fragments thereof, anti-TRAILR1 antibodies or antigen-binding fragments thereof, anti-TRAILR2 antibodies or antigen-binding fragments thereof, anti-TWEAK antibodies or antigen-binding fragments thereof, anti-TWEAKR antibodies or antigen-binding fragments thereof, anti-BRAF antibodies or antigen-binding fragments thereof, anti-MEK antibodies or antigen-binding fragments thereof, anti-CD33 antibodies or antigen-binding fragments thereof, anti-CD20 antibodies or antigen-binding fragments thereof, anti-CD52 antibodies or antigen-binding fragments thereof, anti-A33 antibodies or antigen-binding fragments thereof, anti-GD3 antibodies or antigen-binding fragments thereof, and anti-PSMA antibodies or antigen-binding fragments thereof. an antigen-binding fragment thereof, an anti-Ceacan1 antibody or an antigen-binding fragment thereof, an anti-Galedin9 antibody or an antigen-binding fragment thereof, an anti-HVEM antibody or an antigen-binding fragment thereof, an anti-VISTA antibody or an antigen-binding fragment thereof, an anti-B7H4 antibody or an antigen-binding fragment thereof, an anti-HHLA2 antibody or an antigen-binding fragment thereof, an anti-CD155 antibody or an antigen-binding fragment thereof, an anti-CD80 antibody or an antigen-binding fragment thereof, an anti-BTLA antibody or an antigen-binding fragment thereof, an anti-CD160 antibody or an antigen-binding fragment thereof, an anti-CD28 antibody or an antigen-binding fragment thereof, an anti-CD226 antibody or an antigen-binding fragment thereof, an anti-CEACAM1 antibody or an antigen-binding fragment thereof, an anti-TIM3 antibody or an antigen-binding fragment thereof, an anti-CD96 antibody or an antigen-binding fragment thereof, an anti-CD70 antibody or an antigen-binding fragment thereof, an anti-CD27 antibody or an antigen-binding fragment thereof, an anti-LIGHT antibody or an antigen-binding fragment thereof, an anti-CD137 antibody or an antigen-binding fragment thereof, an anti-DR4 antibody or an antigen-binding fragment thereof, an anti-CR5 antibody or an antigen-binding fragment thereof,These antibodies include an anti-FAS antibody or an antigen-binding fragment thereof, an anti-CD95 antibody or an antigen-binding fragment thereof, an anti-TRAIL antibody or an antigen-binding fragment thereof, an anti-DR6 antibody or an antigen-binding fragment thereof, an anti-EDAR antibody or an antigen-binding fragment thereof, an anti-NGFR antibody or an antigen-binding fragment thereof, an anti-OPG antibody or an antigen-binding fragment thereof, an anti-RANKL antibody or an antigen-binding fragment thereof, an anti-LTβR antibody or an antigen-binding fragment thereof, an anti-BCMA antibody or an antigen-binding fragment thereof, an anti-TACI antibody or an antigen-binding fragment thereof, an anti-BAFFR antibody or an antigen-binding fragment thereof, an anti-EDAR2 antibody or an antigen-binding fragment thereof, an anti-TROY antibody or an antigen-binding fragment thereof, and an anti-RELT antibody or an antigen-binding fragment thereof.

[0237] In an additional aspect, the disclosure features a kit for identifying an individual (e.g., a human) with cancer who may benefit from treatment comprising a PD-L1 axis-binding antagonist. The kit includes one or more reagents for determining the expression level of one or more of the genes CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2 in a sample from the individual. 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, or all 33 of the following: CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2). In some embodiments, the kit further comprises instructions for use of the one or more reagents to identify an individual (e.g., a human) with cancer that may benefit from treatment comprising a PD-L1 axis-binding antagonist according to the method of any of the above aspects or embodiments of the present disclosure.

[0238] In a further aspect, the disclosure features an assay for identifying an individual (e.g., a human) with cancer who is a candidate for treatment comprising a PD-L1 axis-binding antagonist. The assay includes determining the expression level of one or more of the genes CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2 in a sample from the individual. L5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2). An immunoscore expression level of one or more genes in the sample that is greater than a reference immunoscore expression level of the one or more genes identifies the individual as one who may benefit from treatment comprising a PD-L1 axis-binding antagonist.

[0239] In some embodiments of the foregoing aspects, the reference immunoscore expression level is an immunoscore expression level of one or more genes in a reference population. In some embodiments, the reference population is a population of individuals with cancer. In some embodiments, the population of individuals includes a first subset of individuals treated with a PD-L1 axis-binding antagonist and a second subset of individuals treated with a therapy that does not include a PD-L1 axis-binding antagonist. The reference immunoscore expression level significantly separates each of the first and second subsets of individuals, for example, based on a significant difference between the individual's responsiveness to treatment with the PD-L1 axis-binding antagonist and the individual's responsiveness to treatment with the therapy that does not include a PD-L1 axis-binding antagonist that is greater than the reference immunoscore expression level. In such cases, the individual's responsiveness to treatment with the PD-L1 axis-binding antagonist may be significantly improved compared to the individual's responsiveness to treatment with the therapy that does not include a PD-L1 axis-binding antagonist.

[0240] In some embodiments, the reference immunoscore expression level significantly separates each of the first and second subsets of individuals based on a significant difference between the individual's responsiveness to treatment with the PD-L1 axis-binding antagonist and the individual's responsiveness to treatment with a therapy that does not include a PD-L1 axis-binding antagonist that is below the reference immunoscore expression level. In such cases, the individual's responsiveness to treatment with the therapy that does not include a PD-L1 axis-binding antagonist is significantly improved compared to the individual's responsiveness to treatment with the PD-L1 axis-binding antagonist.

[0241] In some embodiments, the PD-L1 axis binding antagonist-free therapy comprises an anti-tumor agent, a chemotherapeutic agent, a growth inhibitory agent, an anti-angiogenic agent, radiation therapy, a cytotoxic agent, or a combination thereof. For example, the PD-L1 axis binding antagonist-free therapy comprises a chemotherapeutic agent, such as docetaxel.

[0242] In some embodiments, responsiveness to treatment comprises an increase in progression-free survival (PFS). In some embodiments, responsiveness to treatment comprises an increase in overall survival (OS).

[0243] In some embodiments, the reference immunoscore expression level is the median of the expression level of one or more genes in the reference population.For example, the immunoscore expression level of one or more genes in the reference population can be the median of the normalized expression level of one or more genes in the reference population.

[0244] In some embodiments, reference immune score expression level is the average of each expression level of one or more genes in reference population.For example, the immune score expression level of one or more genes in reference population can be the average of each normalized expression level of one or more genes in reference population.In some embodiments, each normalized expression level of one or more genes is the expression level of one or more genes normalized to reference gene, for example, CD8A.

[0245] In some embodiments, the reference immune score expression level is a pre-assigned expression level of one or more genes.

[0246] In some embodiments, the benefit of treatment comprising a PD-L1 axis binding antagonist comprises an increase in OS. In some embodiments, the benefit of treatment comprising a PD-L1 axis binding antagonist comprises an increase in PFS. In some embodiments, the benefit of treatment comprising a PD-L1 axis binding antagonist comprises an increase in OS and PFS.

[0247] In some embodiments of either of the preceding two aspects, the one or more genes include more than one of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2. For example, in some embodiments, the one or more genes include two or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2. In some embodiments, the one or more genes include three or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2. In some embodiments, the one or more genes include four or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2.In some embodiments, the one or more genes include five or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2. In some embodiments, the one or more genes include six or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2. In some embodiments, the one or more genes include seven or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2. In some embodiments, the one or more genes include eight or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2.In some embodiments, the one or more genes include nine or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2. In some embodiments, the one or more genes include ten or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2. In some embodiments, the one or more genes include 11 or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2. In some embodiments, the one or more genes include 12 or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2.In some embodiments, the one or more genes include 13 or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2. In some embodiments, the one or more genes include 14 or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2. In some embodiments, the one or more genes include 15 or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2. In some embodiments, the one or more genes include 16 or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2.In some embodiments, the one or more genes include 17 or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2. In some embodiments, the one or more genes include 18 or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2. In some embodiments, the one or more genes include 19 or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2. In some embodiments, the one or more genes include 20 or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2.In some embodiments, the one or more genes include 21 or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2. In some embodiments, the one or more genes comprise 22 or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2. In some embodiments, the one or more genes comprise CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2. 7, including 23 or more of GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2. In some embodiments, the one or more genes include 24 or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2. In some embodiments, the one or more genes include 25 or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2. In some embodiments, the one or more genes include 26 or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2.In some embodiments, the one or more genes include 27 or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2. In some embodiments, the one or more genes include 28 or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2. In some embodiments, the one or more genes include 29 or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2. In some embodiments, the one or more genes include 30 or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2.In some embodiments, the one or more genes include 31 or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2. In some embodiments, the one or more genes include 32 or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2. In some embodiments, the one or more genes include all 33 of: CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2.

[0248] In some embodiments of either of the preceding two aspects, the one or more genes include one or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, and LITAF. For example, the one or more genes can include one, two, three, four, five, six, seven, eight, nine, ten, eleven, twelve, thirteen, fourteen, fifteen, sixteen, seventeen, eighteen, nineteen, or all twenty of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, and LITAF.

[0249] For example, in some embodiments of either of the preceding two aspects, the one or more genes include two or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, and LITAF. In some embodiments, the one or more genes include three or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, and LITAF. In some embodiments, the one or more genes include four or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, and LITAF. In some embodiments, the one or more genes include five or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, and LITAF. In some embodiments, the one or more genes include six or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, and LITAF. In some embodiments, the one or more genes include seven or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, and LITAF.In some embodiments, the one or more genes include eight or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, and LITAF. In some embodiments, the one or more genes include nine or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, and LITAF. In some embodiments, the one or more genes include 10 or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, and LITAF. In some embodiments, the one or more genes include 11 or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, and LITAF. In some embodiments, the one or more genes include 12 or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, and LITAF. In some embodiments, the one or more genes include 13 or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, and LITAF.In some embodiments, the one or more genes include 14 or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, and LITAF. In some embodiments, the one or more genes include 15 or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, and LITAF. In some embodiments, the one or more genes include 16 or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, and LITAF. In some embodiments, the one or more genes include 17 or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, and LITAF. In some embodiments, the one or more genes include 18 or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, and LITAF. In some embodiments, the one or more genes include 19 or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, and LITAF.In some embodiments, the one or more genes include all 20 of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, and LITAF. In some embodiments, the one or more genes consist of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, and LITAF.

[0250] In some embodiments of either of the preceding two aspects, the one or more genes comprise a panel of genes set forth in any one of Tables 1-3 above. In some embodiments, the one or more genes comprise the panel of CST7, NKG7, GZMH, MT-ND4, and HLA-H.

[0251] In some embodiments of either of the preceding two aspects, the one or more genes include one or more of GZMB, KLRD1, FGFBP2, NKG7, CST7, KLRC4-KLRK1, KLRK1, GZMH, B2M, LITAF, CTSW, GZMA, ID2, CX3CR1, PRSS23, CCL5, GNLY, PRF1, PATL2, and CD8A. For example, the one or more genes can include one, two, three, four, five, six, seven, eight, nine, ten, eleven, twelve, thirteen, fourteen, fifteen, sixteen, seventeen, eighteen, nineteen, or all twenty of GZMB, KLRD1, FGFBP2, NKG7, CST7, KLRC4-KLRK1, KLRK1, GZMH, B2M, LITAF, CTSW, GZMA, ID2, CX3CR1, PRSS23, CCL5, GNLY, PRF1, PATL2, and CD8A.

[0252] For example, in some embodiments of either of the preceding two aspects, the one or more genes include two or more of GZMB, KLRD1, FGFBP2, NKG7, CST7, KLRC4-KLRK1, KLRK1, GZMH, B2M, LITAF, CTSW, GZMA, ID2, CX3CR1, PRSS23, CCL5, GNLY, PRF1, PATL2, and CD8A. In some embodiments, the one or more genes include three or more of GZMB, KLRD1, FGFBP2, NKG7, CST7, KLRC4-KLRK1, KLRK1, GZMH, B2M, LITAF, CTSW, GZMA, ID2, CX3CR1, PRSS23, CCL5, GNLY, PRF1, PATL2, and CD8A. In some embodiments, the one or more genes include four or more of GZMB, KLRD1, FGFBP2, NKG7, CST7, KLRC4-KLRK1, KLRK1, GZMH, B2M, LITAF, CTSW, GZMA, ID2, CX3CR1, PRSS23, CCL5, GNLY, PRF1, PATL2, and CD8A. In some embodiments, the one or more genes include five or more of GZMB, KLRD1, FGFBP2, NKG7, CST7, KLRC4-KLRK1, KLRK1, GZMH, B2M, LITAF, CTSW, GZMA, ID2, CX3CR1, PRSS23, CCL5, GNLY, PRF1, PATL2, and CD8A. In some embodiments, the one or more genes include six or more of GZMB, KLRD1, FGFBP2, NKG7, CST7, KLRC4-KLRK1, KLRK1, GZMH, B2M, LITAF, CTSW, GZMA, ID2, CX3CR1, PRSS23, CCL5, GNLY, PRF1, PATL2, and CD8A. In some embodiments, the one or more genes include seven or more of GZMB, KLRD1, FGFBP2, NKG7, CST7, KLRC4-KLRK1, KLRK1, GZMH, B2M, LITAF, CTSW, GZMA, ID2, CX3CR1, PRSS23, CCL5, GNLY, PRF1, PATL2, and CD8A.In some embodiments, the one or more genes include eight or more of GZMB, KLRD1, FGFBP2, NKG7, CST7, KLRC4-KLRK1, KLRK1, GZMH, B2M, LITAF, CTSW, GZMA, ID2, CX3CR1, PRSS23, CCL5, GNLY, PRF1, PATL2, and CD8A. In some embodiments, the one or more genes include nine or more of GZMB, KLRD1, FGFBP2, NKG7, CST7, KLRC4-KLRK1, KLRK1, GZMH, B2M, LITAF, CTSW, GZMA, ID2, CX3CR1, PRSS23, CCL5, GNLY, PRF1, PATL2, and CD8A. In some embodiments, the one or more genes include 10 or more of GZMB, KLRD1, FGFBP2, NKG7, CST7, KLRC4-KLRK1, KLRK1, GZMH, B2M, LITAF, CTSW, GZMA, ID2, CX3CR1, PRSS23, CCL5, GNLY, PRF1, PATL2, and CD8A. In some embodiments, the one or more genes include 11 or more of GZMB, KLRD1, FGFBP2, NKG7, CST7, KLRC4-KLRK1, KLRK1, GZMH, B2M, LITAF, CTSW, GZMA, ID2, CX3CR1, PRSS23, CCL5, GNLY, PRF1, PATL2, and CD8A. In some embodiments, the one or more genes include 12 or more of GZMB, KLRD1, FGFBP2, NKG7, CST7, KLRC4-KLRK1, KLRK1, GZMH, B2M, LITAF, CTSW, GZMA, ID2, CX3CR1, PRSS23, CCL5, GNLY, PRF1, PATL2, and CD8A. In some embodiments, the one or more genes include 13 or more of GZMB, KLRD1, FGFBP2, NKG7, CST7, KLRC4-KLRK1, KLRK1, GZMH, B2M, LITAF, CTSW, GZMA, ID2, CX3CR1, PRSS23, CCL5, GNLY, PRF1, PATL2, and CD8A.In some embodiments, the one or more genes include 14 or more of GZMB, KLRD1, FGFBP2, NKG7, CST7, KLRC4-KLRK1, KLRK1, GZMH, B2M, LITAF, CTSW, GZMA, ID2, CX3CR1, PRSS23, CCL5, GNLY, PRF1, PATL2, and CD8A. In some embodiments, the one or more genes include 15 or more of GZMB, KLRD1, FGFBP2, NKG7, CST7, KLRC4-KLRK1, KLRK1, GZMH, B2M, LITAF, CTSW, GZMA, ID2, CX3CR1, PRSS23, CCL5, GNLY, PRF1, PATL2, and CD8A. In some embodiments, the one or more genes include 16 or more of GZMB, KLRD1, FGFBP2, NKG7, CST7, KLRC4-KLRK1, KLRK1, GZMH, B2M, LITAF, CTSW, GZMA, ID2, CX3CR1, PRSS23, CCL5, GNLY, PRF1, PATL2, and CD8A. In some embodiments, the one or more genes include 17 or more of GZMB, KLRD1, FGFBP2, NKG7, CST7, KLRC4-KLRK1, KLRK1, GZMH, B2M, LITAF, CTSW, GZMA, ID2, CX3CR1, PRSS23, CCL5, GNLY, PRF1, PATL2, and CD8A. In some embodiments, the one or more genes include 18 or more of GZMB, KLRD1, FGFBP2, NKG7, CST7, KLRC4-KLRK1, KLRK1, GZMH, B2M, LITAF, CTSW, GZMA, ID2, CX3CR1, PRSS23, CCL5, GNLY, PRF1, PATL2, and CD8A. In some embodiments, the one or more genes include 19 or more of GZMB, KLRD1, FGFBP2, NKG7, CST7, KLRC4-KLRK1, KLRK1, GZMH, B2M, LITAF, CTSW, GZMA, ID2, CX3CR1, PRSS23, CCL5, GNLY, PRF1, PATL2, and CD8A.In some embodiments, the one or more genes include all 20 of GZMB, KLRD1, FGFBP2, NKG7, CST7, KLRC4-KLRK1, KLRK1, GZMH, B2M, LITAF, CTSW, GZMA, ID2, CX3CR1, PRSS23, CCL5, GNLY, PRF1, PATL2, and CD8A. In some embodiments, the one or more genes consist of GZMB, KLRD1, FGFBP2, NKG7, CST7, KLRC4-KLRK1, KLRK1, GZMH, B2M, LITAF, CTSW, GZMA, ID2, CX3CR1, PRSS23, CCL5, GNLY, PRF1, PATL2, and CD8A.

[0253] In some embodiments of either of the preceding two aspects, the one or more genes include one or more cytotoxic genes, e.g., one or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C. In some embodiments, the one or more genes include two or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C. In some embodiments, the one or more genes include three or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C. In some embodiments, the one or more genes include four or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C. In some embodiments, the one or more genes include five or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C. In some embodiments, the one or more genes include six or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C. In some embodiments, the one or more genes include seven or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C. In some embodiments, the one or more genes include eight or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C. In some embodiments, the one or more genes include nine or more of CST7, PLEK, HCST, CTsSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C. In some embodiments, the one or more genes include ten or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C.In some embodiments, the one or more genes comprise 11 or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C. In some embodiments, the one or more genes consist of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C.

[0254] In some embodiments of either of the preceding two aspects, the one or more genes include one or more mitochondrial genes, for example, one or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6. In some embodiments, the one or more genes include two or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6. In some embodiments, the one or more genes include three or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6. In some embodiments, the one or more genes include four or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6. In some embodiments, the one or more genes include all five of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6. In some embodiments, the one or more genes consist of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6.

[0255] In some embodiments of either of the preceding two aspects, the one or more genes include one or more cytotoxic genes (e.g., one or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C) and one or more mitochondrial genes (e.g., one or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6). For example, in some embodiments, the one or more genes include a cytotoxic gene and one or more mitochondrial genes. In some embodiments, the one or more genes include CST7 and one or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6 (e.g., any one of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; two or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; three or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; four or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; or all five of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6). In some embodiments, the one or more genes include PLEK and one or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6 (e.g., any one of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; two or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; three or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; four or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; or all five of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6).In some embodiments, the one or more genes include HCST and one or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6 (e.g., any one of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; two or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; three or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; four or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; or all five of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6). In some embodiments, the one or more genes include CTSW and one or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6 (e.g., any one of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; two or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; three or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; four or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; or all five of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6). In some embodiments, the one or more genes include KLRG1 and one or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6 (e.g., any one of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; two or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; three or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; four or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; or all five of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6).In some embodiments, the one or more genes include CD8A and one or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6 (e.g., any one of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; two or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; three or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; four or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; or all five of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6). In some embodiments, the one or more genes include CCL5 and one or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6 (e.g., any one of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; two or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; three or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; four or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; or all five of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6). In some embodiments, the one or more genes include CD8B and one or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6 (e.g., any one of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; two or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; three or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; four or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; or all five of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6).In some embodiments, the one or more genes include NKG7 and one or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6 (e.g., any one of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; two or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; three or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; four or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; or all five of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6). In some embodiments, the one or more genes include GZMH and one or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6 (e.g., any one of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; two or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; three or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; four or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; or all five of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6). In some embodiments, the one or more genes include HLA-C and one or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6 (e.g., any one of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; two or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; three or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; four or more of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6; or all five of MT-ND4, MT-CYB, MT-CO2, MT-ND4L, and MT-ATP6).

