Multispecific antibodies in combination therapy for cancer immunotherapy

Abstract

Multispecific antibodies for use in cancer immunotherapy and methods of treating cancer are provided.SOLUTION: Multispecific antibodies are provided that have binding sites for ICOS and for second antigens, e.g., immune checkpoint molecules such as PD - L1. Also provided is a method of treating cancer in a human patient comprising administering to the patient (i) a multispecific antibody that binds to ICOS and PD-L1 and (ii) an anti-CTLA-4 or anti-PD-1 antibody.SELECTED DRAWING: Figure 1

Description

[Technical Field]

[0001] The present invention relates to antigen-binding molecules that bind to cell surface receptors involved in the regulation of immune responses. They relate to antibodies for use in stimulating a patient's immune system, in particular effector T cell responses, and have applications in the field of cancer immunotherapy, particularly tumor treatment. [Background technology]

[0002] The adaptive immune response involves the activation, selection, and clonal expansion of two major classes of lymphocytes, called T cells and B cells. After encountering an antigen, T cells proliferate and differentiate into antigen-specific effector cells, while B cells proliferate and differentiate into antibody-secreting cells. T cell activation is a multistep process that requires several signaling events between T cells and antigen-presenting cells (APCs). For T cell activation to occur, two types of signals must be delivered to resting T cells. The first type is mediated by the antigen-specific T cell receptor (TCR) and confers specificity to the immune response. The second signal, a costimulatory signal, controls the magnitude of the response and is delivered via accessory receptors on the T cell.

[0003] The primary costimulatory signal is delivered via the activating CD28 receptor upon engagement of its ligands, B7-1 or B7-2. In contrast, engagement of the inhibitory CTLA-4 receptor by the same B7-1 or B7-2 ligand results in attenuation of T cell responses. Thus, CTLA-4 signals antagonize CD28-mediated costimulation. At high antigen concentrations, CD28 costimulation negates the inhibitory effect of CTLA-4. Primary control of CD28 and CTLA-4 expression maintains a balance between activating and inhibitory signals, ensuring the development of an effective immune response while protecting against the development of autoimmunity.

[0004] Programmed death 1 (PD-1) is a 50-55 kDa type I transmembrane receptor that is a member of the CD28 family. PD-1 is involved in the regulation of T cell activation and is expressed on T cells, B cells, and myeloid cells. Two ligands for PD-1, PD ligand 1 (PD-L1) and ligand 2 (PD-L2), have been identified and have costimulatory properties.

[0005] Programmed cell death 1 ligand 1 (PD-L1), also known as cluster of differentiation (CD274) or B7 homolog 1 (B7-H1), is a member of the B7 family that regulates the activation or inhibition of the PD-1 receptor. The PD-L1 open reading frame encodes a putative type 1 transmembrane protein of 290 amino acids, containing two extracellular Ig domains (an N-terminal V-like domain and an Ig C-like domain), a hydrophobic transmembrane domain, and a 30-amino acid cytoplasmic tail. The 30-amino acid intracellular (cytoplasmic) domain does not contain any obvious signaling motifs but does contain potential sites for protein kinase C phosphorylation. The complete amino acid sequence of PD-L1 can be found in the NCBI reference sequence NP_054862.1 (SEQ ID NO: 1), which references numerous journal articles [1]. The PD-L1 gene is conserved in chimpanzees, rhesus monkeys, dogs, cattle, mice, rats, chickens, and zebrafish. The murine form of PD-L1 has 69% amino acid identity with the human form of PD-L1 and also shares a conserved structure.

[0006] In humans, PD-L1 is expressed on several T cells, including activated and anergic / exhausted T cells. PD-L1 is expressed on immune cell types, including naive and activated B cells, as well as myeloid dendritic cells (DCs), monocytes, and mast cells. PD-L1 is also expressed on non-immune cells, including pancreatic islets, Kupffer cells of the liver, vascular endothelium, and selected epithelia, such as airway epithelium and renal tubular epithelium, and its expression is enhanced during inflammatory episodes. PD-L1 expression is also associated with breast cancer (e.g., triple myeloma). negative breast cancer and inflammatory breast cancer), ovarian cancer, cervical cancer, colon cancer, colorectal cancer, lung cancer (e.g., non-small cell lung cancer), renal cancer (e.g., renal cell carcinoma), gastric cancer, esophageal cancer, bladder cancer, hepatocellular carcinoma, head and neck cancer. It is found at elevated levels in some tumors, such as squamous cell carcinoma of the nasal cavity (SCCHN), as well as pancreatic cancer, melanoma, and uveal melanoma.