[0256] In some embodiments of either of the preceding two aspects, the one or more genes include a mitochondrial gene and one or more cytotoxic genes. For example, in some embodiments, the one or more genes include MT-ND4 and one or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C (e.g., any one of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C, CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8 B, two or more of NKG7, GZMH, and HLA-C, three or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C, four or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C, CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and 5 or more of HLA-C, CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and 6 or more of HLA-C, CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and 7 or more of HLA-C, CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and 8 or more of HLA-C. nine or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C; ten or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C; or all eleven of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C).In some embodiments, the one or more genes are MT-CYB and one or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C (e.g., any one of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C, ... Two or more of KG7, GZMH, and HLA-C; Three or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C; Four or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C; CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C. Five or more of the following genes: CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C; six or more of the following genes: CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C; seven or more of the following genes: CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C; eight or more of the following genes: CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C nine or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C, ten or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C, or all eleven of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C).In some embodiments, the one or more genes are MT-CO2 and one or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C (e.g., any one of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C, ... Two or more of KG7, GZMH, and HLA-C; Three or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C; Four or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C; CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C. Five or more of the following genes: CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C; six or more of the following genes: CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C; seven or more of the following genes: CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C; eight or more of the following genes: CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C nine or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C, ten or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C, or all eleven of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C).In some embodiments, the one or more genes are MT-ND4L and one or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C (e.g., any one of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C, CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, Two or more of NKG7, GZMH, and HLA-C; Three or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C; Four or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C; CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and Five or more of HLA-C, CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and six or more of HLA-C, CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and seven or more of HLA-C, and eight or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C nine or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C, ten or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C, or all eleven of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C).In some embodiments, the one or more genes are MT-ATP6 and one or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C (e.g., any one of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C, CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, Two or more of NKG7, GZMH, and HLA-C; Three or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C; Four or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C; CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and Five or more of HLA-C, CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and six or more of HLA-C, CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and seven or more of HLA-C, and eight or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C nine or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C, ten or more of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C, or all eleven of CST7, PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, NKG7, GZMH, and HLA-C).

[0257] In some embodiments of either of the preceding two aspects, the one or more genes include a combination of a cytotoxic gene and a mitochondrial gene as set forth in Table 4 above.

[0258] In some embodiments of either of the preceding two aspects, the one or more genes include one or more of NKG7, CST7, GZMH, LITAF, CTSW, CCL5, and CD8A. For example, the one or more genes can include one, two, three, four, five, six, or all seven of NKG7, CST7, GZMH, LITAF, CTSW, CCL5, and CD8A.

[0259] For example, in some embodiments of either of the preceding two aspects, the one or more genes include two or more of NKG7, CST7, GZMH, LITAF, CTSW, CCL5, and CD8A. In some embodiments, the one or more genes include three or more of NKG7, CST7, GZMH, LITAF, CTSW, CCL5, and CD8A. In some embodiments, the one or more genes include four or more of NKG7, CST7, GZMH, LITAF, CTSW, CCL5, and CD8A. In some embodiments, the one or more genes include five or more of NKG7, CST7, GZMH, LITAF, CTSW, CCL5, and CD8A. In some embodiments, the one or more genes include six or more of NKG7, CST7, GZMH, LITAF, CTSW, CCL5, and CD8A. In some embodiments, the one or more genes include all seven of NKG7, CST7, GZMH, LITAF, CTSW, CCL5, and CD8A, hi some embodiments, the one or more genes consist of NKG7, CST7, GZMH, LITAF, CTSW, CCL5, and CD8A.

[0260] In some embodiments of either of the two preceding aspects, the expression level is a nucleic acid expression level, e.g., an mRNA expression level. The mRNA expression level may be determined, for example, by RNA-seq, RT-qPCR, qPCR, multiplex qPCR or RT-qPCR, microarray analysis, SAGE, MassARRAY techniques, FISH, or a combination thereof.

[0261] In some embodiments of either of the two preceding aspects, the expression level is detected in tumor cells, tumor-infiltrating immune cells, stromal cells, normal adjacent tissue (NAT) cells, or a combination thereof. In some embodiments, the sample is a tissue sample, a cell sample, a whole blood sample, a plasma sample, a serum sample, or a combination thereof. The tissue sample can be, for example, a tumor tissue sample, such as a tumor tissue sample containing tumor cells, tumor-infiltrating immune cells, stromal cells, NAT cells, or a combination thereof. In some embodiments, the tumor tissue sample is a formalin-fixed and paraffin-embedded (FFPE) sample, an archival sample, a fresh sample, or a frozen sample. In some embodiments, the tumor tissue sample is an FFPE sample.

[0262] In some embodiments of either of the preceding two aspects, the cancer is lung cancer, kidney cancer, bladder cancer, breast cancer, colorectal cancer, ovarian cancer, pancreatic cancer, gastric cancer, esophageal cancer, mesothelioma, melanoma, head and neck cancer, thyroid cancer, sarcoma, prostate cancer, glioblastoma, cervical cancer, thymic cancer, leukemia, lymphoma, myeloma, mycosis fungoides, Merkel cell carcinoma, or a hematological malignancy. For example, the cancer can be lung cancer, kidney cancer, bladder cancer, or breast cancer. In some embodiments, the lung cancer is non-small cell lung cancer (NSCLC). In some embodiments, the kidney cancer is renal cell carcinoma (RCC). In some embodiments, the bladder cancer is urothelial carcinoma (UC). In some embodiments, the breast cancer is triple-negative breast cancer (TNBC).

[0263] In some embodiments of either of the preceding two aspects, the PD-L1 axis binding antagonist is a PD-L1 binding antagonist, a PD-1 binding antagonist, or a PD-L2 binding antagonist. In some embodiments, the PD-L1 axis binding antagonist is a PD-L1 binding antagonist. The PD-L1 binding antagonist may inhibit the binding of PD-L1 to one or more of its ligand binding partners. In some embodiments, the PD-L1 binding antagonist inhibits the binding of PD-L1 to PD-1. In some embodiments, the PD-L1 binding antagonist inhibits the binding of PD-L1 to B7-1. In some embodiments, the PD-L1 binding antagonist inhibits the binding of PD-L1 to both PD-1 and B7-1.

[0264] In some embodiments, the PD-L1 binding antagonist is an antibody or antigen-binding fragment thereof. The antibody can be, for example, atezolizumab (MPDL3280A), YW243.55.S70, MDX-1105, MEDI4736 (durvalumab), and MSB0010718C (avelumab). In some embodiments, the PD-L1 binding antagonist is atezolizumab (MPDL3280A).

[0265] In some embodiments, the antibody comprises a heavy chain comprising the HVR-H1 sequence of SEQ ID NO: 41, the HVR-H2 sequence of SEQ ID NO: 42, and the HVR-H3 sequence of SEQ ID NO: 43, and a light chain comprising the HVR-L1 sequence of SEQ ID NO: 44, the HVR-L2 sequence of SEQ ID NO: 45, and the HVR-L3 sequence of SEQ ID NO: 46.

[0266] In some embodiments, the antibody comprises a heavy chain variable region comprising the amino acid sequence of SEQ ID NO:47 and a light chain variable region comprising the amino acid sequence of SEQ ID NO:48.

[0267] In some embodiments of either of the preceding two aspects, the PD-L1 axis binding antagonist is a PD-1 binding antagonist. The PD-1 binding antagonist may, for example, inhibit the binding of PD-1 to one or more of its ligand binding partners. In some embodiments, the PD-1 binding antagonist inhibits the binding of PD-1 to PD-L1. In some embodiments, the PD-1 binding antagonist inhibits the binding of PD-1 to PD-L2. In some embodiments, the PD-1 binding antagonist inhibits the binding of PD-1 to both PD-L1 and PD-L2.

[0268] In some embodiments, the PD-1 binding antagonist is an antibody or antigen-binding fragment thereof. The antibody can be, for example, MDX-1106 (nivolumab), MK-3475 (pembrolizumab), MEDI-0680 (AMP-514), PDR001, REGN2810, and BGB-108.

[0269] In some embodiments, the PD-1 binding antagonist is an Fc fusion protein, such as AMP-224. [Brief explanation of the drawings]

[0270] [Figure 1A] 1 is a chart showing statistics for the six-sample dataset described in Example 1 after bioinformatics filtering. Samples were obtained from the tumor (T) and adjacent normal (N) compartments of three patients, designated by labels T1 through N3 as described in Example 1. Total counts are provided for the number of sequenced transcripts based on RNA sequencing (column labeled "Transcripts"), cells with gene expression data based on RNA sequencing ("Cells"), and cells with clonotype assignments based on TCR sequencing ("Types"). The right-most column displays a Venn diagram showing the number of distinct clonotypes unique to and shared between each patient's tumor and NAT compartments. [Figure 1B]1 is a graph showing the proportion of TCR clonotypes in samples obtained from patients described in Example 1 by clonal residency pattern. Percentages are normalized separately for each patient ("Pat") 1 through 3. Each proportion is represented by the length of the bar region, which is stacked from left to right and shaded according to the legend above to show the proportion of sequenced TCR clonotypes with a given observed cell number (singleton or multiplet) and the location of that cell. "NAT" and "tumor" indicate all cell residency in their corresponding compartment only, while "dual residency" indicates cells found in both compartments. [Figure 1C] Scatter plot of T cell clones by clonal residency pattern. Each symbol represents a distinct clonotype, with a different symbol used for each patient. The x and y coordinates of each clonotype are plotted on a logarithmic scale corresponding to the observed clone count (number of cells with a given clonotype) in the NAT and tumor, respectively, with random jitter added to represent the number of dots at each individual coordinate. Various locations on the graph are shaded using a two-dimensional palette based on the NAT and tumor clone counts shown in the upper right. Rounded rectangles and polygons indicate grouping of clones as NAT multiplets, tumor multiplets, and dual-resident clones based on the number and location of clone cells. Labels and arrows indicate clones that are NAT or tumor singletons. [Figure 1D] 1A is a bar graph showing the proportion of cells by clonal residency pattern. The bar graph follows the same format as FIG. 1A, but the proportions are based on cells rather than distinct clonotypes. Thus, expanded clones with multiple cells have a proportionally larger proportion. Cells derived from dual-resident clones can be further distinguished by their location in the NAT or tumor, giving rise to two distinct categories. [Figure 2A]Figure 1C shows a chart illustrating the association between clonal residency patterns and gene expression. Cells from each clonal residency pattern are plotted separately on a common tSNE map calculated from gene expression data for all T cells in the dataset. The tSNE map represents each T cell as a dot, locating cells with similar gene expression and cells with significantly different gene expression. Each clonal residency pattern is represented by a separate plot organized into a 3x2 grid based on cell location and clone number. For each of the six plots, all cells from the dataset are plotted against the same background, and cells from each clonal residency pattern are shaded using the two-dimensional palette in Figure 1C. These plots show that clonal residency patterns occupy distinct regions of the tSNE map, indicating potential gene expression associations. [Figure 2B] Figure 2B shows a series of plots depicting the expression of selected marker genes for T cell subsets. Sixteen plots are shown, each representing the expression of a gene on the same tSNE map as Figure 2A. After log-transformation and normalization by Seurat, each cell is shaded according to the expression of a given gene on a continuous quantile-based scale, with light gray indicating the lowest expression and darker gray indicating higher expression in the dataset. The shaded ellipses and black lines on each tSNE map provide uniform landmarks for the T cell subsets and cluster divisions by CD8 and CD4 expression. These landmarks and the T cell subsets indicated are shown in more detail in Figure 2C. [Figure 2C]Figure 1 shows a plot showing the clustering of T cells into subsets by gene expression. The clustering of cells is indicated on the tSNE map by an arbitrary shading assigned to each cluster and a shaded ellipse approximating each cluster's area. Each cluster is named according to its overall gene expression or characteristic marker genes shown in the labels, which are also shown in some labels. Labels are shown to indicate their designation and provide a name for each cluster based on its gene expression. Additional labels and lines indicate the grouping of subclusters for natural killer T (NKT) cells and T-resident memory (Trm) cells. Black lines indicate the division of clusters based on CD8 or CD4 expression. Some clusters contain both CD8+ and CD4+ cells and are designated CD3+. [Figure 2D] 2C is a graph showing the clonal residency patterns of each T cell subset. Each T cell subset in FIG. 2C is characterized by the proportion of its cells that belong to approximately each clonal residency pattern. The proportion of cells is represented by the length of each bar area and is shaded according to the legend above. T cell subsets are shown to differ in the distribution of their clonal residency patterns. [Figure 2E] Figure 2 shows plots depicting various clonal residency patterns. The same tSNE plots as in Figure 2D are shown, but each cell is shaded according to its tumor and NAT clone size using the two-dimensional palette in Figure 1C, also shown in the upper right of this figure. The shaded ovals, black lines, and cluster labels in Figure 2D are also plotted to provide landmarks. [Figure 3A]Figure 1 shows a series of scatter plots of clonal numbers and residency for each T cell cluster described in Example 1. Each plot is a subset of the data from Figure 1C for each T cell cluster in Figure 2C. The space of tumor and NAT clone numbers is divided into four regions indicated by gray lines: a tumor-only region containing clones with zero NAT clone numbers; a NAT-only region with zero tumor clone numbers; a tumor-dominant region with non-zero tumor and NAT numbers and tumor numbers equal to or greater than the NAT numbers; and a NAT-dominant region with non-zero tumor and NAT numbers and NAT numbers greater than the tumor numbers. The numbers in each region indicate p-values ​​based on a one-sided Poisson test, where the alternative hypothesis is that the number of clones observed in a given T cell subset is greater than that predicted from the mean values ​​observed across all T cell subsets. p-values ​​less than 0.05 are shown in red and are used to classify T cell subsets with different clonal expansion behaviors. [Figure 3B] Schematic of clonal expansion behavior. This plot summarizes the clonal expansion behaviors shown in Figure 3A. Clones are represented by dots using the same technique as Figure 1C. Each clonal expansion behavior is represented by a label indicating the T cell subset involved, and the shaded areas indicate the clones for each behavior. Lines indicate the boundaries between different types of clonal expansion: tumor-only, NAT-only, tumor-dominant, and NAT-dominant (defined in Figure 3A). [Figure 4A-B]Figure 4A is a plot showing the matching of cellular TCRs to known virus-reactive CDR3 sequences from the VDJdb, as described in Example 1. Each T cell cluster in Figure 2C is characterized by the percentage of cells with CDR3 amino acid sequences that match CDR3 sequences in the VDJdb that react to known epitopes from commonly occurring viruses (cytomegalovirus, Epstein-Barr virus, and influenza A). For each T cell subset, the number of cells assigned to that T cell subset is divided by the total number of cells with a clonotype. Cells are further divided according to their clonal number and residency, as indicated by the shaded bar regions according to the legend above. Figure 4B is a plot showing the matching of cellular TCRs to CDR3 sequences from multiple cancers from The Cancer Genome Atlas (TCGA). Matching was performed between the CDR3 amino acid sequence from the dataset obtained in Example 1 and a set of 716,720 TCR sequences collected from TCGA as described in Li, B. et al. Nat. Genet. 48,725-732,2016 (the entire disclosure of which is incorporated herein by reference). Matches were counted if the CDR3 amino acid sequence occurred in three or more different tumor types in TCGA, suggesting reactivity to viral antigens rather than tumor antigens. The bar graph follows the same method as in Figure 4A. [Figure 4C]This is a heatmap showing the T cell composition of clonotypes. Each column in this heatmap represents distinct clones with at least 10 total observations. The shading within the heatmap indicates the proportion of cells of that clone relative to the total T cell subtype. Some T cell subtypes are divided by their location in the NAT (N) or tumor (T) to reveal additional information. Clones are grouped by similarity to each other to show distinct mixtures of T cell clusters, highlighted by bold rectangles, and labeled at the bottom with the seven mixtures of T cell subtypes observed. The shading at the top of the heatmap indicates the number and persistence of clones for each clonotype, using the scheme in Figure 1C. The mixture shading and corresponding legend at the bottom of the heatmap indicate the clonal expansion behavior within each mixture, supported by the range of patterns of clonal number and persistence at the top of the heatmap. [Figure 4D-F] Figure 4D shows a graph depicting the relationship between clonal residency of 8.2-Tem cells and their differentiation into 8.3-Trm cells. Each circle represents a highly expanded clonotype containing at least one 8.2-Tem cell, with x and y coordinates corresponding to the number of 8.2-Tem cells in the NAT and tumor, respectively, and is shaded according to the two-dimensional palette in Figure 1C. Random jitter has been added to the integer values ​​of the x and y coordinates to indicate multiple occurrences of the same number of points. The size of each circle indicates the total number of 8.3-Trm cells (8.3a-Trm + 8.3b-Trm + 8.3c-Trm) in the clonotype, and a scale is provided below the plot. Figure 4E shows a graph depicting the relationship between clonal residency of 4.3-activated cells and the expansion of 4.1-Trm cells. The same plot as Figure 4D is shown, except for clones containing at least one 4.3-activated cell, and the size of the circle indicates the total number of 4.1-Trm cells for each clone. Figure 4F shows the relationship between clonal residency of 4.2-activated cells and the number of 4.4 and 4.5 cells. The same plot as in Figure 4D is shown, except for clones containing at least one 4.3-activated cell, with the size of the circle indicating the total number of 4.4-HSP, 4.5t-Ribo, and 4.5n-Ribo cells. [Figure 4G] 4C is a graph showing mitotic activity in highly expanded clonotypes. Each mixture of T cell subtypes shown in the bottom of Figure 4C is characterized by the proportion of highly expanded clones with at least one detail assigned to the 3.2-mitotic cluster in tumor (T), NAT (N), or both. [Figure 5A] 2C is a plot showing the proportion of cells across T cell subtypes as described in Examples 1 and 2. Each patient ("Pat") is characterized by the proportion of its cells assigned to the different T cell subtypes in FIG. 2C. The proportions are represented by the length of each bar area and are shaded according to the legend above. The T cell subtypes with the greatest proportions are also labeled, space permitting. [Figure 5B]Figure 1 shows a plot of gene set enrichment analysis (GSEA) applied to clinical trial data. A signed GSEA analysis was performed on the 1,101 most variable genes from the Seurat-based analysis described in Example 2. These genes were ranked for each T cell cluster, as indicated by a white-to-black gradient, with up-regulated genes (fold change >1) on the left and down-regulated genes (fold change <1) on the right, with genes with the smallest (most significant) p-values ​​at the extremes of the ranking. Highly variable genes were also evaluated for positive and negative associations with overall survival based on bulk RNA-seq expression from the POPLAR phase II clinical trial of patients with non-small cell lung cancer (NSCLC). For each T cell cluster, genes with a positive association with clinical response genes are indicated by a red checkmark (atezolizumab response) and a blue checkmark (docetaxel response) at the corresponding rank among the up-regulated genes. Genes negatively associated with clinical response are indicated by a black check mark at the corresponding rank among down-regulated genes. Statistical tests for distribution differences were performed separately for atezolizumab and docetaxel using the enrichment score method from Connectivity Map (Lamb, J. et al. Science 313, 1929-1935, 2006) compared to distributions based on 100,000 random permutations, and the results are presented as p- and q-values ​​corrected for multiple hypothesis testing, or local false discovery rates. The sample size is given as "n1" for each statistic and indicates the crossover number of genes associated with clinical response. In both cases, n2 = 1101 genes associated with cluster membership. T cell clusters with statistically significant results (q<0.05) for atezolizumab are highlighted with gray boxes. [Figure 5C]5B shows a plot of the ranking of clinically relevant genes in each T cell subset. This heat map shows genes and their ranks in each T cell subset calculated in the gene set enrichment analysis shown in FIG. 5B. These values ​​correspond to the check marks in FIG. 5B, which represent genes that showed a statistically significant association with response to atezolizumab treatment. Each column represents a distinct T cell subtype, and the shading indicates the rank of the gene, which was based on the p-value of the differential expression of the gene in cells in that T cell subtype compared to all other cells. Gray cells indicate that the gene did not show a positive log-fold change for that T cell subset. The average rank was calculated for each gene in that T cell subset with a positive log-fold change, and the genes were sorted from top to bottom by ascending average rank. The average rank is also shown by the plot to the right of the heat map. [Figure 5D-I] 5A and 5B are plots showing the association between expansion mode and clinical response in NSCLC. Kaplan-Meier curves are shown for both overall survival (OS) and progression-free survival (PFS) in the POPLAR clinical trial by treatment group (atezolizumab, upper limit hazard ratio and p-value, n=93 patients; docetaxel, lower limit hazard ratio and p-value, n=100 patients) and signature expression (solid line: high; dashed line: low; split at median) for gene signatures of tumor-resident multiplets (Figures 5F and 5G), NAT-resident multiplets (Figures 5H and 5I), and dual-resident clones (Figures 5D and 5E). [Figure 5J-O] Forest plots showing hazard ratios for OS (top panel) and PFS (bottom panel) at various cutoffs for the double (Figures 5J and 5K), tumor (Figures 5L and 5M), and NAT (Figures 5N and 5O) expansion signatures in the single-arm IMvigor210 clinical trial of atezolizumab in urothelial carcinoma (n=354 patients). Whiskers represent 95% confidence intervals. [Figure 6]This is a heatmap of genes and their expression in highly expanded clones grouped according to clonal expansion mode. Each column in this heatmap represents the average gene expression of a single clonotype, using highly expanded clones with a count of 10 or more in the NAT (NAT-resident multiplets), tumor (tumor-resident multiplets), or both (dual-resident clones). Dual-resident clones are further divided into cells located in the NAT and cells located in the tumor. Columns for NAT and tumor singleton modes were constructed by randomly pooling 20 singleton cells and calculating their average gene expression. Regions of high gene expression are enclosed in boxes and labeled categories A through H with a brief description. [Figure 7] 1 is a heat map showing the intercorrelation of expression levels of the genes shown in Table 7 of Example 2. The heat map indicates the presence of two subsets of genes within the panel shown in Table 7 that exhibit particularly high intercorrelation. The first subset includes five mitochondrial genes: MT-ND4L, MT-ND4, MT-ATP6, MT-CO2, and MT-CYB. The second subset includes 11 cytotoxic genes: PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, CST7, NKG7, GZMH, and HLA-C. Correlation coefficients are indicated by a continuous shading ranging from 0 to approximately 1.00. [Figure 8A-N]8A and 8B , respectively. Figure 8B shows the association of expression levels of specific cytotoxic and mitochondrial genes with clinical response in NSCLC. For both OS and PFS in the POPLAR clinical trial, Plan-Meier curves are shown for the following genes by treatment group (atezolizumab, upper bound hazard ratio and p-value, n = 93 patients; docetaxel, lower bound hazard ratio and p-value, n = 100 patients) and expression level (solid line: high; dashed line: low; split at median): CST7 ( Figures 8A and 8B ), MT-ND4 ( Figures 8C and 8D ), combined expression of CST7 and MT-ND4 ( Figures 8E and 8F ), MT-CYB ( Figures 8G and 8H ), combined expression of CST7 and MT-CYB ( Figures 8I and 8J ), MT-CO2 ( Figures 8K and 8L ), and combined expression of CST7 and MT-CO2 ( Figures 8M and 8N ). [Figure 9A-F] 1 is a plot showing the association of expression levels of cytotoxic and mitochondrial gene panels with clinical response in NSCLC. Kaplan-Meier curves are shown for the following gene panels by treatment group (atezolizumab, upper bound hazard ratio and p-value, n=93 patients; docetaxel, lower bound hazard ratio and p-value, n=100 patients) and expression level (solid line: high; dashed line: low; split at median) for both OS and PFS in the POPLAR clinical trial: cytotoxic genes (PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, CST7, NKG7, GZMH, and and HLA-C, Figures 9A and 9B), mitochondrial genes (MT-ND4L, MT-ND4, MT-ATP6, MT-CO2, and MT-CYB, Figures 9C and 9D), and combined expression of cytotoxic and mitochondrial genes (PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, CST7, NKG7, GZMH, HLA-C, MT-ND4L, MT-ND4, MT-ATP6, MT-CO2, and MT-CYB, Figures 9E and 9F). [Figure 10A-J]Figure 1 shows plots showing the clonal numbers and resident types of T cell clones, as described in a recent report by Guo, X. et al. Nat. Med. 24,978 (2018), the disclosure of which is incorporated herein by reference. The clustering analysis described for the three patients analyzed in Example 1 below was applied to the Guo et al. dataset, which included various samples from tumors, NATs, and peripheral blood from 14 patients. Each plot represents a T cell subset from this framework, determined by representative genes, along with corresponding clusters from other datasets. This plot is similar to the plots in Figures 1C and 3A, with NAT and tumor clone numbers represented by coordinates on the x and y axes, respectively, and shaded according to the two-dimensional palette shown at the bottom. Peripheral blood clone numbers are represented herein by the size of each dot, according to the respective legends. 4 [Figure 11A-D] Figure 11 shows the association between expansion mode and clinical response in NSCLC using T cell gene signatures derived from the dataset of Guo, X. et al. Nat. Med. 24,978 (2018). Kaplan-Meier curves are shown for both overall survival (OS) and progression-free survival (PFS) in the POPLAR clinical trial by treatment group (atezolizumab, upper limit hazard ratio and p-value, n = 93 patients; docetaxel, lower limit hazard ratio and p-value, n = 100 patients) and signature expression (solid line: high; dashed line: low; split at median) for the gene signatures of dual resident clones (Figures 11A and 11B) and tumor-resident clones (Figures 11C and 11D). [Figure 11E-H]Forest plots showing OS (top panel) and PFS (bottom panel) hazard ratios at various cutoffs using T cell gene signatures derived from the dataset from Guo, X. et al. Nat. Med. 24,978 (2018). The plots show threshold-dependent hazard ratios for the dual resident T cell gene signature (Figures 11E and 11F) and tumor resident T cell gene signature (Figures 11G and 11H) in the single-arm IMvigor210 clinical trial of atezolizumab in urothelial carcinoma (n = 354 patients). Whiskers represent 95% confidence intervals. [Figure 12A-I]Plots showing that dual clonal expansion is associated with peripheral clonal expansion. (Figure 12A) Clonal expansion scatter plots for the NAT and tumor compartments. Each patient is represented by a separate scatter plot, with each dot representing a distinct clonotype. The x and y coordinates of the scale correspond to the logarithm ((1 + clonotype cell count) / (total cells)) in the NAT and tumor compartments, respectively, allowing for the representation of zero counts. These coordinates indicate the relative proportion of cells of that clonotype in each compartment. Dots are colored using the two-dimensional palette at the bottom right, with blue increasing with increasing NAT count, pink increasing with increasing tumor count, and purple increasing with increasing counts in both compartments. Clonotypes with only one observed clone, i.e., singletons, are colored yellow (1 NAT clone) or orange (1 tumor clone). Vertical and horizontal gray lines indicate the boundary between absence (0 cells of that clonotype) and presence (1 or more cells) within each compartment, which is used to define clonal expansion patterns. The gray diagonal line indicates equal cell proportions in the two compartments. The numbers in each title indicate the degree of parallel expansion as measured by the Pearson correlation coefficient of doubly expanded clones on a log-log scale; patients are ordered by decreasing coefficient. (Figure 12B) Clonal expansion scatter plots for the tumor, NAT, and blood compartments. These plots are similar to those in (Figure 12A), but provide data for four patients with blood scTCR-seq samples. Green and brown colors have been added to indicate blood singletons and multiplets, respectively; however, the predominance of blood singletons means that blood multiplets are rarely observed. The number of blood cells observed for each clone is represented by the size of each dot, according to the legend at the bottom. As in (Figure 12A), the numbers in each title indicate the degree of parallel expansion, and patients are ordered by decreasing coefficient. (Figure 12C) Venn diagram of cells by compartment. Each patient with a blood sample is represented by a Venn diagram that matches the corresponding scatter plot in (Figure 12B).Each Venn diagram contains a central ellipse, colored according to the legend on the right, for each of the five tissue-based clonal expansion patterns. These ellipses are further divided by larger upper and lower ellipses to represent clonotypes present in the blood, with single cells in the blood (blood non-clones) indicated by the upper dark green ellipse and multiple cells in the blood (blood-expanded clones) indicated by the lower brown ellipse. Thus, cells without representative cells are represented by the number in the central section, which does not overlap with the larger ellipse. Additionally, cells from clonotypes present only in the blood, but not in tumor or NAT, are represented by the area within the larger ellipses for blood singletons (light green areas) and blood multiplets (light brown areas). The numbers to the right of each Venn diagram indicate the sum of the upper and lower ellipses, which is the total number of cells from non-blood-expanded and expanded clonotypes. (Figure 12D) Blood expansion. Each bar corresponds to the Venn diagram shown above, showing the percentage of cells representing non-blood-expanded clonotypes (upper oval) and expanded clonotypes (lower oval) as green and brown bars, respectively. The numbers above each bar indicate the number to the right of each Venn diagram used to calculate the percentage. (Figure 12E) Infiltration of tumor / NAT by blood-expanded clones. Each bar shows the distribution of blood-expanded clonotypes (lower oval) into five tissue-based clonal expansion patterns, plus blood multiplets colored according to the legend to the right of (Figure 12C). The numbers above each bar indicate the number from the Venn diagram used to calculate the percentage, as space permits. (Figure 12F) Presence of tumor / NAT TCRs in blood-expanded clones. Each bar shows the percentage of cells from each tissue-based clonal expansion pattern that share a TCR with the blood-expanded clonotype, i.e., the lower oval in (Figure 12C). Bars are color-coded according to the legend on the right (Figure 12C) and are further labeled with the following abbreviations: n = NAT singleton, N = NAT multiplet, D = double expansion, T = tumor multiplet, and t = tumor singleton. The numbers above the double expansion pattern in each bar indicate the number from the Venn diagram used to calculate the percentage. (Figure 12G) Correlation of TCR repertoires between blood, tumor, and NAT.Each dot represents a clone from four patients with blood samples. Absolute clone size in blood relative to clone size in tumor (left) or NAT (right) is plotted on a log-log scale and color-coded by clonal expansion pattern using the two-dimensional palette on the right. To allow for the display of zeros, a value of 1 is added to each clone size. Gray horizontal and vertical lines separate zeros from ones. (Figure 12H) Confirmation of clone size correlation with an external dataset. The same plot as (Figure 12G) is presented using the dataset from Guo et al., 2018. (Figure 12I) Association between blood expansion and parallel tissue clonal expansion. Each dot represents a patient from the Guo et al., 2018 dataset, and the analysis is summarized in Figure 21. The x-coordinate represents the degree of blood expansion, calculated as the percentage of cells in blood-expanded versus non-expanded clonotypes. The y-coordinate is measured as the Pearson correlation coefficient of the double-expanded clonotypes, which is the degree of parallel tumor / NAT clonal expansion weighted by (1 + clone size in blood). Only 10 of the 14 patients in the dataset had sufficient sampling to calculate the y-coordinate; two patients had no sampling in the NAT, and two patients had only one double-expanded clone. Statistical testing using Spearman correlation yielded a p-value of 0.04, shown in the lower right corner. [Figures 13A-J]Plots demonstrating that T cell subtypes are associated with clonal expansion patterns. (Figure 13A) Two-dimensional mapping of gene expression. This plot shows each T cell from the dataset plotted according to the first two components, x and y coordinates, respectively, from the dimensionality reduction of single-cell gene expression data. A distinct color was selected for each cluster, and cells are colored according to their cluster assignment. Ellipses and labels were manually added to approximate cluster areas and provide biological meaning. The curve through the center of the figure indicates the boundary between CD8+ (left) and CD4+ (right) T cells. (Figure 13B) Comparison with external gene signatures of T cell subtypes. A normalized cross-tabulation of cells between cluster assignments from this study (shown in columns) and those provided by external gene signatures from Guo et al., 2018 (top panel) and Yost et al., 2019 (bottom panel) (shown in rows) is shown. The intensity of the displayed heatmap indicates the frequency with which cells fell into both sets of clusters. Columns represent clusters from this study and are colored according to the scheme in (Figure 13A). (Figure 13C) Selected biomarkers. Box plots display the distribution of expression of selected genes across clusters obtained from scaled data from the integrated assay calculated using Seurat. Boxes indicate the interquartile range, with the median indicated by a line across the middle. Whiskers indicate an additional 1.5-fold interquartile range from the median, limited by the range of the data. The range for each gene was selected to include the interquartile range of all box plots. Each box plot is colored according to the scheme in (Figure 13A). (Figure 13D) Stem-like exhaustion versus terminal exhaustion. The top figure shows gene expression of PDCD1 (PD-1) overlaid on the map in (Figure 13A) using a continuous coloring scheme for number quantiles from the SCT assay calculated using Seurat, along with the source of the cell: tumor (red), NAT (blue), or blood (green). The bottom figure shows published gene signature score values ​​for stem-like exhaustion vs. terminal exhaustion (Im 2015). The cluster ellipses in (Figure 13A) are provided as reference. (Figure 13E) Tumor and NAT origin.Cells from the tumor and NAT compartments are plotted on the map from (Figure 13A) and colored red and blue, respectively. The cluster ellipses from (Figure 13A) are provided as a reference. The overall percentage of tumor cells is indicated in the legend, and the percentage of tumor cells in each cluster is provided for all 14 patients. (Figure 13F) Comparison of resident memory T cells. Simple statistics are provided for clusters 8.3a, b, and c, representing resident memory T cells, showing the numbers from (Figure 13D) and (Figure 13E) for each cluster. The bar graph is colored using the scheme from (Figure 13A). (Figure 13G) Blood expansion. Cells from the blood compartment are plotted on top from (Figure 13A) and are colored in a green gradient for a blood count of 1 and a brown gradient for blood counts increasing above 1. Cells from the tumor and NAT compartments are not plotted. Percentages are not provided due to the high degree of variability among the four patients, as shown in Figure 7D. (Figure 13H) Clonal expansion patterns by cell. Each cell derived from the tumor or NAT is plotted on the map in (Figure 13A) and colored according to its parent clone's tissue-based clonal expansion pattern using the two-dimensional palette in the lower left. The oval in (Figure 13A) is provided as a reference. (Figure 13I) Clonal expansion patterns by subtype. Each dot represents a clone synthesized from four patients with blood samples plotted according to its absolute cell number in the NAT and tumor using a logarithmic scale. Clone size in the blood is indicated by the size of the dot according to the legend at the bottom. Clones were classified according to their major cell cluster and placed on separate plots colored using the scheme in (Figure 13A). Clones from the 8.3a, 8.3b, and 8.3c clusters were synthesized on a single plot but colored separately according to their cluster. (Figure 13J) Clonal expansion patterns by patient. Clones derived from four patients with blood samples are shown using the same representation as in Figure 12B. As shown in the figure, the x and y coordinates indicate the percentage of cells in the NAT and tumor, respectively, and the size of the dot represents its absolute cell number in the blood using the legend at the bottom.However, in this case the clones are colored by their predominant cell cluster instead of their clonal expansion pattern, using the scheme in (Figure 13A). [Figure 14A-D]Plots showing accumulation of non-exhausted CD8+ T cells in tumors from peripheral blood (Figure 14A). TCR plots correlate between novel post-treatment CD8+ clonotypes and pre-treatment blood. Scatter plots are shown for six patients from Yost et al., 2019, with matching clones in pre-treatment and post-treatment tumors and blood-derived pre- or post-treatment bulk TCR-seq samples. Novel post-treatment clonotypes with the CD8+ subtype as their predominant post-treatment cell cluster were identified by their occurrence in the post-treatment scTCR-seq assay (but not in the pre-treatment scTCR-seq assay) and plotted according to their clonal size in pre-treatment blood from bulk TCR-seq (post-treatment blood was used for patient bcc.su002 out of necessity) and post-treatment tumor from scTCR-seq. Clones are colored according to their predominant cell cluster in the post-treatment tumor as determined in external analyses. A vertical gray line separates clones with a clone size of 0 in the blood from clones with a clone size of 1 or greater, thereby separating novel clones into non-blood-associated clones on the left and blood-associated clones on the right. Patients are ordered according to the total number of cells from novel CD8+ clonotypes that share TCRs with the blood, as indicated by the numbers at the bottom. (Figure 14B) TCR repertoires in blood before and after treatment. Scatter plots of pre- and post-treatment clone sizes in blood are shown for the six patients in (Figure 14A). The coordinates represent the number of templates observed for each TCR beta chain. For patient bcc.su002, who did not have a pre-treatment blood sample, a one-dimensional strip chart is shown instead, with horizontal jitter added to display all points. The numbers below each plot indicate the pre- and post-treatment clonal diversity of each distribution of clone sizes, measured as the Shannon entropy for all clone sizes ≥ 10; the threshold was chosen to reduce the effect of noise from low-level sampling. Additionally, increasing amounts of clonal diversity can be visually observed as increasing presence of clones along the main diagonal line from lower left to upper right.(Figure 14C) Tracking of CD8+ T cell clones in tumors. For each patient (Figure 14A), clones were selected for the CD8+ subtype as their predominant cell cluster in the tumor before or after treatment. These clones were separated according to whether they shared a TCR in pretreatment blood (or posttreatment blood for patient bcc.su002, who lacked a pretreatment blood sample). Blood-associated novel clones, i.e., those that shared a TCR with the blood, are plotted in the upper panel by a plot titled "Blood+." Non-blood-associated novel clones, i.e., those that did not share a TCR with the blood, are plotted in the lower panel by a plot titled "Blood-." Within each plot, clones were further classified according to their pretreatment ("Pre-rx") or posttreatment ("Post-rx") measurements as lost (present before treatment but absent in posttreatment scRNA-seq), residual (present in both), or novel (present after treatment but absent in pretreatment scRNA-seq), abbreviated as "lost," "residual," and "novel," respectively. The y-coordinate indicates the size of each clone in the tumor before or after treatment, and is colored according to its pre- or post-treatment cluster assignment, as appropriate, as determined by the original paper. Matching between pre- and post-treatment clonotypes in the remaining category is connected by a gray line. The average ratio of clone size in the post-treatment condition to the pre-treatment condition in the remaining category was calculated for the blood-matched and non-blood-matched categories and is shown as a number below each pair of plots. (Figure 14D) Distribution of cell clusters for novel CD8+ clonotypes due to tumor expansion and TCR sharing with blood. Blood-associated and non-blood-associated novel clones were identified in tumors with CD8+ clonotypes and were further separated into non-expanding (tumor clone size = 1) or expanding (tumor clone size > 1) clones, abbreviated as "non-expanding" and "expanding," respectively, in the figure. For each category, cells belonging to the clones were tabulated according to their cluster assignment based on the original analysis, and the distribution is shown in a bar graph and colored according to the legend below. The legend also indicates the grouping of clusters into exhausted and non-exhausted categories. [Figure 15A-D]Plots showing the clinical association of CD8+ clusters. (Figure 15A) Gene expression profiles of CD8+ clusters across patients. Heat maps show the gene expression profiles of each gene (rows) in each patient cluster (columns) for the CD8+ clusters based on the unsupervised analysis in Figure 13A. Genes in each signature are grouped into rows and selected by logistic regression as the 20 with the highest signed deviance. Genes replicated in multiple signatures were assigned lower ranks. Cells from each patient were grouped by their cluster, and the average log fold change was calculated for each group relative to all other CD8+ clusters for that patient. Each cell in the heat map shows the resulting log fold change, colored using the palette below. (Figure 15B) Gene expression profiles of CD8+ clusters across clonal expansion patterns. Same representation as (Figure 15A), except that each patient's cells were grouped according to the clonal expansion patterns of their parent clones. (Figure 15C) Forest plot of gene signatures for CD8+ clusters. Each dot represents a gene from a 20-gene signature for the CD8+ cluster, and the x-coordinate indicates the hazard ratio for progression-free survival (PFS) in each treatment arm of the clinical trial, shown as a separate table. Vertical jitter was added to allow visualization of all dots. A hazard ratio of less than 1 indicates a favorable association with survival. The numbers to the right of each signature indicate the gene's average hazard ratio (HR) and one-sided p-value for the null hypothesis that the hazard ratio is 1 or greater, adjusted for multiple hypothesis testing using the Benjamini-Hochberg procedure. Statistical results yielding an adjusted p-value <0.05 are colored red. (Figure 15D) Kaplan-Meier survival curves. Kaplan-Meier curves are plotted for the 8.1-Eff (left) and 8.2-Tem (right) signatures, with patients dichotomized by signature scores above (red) or below (black) their corresponding median values ​​in each trial. Each curve shows progression-free survival (PFS) on the x-axis and the proportion of patients surviving on the y-axis. A plus sign on each survival curve indicates a censored event.The legend for each plot indicates the hazard ratio (HR) and p-value for the Cox proportional hazards model comparing the two patient populations, with p-values ​​<0.05 marked with an asterisk. [Figures 16A-F]Plots showing sample statistics and clonal expansion patterns. (Figure 16A) Represents CD3- and CD45-selected samples across clusters. Because the immune cell samples in the study were selected for CD3 (expected to contain only T cells) or CD45 (including both T and non-T cells), computational separation of T and non-T cells was necessary. The T cell separation process should result in clusters containing cells from both CD3- and CD45-selected samples. The bar graph shows the distribution of samples among the T cell clusters obtained from the unsupervised clustering analysis, with expected representation by CD3- and CD45-selected samples colored red and blue, respectively. (Figure 16B) T cell mixing across patients. A two-dimensional map of all cells corresponding to the dimensionality reduction calculation in Figure 13A is shown. Each cell is colored by its patient of origin. Patients are observed to be well mixed across the map, indicating that the individual datasets are well integrated. (Figure 16C) Sample statistics. Statistics are provided for cells in the dataset after separation into T and non-T categories. The presence of CD45-selected samples allowed us to identify contaminating non-T cells in CD3-selected samples and explain the number of non-T cells in those samples. Patients were derived from four cancer types: non-small cell lung cancer (Lung), endometrial adenocarcinoma (Endo), colorectal adenocarcinoma (Colon), and renal clear cell carcinoma (Renal). Numbers indicate the total number of transcripts and cells from scRNA-seq and the number of cells with clonotypes from scTCR-seq. Cells were grouped into distinct clonotypes, and the number of distinct clonotypes is shown for each sample. (Figure 16D) TCR sharing between compartments. Clonotypes were matched between compartments for each patient. Venn diagrams for each sample are presented adjacent to the corresponding column in (Figure 16C). Each Venn diagram shows the number of distinct clonotypes unique to each compartment or shared between compartments (N = NAT, T = tumor, B = blood). (Figure 16E) Distribution of clones by clonal expansion pattern. Each clone was classified into one clonal expansion pattern based on its clone size in the tumor and NAT compartments.NAT singletons were clones with a NAT cell count (or clone size) of 1 and a tumor clone size of 0; NAT multiplets had a NAT clone size of 2 or greater and a tumor clone size of 0. Conversely, tumor singletons were clones with a tumor clone size of 1 and a NAT clone size of 0; tumor multiplets had a tumor clone size of 2 or greater and a NAT clone size of 0. Doubly expanded clones had a clone size of 1 or greater in both the tumor and NAT. The bar graphs show the distribution of clones by this classification in each patient and are colored according to the legend below. While the majority of clonotypes were singletons, 9-18% of clonotypes in each patient were multiplets or doubly expanded clones. (Figure 16F) Distribution of cells by clonal expansion pattern. Each T cell in the NAT (left) and tumor (right) was classified according to the clonal expansion pattern of its parent clone. The bar graphs show the distribution of cells by this classification in each patient and are colored according to the legend below. In some cases, cells from double-expanded clones account for a proportion of cells in the tumor and NAT, constituting the majority of T cells. [Figure 17]This plot shows the association between parallel tumor / NAT expansion and peripheral clonal expansion in an external dataset. Each of the 14 non-small cell lung cancer patients from Guo et al., 2018, is shown in a scatterplot, with each clonotype represented by a dot plotted by clone size in the NAT and tumor. The size of the dot represents the clone size in the blood according to the legend in the lower right. The color of each dot is based on a two-dimensional palette in the lower right. Each patient is also characterized by the degree of blood expansion, the infiltration of blood-expanded clones into the tumor / NAT, and the presence of tumor / NAT TCRs in the blood-expanded clones, using the same representation and color scheme as in Figures 12D-12F. Patients are ordered by the amount of blood expansion. We can see a tendency for dually expanded clones to expand more highly in the blood and for these clones to be located along the main diagonal, reflecting the parallel mode of dual expansion. Patients with higher levels of blood expansion have a tendency for overall parallel clonal expansion in the tumor and NAT, summarized in Figure 12I. [Figures 18A-E]Plots showing T cell subsets and clonal expansion behavior. (Figure 18A) Comparison of T cell subtypes with external gene signatures from Zhang et al., 2018. A normalized cross-tabulation of cells between cluster assignments from this study (shown in the columns) and those provided by external gene signatures from Zhang et al., 2018 is shown. The intensity of the displayed heatmap indicates the frequency with which cells were classified into clusters from both sets. Columns represent clusters from this study and are colored according to the scheme in (Figure 18A). The calculation and representation are the same as in Figure 13B, except for the different sets of reference gene signatures. It can be seen that there is a relationship between the 8.1-Eff cluster from this study and the CD8_C3-CXCR1 cluster from the external dataset, and between 8.2-Tem and CD8-C4-GZMK. (Figure 18B) Selected gene expression for CD4+ subsets. Expression for selected genes indicates differences between CD4+ subsets. The representation is the same as in Figure 13C, except that genes were selected for high expression in the CD4+ subset. (Figure 18C) Exhaustion biomarkers. Gene expression values ​​are plotted on the map in Figure 13A for several genes associated with T cell exhaustion. The representation is the same as in Figure 13D (top panel), with colors based on count-based gene expression quantiles and cell source: tumor (red), NAT (blue), or blood (green). For each gene, a gradient of increasing expression can be observed from cluster 8.3c to clusters 8.3b and 8.3a. The high expression of ITGAE in cluster 4.1-S100A6 suggests that it represents CD4+ resident memory T cells. (Figure 18D) Clonal expansion patterns by T cell cluster. Clonal expansion patterns are tabulated for clones and cells in each cluster based on the unsupervised analysis in Figure 13A. The left bar graph shows the distribution of clones and their major clusters, and the four middle bar graphs show the distribution of cells and their parental clones. The cell bars are further divided according to their origin in the NAT (left of each pair) and tumor (right of each pair). Separate bar graphs are provided for cells from double-expanded clones.Bar graphs are colored according to the legend below. An asterisk next to each bar indicates a clonal expansion pattern with statistically significant overrepresentation relative to the overall distribution, as determined by a Bonferroni-adjusted p-value of <0.01. Asterisks are labeled by column using the following abbreviations: n = NAT singleton, N = NAT multiplet, T = tumor multiplet, t = tumor singleton, D = double-expanded clone. The four bar graphs on the right show the distribution of non-blood-expanded and blood-expanded cells separately for the four patients in this study who had blood samples; the bar graphs are colored according to the legend below. (Figure 18E) T cell clusters exhibit distinct clonal expansion patterns. Clones from all 14 patients were composited and grouped by their major cell clusters. Each scatterplot is plotted on a linear scale according to size in the NAT and tumor, with clones truncated at 50 on each axis to display the expansion patterns of the majority of clones with relatively low levels of expansion. Dots are colored according to the two-dimensional palette in the upper right. [Figure 19A-B]Plots showing clonal expansion patterns from external datasets. (Figure 19A) Analysis of data from Guo et al., 2018, on 14 patients with non-small cell carcinoma. (Figure 19B) Analysis of data from Zhang et al., 2018, on 12 patients with colorectal adenocarcinoma. Both datasets were analyzed using the same methodology as Figure 12B. Each scatterplot shows individual clonotypes plotted according to their size in the NAT and tumor; clone size in the blood is represented by the size of the dot according to the legend in the lower left. Clones are colored according to their clonal expansion pattern using the two-dimensional palette in the lower left. Clones were classified according to their major cell cluster, as determined in the external analysis, and plotted on each scatterplot. Results show that dual tumor / NAT expansion and corresponding peripheral clonal expansion are specific to the CD8-C3-CX3CR1 and CD4-C3-GNLY clusters, which correlate with the 8.1-Eff cluster in the dataset (Figure 13B). Dual tumor / NAT expansion with low level blood expansion was also observed in the CD8-C4-GZMK cluster, which correlates with the 8.2-Tem cluster in the dataset. [Figure 20A-C]Figure 20A shows plots of pre- and post-treatment CD4+ clonotypes in tumors with and without TCR sharing in blood. Analysis was performed on CD4+ clonotypes from Yost et al., 2019, in the same manner as for CD8+ clonotypes in Figure 14. Patients are the same as those in Figure 14 and are shown in the same order. (Figure 20A) TCR representation is not correlated between novel post-treatment CD8+ clonotypes and pre-treatment blood. The analysis in Figure 14A was performed on novel clones with CD4+ clonotypes as the major post-treatment cell cluster. Each dot represents a clonotype plotted according to clone size in pre-treatment blood (or post-treatment blood for patient bcc.su002, who did not have a pre-treatment blood sample) and post-treatment tumor. In contrast to CD8+ clonotypes, CD4+ clone size did not show the same degree of association with matching clonotypes in blood. The total number of cells from novel CD4+ clonotypes sharing TCRs in the blood is provided in each scatterplot below, and they do not correlate with peripheral clonal diversity between patients (Figure 20B). Tracking CD4+ T cell clones in tumors. The analysis in Figure 14C was performed on clones with the CD4+ subtype as the predominant cell cluster in tumors before or after treatment. Under these comprehensive criteria, the predominant cell cluster included several clones that showed a shift from the CD4+ naive subtype before treatment to the CD8_act subtype before treatment. In contrast to CD8+ clonotypes, whose residual clones showed higher clonal expansion in blood-associated conditions (Figure 14C), CD4+ clonotypes generally showed higher clonal expansion in non-blood-associated conditions. Another difference was that patients had more lapsed CD4+ clones and larger clone sizes than lapsed CD8+ clones, suggesting that CD4+ clones may respond more dynamically to antigenic changes than CD8+ clones. (FIG. 20C) Distribution of cell clusters for novel CD4+ clonotypes by tumor expansion and TCR sharing with blood. The analysis in FIG. 14D was performed on novel clones with the CD4+ subtype as the predominant cell cluster after treatment.Differences could be observed in the association of T follicular helper (Tfh) cells with non-blood-associated clones, suggesting that these novel clones are primary or locally expanding rather than peripheral blood-derived. [Figure 21A-C]Plots show a clonotype matching database of known and putative virus-reactive TCRs. (Figure 21A) Diversity of clonotypes reactive to viral antigens. Each patient's clonotype was matched against TCRs listed as reactive to common viral antigens from the VDJdb database (Shugay 2018). Viruses included cytomegalovirus (p65 antigen), Epstein-Barr virus (BMLF1, EBNA3), and influenza A (M1). Other antigens from these viruses are classified as "other." Each matching clone is represented by a rectangular area above its corresponding antigen. The color corresponds to its major cell cluster using the color scheme in Figure 13A, also shown in the legend in the lower right. The height indicates its total clonal size in the tumor + NAT. Clones are stacked one on top of the other in random order. When adjacent clones have the same color, a black line is used to separate them when both clones have a clonal count greater than 5, indicating that they should be resolved. These plots show that while the majority of matching clonotypes are singletons, patients often harbor a few highly expanded, virally reactive clones. (Figure 21B) Association of doubly expanded and blood multiplet clones with viral reactivity. Clones matching VDJdbTCRs and multiple cancer TCRs from The Cancer Genome Atlas (TCGA), suggesting reactivity to viral antigens, were grouped according to their clonal expansion patterns (n ​​= NAT singleton, N = NAT multiplet, D = doubly expanded, T = tumor multiplet, t = tumor singleton, b = blood singleton, B = blood multiplet). The frequency of matches is shown for each database and as a union. Doubly expanded clones were most likely to match virally reactive TCRs. Statistical testing using a chi-square test showed a significant dependence of viral reactivity in each case on clonal expansion pattern; p values ​​are shown in the legend. (Figure 21C) T cell clusters are associated with viral reactivity. Clones were grouped according to their predominant cell cluster using the unsupervised analysis of Figure 13A.The percentage of clones matching VDJdbTCR (left) and multi-cancer TCRs from TCGA (right) is plotted for each cluster, with the average value across all clones shown as a vertical line. The color of each bar corresponds to the scheme in Figure 13A and is indicated in the legend of (Figure 21A). Adjusted p-values ​​from chi-square tests after multiple hypothesis testing using the Benjamini-Hochberg procedure are shown for clusters with values ​​<0.05. [Figure 22A-B] Figure 22A shows plots showing clinical associations with the CD8+ cluster gene signature. (Figure 22A) Correlation of gene signatures with CD8A expression. Each dot represents a pretreatment patient sample from one of three clinical trials of the anti-PD-L1 therapeutic atezolizumab: IMvigor210 (top row), POPLAR (middle row), and IMmotion150 (bottom row). Dots are colored according to randomized treatment group, if present, using the legend in the lower right corner of each row. CD8A expression is compared with the scores of each of the five CD8+ gene signatures, each in a separate column. Numbers in the titles indicate the Pearson correlation coefficients of the correlations between scores. The 8.1-Eff and 8.2-Tem signature scores showed the highest correlation with CD8A expression, an indicator of the presence of CD8+ T cells in the tumor, suggesting their relative importance in explaining variability in CD8+ T cell prevalence among patients. (Figure 22B) Kaplan-Meier survival curves. Kaplan-Meier curves are plotted for CD8A, 8.3a-Trm, 8.3b-Trm, and 8.3c-Trm signature scores to complement those provided in Figure 15D. The presentation is the same as in Figure 15D, dichotomizing patients by signature scores above (red) and below (black) the median. Each curve shows progression-free survival (PFS) on the x-axis and the proportion of patient survival on the y-axis. A plus sign on each survival curve indicates a censored event. The legend for each plot shows the hazard ratio (HR) and p-value for the Cox proportional hazards model comparing the two patient populations; p-values ​​<0.05 are marked with an asterisk. The CD8A curve is included because the incidence of CD8+ T cells in tumors is known to be associated with survival in cancer immunotherapy. DETAILED DESCRIPTION OF THE INVENTION