[0007] PD-1 / PD-L1 signaling negatively regulates T cell responses, thereby affecting the immune system. PD-L1 is thought to play critical, non-redundant functions within the system. This regulation is involved in T cell development in the thymus, control of chronic inflammatory responses, and maintenance of both peripheral tolerance and immune privilege. Upregulation of PD-L1 may enable cancer to evade the host immune system, and in many cancers, PD-L1 expression is associated with reduced survival and poor prognosis. Therapeutic monoclonal antibodies capable of blocking the PD-1 / PD-L1 pathway offer promise for the treatment of cancer patients. It may enhance anti-tumor immune responses. Published clinical data show a correlation between clinical response and tumor membrane expression of PD-L1, and an even stronger correlation between lack of clinical response and lack of membrane-localized PD-L1 protein [2, 3]. PD-L1 expression on tumors or tumor-infiltrating leukocytes is therefore a candidate molecular marker for use in patient selection for immunotherapy, e.g., immunotherapy using anti-PD-L1 antibodies [4]. Enrichment of patients based on PD-L1 surface expression may support treatment with drugs targeting the PD-1 / PD-L1 pathway. There is also evidence for the presence of an ongoing immune response, such as tumor-infiltrating CD8+ T cells, or signs of cytokine activation, such as IFNγ.

[0008] Further evidence of PD-L1 expression and its correlation with disease will emerge from numerous ongoing clinical trials. Atezolizumab is the most advanced anti-PD-L1 antibody in development, and phase II trials have demonstrated its role in metastatic urothelial carcinoma and NSCLC, particularly in the PD-L1 expression in the tumor microenvironment. + The study showed a therapeutic effect of immune cells [5, 6]. Recent results from a phase III trial showed improved survival in patients receiving azelizumab compared with chemotherapy, regardless of tumor expression of PD-L1 (Rittmeye r et al., 2017, The Lancet, 389(10066), 255-265).

[0009] Another member of the CD28 gene family, ICOS (inducible T-cell costimulator), was identified in 1999 [7]. It is a 55-kDa transmembrane protein that exists as a disulfide-linked homodimer with two differentially glycosylated subunits. ICOS is exclusively expressed on T lymphocytes and is found in various T-cell subsets. It is present at low levels on naive T lymphocytes, but its expression is rapidly induced upon immune activation and upregulated in response to proinflammatory stimuli, such as TCR engagement and costimulation with CD28 [8, 9]. ICOS plays a role in the late stages of T-cell activation, the late stages of memory T-cell formation, and, importantly, in the regulation of humoral responses via T-cell-dependent B-cell responses [10, 11]. Intracellularly, ICOS binds to PI3K and activates the kinase phophoinositide-dependent kinase 1 (PDK1). 1) and protein kinase B (PKB). Activation of ICOS leads to cell death. In the absence of ICOS (ICOS knockout), the cell metabolism is upregulated. In the presence of anti-ICOS (anti-ICOS) or anti-ICOS neutralizing antibodies, there is suppression of the pro-inflammatory response.

[0010] ICOS is expressed on B cells and antigen-presenting cells (APCs) by the ICOS ligand (IC It binds to the TCR mediator of antigen-mediated responses [12, 13]. ICOS expression on regulatory T cells is important because this cell type has been suggested to play a negative role in the immune surveillance of cancer cells, and there is emerging evidence for this in ovarian cancer

[14] . Importantly, ICOS expression has been reported to be higher on intratumoral regulatory T cells (TReg) compared to CD4+ and CD8+ effector cells present in the tumor microenvironment. Fc-mediated cellular effectors Depletion of Tregs using antibodies with inhibitory function has shown strong antitumor efficacy in preclinical models

[15] . Growing evidence indicates that ICOS contributes to antitumor efficacy in both animal models and patients treated with immune checkpoint inhibitors. While the antitumor efficacy of anti-CTLA4 therapy is reduced in ICOS- or ICOSL-depleted mice

[16] , in normal mice, ICOS ligand increases the efficacy of anti-CTLA4 treatment in melanoma and prostate cancer

[17] . Furthermore, in humans, a previous study of patients with advanced melanoma showed elevated ICOS levels after ipilimumab (anti-CTLA4) treatment

[18] . Additionally, ICOS expression is upregulated in patients with bladder cancer treated with anti-CTLA4

[19] . In cancer patients treated with anti-CTLA4 therapy, the majority of tumor-specific IFNγ-producing CD4 T cells are ICOS-positive, and it has also been observed that a sustained increase in ICOS-positive CD4 T cells correlates with survival [18, 19, 20].