[0271] The present disclosure provides diagnostic methods, therapeutic methods, and compositions for the treatment of cancer (e.g., lung cancer (e.g., non-small cell lung cancer (NSCLC)), bladder cancer (e.g., urothelial carcinoma (UC)), kidney cancer (e.g., renal cell carcinoma (RCC)), and breast cancer (e.g., triple-negative breast cancer (TNBC))). The present disclosure relates to a method for detecting a cancer in an individual, the method comprising ... peripheral The present invention is based on the discovery that the presence of clonally expanded T cells in the blood can be used as a biomarker (e.g., a predictive biomarker) in methods for identifying whether an individual is likely to respond to a treatment comprising a PD-L1 axis-binding antagonist (e.g., a PD-L1-binding antagonist (e.g., an anti-PD-L1 antibody, e.g., atezolizumab (MPDL3280A)) or a PD-1-binding antagonist (e.g., an anti-PD-1 antibody)); selecting a therapy for treating said individual; optimizing the therapeutic effect of a treatment comprising a PD-L1 axis-binding antagonist; and / or monitoring an individual's response to a treatment comprising a PD-L1 axis-binding antagonist.

[0272] I. Definition The term "about" as used herein refers to a normal error range for the respective value, which is readily understood by a person skilled in the art. Reference to a value or parameter preceded by "about" herein includes (describes) an embodiment that is oriented around the value or parameter itself.

[0273] As used herein, "administering" refers to a method of providing a subject with a dosage of a compound (e.g., a PD-L1 axis binding antagonist (e.g., a PD-L1 binding antagonist (e.g., an anti-PD-L1 antibody, e.g., atezolizumab (MPDL3280A)), or a PD-1 binding antagonist (e.g., an anti-PD-1 antibody)) or composition (e.g., a pharmaceutical composition, e.g., a pharmaceutical composition comprising a PD-L1 axis binding antagonist). The compounds and / or compositions used in the methods described herein can be administered, for example, intravenously (e.g., by intravenous infusion), subcutaneously, intramuscularly, or intravenously. The compound may be administered intradermally, transcutaneously, intra-arterially, intraperitoneally, intralesionally, intracranially, intra-articularly, intraprostatically, intrapleurally, intratracheally, intranasally, intravitreally, intravaginally, intrarectally, topically (locally), intratumorally, intraperitoneally, subconjunctivally, intravesicularly, transmucosally, intrapericardially, intraumbilically, intraocularly, orally, topically, locally, by inhalation, injection, infusion, continuous infusion, localized perfusion of target cells directly, by catheter, lavage, cream, or lipid composition. The method of administration may vary depending on various factors (e.g., the compound or composition being administered and the severity of the condition, disease, or disorder being treated).

[0274] "Affinity" refers to the strength of the total non-covalent interactions between a single binding site of a molecule (e.g., an antibody) and its binding partner (e.g., an antigen). Unless otherwise specified, as used herein, "binding affinity" refers to the intrinsic binding affinity that reflects a 1:1 interaction between members of a binding pair (e.g., an antibody and an antigen). The affinity of a molecule X for its partner Y is generally determined by the dissociation constant (K D ) Affinity can be measured by methods common in the art, including those described herein. Specific illustrative examples and exemplary embodiments for measuring binding affinity are described below.

[0275] An "affinity matured" antibody refers to an antibody that has one or more alterations in one or more hypervariable regions (HVRs) compared to a parent antibody that does not have such alterations, which improve the affinity of the antibody for its antigen.

[0276] As used herein, "amplification" generally refers to the process of generating multiple copies of a desired sequence. "Multiple copies" means at least two copies. A "copy" does not necessarily imply perfect sequence complementarity or identity to the template sequence. For example, a copy may contain nucleotide analogs such as deoxyinosine, intentional sequence modifications (such as those introduced by primers containing sequences that are hybridizable to, but not complementary to, the template), and / or sequence errors that occur during amplification.

[0277] The term "antibody" is used herein in the broadest sense and encompasses a variety of antibody structures, including, but not limited to, monoclonal antibodies, polyclonal antibodies, multispecific antibodies (e.g., bispecific antibodies), and antibody fragments, so long as they exhibit the desired antigen-binding activity.

[0278] "Antibody fragment" refers to a molecule other than an intact antibody that contains a portion of an intact antibody that binds to the antigen to which the intact antibody binds. Examples of antibody fragments include, but are not limited to, Fv, Fab, Fab', Fab'-SH, F(ab')2, diabodies, linear antibodies, single-chain antibody molecules (e.g., scFv), and multispecific antibodies formed from antibody fragments.

[0279] An "antibody that binds to the same epitope" as a reference antibody refers to an antibody that blocks the reference antibody from binding to its antigen by 50% or more in a competition assay, and conversely, a reference antibody that blocks the antibody from binding to its antigen by 50% or more in a competition assay. Exemplary competition assays are provided herein.

[0280] The terms "anti-PD-L1 antibody" and "antibody that binds to PD-L1" refer to an antibody that is capable of binding to PD-L1 with sufficient affinity such that the antibody is useful as a diagnostic and / or therapeutic agent in targeting PD-L1. In one embodiment, the extent to which the anti-PD-L1 antibody binds to an unrelated, non-PD-L1 protein is less than about 10% of the binding of the antibody to PD-L1, as measured, for example, by radioimmunoassay (RIA). In a specific embodiment, the anti-PD-L1 antibody binds to an epitope of PD-L1 that is conserved among PD-L1 from different species. In a specific embodiment, the anti-PD-L1 antibody is atezolizumab (MPDL3280A). PD-L1 (programmed death ligand 1) is also referred to in the art as "programmed cell death 1 ligand 1," "PDCD1LG1," "CD274," "B7-H," and "PDL1." An exemplary human PD-L1 is shown in UniProtKB / Swiss-Prot Accession No. Q9NZQ7.1.

[0281] The term "anti-cancer therapy" refers to a therapy useful for treating cancer (e.g., lung cancer (e.g., non-small cell lung cancer (NSCLC)), bladder cancer (e.g., urothelial carcinoma (UC)), kidney cancer (e.g., renal cell carcinoma (RCC)), or breast cancer (e.g., triple-negative breast cancer (TNBC))). Examples of anti-cancer therapeutic agents include, but are not limited to, chemotherapeutic agents, growth inhibitory agents, cytotoxic agents, agents used in radiation therapy, anti-angiogenic agents, apoptotic agents, anti-tubulin agents, and other agents for treating cancer, such as, but not limited to, anti-CD20 antibodies, platelet-derived growth factor inhibitors (e.g., GLEEVEC™ (imatinib mesylate)), COX-2 inhibitors (e.g., celecoxib), interferons, cytokines, antagonists (e.g., neutralizing antibodies) that bind to one or more of the following targets: PDGFR-β, BlγS, APRIL, BCMA receptor(s), TRAIL / Apo2, other bioactive and organic chemical agents, and the like. Combinations of these are also encompassed by the present invention.

[0282] An "article of manufacture" is any manufacture (e.g., package or container) or kit that includes at least one reagent, e.g., a pharmaceutical agent for the treatment of a disease or disorder (e.g., cancer, e.g., lung cancer (e.g., NSCLC), bladder cancer (e.g., UC), kidney cancer (e.g., RCC), or breast cancer (e.g., TNBC)), or a probe for specifically detecting a biomarker described herein. In certain embodiments, the article of manufacture or kit is promoted, distributed, or sold as a unit for performing a method described herein.

[0283] The phrase "based on," as used herein, means that information about one or more biomarkers is used to inform, for example, treatment decisions, information provided in package inserts, or marketing / promotional guidelines.

[0284] A "blocking" or "antagonist" antibody is one that inhibits or reduces the biological activity of the antigen to which it binds. Preferred blocking or antagonist antibodies substantially or completely inhibit the biological activity of the antigen.

[0285] "Binding domain" refers to a portion of a compound or molecule that specifically binds to a target epitope, antigen, ligand, or receptor. Binding domains include, but are not limited to, antibodies (e.g., monoclonal, polyclonal, recombinant, humanized, and chimeric antibodies), antibody fragments or portions thereof (e.g., Fab fragments, Fab'2, scFv antibodies, SMIPs, domain antibodies, diabodies, minibodies, scFv-Fc, affibodies, nanobodies, and antibody VH and / or VL domains), receptors, ligands, aptamers, and other molecules with identified binding partners.

[0286] The term "biomarker" as used herein refers to an indicator, e.g., a predictive, diagnostic, and / or prognostic indicator, that can be detected in a sample (e.g., one or more of CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, PATL2, GPR56, IFITM2, IFITM1, TMSB10, CD247, COX2, COX1, CLIC3, S100A4, and CYBA). Biomarkers serve as indicators of particular subtypes of diseases or disorders (e.g., cancer, e.g., lung cancer (e.g., NSCLC), bladder cancer (e.g., UC), kidney cancer (e.g., RCC), or breast cancer (e.g., TNBC)), characterized by specific molecular, pathological, histological, and / or clinical features. In some embodiments, a biomarker is a gene. Biomarkers include, but are not limited to, polynucleotide (e.g., DNA and / or RNA), polynucleotide copy number changes (e.g., DNA copy number), polypeptide, polypeptide and polynucleotide modifications (e.g., post-translational modifications), carbohydrate, and / or glycolipid-based molecular markers.

[0287] The terms "biomarker signature," "signature," "biomarker expression signature," or "expression signature" are used interchangeably herein to refer to a biomarker whose expression is an indicator, e.g., a predictive, diagnostic, and / or prognostic indicator (e.g., CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, C A biomarker signature refers to one or a combination of biomarkers that result in an immune score (expression levels of one or more of TSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, PATL2, GPR56, IFITM2, IFITM1, TMSB10, CD247, COX2, COX1, CLIC3, S100A4, and CYBA). A biomarker signature serves as an indicator of a particular subtype of a disease or disorder (e.g., cancer, e.g., lung cancer (e.g., NSCLC), bladder cancer (e.g., UC), kidney cancer (e.g., RCC), or breast cancer (e.g., TNBC)) characterized by specific molecular, pathological, histological, and / or clinical features. In some embodiments, a biomarker signature is a "gene signature." The term "gene signature" is used interchangeably with "gene expression signature" and refers to a combination of polynucleotides whose expression is an indicator, e.g., a predictive, diagnostic, and / or prognostic indicator. In some embodiments, a biomarker signature is a "protein signature." The term "protein signature" is used interchangeably with "protein expression signature" and refers to one or a combination of polypeptides whose expression is an indicator, e.g., a predictive, diagnostic, and / or prognostic indicator.

[0288] The term "clonally expanded T cells" refers to T cells that share a common antigen specificity, for example, as assessed by T cell receptor (TCR) sequence homology. For example, clonally expanded T cells can share at least 85% sequence identity (e.g., 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, 99.9%, or 100% sequence identity) across their TCR alpha and / or beta chains (e.g., 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, 99.9%, or 100% sequence identity across the CDR1, CDR2, and / or CDR3 regions of their TCR alpha and / or beta chains).

[0289] The term "corresponding" complementarity determining regions (CDRs), as used herein in the context of two or more TCRs, refers to the equivalent CDRs present in each TCR; i.e., CDR-1 of a particular TCR alpha chain is said to "correspond" to CDR-1 of another TCR alpha chain. Similarly, CDR-2 of a particular TCR alpha chain is said to "correspond" to CDR-3 of another TCR alpha chain; CDR-1 of a particular TCR beta chain is said to "correspond" to CDR-1 of another TCR beta chain; CDR-2 of a particular TCR beta chain is said to "correspond" to CDR-2 of another TCR beta chain, and CDR-3 of a particular TCR beta chain is said to "correspond" to CDR-3 of another TCR beta chain.

[0290] As used herein, the term "chimeric antigen receptor" ("CAR") refers to a recombinant polypeptide comprising one or more antigen-binding regions (e.g., one or more CDRs) that recognize and specifically bind to a given antigen (e.g., an antigen expressed by a tumor cell). A CAR can be expressed by binding to an endogenous TCR (e.g., a TCR of a patient with cancer, e.g., a cancer described herein). peripheralThe CAR may comprise one or more antigen-binding regions of a TCR (a T cell expressed by a T cell in the blood). However, the CAR may not have the molecular composition of a naturally occurring TCR.

[0291] As described herein, a CAR typically comprises at least an extracellular antigen-binding domain, a transmembrane domain, and a cytoplasmic signaling domain (also referred to herein as an "intracellular signaling domain") comprising a functional signaling domain derived from a stimulatory molecule as defined herein. The stimulatory molecule may be the zeta chain associated with the T cell receptor complex. In some embodiments, the intracellular signaling domain further comprises one or more functional signaling domains derived from at least one costimulatory molecule, as described below. The costimulatory molecule may comprise, for example, 4-1BB (i.e., CD137), CD27, and / or CD28. In some embodiments, a CAR comprises a chimeric fusion protein having an extracellular antigen-recognition domain, a transmembrane domain, and a cytoplasmic signaling domain comprising a functional signaling domain derived from a stimulatory molecule. A CAR may comprise, for example, a chimeric fusion protein having an extracellular antigen-recognition domain, a transmembrane domain, and a cytoplasmic signaling domain comprising a functional signaling domain derived from a costimulatory molecule and a functional signaling domain derived from a stimulatory molecule. In some embodiments, a CAR comprises a chimeric fusion protein having an extracellular antigen recognition domain, a transmembrane domain, and an intracellular signaling domain comprising two functional signaling domains derived from one or more costimulatory molecule(s) and one functional signaling domain derived from a stimulatory molecule. In some embodiments, a CAR comprises a chimeric fusion protein having an extracellular antigen recognition domain, a transmembrane domain, and an intracellular signaling domain comprising at least two functional signaling domains derived from one or more costimulatory molecule(s) and one functional signaling domain derived from a stimulatory molecule. A CAR may comprise a leader sequence at the amino terminus of the CAR fusion protein. In some embodiments, a CAR further comprises a leader sequence at the N-terminus of the extracellular antigen recognition domain that may be cleaved from the antigen recognition domain, e.g., (scFv), during cellular processing and localization of the CAR to the cellular membrane. For the avoidance of doubt, the terms "intracellular domain" and "cytoplasmic domain" are used interchangeably as used herein.

[0292] The term "signaling domain" refers to a functional portion of a protein that acts by transmitting intracellular information through defined signaling pathways to control cellular activity, either by generating second messengers or by functioning as an effector by responding to such messengers.

[0293] The CARs described herein can comprise antibodies or antibody fragments thereof, which can exist in a variety of forms. For example, the antigen-binding domain can be expressed as part of a contiguous polypeptide chain, including, for example, single-domain antibody fragments (sdAbs), single-chain antibodies (scFvs), and humanized antibodies (Harlow et al., 1999, In: Using Antibodies: A Laboratory Manual, Cold Spring Harbor Laboratory Press, NY; Harlow et al., 1989, In: Antibodies: A Laboratory Manual, Cold Spring Harbor, NY; Houston et al., 1988, Proc. Natl. Acad. Sci. USA 85:5879-5883; Bird et al., 1988, Science 242:423-426).

[0294] As used herein, the term "stimulatory molecule" refers to a molecule expressed by a T cell that provides a primary cytoplasmic signaling sequence(s) that regulates primary activation of the TCR complex in a stimulatory manner with respect to at least some aspects of the T cell signaling pathway. In one embodiment, the primary signal is initiated, for example, by binding of the TCR / CD3 complex to a peptide-loaded MHC molecule and results in mediation of a T cell response, including, but not limited to, proliferation, activation, and differentiation. Primary cytoplasmic signaling sequences (also referred to as "primary signaling domains") that act in a stimulatory manner can contain signaling motifs known as immunoreceptor tyrosine-based activation motifs, or ITAMs. Examples of ITAMs containing primary cytoplasmic signaling sequences that are particularly useful in the present invention include, but are not limited to, those derived from TCR zeta, FcR gamma, FcR beta, CD3 gamma, CD3 delta, CD3 epsilon, CD5, CD22, CD79a, CD79b, CD278 (also known as "ICOS"), and CD66d. In certain CAR molecules of the invention, the intracellular signaling domain of any one or more CAR molecules of the invention comprises an intracellular signaling sequence, e.g., the primary signaling sequence of CD3-zeta. In certain CARs of the invention, the primary signaling sequence of CD3-zeta is the human sequence (SEQ ID NO: 99) or equivalent residues from a non-human species, such as mouse, rodent, monkey, and ape.

[0295] As used herein, the term "intracellular signaling domain" refers to the intracellular portion of a molecule. The intracellular signaling domain generates a signal that promotes immune effector function of the CAR-containing cell, e.g., a CAR T cell. Examples of immune effector function, e.g., in a CAR T cell, include cytolytic responses and helper activity, including cytokine secretion.

[0296] In some embodiments, the intracellular signaling domain can comprise a primary intracellular signaling domain. Exemplary primary intracellular signaling domains include those derived from molecules responsible for primary stimulation or antigen-dependent stimulation. In one embodiment, the intracellular signaling domain can comprise a costimulatory intracellular domain. Exemplary costimulatory intracellular signaling domains include those derived from molecules responsible for costimulatory signals or antigen-dependent stimulation. For example, in the case of a CAR T, the primary intracellular signaling domain can comprise a cytoplasmic sequence of a T cell receptor, and the costimulatory intracellular signaling domain can comprise a cytoplasmic sequence derived from a co-receptor or costimulatory molecule.

[0297] The primary intracellular signaling domain can comprise a signaling motif known as an immunoreceptor tyrosine-based activation motif, or ITAM. Examples of ITAMs that contain primary cytoplasmic signaling sequences include, but are not limited to, those derived from CD3 zeta, FcR gamma, FcR beta, CD3 gamma, CD3 delta, CD3 epsilon, CD5, CD22, CD79a, CD79b, and CD66d DAP10 and DAP12.

[0298] As used herein, "zeta" or alternatively "zeta chain," "CD3-zeta," or "TCR-zeta" is defined as the protein provided as GenBank Acc. No. BAG36664.1, or equivalent residues from non-human species, such as mouse, rodent, monkey, and ape, and a "zeta stimulatory domain" or alternatively "CD3-zeta stimulatory domain" or "TCR-zeta stimulatory domain" is defined as the amino acid residues from the cytoplasmic domain of the zeta chain sufficient to functionally transmit the initial signal required for T cell activation. In one embodiment, the zeta cytoplasmic domain comprises residues 52 to 164 of GenBank Acc. No. BAG36664.1, or equivalent residues from non-human species, such as mouse, rodent, monkey, and ape, that are functional orthologs thereof. In one embodiment, a "zeta stimulatory domain" or "CD3-zeta stimulatory domain" is the sequence provided as SEQ ID NO: 100 or SEQ ID NO: 99.

[0299] A "costimulatory molecule" refers to the cognate binding partner on a T cell that specifically binds to a costimulatory ligand and thereby mediates a costimulatory response by the T cell, such as, but not limited to, proliferation. A costimulatory molecule is a surface molecule other than an antigen receptor or its ligand that is required for an efficient immune response. Costimulatory molecules include, but are not limited to, MHC class I molecules, BTLA and Toll ligand receptors, as well as OX40, CD27, CD28, CDS, ICAM-1, LFA-1 (CD11a / CD18), and 4-1BB (CD137).

[0300] The costimulatory intracellular signaling domain can be derived from the intracellular portion of a costimulatory molecule. Costimulatory molecules can be represented by the following protein families: TNF receptor proteins, immunoglobulin-like proteins, cytokine receptors, integrins, signaling lymphocyte activation molecules (SLAM proteins), and activating NK cell receptors. Examples of such molecules include ligands that specifically bind to CD27, CD28, 4-1BB (CD137), OX40, GITR, CD30, CD40, ICOS, BAFFR, HVEM, lymphocyte function-associated antigen-1 (LFA-1), CD2, CD7, LIGHT, NKG2C, SLAMF7, NKp80, CD160, B7-H3, and CD83. The intracellular signaling domain can comprise the entire intracellular portion or the entire native intracellular signaling domain of the molecule from which it is derived, or a functional fragment thereof.

[0301] As used herein, "4-1BB" refers to a member of the TNFR superfamily having the amino acid sequence provided as GenBank Acc. No. AAA62478.2, or equivalent residues from a non-human species, such as mouse, rodent, monkey, and ape; a "4-1BB costimulatory domain" is defined as amino acid residues 214-255 of GenBank Acc. No. AAA62478.2, or equivalent residues from a non-human species, such as mouse, rodent, monkey, and ape. In one embodiment, the "4-1BB costimulatory domain" is a human sequence or equivalent residues from a non-human species, such as mouse, rodent, monkey, and ape. In one embodiment, the "4-1BB costimulatory domain" is the sequence provided as SEQ ID NO: 102, or equivalent residues from a non-human species, such as mouse, rodent, monkey, and ape.

[0302] The term "CST7," as used herein, unless otherwise indicated, refers to any native CST7-containing cystatin F gene from any vertebrate source, including mammals such as primates (e.g., humans) and rodents (e.g., mice and rats). The term encompasses "full-length," unprocessed CST7, and any form of CST7 that results from processing in cells. The term also encompasses naturally occurring variants of CST7, such as splice variants or allelic variants. An exemplary nucleic acid sequence of human CST7 is set forth in SEQ ID NO: 1 (GENBANK). TM The amino acid sequence of an exemplary protein encoded by human CST7 is listed in SEQ ID NO: 2 (GENBANK). TM Accession No. CAD52872.1).

[0303] The term "NKG7," as used herein, unless otherwise indicated, refers to the natural killer cell granule protein 7 gene, including any native NKG7 from any vertebrate source, including mammals such as primates (e.g., humans) and rodents (e.g., mice and rats). The term encompasses "full-length," unprocessed NKG7, and any form of NKG7 that results from processing in the cell. The term also encompasses naturally occurring variants of NKG7, such as splice variants or allelic variants. An exemplary nucleic acid sequence for human NKG7 is set forth in SEQ ID NO:3 (GENBANK TM The amino acid sequence of an exemplary protein encoded by human NKG7 is listed in SEQ ID NO: 4 (Uniprot TM Accession No. Q16617-1).

[0304] As used herein, the term "GZMH," unless otherwise indicated, refers to any native GZMH-containing granzyme H gene from any vertebrate source, including mammals such as primates (e.g., humans) and rodents (e.g., mice and rats). The term encompasses "full-length," unprocessed GZMH and any form of GZMH that results from processing in the cell. The term also encompasses naturally occurring variants of GZMH, such as splice variants or allelic variants. An exemplary nucleic acid sequence for human GZMH is set forth in SEQ ID NO:5 (GENBANK). TM The amino acid sequence of an exemplary protein encoded by human GZMH is listed in SEQ ID NO: 6 (Uniprot TM Accession No. P20718-1).

[0305] The term "MT-ND4," as used herein, unless otherwise indicated, refers to the mitochondrially encoded NADH:ubiquinone oxidoreductase core subunit 4 gene, including any native MT-ND4 from any vertebrate source, including mammals such as primates (e.g., humans) and rodents (e.g., mice and rats). The term encompasses "full-length," unprocessed MT-ND4, and any form of MT-ND4 that results from processing in the cell. The term also encompasses naturally occurring variants of MT-ND4, such as splice variants or allelic variants. An exemplary nucleic acid sequence of human MT-ND4 is set forth in SEQ ID NO:7 (GENBANK TM The amino acid sequence of an exemplary protein encoded by human MT-ND4 is listed in SEQ ID NO: 8 (Uniprot TM Accession No. P03905-1).

[0306] The term "HLA-H," as used herein, unless otherwise indicated, refers to the major histocompatibility complex, class I, H gene comprising any native HLA-H from any vertebrate source, including mammals such as primates (e.g., humans) and rodents (e.g., mice and rats). The term encompasses "full-length," unprocessed HLA-H and any form of HLA-H resulting from processing in cells. The term also encompasses naturally occurring variants of HLA-H, such as splice variants or allelic variants. The nucleic acid sequence of an exemplary human HLA-H is listed in SEQ ID NO: 9 (European Nucleotide Archive Accession No. AAA36218.1). The amino acid sequence of an exemplary protein encoded by human HLA-H is listed in SEQ ID NO: 10 (Uniprot TM Accession No. P01893-1).

[0307] As used herein, the term "CCL5," unless otherwise indicated, refers to the CC motif chemokine ligand 5 gene, including any native CCL5, from any vertebrate source, including mammals such as primates (e.g., humans) and rodents (e.g., mice and rats). The term encompasses "full-length," unprocessed CCL5, and any form of CCL5 that results from processing in cells. The term also encompasses naturally occurring variants of CCL5, such as splice variants or allelic variants. An exemplary nucleic acid sequence of human CCL5 is listed in SEQ ID NO: 11 (European Nucleotide Archive Accession No. AF043341.1). The amino acid sequence of an exemplary protein encoded by human CCL5 is listed in SEQ ID NO: 12 (Uniprot TM Accession No. P13501-1).

[0308] The term "CD8A," as used herein, unless otherwise indicated, refers to the CD8A molecule gene, including any native CD8A from any vertebrate source, including mammals such as primates (e.g., humans) and rodents (e.g., mice and rats). The term encompasses "full-length," unprocessed CD8A, and any form of CD8A that results from processing in the cell. The term also encompasses naturally occurring variants of CD8A, such as splice variants or allelic variants. An exemplary nucleic acid sequence for human CD8A is set forth in SEQ ID NO: 13 (GENBANK TM The amino acid sequence of an exemplary protein encoded by human CD8A is listed in SEQ ID NO: 14 (Uniprot TM Accession No. P01732-1).

[0309] As used herein, the term "CMC1," unless otherwise indicated, refers to a C-X9-C motif-containing 1 gene, including any native CMC1 from any vertebrate source, including mammals such as primates (e.g., humans) and rodents (e.g., mice and rats). The term encompasses "full-length," unprocessed CMC1, and any form of CMC1 that results from processing in a cell. The term also encompasses naturally occurring variants of CMC1, such as splice variants or allelic variants. An exemplary nucleic acid sequence of human CMC1 is listed in SEQ ID NO: 15 (RefSeq Accession No. NM_182523.1). The amino acid sequence of an exemplary protein encoded by human CMC1 is listed in SEQ ID NO: 16 (Uniprot TM Accession No. Q7Z7K0-1).

[0310] The term "CD8B," as used herein, unless otherwise indicated, refers to the CD8B molecule gene, including any native CD8B from any vertebrate source, including mammals such as primates (e.g., humans) and rodents (e.g., mice and rats). The term encompasses "full-length," unprocessed CD8B, and any form of CD8B that results from processing in cells. The term also encompasses naturally occurring variants of CD8B, such as splice variants or allelic variants. An exemplary nucleic acid sequence of human CD8B is listed in SEQ ID NO: 17 (European Nucleotide Archive Accession No. BC100911.1). The amino acid sequence of an exemplary protein encoded by human CD8B is listed in SEQ ID NO: 18 (Uniprot TM Accession No. P10966-1).

[0311] The term "HCST," as used herein, unless otherwise indicated, refers to any native HCST-containing hematopoietic cell signaling gene from any vertebrate source, including mammals such as primates (e.g., humans) and rodents (e.g., mice and rats). The term encompasses "full-length," unprocessed HCST, and any form of HCST that results from processing in cells. The term also encompasses naturally occurring variants of HCST, such as splice variants or allelic variants. An exemplary nucleic acid sequence of human HCST is listed in SEQ ID NO: 19 (European Nucleotide Archive Accession No. AF285447.1). The amino acid sequence of an exemplary protein encoded by human HCST is listed in SEQ ID NO: 20 (Uniprot TM Accession No. Q9UBK5-1).

[0312] The term "MT-CYB," as used herein, unless otherwise indicated, refers to the mitochondrially encoded cytochrome B gene, including any native MT-CYB from any vertebrate source, including mammals such as primates (e.g., humans) and rodents (e.g., mice and rats). The term encompasses "full-length," unprocessed MT-CYB, and any form of MT-CYB that results from processing in the cell. The term also encompasses naturally occurring variants of MT-CYB, such as splice variants or allelic variants. The nucleic acid sequence of an exemplary human MT-CYB is listed in SEQ ID NO: 21 (European Nucleotide Archive Accession No. M28016.1). The amino acid sequence of an exemplary protein encoded by human MT-CYB is listed in SEQ ID NO: 22 (Uniprot TM Accession No. P00156-1).

[0313] As used herein, the term "MT-ND4L," unless otherwise indicated, refers to the mitochondrially encoded NADH:ubiquinone oxidoreductase core subunit 4L gene, including any native MT-ND4L from any vertebrate source, including mammals such as primates (e.g., humans) and rodents (e.g., mice and rats). The term encompasses "full-length," unprocessed MT-ND4L and any form of MT-ND4L that results from processing in the cell. The term also encompasses naturally occurring variants of MT-ND4L, such as splice variants or allelic variants. The nucleic acid sequence of an exemplary human MT-ND4L is listed in SEQ ID NO:23 (RefSeq Accession No. NC_012920.1). The amino acid sequence of an exemplary protein encoded by human MT-ND4L is listed in SEQ ID NO:24 (Uniprot TM Accession No. P03901-1).

[0314] The term "KLRG1," as used herein, unless otherwise indicated, refers to the killer cell lectin-like G1 gene, including any native KLRG1 from any vertebrate source, including mammals such as primates (e.g., humans) and rodents (e.g., mice and rats). The term encompasses "full-length," unprocessed KLRG1, and any form of KLRG1 that results from processing in a cell. The term also encompasses naturally occurring variants of KLRG1, such as splice variants or allelic variants. An exemplary nucleic acid sequence of human KLRG1 is listed in SEQ ID NO: 25 (European Nucleotide Archive Accession No. BC012621.1). The amino acid sequence of an exemplary protein encoded by human KLRG1 is listed in SEQ ID NO: 26 (Uniprot TM Accession No. Q96E93-1).

[0315] As used herein, the term "MT-CO2," unless otherwise indicated, refers to the mitochondrial-encoded cytochrome C oxidase II gene, including any native MT-CO2 from any vertebrate source, including mammals such as primates (e.g., humans) and rodents (e.g., mice and rats). The term encompasses "full-length," unprocessed MT-CO2 and any form of MT-CO2 resulting from processing in the cell. The term also encompasses naturally occurring variants of MT-CO2, such as splice variants or allelic variants. An exemplary nucleic acid sequence for human MT-CO2 is listed in SEQ ID NO: 27 (European Nucleotide Archive Accession No. X55654.1). The amino acid sequence of an exemplary protein encoded by human MT-CO2 is listed in SEQ ID NO: 28 (Uniprot TM Accession No. P00403-1).

[0316] As used herein, the term "MT-ATP6," unless otherwise indicated, refers to the mitochondrially encoded ATP synthase membrane subunit 6 gene, including any native MT-ATP6 from any vertebrate source, including mammals such as primates (e.g., humans) and rodents (e.g., mice and rats). The term encompasses "full-length," unprocessed MT-ATP6, and any form of MT-ATP6 resulting from processing in the cell. The term also encompasses naturally occurring variants of MT-ATP6, such as splice variants or allelic variants. An exemplary nucleic acid sequence of human MT-ATP6 is listed in SEQ ID NO:29 (RefSeq Accession No. NC_012920.1). The amino acid sequence of an exemplary protein encoded by human MT-ATP6 is listed in SEQ ID NO:30 (Uniprot TM Accession No. P00846-1).

[0317] As used herein, the term "PLEK," unless otherwise indicated, refers to the pleckstrin gene, including any native PLEK, from any vertebrate source, including mammals such as primates (e.g., humans) and rodents (e.g., mice and rats). The term encompasses "full-length," unprocessed PLEK, and any form of PLEK that results from processing in a cell. The term also encompasses naturally occurring variants of PLEK, such as splice variants or allelic variants. An exemplary nucleic acid sequence of human PLEK is listed in SEQ ID NO: 31 (European Nucleotide Archive Accession No. AK313756.1). The amino acid sequence of an exemplary protein encoded by human PLEK is listed in SEQ ID NO: 32 (Uniprot TM Accession No. P08567-1).

[0318] As used herein, the term "CTSW," unless otherwise indicated, refers to any native CTSW-containing cathepsin W gene from any vertebrate source, including mammals such as primates (e.g., humans) and rodents (e.g., mice and rats). The term encompasses "full-length," unprocessed CTSW and any form of CTSW that results from processing in cells. The term also encompasses naturally occurring variants of CTSW, such as splice variants or allelic variants. An exemplary nucleic acid sequence of human CTSW is listed in SEQ ID NO: 33 (European Nucleotide Archive Accession No. BC048255.1). The amino acid sequence of an exemplary protein encoded by human CTSW is listed in SEQ ID NO: 34 (Uniprot TM Accession No. P56202-1).

[0319] The term "HLA-C," as used herein, unless otherwise indicated, refers to the major histocompatibility complex, class I, C gene comprising any native HLA-C from any vertebrate source, including mammals such as primates (e.g., humans) and rodents (e.g., mice and rats). The term encompasses "full-length," unprocessed HLA-C and any form of HLA-C resulting from processing in a cell. The term also encompasses naturally occurring variants of HLA-C, such as splice variants or allelic variants. The nucleic acid sequence of an exemplary human HLA-C is listed in SEQ ID NO: 35 (European Nucleotide Archive Accession No. D64152.1). The amino acid sequence of an exemplary protein encoded by human HLA-C is listed in SEQ ID NO: 36 (Uniprot TM Accession No. P30508-1).

[0320] As used herein, the term "LYAR," unless otherwise indicated, refers to the Ly1 antibody-reactive gene, including any native LYAR, from any vertebrate source, including mammals such as primates (e.g., humans) and rodents (e.g., mice and rats). The term encompasses "full-length," unprocessed LYAR, and any form of LYAR resulting from processing in cells. The term also encompasses naturally occurring variants of LYAR, such as splice variants or allelic variants. An exemplary nucleic acid sequence of human LYAR is listed in SEQ ID NO: 37 (European Nucleotide Archive Accession No. BC015796.2). The amino acid sequence of an exemplary protein encoded by human LYAR is listed in SEQ ID NO: 38 (Uniprot TM Accession No. Q9NX58-1).

[0321] The term "LITAF," as used herein, unless otherwise indicated, refers to a lipopolysaccharide-inducible TNF factor gene, including any native LITAF, from any vertebrate source, including mammals such as primates (e.g., humans) and rodents (e.g., mice and rats). The term encompasses "full-length," unprocessed LITAF, and any form of LITAF that results from processing in the cell. The term also encompasses naturally occurring variants of LITAF, such as splice variants or allelic variants. An exemplary nucleic acid sequence for human LITAF is set forth in SEQ ID NO: 39 (GENBANK TM The amino acid sequence of an exemplary protein encoded by human LITAF is listed in SEQ ID NO: 40 (Uniprot TM Accession No. Q99732-1).

[0322] As used herein, the term "GZMB," unless otherwise indicated, refers to the granzyme B gene, including any native GZMB, from any vertebrate source, including mammals such as primates (e.g., humans) and rodents (e.g., mice and rats). The term encompasses "full-length," unprocessed GZMB, and any form of GZMB that results from processing in the cell. The term also encompasses naturally occurring variants of GZMB, such as splice variants or allelic variants. An exemplary nucleic acid sequence for human GZMB is SEQ ID NO: 53 (GENBANK TM The amino acid sequence of an exemplary protein encoded by human GZMB is listed in SEQ ID NO: 54 (Uniprot TM Accession No. P10144-1).

[0323] The term "KLRD1," as used herein, unless otherwise indicated, refers to the killer cell lectin-like D1 gene, including any native KLRD1 from any vertebrate source, including mammals such as primates (e.g., humans) and rodents (e.g., mice and rats). The term encompasses "full-length," unprocessed KLRD1, and any form of KLRD1 that results from processing in the cell. The term also encompasses naturally occurring variants of KLRD1, such as splice variants or allelic variants. An exemplary nucleic acid sequence for human KLRD1 is set forth in SEQ ID NO: 55 (GENBANK TM The amino acid sequence of an exemplary protein encoded by human KLRD1 is listed in SEQ ID NO: 56 (Uniprot TM Accession No. Q13241-1).

[0324] The term "FGFBP2," as used herein, unless otherwise indicated, refers to any fibroblast growth factor binding protein 2 gene, including any FGFBP2, from any vertebrate source, including mammals such as primates (e.g., humans) and rodents (e.g., mice and rats). The term encompasses "full-length," unprocessed FGFBP2, and any form of FGFBP2 that results from processing in the cell. The term also encompasses naturally occurring variants of FGFBP2, such as splice variants or allelic variants. An exemplary nucleic acid sequence for human FGFBP2 is set forth in SEQ ID NO: 57 (GENBANK TM The amino acid sequence of an exemplary protein encoded by human FGFBP2 is listed in SEQ ID NO: 58 (Uniprot TM Accession No. Q9BYJ0-1).

[0325] As used herein, the term "KLRC4-KLRK1," unless otherwise indicated, refers to the KLRC4-KLRK1 readthrough gene, including any native KLRC4-KLRK1 from any vertebrate source, including mammals such as primates (e.g., humans) and rodents (e.g., mice and rats). The term encompasses "full-length," unprocessed KLRC4-KLRK1, and any form of KLRC4-KLRK1 that results from processing in a cell. The term also encompasses naturally occurring variants of KLRC4-KLRK1, such as splice variants or allelic variants. The nucleic acid sequence of an exemplary human KLRC4-KLRK1 is listed in SEQ ID NO: 59 (RefSeq Accession No. NM_001199805.1). The amino acid sequence of an exemplary protein encoded by human KLRC4-KLRK1 is listed in SEQ ID NO: 60 (Uniprot TM Accession No. H3BQV0-1).

[0326] The term "KLRK1," as used herein, unless otherwise indicated, refers to the killer cell lectin-like K1 gene, including any native KLRK1 from any vertebrate source, including mammals such as primates (e.g., humans) and rodents (e.g., mice and rats). The term encompasses "full-length," unprocessed KLRK1, and any form of KLRK1 that results from processing in the cell. The term also encompasses naturally occurring variants of KLRK1, such as splice variants or allelic variants. An exemplary nucleic acid sequence for human KLRK1 is set forth in SEQ ID NO: 61 (GENBANK). TM The amino acid sequence of an exemplary protein encoded by human KLRK1 is listed in SEQ ID NO: 62 (Uniprot TM Accession No. P26718-1).

[0327] The term "B2M," as used herein, unless otherwise indicated, refers to any native B2M-containing beta-2-microglobulin gene from any vertebrate source, including mammals such as primates (e.g., humans) and rodents (e.g., mice and rats). The term encompasses "full-length," unprocessed B2M, and any form of B2M resulting from processing in the cell. The term also encompasses naturally occurring variants of B2M, such as splice variants or allelic variants. An exemplary human B2M nucleic acid sequence is listed in SEQ ID NO: 63 (European Nucleotide Archive Accession No. AF072097.1). The amino acid sequence of an exemplary protein encoded by human B2M is listed in SEQ ID NO: 64 (Uniprot TM Accession No. P61769-1).

[0328] The term "GZMA," as used herein, unless otherwise indicated, refers to any native GZMA-containing granzyme A gene from any vertebrate source, including mammals such as primates (e.g., humans) and rodents (e.g., mice and rats). The term encompasses "full-length," unprocessed GZMA and any form of GZMA that results from processing in the cell. The term also encompasses naturally occurring variants of GZMA, such as splice variants or allelic variants. An exemplary nucleic acid sequence for human GZMA is SEQ ID NO: 65 (GENBANK TM The amino acid sequence of an exemplary protein encoded by human GZMA is listed in SEQ ID NO: 66 (Uniprot TM Accession No. P12544-1).

[0329] The term "ID2," as used herein, unless otherwise indicated, refers to the inhibitor of DNA binding 2 gene, including any native ID2 from any vertebrate source, including mammals such as primates (e.g., humans) and rodents (e.g., mice and rats). The term encompasses "full-length," unprocessed ID2, and any form of ID2 that results from processing in the cell. The term also encompasses naturally occurring variants of ID2, such as splice variants or allelic variants. An exemplary nucleic acid sequence of human ID2 is listed in SEQ ID NO: 67 (European Nucleotide Archive Accession No. BC030639). The amino acid sequence of an exemplary protein encoded by human ID2 is listed in SEQ ID NO: 68 (Uniprot TM Accession No. Q02363-1).

[0330] The term "CX3CR1," as used herein, unless otherwise indicated, refers to any native CX3CR1-containing C-X3-C motif chemokine receptor 1 gene from any vertebrate source, including mammals such as primates (e.g., humans) and rodents (e.g., mice and rats). The term encompasses "full-length," unprocessed CX3CR1, and any form of CX3CR1 that results from processing in cells. The term also encompasses naturally occurring variants of CX3CR1, such as splice variants or allelic variants. An exemplary nucleic acid sequence for human CX3CR1 is set forth in SEQ ID NO: 69 (GENBANK TM The amino acid sequence of an exemplary protein encoded by human CX3CR1 is listed in SEQ ID NO: 70 (Uniprot TM Accession No. P49238-1).

[0331] As used herein, the term "PRSS23," unless otherwise indicated, refers to any native serine protease 23 gene, including any native PRSS23, from any vertebrate source, including mammals such as primates (e.g., humans) and rodents (e.g., mice and rats). The term encompasses "full-length," unprocessed PRSS23, and any form of PRSS23 that results from processing in the cell. The term also encompasses naturally occurring variants of PRSS23, such as splice variants or allelic variants. An exemplary nucleic acid sequence for human PRSS23 is set forth in SEQ ID NO: 71 (GENBANK TM The amino acid sequence of an exemplary protein encoded by human PRSS23 is listed in SEQ ID NO: 72 (Uniprot TM Accession No. O95084-1).

[0332] As used herein, the term "GNLY," unless otherwise indicated, refers to any native granulysin gene, including any native GNLY, from any vertebrate source, including mammals such as primates (e.g., humans) and rodents (e.g., mice and rats). The term encompasses "full-length," unprocessed GNLY, and any form of GNLY that results from processing in a cell. The term also encompasses naturally occurring variants of GNLY, such as splice variants or allelic variants. An exemplary nucleic acid sequence for human GNLY is set forth in SEQ ID NO: 73 (GENBANK TM The amino acid sequence of an exemplary protein encoded by human GNLY is listed in SEQ ID NO: 74 (Uniprot TM Accession No. P22749-1).

[0333] The term "PRF1," as used herein, unless otherwise indicated, refers to the perforin 1 gene, including any native PRF1, from any vertebrate source, including mammals such as primates (e.g., humans) and rodents (e.g., mice and rats). The term encompasses "full-length," unprocessed PRF1, and any form of PRF1 that results from processing in a cell. The term also encompasses naturally occurring variants of PRF1, such as splice variants or allelic variants. An exemplary nucleic acid sequence for human PRF1 is set forth in SEQ ID NO: 75 (GENBANK TM The amino acid sequence of an exemplary protein encoded by human PRF1 is listed in SEQ ID NO: 76 (Uniprot TM Accession No. P14222-1).

[0334] As used herein, the term "PATL2," unless otherwise indicated, refers to the PAT1 homolog 2 gene, including any native PATL2, from any vertebrate source, including mammals such as primates (e.g., humans) and rodents (e.g., mice and rats). The term encompasses "full-length," unprocessed PATL2, and any form of PATL2 that results from processing in a cell. The term also encompasses naturally occurring variants of PATL2, such as splice variants or allelic variants. An exemplary nucleic acid sequence of human PATL2 is listed in SEQ ID NO: 77 (RefSeq Accession No. NM_001145112.1). The amino acid sequence of an exemplary protein encoded by human PATL2 is listed in SEQ ID NO: 78 (Uniprot TM Accession No. C9JE40-1).

[0335] The term "GPR56," as used herein, unless otherwise indicated, refers to the adhesion G protein-coupled receptor G1 gene, including any native GPR56, from any vertebrate source, including mammals such as primates (e.g., humans) and rodents (e.g., mice and rats). The term encompasses "full-length," unprocessed GPR56, and any form of GPR56 that results from processing in a cell. The term also encompasses naturally occurring variants of GPR56, such as splice variants or allelic variants. An exemplary nucleic acid sequence of human GPR56 is listed in SEQ ID NO: 79 (RefSeq Accession No. NM_001145770.2). The amino acid sequence of an exemplary protein encoded by human GPR56 is listed in SEQ ID NO: 80 (Uniprot TM Accession No. Q9Y653-1).

[0336] The term "IFITM2," as used herein, unless otherwise indicated, refers to the interferon-inducible transmembrane protein 2 gene, including any native IFITM2 from any vertebrate source, including mammals such as primates (e.g., humans) and rodents (e.g., mice and rats). The term encompasses "full-length," unprocessed IFITM2, and any form of IFITM2 that results from processing in a cell. The term also encompasses naturally occurring variants of IFITM2, such as splice variants or allelic variants. An exemplary nucleic acid sequence of human IFITM2 is listed in SEQ ID NO: 81 (RefSeq Accession No. NM_003641.3). The amino acid sequence of an exemplary protein encoded by human IFITM2 is listed in SEQ ID NO: 82 (Uniprot TM Accession No. Q01629-1).

[0337] The term "IFITM1," as used herein, unless otherwise indicated, refers to the interferon-inducible transmembrane protein 1 gene, including any native IFITM1 from any vertebrate source, including mammals such as primates (e.g., humans) and rodents (e.g., mice and rats). The term encompasses "full-length," unprocessed IFITM1, and any form of IFITM1 that results from processing in a cell. The term also encompasses naturally occurring variants of IFITM1, such as splice variants or allelic variants. An exemplary nucleic acid sequence of human IFITM1 is listed in SEQ ID NO: 83 (RefSeq Accession No. NM_001145112.1). The amino acid sequence of an exemplary protein encoded by human PATL2 is listed in SEQ ID NO: 84 (Uniprot TM Accession No. P13164-1).

[0338] As used herein, the term "TMSB10," unless otherwise indicated, refers to the thymosin beta-10 gene, including any native TMSB10, from any vertebrate source, including mammals such as primates (e.g., humans) and rodents (e.g., mice and rats). The term encompasses "full-length," unprocessed TMSB10, and any form of TMSB10 that results from processing in a cell. The term also encompasses naturally occurring variants of TMSB10, such as splice variants or allelic variants. An exemplary nucleic acid sequence of human TMSB10 is listed in SEQ ID NO: 85 (RefSeq Accession No. NM_021103.3). The amino acid sequence of an exemplary protein encoded by human TMSB10 is listed in SEQ ID NO: 86 (Uniprot TM Accession No. P63313-1).

[0339] The term "CD247," as used herein, unless otherwise indicated, refers to the T-cell surface glycoprotein CD3 zeta chain gene, including any native CD247, from any vertebrate source, including mammals such as primates (e.g., humans) and rodents (e.g., mice and rats). The term encompasses "full-length," unprocessed CD247, and any form of CD247 that results from processing in the cell. The term also encompasses naturally occurring variants of CD247, such as splice variants or allelic variants. An exemplary nucleic acid sequence of human CD247 is listed in SEQ ID NO: 87 (RefSeq Accession No. NM_000734.3). The amino acid sequence of an exemplary protein encoded by human CD247 is listed in SEQ ID NO: 88 (Uniprot TM Accession No. P20963-1).

[0340] As used herein, the term "COX2," unless otherwise indicated, refers to the prostaglandin G / H synthase 2 gene, including any native COX2, from any vertebrate source, including mammals such as primates (e.g., humans) and rodents (e.g., mice and rats). The term encompasses "full-length," unprocessed COX2, and any form of COX2 that results from processing in a cell. The term also encompasses naturally occurring variants of COX2, such as splice variants or allelic variants. An exemplary human COX2 nucleic acid sequence is listed in SEQ ID NO: 89 (RefSeq Accession No. NM_000963.3). The amino acid sequence of an exemplary protein encoded by human COX2 is listed in SEQ ID NO: 90 (Uniprot TM Accession No. P35354-1).

[0341] The term "COX1," as used herein, unless otherwise indicated, refers to the prostaglandin G / H synthase 1 gene, including any native COX1, from any vertebrate source, including mammals such as primates (e.g., humans) and rodents (e.g., mice and rats). The term encompasses "full-length," unprocessed COX1, and any form of COX1 that results from processing in a cell. The term also encompasses naturally occurring variants of COX1, such as splice variants or allelic variants. An exemplary nucleic acid sequence of human COX1 is listed in SEQ ID NO: 91 (RefSeq Accession No. NM_000962.3). The amino acid sequence of an exemplary protein encoded by human COX1 is listed in SEQ ID NO: 92 (Uniprot TM Accession No. P23219-1).

[0342] As used herein, the term "CLIC3," unless otherwise indicated, refers to any native chloride intracellular channel protein 3 gene, including any native CLIC3, from any vertebrate source, including mammals such as primates (e.g., humans) and rodents (e.g., mice and rats). The term encompasses "full-length," unprocessed CLIC3, and any form of CLIC3 that results from processing in the cell. The term also encompasses naturally occurring variants of CLIC3, such as splice variants or allelic variants. An exemplary nucleic acid sequence of human CLIC3 is listed in SEQ ID NO: 93 (RefSeq Accession No. NM_004669.2). The amino acid sequence of an exemplary protein encoded by human CLIC3 is listed in SEQ ID NO: 94 (Uniprot TM Accession No. O95833-1).

[0343] The term "S100A4," as used herein, unless otherwise indicated, refers to any native S100A4-containing protein S100-A4 gene from any vertebrate source, including mammals such as primates (e.g., humans) and rodents (e.g., mice and rats). The term encompasses "full-length," unprocessed S100A4, and any form of S100A4 resulting from processing in the cell. The term also encompasses naturally occurring variants of S100A4, such as splice variants or allelic variants. An exemplary human S100A4 nucleic acid sequence is listed in SEQ ID NO: 95 (RefSeq Accession No. NM_002961.2). The amino acid sequence of an exemplary protein encoded by human S100A4 is listed in SEQ ID NO: 96 (Uniprot TM Accession No. P26447-1).

[0344] The term "CYBA," as used herein, unless otherwise indicated, refers to the cytochrome b-245 light chain gene, including any native CYBA, from any vertebrate source, including mammals such as primates (e.g., humans) and rodents (e.g., mice and rats). The term encompasses "full-length," unprocessed CYBA, and any form of CYBA resulting from processing in the cell. The term also encompasses naturally occurring variants of CYBA, such as splice variants or allelic variants. An exemplary nucleic acid sequence of human CYBA is listed in SEQ ID NO: 97 (RefSeq Accession No. NM_000101.3). The amino acid sequence of an exemplary protein encoded by human CYBA is listed in SEQ ID NO: 98 (Uniprot TM Accession No. P13498-1).

[0345] The terms "cancer" and "cancerous" refer to or describe the physiological condition in mammals that is typically characterized by uncontrolled cell growth. Examples of cancer include, but are not limited to, carcinoma, lymphoma, blastoma, sarcoma, and leukemia or lymphoid malignancies. More specific examples of such cancers include, but are not limited to, lung cancer, including small cell lung cancer, non-small cell lung cancer, adenocarcinoma of the lung, and squamous cell carcinoma of the lung; bladder cancer (e.g., urothelial carcinoma (UC), muscle-invasive bladder cancer (MIBC), and BCG-refractory non-muscle-invasive bladder cancer (NMIBC)); kidney or renal cancer (e.g., renal cell carcinoma (RCC)); cancers of the urinary tract; breast cancer (e.g., HER2+ breast cancer and triple-negative breast cancer (TNBC)), which They are estrogen receptor (ER-), progesterone receptor (PR-), and HER2 (HER2-) negative); prostate cancer, e.g., castration-resistant prostate cancer (CRPC); cancer of the peritoneum; hepatocellular carcinoma; gastric cancer or cancer of the stomach, e.g., gastrointestinal cancer and gastrointestinal stromal cancer; pancreatic cancer; glioblastoma; cervical cancer; ovarian cancer; liver cancer; hepatic cancer; colon cancer; rectal cancer; colorectal cancer; endometrial or uterine cancer; salivary gland cancer; prostate cancer; vulvar cancer; thyroid cancer; liver cancer; anal Cancer; penile cancer; melanoma, including superficial spreading melanoma, lentigo maligna melanoma, acral lentiginous melanoma, and nodular melanoma; multiple myeloma and B-cell lymphomas (including low-grade / follicular non-Hodgkin's lymphoma (NHL); small lymphocytic (SL) NHL, intermediate-grade / follicular NHL; intermediate-grade diffuse NHL; high-grade immunoblastic NHL; high-grade lymphoblastic NHL; high-grade small noncleaved cell NHL; bulky disease NHL; mantle cell lymphoma; AIDS-related and Waldenstrom's macroglobulinemia); chronic lymphocytic leukemia (CLL); acute lymphoblastic leukemia (ALL); acute myeloid leukemia (AML); hairy cell leukemia; chronic myeloblastic leukemia (CML); post-transplant lymphoproliferative disorder (PTLD); and myelodysplastic syndromes (MDS), as well as abnormal blood vessel proliferation associated with phakomatosis, edema (such as that associated with brain tumors), Meigs syndrome, brain tumors, head and neck cancers, and associated metastases.

[0346] The terms "cell proliferative disorder" and "proliferative disorder" refer to disorders associated with some degree of abnormal cell proliferation. In one embodiment, the cell proliferative disorder is cancer (e.g., lung cancer (e.g., NSCLC), bladder cancer (e.g., UC), kidney cancer (e.g., RCC), or breast cancer (e.g., TNBC)). In another embodiment, the cell proliferative disorder is a tumor.

[0347] A "chemotherapeutic agent" is a chemical agent useful in the treatment of cancer (e.g., cancer, e.g., lung cancer (e.g., NSCLC), bladder cancer (e.g., UC), kidney cancer (e.g., RCC), or breast cancer (e.g., TNBC)). Examples of chemotherapeutic agents include alkylating agents such as thiotepa and cyclophosphamide (CYTOXAN®); alkyl sulfonates such as busulfan, improsulfan, and piposulfan; aziridines such as benzodopa, carboquone, meturedopa, and uredopa; ethylenimines and methylamelamines, including altrateamine, triethylenemelamine, triethylenephosphoramide, triethylenethiophosphoramide, and trimethylolmelamine; acetogenins (particularly bullatacin and bullatacinone); delta-9-tetrahydrocannabinol (dronabinol, MARINOL®); beta-lapachone; lapachol; colchicine; betulinic acid; camptothecin (synthetic analog topotecan (HYCAMTIN®), CPT-11 (irinotecan, CAMPTOSAR®), acetylcamptothecin, scopoletin, and 9-aminomethyl-2-methyl-2-methyl-1 ... including camptothecin; bryostatin; kallistatin; CC-1065 (including its adozelesin, carzelesin, and bizelesin synthetic analogs); podophyllotoxin; podophyllic acid; teniposide; cryptophycins (especially cryptophycin 1 and cryptophycin 8); dolastatins; duocarmycins (including synthetic analogs, KW-2189 and CB1-TM1); eluterobin; pancratistatin; sarcodictin; sucralose Ponzistatin; chlorambucil, chlornaphazine, chlorophosphamide, estramustine, ifosfamide, mechlorethamine, mechlorethamine oxide hydrochloride, melphalan, nobembine, phenesterine, prednimustine, trofosfamide, nitrogen mustards such as uracil mustard; nitrosoureas such as carmustine, chlorozotocin, fotemustine, lomustine, nimustine, and ranimustine;Antibiotics, such as enediyne antibiotics (e.g., calicheamicins, particularly calicheamicin γ1I and calicheamicin ω1I (e.g., Nicolaou et al., Angew. Chem Intl. Ed. Engl., 33:183-186 (1994); CDP323, an oral alpha-4 integrin inhibitor; dynemicins, including dynemicin A; esperamicin; and neocarzinostatin chromophores and related chromoprotein enediyne antibiotic chromophores), aclacinomycins, actinomycin, ausramycin, azaserine, bleomycin, cactinomycin, carabicin, carminomycin, carzinophilin, chromomycins, dactinomycin, daunorubicin, detorubicin, 6-diazo-5-oxo-L-norleucine, doxorubicin (ADRIAMYCIN®, morpholino-doxorubicin, cyanomorpholino-doxorubicin, 2-pyrrolino-doxorubicin, doxorubicin HCl liposome injection (DOXIL®), liposomal doxorubicin TLC D-99 (MYOCET®), pegylated liposomal doxorubicin (including CAELYX® and deoxydoxorubicin), epirubicin, esorubicin, idarubicin, marcellomycin, mitomycins such as mitomycin C, mycophenolic acid, nogalamycin, olivomycin, peplomycin, porfiromycin, puromycin, chelamycin, rodorubicin, streptonigrin, streptozocin, tubercidin, ubenimex, zinostatin, zorubicin; antimetabolites, e.g., methotrexate, gemcitabine (GEM ZAR®), tegafur (UFTORAL®), capecitabine (XELODA®), epothilones and 5-fluorouracil (5-FU); combretastatins; folic acid analogs such as denopterin, methotrexate, pteropterin, trimetrexate; purine analogs such as fludarabine, 6-mercaptopurine, thiamiprine, thioguanine; pyrimidine analogs such as ancitabine, azacitidine, 6-azauridine, carmofur, cytarabine, dideoxyuridine, doxifluridine, enocitabine, floxuridine;Androgens, such as calsterone, dromostanolone propionate, epitiostanol, mepitiostane, testolactone; antiadrenal agents, such as aminoglutethimide, mitotane, trilostane; folic acid replenishers, such as furoic acid; aceglatone; aldophosphamide glycosides; aminolevulinic acid; eniluracil; amsacrine; bestravcil; bisantrene; edatraxate; defofamine; demecolcine; Diazicon; Elformitin; Elliptinium acetate; Epothilone; Etoglucide; Gallium nitrate; Hydroxyurea; Lentinan; Lonidynin; Maytansinoids, such as maytansine and ansamitocin; Mitoguazone; Mitoxantrone; Mopidanmol; Nitraerin; Pentostatin; Fenamet; Pirarubicin; Rosoxantrone; 2-Ethylhydrazide; Procarbazine; PSK® Polysaccharide Complex (JHS Natural Products, Eugene, Oreg.); razoxane; rhizoxin; schizofuran; spirogermanium; tenuazonic acid; triazicon; 2,2',2'-trichlorotriethylamine; trichothecenes (especially T-2 toxin, veracrine A, roridin A, and anguidine); urethane; vindesine (ELDISINE®, FILDESIN®); dacarbazine; mannomustine; mitobronitol; mitolactol; pipobroman; gacytosine; arabinoside ("Ara-C"); thiotepa; taxoids, such as paclitaxel (TAXOL®, Bristol-Myers Squibb Oncology, Princeton, NJ), albumin-engineered nanoparticle formulations of paclitaxel (ABRAXANE; TM) and docetaxel (TAXOTERE®, Rhome-Poulene Rorer, Antony, France); chlorambucil; 6-thioguanine; mercaptopurine; methotrexate; platinum drugs such as cisplatin, oxaliplatin (e.g., ELOXATIN®), and carboplatin; vincas that prevent tubulin polymerization to form microtubules, such as vinblastine (VELBAN®), vincristine (ONCOVIN®), vindesine (ELDISINE®, FILDESIN®), and vinorelbine (NAVELBINE®); etoposide (VP-16); ifosfamide; mitoxantrone; leucovorin; novantrone; edatrexate; daunomycin; aminopterin; ibandronate; the topoisomerase inhibitor RFS2000; difluoromethylmethylnitine (DMFO); retinoids such as retinoic acid (VELBAN®). xarotene (TARGRETIN®); bisphosphonates, including clodronate (e.g., BONEFOS® or OSTAC®), etidronic acid (DIDROCAL®), NE-58095, zoledronic acid / zoledronate (ZOMETA®), alendronate (FOSAMAX®), pamidronate (AREDIA®), tiludronate (SKELID®), or risedronate (ACTONEL®); troxacitabine (1,3-dioxolane nucleoside cytosine analog); antisense oligonucleotides, particularly those that inhibit the expression of genes in signal transduction pathways involved in abnormal cell proliferation, such as PKC-alpha, Raf, H-Ras, and epidermal growth factor receptor (EGF-R) (e.g., erlotinib (TARCEVA®)). TM)); and VEGF-A, which reduces cell proliferation; vaccines, such as the THERATOPE® vaccine and gene therapy vaccines, such as the ALLOVECTIN® vaccine, the LEUVECTIN® vaccine, and the VAXID® vaccine; topoisomerase 1 inhibitors (e.g., LURTOTECAN®); rmRH (e.g., ABARELIX®); BAY439006 (sorafenib; Bayer); SU-11248 (sunitinib, SUTENT®, Pfizer); perifosine, COX-2 inhibitors (e.g., Celecoxib, orafenib, ABT510; BCL-2 inhibitors, such as oblimersen sodium (GENASENSE®); pixantrone; EGFR inhibitors; tyrosine kinase inhibitors; serine-threonine kinase inhibitors, such as rapamycin (sirolimus, RAPAMUNE®); farnesyltransferase inhibitors, such as lonafarnib (SCH6636, SARASAR®). TM and pharmaceutically acceptable salts, acids, or derivatives of the foregoing, as well as combinations of two or more of the foregoing, such as CHOP (an abbreviation for cyclophosphamide, doxorubicin, vincristine, and prednisolone combination therapy), and FOLFOX (oxaliplatin in combination with 5-FU and leucovorin (ELOXATIN TM ), and pharmaceutically acceptable salts, acids, or derivatives of any of the above; and combinations of two or more of the above.

[0348] Chemotherapeutic agents, as defined herein, also include "anti-hormonal agents" or "endocrine therapies" that act to regulate, reduce, block, or inhibit the effects of hormones that may promote the growth of cancer (e.g., lung cancer (e.g., NSCLC), bladder cancer (e.g., UC), kidney cancer (e.g., RCC), or breast cancer (e.g., TNBC)). They may themselves be hormones, including, but not limited to, antiestrogens and selective estrogen receptor modulators (SERMs), such as tamoxifen (including NOLVADEX® tamoxifen), raloxifene, droloxifene, 4-hydroxytamoxifen, trioxifene, ketoxifene, LY117018, onapristone, and FARESTON.cndot. toremifene; aromatase inhibitors that inhibit the enzyme aromatase, which regulates estrogen production in the adrenal glands, such as 4(5)-imidazole, aminoglutethimide, MEGASE® megestrol acetate, AROMASIN® exemestane, formestany, fadrozole, RIVISOR® vorozole, FEMARA® letrozole, and ARIMIDEX® anastrozole; and antiandrogens, such as flutamide, nilutamide, bicalutamide, leuprozole, and ARIMIDEX® anastrozole. antisense oligonucleotides, particularly those that inhibit the expression of genes in signal transduction pathways involved in the proliferation of adherent cells, such as PKC-alpha, Raf, and H-Ras; ribozymes, such as VEGF expression inhibitors (e.g., ANGIOZYME® ribozyme) and HER2 expression inhibitors; gene therapy vaccines, such as ALLOVECTIN® vaccine, LEUVECTIN® vaccine, and VAXID® vaccine; PROLEUKIN® rIL-2; LURTOTECAN® topoisomerase 1 inhibitors; ABARELIX® rmRH; vinorelbine and esperamicin (see U.S. Pat. No. 4,675,187), and pharmaceutically acceptable salts, acids, and derivatives of any of the above; and combinations of two or more of the above.

[0349] The term "chimeric" antibody refers to an antibody in which a portion of the heavy and / or light chain is derived from a particular source or species, and the remainder of the heavy and / or light chain is derived from a different source or species.

[0350] The "class" of an antibody refers to the type of constant domain or constant region carried by its heavy chain. There are five major classes of antibodies: IgA, IgD, IgE, IgG, and IgM, and some of these can be further divided into subclasses (isotypes), e.g., IgG1, IgG2, IgG3, IgG4, IgA1, and IgA2. The heavy-chain constant domains that correspond to the different classes of immunoglobulins are called α, σ, ε, γ, and μ, respectively.

[0351] As used herein, the term "cytotoxic gene" refers to a gene whose expression is indicative of the presence and / or activity of a cytotoxic T lymphocyte (CTL). Cytotoxic genes include those that encode proteins present on the surface of CTLs, such as CD8A, as well as those that encode secreted proteins associated with CTL function, such as CCL5. Exemplary cytotoxic genes for use in combination with the compositions and methods described herein include PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, CST7, NKG7, GZMH, and HLA-C.

[0352] As used herein, the term "mitochondrial gene" refers to a gene encoding RNA and / or proteins involved in mitochondrial function, such as the oxidative production of adenosine triphosphate. Exemplary mitochondrial genes for use in combination with the compositions and methods described herein include MT-ND4, MT-ND4L, MT-ATP6, MT-CO2, and MT-CYB.

[0353] The term "cytotoxic agent," as used herein, refers to a substance that inhibits or prevents the function of cells and / or causes cell death or destruction. Cytotoxic agents include radioisotopes (e.g., At 211 , I131 , I 125 , Y 90 ,Re 186 ,Re 188 , Sm 153 , Bi 212 , P 32 , Pb 212 , and radioactive isotopes of Lu); chemotherapeutic agents or drugs (e.g., methotrexate, adriamycin, vinca alkaloids (vincristine, vinblastine, etoposide), doxorubicin, melphalan, mitomycin C, chlorambucil, daunorubicin or other intercalating agents); growth inhibitory agents; enzymes and fragments thereof, e.g., nucleases; antibiotics; toxins, such as small molecule toxins or enzymatically active toxins of bacterial, fungal, plant or animal origin (including fragments and / or variants thereof); and various anti-tumor or anti-cancer agents disclosed below.

[0354] The term "concurrently" is used herein to refer to the administration of two or more therapeutic agents where at least some of the administration overlaps in time. Thus, concurrent administration includes a dosing regimen in which the administration of one or more agents continues after the administration of one or more other agents has ceased.

[0355] As used herein, "delaying the progression" of a disorder or disease means to postpone, prevent, delay, slow, stabilize, and / or postpone the onset of the disease or disorder (e.g., cancer, e.g., lung cancer (e.g., NSCLC), bladder cancer (e.g., UC), kidney cancer (e.g., RCC), or breast cancer (e.g., TNBC)). The delay can be of varying duration depending on the history of the disease and / or the subject being treated. As will be apparent to one of skill in the art, a sufficient or significant delay can, in effect, encompass prevention, in that the subject does not develop the disease.

[0356] The terms "determination," "determining," "detection," "detecting," and grammatical variations thereof include any means of determining or detecting, including direct and indirect determination or detection.

[0357] A "disorder" or "disease" is any condition that would benefit from treatment, including, but not limited to, chronic and acute disorders or diseases, including conditions that predispose a mammal to the disorder in question (e.g., cancer, e.g., lung cancer (e.g., NSCLC), bladder cancer (e.g., UC), kidney cancer (e.g., RCC), or breast cancer (e.g., TNBC)).

[0358] As used herein, the term "diagnosis" refers to the identification or classification of a molecular state or pathology, disease or condition (e.g., cancer, e.g., lung cancer (e.g., NSCLC), bladder cancer (e.g., UC), kidney cancer (e.g., RCC), or breast cancer (e.g., TNBC)). For example, "diagnosis" can refer to the identification of a particular type of cancer. "Diagnosis" can also refer to the classification of a cancer into a particular subtype, e.g., by histopathological criteria or by molecular features (e.g., a subtype characterized by the expression of a biomarker or combination of biomarkers (e.g., a particular gene or the protein encoded by that gene)).

[0359] "Effector function" refers to a biological activity attributable to the Fc region of an antibody, which differs depending on the antibody isotype. Examples of antibody effector functions include C1q binding and complement-dependent cytotoxicity (CDC), Fc receptor binding, antibody-dependent cell-mediated cytotoxicity (ADCC), phagocytosis, downregulation of cell surface receptors (e.g., PD-L1), and B-cell activation.

[0360] An "effective amount" of a compound, e.g., a PD-L1 axis binding antagonist (e.g., a PD-L1 binding antagonist (e.g., an anti-PD-L1 antibody, e.g., atezolizumab (MPDL3280A)) or a PD-1 binding antagonist (e.g., an anti-PD-1 antibody) The antagonist (e.g., an anti-PD-1 antibody), or composition thereof (e.g., a pharmaceutical composition), is at least the minimum amount necessary to achieve a desired therapeutic or prophylactic result, such as a measurable increase in overall survival (OS) or progression-free survival (PFS) of a particular disease or disorder (e.g., cancer, e.g., lung cancer (e.g., NSCLC), bladder cancer (e.g., UC), kidney cancer (e.g., RCC), or breast cancer (e.g., TNBC)). An effective amount herein may vary depending on factors such as the individual's condition, age, sex, and weight, as well as the ability of the antibody to elicit a desired response in the subject. An effective amount is also one in which any toxic or adverse effects of the treatment are outweighed by the therapeutically beneficial effects. Beneficial or desired results for prophylactic use include results such as elimination or reduction of the risk, reduction in severity, or delay in the onset of disease, including biochemical, histological, and / or behavioral symptoms of the disease, its complications, and intermediate pathological phenotypes manifesting during the development of the disease. An effective amount may be administered in one or more administrations. In the present invention, an effective amount of a drug, compound, or pharmaceutical composition is an amount sufficient to directly or indirectly achieve prophylactic or therapeutic treatment. As clinically understood, an effective amount of a drug, compound, or pharmaceutical composition may optionally be achieved in combination with another drug, compound, or pharmaceutical composition. Thus, an "effective amount" can be considered in the context of the administration of one or more therapeutic agents, and a single agent can be considered to be given in an effective amount if, in combination with one or more other agents, a desired result can be achieved or is achieved.For example, an effective amount of a PD-L1 axis binding antagonist (e.g., a PD-L1 binding antagonist (e.g., an anti-PD-L1 antibody, e.g., atezolizumab (MPDL3280A)) or a PD-1 binding antagonist (e.g., an anti-PD-1 antibody)) as a cancer treatment reduces the number of cancer cells; shrinks primary tumor size; peripheral The drug may inhibit (i.e., slow to some extent, and preferably stop) cancer cell invasion into organs; inhibit (i.e., slow to some extent, and preferably stop) tumor metastasis; inhibit to some extent tumor growth; and / or alleviate to some extent one or more symptoms associated with the disorder. To the extent that the drug may prevent the growth of and / or kill existing cancer cells, the drug may be cytostatic and / or cytotoxic. For cancer therapy, in vivo efficacy can be measured, for example, by assessing survival time, time to disease progression (TTP), response rate (RR), duration of response, and / or quality of life.

[0361] The term "Fc region" is used herein to define the C-terminal region of an immunoglobulin heavy chain containing at least a portion of the constant region. This term includes native-sequence Fc regions and variant Fc regions. In one embodiment, a human IgG heavy chain Fc region extends from Cys226, or from Pro230, to the carboxyl terminus of the heavy chain. However, the C-terminal lysine (Lys447) of the Fc region may or may not be present. Unless otherwise specified herein, numbering of amino acid residues within the Fc region or constant region follows the EU numbering system, also known as the EU index, as described in Kabat et al., Sequences of Proteins of Immunological Interest, 5th Ed. Public Health Service, National Institutes of Health, Bethesda, MD, 1991.

[0362] "Framework" or "FR" refers to variable domain residues other than hypervariable region (HVR) residues. The FR of a variable domain generally consists of four FR domains, FR1, FR2, FR3, and FR4. Thus, the HVR and FR sequences are commonly represented in VH (or VL) as follows: FR1-H1(L1)-FR2-H2(L2)-FR3-H3(L3)-FR4.

[0363] The terms "full-length antibody," "intact antibody," and "whole antibody" are used interchangeably herein to refer to an antibody having a structure substantially similar to a native antibody structure or having a heavy chain that includes an Fc region as defined herein.

[0364] A "human antibody" is one that possesses an amino acid sequence corresponding to the amino acid sequence of an antibody derived from a non-human source produced by a human or human cell, or utilizing the human antibody repertoire, or other human antibody coding sequence. This definition of a human antibody specifically excludes humanized antibodies containing non-human antigen-binding residues. Human antibodies can be generated using various techniques known in the art, including phage display libraries. The methods described in Hoogenboom and Winter, J. Mol. Biol., 227:381 (1991); Marks et al., J. Mol. Biol., 222:581 (1991). Cole et al., Monoclonal Antibodies and Cancer Therapy, Alan R. Liss, p. 77 (1985); Boerner et al., J. Immunol., 147(1):86-95 (1991) are also available for preparing human monoclonal antibodies. See also van Dijk and van de Winkel, Curr. Opin. Pharmacol., 5:368-74 (2001). Human antibodies can be prepared by administering antigen to transgenic animals, e.g., immunized xenomouse, that have been engineered to produce such antibodies in response to antigen challenge, but whose endogenous gene loci have been disabled (see, e.g., U.S. Patent Nos. 6,075,181 and 6,150,584 regarding XENOMOUSE™ technology). See also, e.g., Li et al., Proc. Natl. Acad. Sci. USA, 103:3557-3562 (2006), regarding human antibodies produced by human B-cell hybridoma technology.

[0365] A "humanized" antibody refers to a chimeric antibody comprising amino acid residues from non-human HVRs and amino acid residues from human FRs. In certain embodiments, a humanized antibody comprises substantially all of at least one, and typically two, variable domains, in which all or substantially all of the HVRs (e.g., CDRs) correspond to those of a non-human antibody and all or substantially all of the FRs correspond to those of a human antibody. A humanized antibody may optionally comprise at least a portion of an antibody constant region derived from a human antibody. A "humanized form" of an antibody, e.g., a non-human antibody, refers to an antibody that has undergone humanization.

[0366] The term "hypervariable region" or "HVR", as used herein, refers to each region of an antibody variable domain that is hypervariable in sequence ("complementarity determining region" or "CDR") and / or structurally forms defined loops ("hypervariable loops") and / or contains residues that contact the antigen ("antigen contacts"). Generally, antibodies contain six HVRs, three in the VH (H1, H2, H3) and three in the VL (L1, L2, L3). Exemplary HVRs of the invention include the following: (a) hypervariable loops occurring at amino acid residues 26-32 (L1), 50-52 (L2), 91-96 (L3), 26-32 (H1), 53-55 (H2), and 96-101 (H3) (Chothia and Lesk, J. Mol. Biol. 196:901-917 (1987)); (b) CDRs occurring at amino acid residues 24-34 (L1), 50-56 (L2), 89-97 (L3), 31-35b (H1), 50-65 (H2), and 95-102 (H3) (Kabat et al., Sequences of Proteins of Immunological Interest, 5th Ed. Public Health Service, National Institutes of Health, Bethesda, MD (1991)); (c) antigen contacts occurring at amino acid residues 27c-36 (L1), 46-55 (L2), 89-96 (L3), 30-35b (H1), 47-58 (H2), and 93-101 (H3) (MacCallum et al., J. Mol. Biol., 262:732-745 (1996)); and Combinations of (a), (b) and / or (c), HVR amino acid residues 46-56 (L2), 47-56 (L2), 48-56 (L2), 49-56 (L2), 26-35 (H1), 26-35b (H1), 49-65 (H2), 93-102 (H3) and 94-102 (H3).

[0367] Unless otherwise indicated herein, HVR residues and other residues in the variable domain (e.g., FR residues) are numbered herein according to Kabat et al., supra.

[0368] An "isolated" antibody is one that has been separated from a component of its natural environment. In some embodiments, the antibody is purified to greater than 95% or greater than 99% purity, for example, as determined by electrophoresis (e.g., SDS-PAGE, isoelectric focusing (IEF), capillary electrophoresis) or chromatography (e.g., ion exchange or reverse-phase HPLC). For a review of testing methods for antibody purification, see, e.g., Flatman et al., J. Chromatogr. B 848:79-87 (2007).

[0369] An "isolated" nucleic acid refers to a nucleic acid molecule that has been separated from a component of its natural environment. Isolated nucleic acid includes nucleic acid molecules that are contained in cells that originally contained the nucleic acid molecule, but where the nucleic acid molecule is present extrachromosomally or at a chromosomal location that is different from its natural chromosomal location.

[0370] As used herein, the term "label" refers to a detectable compound or composition. Labels are typically directly or indirectly conjugated or condensed with a reagent, such as a polynucleotide probe or antibody, to facilitate detection of the conjugated or condensed reagent. The label may itself be detectable (e.g., a radioisotope label or a fluorescent label) or, in the case of an enzymatic label, may catalyze chemical alteration of a substrate compound or composition that results in a detectable product.

[0371] The terms "level of expression" or "expression level" are generally used interchangeably and generally refer to the amount of a biomarker in a biological sample. "Expression" generally refers to the process by which information (e.g., genetic coding information and / or epigenetic information) is converted into structures present and functioning in a cell. Thus, as used herein, "expression" can refer to transcription into a polynucleotide, translation into a polypeptide, or even polynucleotide and / or polypeptide modification (e.g., post-translational modification of a polypeptide). Fragments of a transcribed polynucleotide, a translated polypeptide, or a polynucleotide and / or polypeptide modification (e.g., post-translational modification of a polypeptide) should also be considered expressed, regardless of whether they are derived from a transcript generated by alternative splicing or a degraded transcript, or from post-translational processing of a polypeptide, for example, by proteolysis. "Expressed genes" include those that are transcribed into a polynucleotide as mRNA and then translated into a polypeptide, and also those that are transcribed into RNA but not translated into a polypeptide (e.g., transfer and ribosomal RNA). Expression levels can be measured by methods known to those skilled in the art, including, for example, RT-qPCR and RNA-seq, and can also be measured by methods disclosed herein. The assessed expression levels can be used to determine response to treatment.

[0372] The term "immunoscore expression level" refers to a numerical value reflecting the expression level (e.g., normalized expression level) of a single gene of interest or the aggregated expression levels of multiple genes of interest (e.g., at least two, at least three, at least four, at least five, or more genes of interest) in relation to an immune response. The immunoscore expression level for multiple genes of interest can be determined by aggregation methods known to those of skill in the art and disclosed herein, including, for example, calculating the median or mean expression levels of all genes of interest. Prior to aggregation, the expression level of each gene of interest may be normalized using statistical methods known to those of skill in the art and disclosed herein, including, for example, normalizing to the expression level of one or more housekeeping genes, or may be normalized to the total library size, or may be normalized to the median or mean expression levels of all genes measured. In some cases, prior to aggregation of multiple genes of interest, the normalized expression level of each gene of interest can be standardized by calculating a Z-score of the normalized expression level of each gene of interest. In some cases, each gene of interest may be assigned a weighted score, and the immunoscore expression levels of multiple genes of interest can be calculated by determining the average weighted expression levels of all genes of interest by incorporating the weighted scores. For example, an Immunoscore expression level may refer to a number reflecting the normalized expression level of a single gene selected from CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, PATL2, GPR56, IFITM2, IFITM1, TMSB10, CD247, COX2, COX1, CLIC3, S100A4, and CYBA.Alternatively, the immune score expression levels can be determined using, for example, the following genes: CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, PATL2, GPR56, IFITM1, IFITM2, IFITM3, IFITM4, IFITM5, IFITM6, IFITM7, IFITM8, IFITM9, IFITM10, IFITM11, IFITM12, IFITM13, IFITM14, IFITM15, IFITM16, IFITM17, IFITM18, IFITM19 ... It refers to a numerical value reflecting the aggregated normalized expression levels (e.g., median normalized expression levels or mean normalized expression levels) for at least two, at least three, at least four, at least five, at least six, at least seven, at least eight, at least nine, at least ten, at least eleven, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, or more of TMSB10, CD247, COX2, COX1, CLIC3, S100A4, and CYBA, or a combination thereof.In some cases, the immunoscore expression levels were measured using the following genes: CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, PATL2, GPR56, IFITM2, IFITM1, TMSB10, CD247, COX2, COX1, CLIC3, and TMSB10. It refers to a number reflecting the aggregated Z-score expression levels (e.g., the mean Z-score normalized expression levels or the median Z-score normalized expression levels) for at least two, at least three, at least four, at least five, at least six, at least seven, at least eight, at least nine, at least ten, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, or more of S100A4, and CYBA, or a combination thereof.

[0373] As used herein, the term "reference immunoscore expression level" refers to a reference immunoscore expression level that is compared to another immunoscore expression level (e.g., a reference immunoscore expression level for genes CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, IL-1, IL-2, IL-3, IL-4, IL-5, IL-6, IL-7, IL-8, IL-9, IL-11, IL-12, IL-13, IL-14, IL-15, IL-16, IL-17, IL-18, IL-19, IL-20, IL-21, IL-22, IL-23, IL-24, IL-25, IL-26, IL-27, IL-28, IL-29, IL-30, IL-31, IL-32, IL-33, IL-34, IL-35, IL-36, IL-37, IL-38, IL-39, IL-40, IL-41, IL-42, IL-43, IL-44, IL-45, IL-46, IL-47, IL-48, IL-49, IL-59, IL-50, IL-51, IL-52, IL-53, IL-54, IL-55, IL-56, IL-57, IL-58, IL-59, IL-59, IL-59, IL-59, IL-59, IL-59, , LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, PATL2, GPR56, IFITM2, IFITM1, TMSB10, CD247, COX2, COX1, CLIC3, S100A4, and CYBA).For example, a reference immune score expression level can be the expression level in a reference sample (e.g., genes CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK). 1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, PATL2, GPR56, IFITM2, IFITM1, TMSB10, CD247, COX2, COX1, CLIC3, S100A4, and CYBA), a reference population, and / or a pre-assigned value (e.g., a reference population) for a PD-L1 axis binding antagonist (e.g., PD-L1 A first subset of individuals treated with a therapy with a binding antagonist (e.g., an anti-PD-L1 antibody, e.g., atezolizumab (MPDL3280A)) or a PD-1 binding antagonist (e.g., an anti-PD-1 antibody)) and a second subset of individuals in the same reference population treated with a therapy with a non-PD-L1 axis binding antagonist that does not include a PD-L1 axis binding antagonist are compared based on a significant difference between the responsiveness of the individuals to treatment with the PD-L1 axis binding antagonist therapy and the responsiveness of the individuals to treatment with the non-PD-L1 axis binding antagonist therapy above and / or below a cutoff value. The PD-L1 axis binding antagonist therapy may be derived from a cutoff value previously determined to result in a significant (e.g., statistically significant) separation between the two PD-L1 axis binding antagonist therapies, where the individual's responsiveness to treatment with the PD-L1 axis binding antagonist therapy is significantly (e.g., statistically significantly) improved compared to the individual's responsiveness to treatment with a PD-L1 axis binding antagonist therapy above the cutoff value and / or is significantly (e.g., statistically significantly) improved compared to the individual's responsiveness to treatment with a non-PD-L1 axis binding antagonist therapy below the cutoff value.One of skill in the art will understand that the numerical values ​​for reference immunoscore expression levels may vary depending on the indication (e.g., cancer (e.g., breast cancer, lung cancer, kidney cancer, or bladder cancer), the methodology used to detect expression levels (e.g., RNA-seq or RT-qPCR), the statistical method used to generate the immunoscore, and / or the particular combination of genes tested.

[0374] "Elevated expression," "elevated expression level," or "elevated level" refers to elevated expression or levels of a gene or combination of genes of interest relative to a control, e.g., one or more subjects not afflicted with a disease or disorder (e.g., cancer, e.g., lung cancer (e.g., NSCLC), bladder cancer (e.g., UC), kidney cancer (e.g., RCC), or breast cancer (e.g., TNBC)), or relative to an internal standard (e.g., housekeeping gene), or reference level, e.g., a reference immune score expression level (e.g., a reference immune score expression level). , CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, PATL2, GPR56, IFITM2, IFITM1, TMSB10, CD247, COX2, COX1, CLIC3, S100A4, and CYBA).

[0375] "Decreased expression," "reduced expression level," or "reduced level" refers to decreased expression or level of a gene or combination of genes in a subject relative to a control, e.g., one or more subjects not afflicted with a disease or disorder (e.g., cancer, e.g., lung cancer (e.g., NSCLC), bladder cancer (e.g., UC), kidney cancer (e.g., RCC), or breast cancer (e.g., TNBC)), or relative to an internal standard (e.g., housekeeping gene), or reference level, e.g., a reference immune score expression level (e.g., reference immune score expression level). , CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, PATL2, GPR56, IFITM2, IFITM1, TMSB10, CD247, COX2, COX1, CLIC3, S100A4, and CYBA). In some embodiments, reduced expression refers to little or no expression.

[0376] As used herein, a "reference gene" refers to a gene or group of genes (e.g., one, two, three, four, five, six, or more genes) used for comparison purposes, such as housekeeping genes. "Housekeeping gene," as used herein, refers to a gene or group of genes (e.g., one, two, three, four, five, six, or more genes) that encode proteins whose activity is essential for maintaining cellular function and that are typically present similarly in all cell types.

[0377] The term "monoclonal antibody," as used herein, refers to an antibody obtained from a substantially homogeneous population of antibodies. That is, each individual antibody in the population is identical and / or binds to the same epitope, excluding possible variant antibodies, including, for example, naturally occurring mutations or mutations that arise during production of the monoclonal antibody preparation. Such variants are generally present in small amounts. In contrast to polyclonal antibody preparations, which typically include different antibodies directed against different determinants (epitopes), each monoclonal antibody of a monoclonal antibody preparation is directed against a single determinant on an antigen. Thus, the modifier "monoclonal" indicates the character of the antibody as being obtained from a substantially homogeneous population of antibodies and is not constructed to require production of the antibody by any particular method. For example, monoclonal antibodies used in accordance with the present invention can be produced by a variety of techniques, including, but not limited to, hybridoma methods, recombinant DNA methods, phage display methods, and methods utilizing transgenic animals containing all or part of the human immunoglobulin loci; such methods and other exemplary methods for producing monoclonal antibodies are described herein.

[0378] A "naked antibody" refers to an antibody that is not conjugated to a heterologous moiety (e.g., a cytotoxic moiety) or radiolabel. The naked antibody may be present in a pharmaceutical formulation.

[0379] "Native antibodies" refer to naturally occurring immunoglobulin molecules with various structures. For example, native IgG antibodies are heterotetrameric glycoproteins of approximately 150,000 daltons, consisting of two identical light chains and two identical heavy chains that are disulfide-bonded. From the N-terminus to the C-terminus, each heavy chain has a variable region (VH), also called a variable heavy domain or a heavy chain variable domain, followed by three constant domains (CH1, CH2, and CH3). Similarly, from the N-terminus to the C-terminus, each light chain has a variable region (VL), also called a variable light domain or a light chain variable domain, followed by a constant light (CL) domain. Based on the amino acid sequence of its constant domain, the light chain of an antibody may be divided into two types, called kappa (κ) and lambda (λ).

[0380] The term "oligonucleotide" refers to a relatively short polynucleotide (e.g., less than about 250 nucleotides in length), including, but not limited to, single-stranded deoxyribonucleotides, single- or double-stranded ribonucleotides, RNA:DNA hybrids, and double-stranded DNA. Oligonucleotides, such as single-stranded DNA probe oligonucleotides, are often synthesized by chemical methods, for example, using commercially available automated oligonucleotide synthesizers. However, oligonucleotides can be produced by a variety of other methods, including in vitro recombinant DNA-mediated techniques, and by expression of DNA in cells and organisms.

[0381] The term "package insert" is used to refer to instructions customarily included in commercial packaging of therapeutic products that contain information about the indications, uses, dosages, administration, concomitant therapies, contraindications, and / or warnings regarding the use of such therapeutic product.

[0382] The term "pharmaceutical formulation" refers to a preparation that is in a form such that the biological activity of the active ingredients contained in the preparation is effective and that does not contain additional components that are unacceptably toxic to the subject to which the formulation is administered.

[0383] A "pharmaceutically acceptable carrier" refers to an ingredient in a pharmaceutical formulation, other than an active ingredient, that is non-toxic to a subject. Pharmaceutically acceptable carriers include, but are not limited to, buffers, excipients, stabilizers, or preservatives.

[0384] As used herein, the term "protein," unless otherwise indicated, refers to any naturally occurring protein from any vertebrate source, including mammals such as primates (e.g., humans) and rodents (e.g., mice and rats). The term encompasses "full-length," unprocessed proteins, and any form of a protein that results from processing within a cell. The term also encompasses naturally occurring variants of the protein, such as splice variants or allelic variants.

[0385] "Percent (%) amino acid sequence identity" with respect to a reference polypeptide sequence is defined as the percentage of amino acid residues in a candidate sequence that are identical to those in the reference polypeptide sequence after aligning the sequences and introducing gaps, if necessary, to achieve the maximum percent sequence identity, without considering any conservative substitutions as part of the sequence identity. Alignment for purposes of determining percent amino acid sequence identity can be achieved in a variety of ways that are within the skill of the art, for example, using known computer software such as BLAST, BLAST-2, ALIGN, or Megalign (DNASTAR) software. Those skilled in the art can determine appropriate parameters for aligning sequences, including any algorithms needed to achieve maximum alignment over the entire length of the sequences being compared. However, for purposes herein, percent amino acid sequence identity values ​​are generated using the sequence comparison computer program ALIGN-2. The ALIGN-2 sequence comparison computer program was written by Genentech, Inc., and the source code is on file in the user documentation of the U.S. Copyright Office, Washington, DC, 20559, and is registered under U.S. Copyright Registration No. TXU510087. The ALIGN-2 program is publicly available from Genentech, Inc., South San Francisco, California, or may be compiled from its source code. The ALIGN-2 program should be compiled for use on UNIX operating systems, including Digital UNIX V4.0D. All sequence comparison parameters are set by the ALIGN-2 program and do not vary.

[0386] In situations where ALIGN-2 is employed for amino acid sequence comparison, the % amino acid sequence identity of a given amino acid sequence A to, against, or opposite a given amino acid sequenc...

Claims

1. 1. A kit for selecting a therapy for an individual having non-small cell lung cancer (NSCLC), renal cell carcinoma (RCC), urothelial carcinoma (UC), or triple-negative breast cancer (TNBC), comprising one or more reagents for determining whether the individual has clonally expanded T cells in a sample of peripheral blood from the individual; (a) Whether an individual has clonally expanded T cells in a sample of peripheral blood from the individual is determined by the following: CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRK1, B2M, GZMA, KLRC4-KLRK1, KLRK1, B2M, GZMA, KLRD1, FGFBP2 ...D1, FGFBP2, KLRD1, FGFBP2, KLRD1, FGFBP2, KLRD1, FGFBP2, KLRD , ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2, and the genes include one or more genes selected from PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, CST7, NKG7, GZMH, and HLA-C, and one or more genes selected from MT-ND4L, MT-ND4, MT-ATP6, MT-CO2, and MT-CYB; an immunoscore expression level of the gene in the sample that is greater than a reference immunoscore expression level of the gene is indicative of clonally expanded T cells in the sample; clonally expanded T cells in the sample identify the individual as one who may benefit from treatment comprising atezolizumab; and (b) a therapy comprising atezolizumab is selected for the individual based on the presence of clonally expanded T cells in the sample.

2. 2. The kit of claim 1, wherein the sample comprises clonally expanded T cells and an effective amount of atezolizumab is administered to the individual.

3. 1. A medicament for treating non-small cell lung cancer (NSCLC), renal cell carcinoma (RCC), urothelial carcinoma (UC), or triple-negative breast cancer (TNBC) in an individual who has been determined to have clonally expanded T cells in a sample of peripheral blood from the individual, indicating that the individual would benefit from treatment comprising atezolizumab, comprising an effective amount of atezolizumab; The presence of clonally expanded T cells was associated with the following transcription factors: CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, CX3CR2, CX3CR3, CX3CR4, CX3CR5, CX3CR6, CX3CR7, CX3CR8, CX3CR9, CX3CR1, CX3CR2 ... The immune score is determined based on expression levels of genes selected from PRSS23, GNLY, PRF1, and PATL2, and the genes include one or more genes selected from PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, CST7, NKG7, GZMH, and HLA-C, and one or more genes selected from MT-ND4L, MT-ND4, MT-ATP6, MT-CO2, and MT-CYB; The medicament, wherein an immunoscore expression level of the gene in the sample that is greater than a reference immunoscore expression level of the gene is indicative of clonally expanded T cells in the sample.

4. The pharmaceutical composition according to claim 3, which is for treating NSCLC.

5. The kit of claim 1 , wherein the clonally expanded T cells are CD8+ T cells.

6. The kit according to claim 5 , wherein the CD8+ T cells are CTLs.

7. 1. A kit for selecting a therapy for an individual having non-small cell lung cancer (NSCLC), renal cell carcinoma (RCC), urothelial carcinoma (UC), or triple-negative breast cancer (TNBC), comprising reagents for determining expression levels of the genes NKG7, GNLY, PRF1, GZMH, GPR56, IFITM2, IFITM1, GZMB, TMSB10, PLAC8, LITAF, CD247, CST7, COX2, KLRD1, CLIC3, S100A4, CYBA, and COX1 in a sample of peripheral blood from the individual; (a) an expression level of the gene in a sample of peripheral blood from the individual is determined, and an immunoscore expression level of the gene in the sample that exceeds a reference immunoscore expression level of the gene identifies the individual as one who may benefit from a treatment comprising atezolizumab; and (b) a treatment comprising atezolizumab is selected for the individual based on an immunoscore expression level of the gene in the sample that is greater than a reference immunoscore expression level.

8. The kit of claim 7, wherein the immunoscore expression level of the gene in the sample exceeds the reference immunoscore expression level and an effective amount of atezolizumab is administered to the individual.

9. The kit of claim 7 or 8 for selecting a therapy for an individual with NSCLC.

10. A medicament for treating non-small cell lung cancer (NSCLC), renal cell carcinoma (RCC), urothelial carcinoma (UC), or triple negative breast cancer (TNBC) in an individual, where the Immunoscore expression levels of the genes NKG7, GNLY, PRF1, GZMH, GPR56, IFITM2, IFITM1, GZMB, TMSB10, PLAC8, LITAF, CD247, CST7, COX2, KLRD1, CLIC3, S100A4, CYBA, and COX1 in a sample of peripheral blood from the individual have been determined to be greater than a reference Immunoscore expression level of the genes, indicating that the individual would benefit from a treatment comprising atezolizumab, the medicament comprising an effective amount of atezolizumab.

11. The pharmaceutical of claim 10 , wherein the reference immune score expression level is an immune score expression level of a gene of a reference population.

12. (a) the reference population is a population of individuals with NSCLC, RCC, UC, or TNBC, comprising a first subset of individuals treated with atezolizumab and a second subset of individuals treated with a therapy that does not include atezolizumab; and (b) the reference Immunoscore expression level significantly separates each of the first and second subsets of individuals based on a significant difference between the individual's responsiveness to treatment with atezolizumab and the individual's responsiveness to treatment with a therapy that does not include atezolizumab above the reference Immunoscore expression level, and the individual's responsiveness to treatment with atezolizumab is significantly improved compared to the individual's responsiveness to treatment with a therapy that does not include atezolizumab.

13. The pharmaceutical agent according to any one of claims 10 to 12, which is for treating NSCLC.

14. 2. The kit of claim 1, wherein the benefit of the treatment comprising atezolizumab comprises an extension of OS and / or an extension of PFS.

15. 15. The kit of any one of claims 1, 2, 5, 6, and 14, for selecting a therapy for an individual with NSCLC.

16. 1. A kit for selecting a therapy for an individual having non-small cell lung cancer (NSCLC), renal cell carcinoma (RCC), urothelial carcinoma (UC), or triple-negative breast cancer (TNBC), comprising reagents for determining the expression level of a gene selected from CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2 in a sample from the individual; the reagents comprising one or more reagents for determining the expression level of a gene selected from PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, CST7, NKG7, GZMH, and HLA-C, and one or more reagents for determining the expression level of a gene selected from MT-ND4L, MT-ND4, MT-ATP6, MT-CO2, and MT-CYB; (a) an expression level of the gene in a sample from the individual is determined, and an immunoscore expression level of the gene in the sample that is greater than a reference immunoscore expression level of the gene identifies the individual as one who may benefit from a treatment comprising atezolizumab; and (b) a treatment comprising atezolizumab is selected for the individual based on an immunoscore expression level of the gene in the sample that is greater than a reference immunoscore expression level.

17. The kit of claim 16, wherein the immunoscore expression level of the gene in the sample exceeds the reference immunoscore expression level and an effective amount of atezolizumab is administered to the individual.

18. The kit of claim 16, wherein the reference immune score expression level is an immune score expression level of a gene of a reference population.

19. 17. The kit of claim 16, comprising reagents for determining the expression levels of the genes CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2.

20. The kit according to any one of claims 16 to 19, for selecting a therapy for an individual with NSCLC.

21. 1. A medicament for treating non-small cell lung cancer (NSCLC), renal cell carcinoma (RCC), urothelial carcinoma (UC), or triple-negative breast cancer (TNBC) in an individual, where an immunoscore expression level of a gene in a sample from the individual is determined to be greater than a reference immunoscore expression level of the gene, thereby indicating that the individual would benefit from a treatment comprising atezolizumab, comprising: the gene is selected from CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2; the genes include one or more genes selected from PLEK, HCST, CTSW, KLRG1, CD8A, CCL5, CD8B, CST7, NKG7, GZMH, and HLA-C, and one or more genes selected from MT-ND4L, MT-ND4, MT-ATP6, MT-CO2, and MT-CYB; The medicament comprises an effective amount of atezolizumab.

22. The pharmaceutical composition of claim 21, wherein the genes include CST7, NKG7, GZMH, MT-ND4, HLA-H, CCL5, CD8A, CMC1, CD8B, HCST, MT-CYB, MT-ND4L, KLRG1, MT-CO2, MT-ATP6, PLEK, CTSW, HLA-C, LYAR, LITAF, GZMB, KLRD1, FGFBP2, KLRC4-KLRK1, KLRK1, B2M, GZMA, ID2, CX3CR1, PRSS23, GNLY, PRF1, and PATL2.

23. The medicament according to claim 21 or 22, which is for treating NSCLC.

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