[0011] WO2016 / 120789 describes anti-ICOS antibodies that activate T cells and their use for treating cancer, infections, and / or sepsis. Several murine anti-ICOS antibodies have been generated, and a subset of them have been reported to be agonists of the human ICOS receptor. Antibody "422.2" was selected as the lead anti-ICOS antibody and humanized to produce a human "IgG4PE" antibody designated "H2L5." H2L5 has an affinity of 1.34 nM for human ICOS and 0.95 nM for cynomolgus monkey ICOS, and has been reported to induce cytokine production in T cells and upregulate T cell activation markers in conjunction with CD3 stimulation. However, mice bearing transplanted human melanoma cells were reported to show only minimal tumor growth delay or increased survival when treated with H2L5 hIgG4PE compared to control-treated groups. Furthermore, in combination experiments with ipilimumab (anti-CTLA-4) or pembrolizumab (anti-PD-1), this antibody did not significantly further inhibit tumor growth compared to ipilimumab or pembrolizumab monotherapy. Finally, in mice bearing transplanted colon cancer cells (CT26), a low dose of a murine cross-reactive H2L5 surrogate combined with a murine surrogate of ipilimumab or pembrolizumab only modestly improved overall survival compared with anti-CTL4 and anti-PD1 therapy alone. A similar lack of robust therapeutic effect was observed in mice bearing transplanted EMT6 cells.

[0012] Further examples of anti-ICOS antibodies are described in WO2016 / 154177. These antibodies have been reported to be agonists of CD4+ T cells, including effector CD8+ T cells (TEff), and to deplete regulatory T cells (TReg). The selective effect of the antibodies on TEff cells versus TReg cells has been described, with these antibodies preferentially depleting TRegs but having minimal effect on TEff cells, which express lower levels of ICOS. Anti-ICOS antibodies have been proposed for use in cancer treatment, and combination therapy with anti-PD-1 or anti-PD-L1 antibodies has been described.

[0013] Although significant progress has been made in recent years in the field of cancer immunotherapy, current response rates of cancer immunotherapy drugs remain low. For example, the response rate of the anti-PD-1 antibody nivolumab in melanoma is approximately 30%, while the response rate of the anti-PD-L1 atezolizumab in its Phase II clinical trial for urothelial carcinoma was approximately 15% overall, regardless of PD-L1 expression, or 26% in patients with PD-L1-expressing tumors. Attempts to enhance the efficacy of cancer immunotherapy include combining multiple drugs, such as antibodies with traditional chemotherapy or radiation, and drugs targeting different immune checkpoint inhibitors. The combination of nivolumab (anti-PD-1) and ipilimumab (anti-CTLA-4) has shown efficacy in previously untreated cases of melanoma, with promising headline response rates and overall survival

[21] . However, while combination therapy may produce new or enhanced biological effects in vivo, this carries the associated risks of negative drug interactions and new or worsening side effects. Immune checkpoint inhibitor therapy It is already associated with immune-related adverse events, including neurological events ranging from mild headache to life-threatening encephalitis.

[22] Furthermore, at a practical level, treatment regimens involving the combination of multiple therapeutic agents have the disadvantages of complex dosing schedules and high costs. Summary of the Invention

[0014] The present invention relates to antigen-binding molecules that contain multiple antigen-binding sites ("multispecific antigen-binding molecules"), including an antigen-binding site for ICOS and an antigen-binding site for another target antigen, for example, PD-L1. This relates to the "combined molecule."

[0015] Both ICOS and PD-L1 are expressed after primary T cell activation. PD-L1 negatively regulates T cell activation, and inhibition of PD-L1 signaling has been clinically validated as an approach to upregulate T cell immune responses against tumor cells. In this context, the parallel depletion of ICOS-high Tregs and stimulation of ICOS-low effector T cells can enhance T cell activation to promote antitumor activity.

[0016] Multispecific antigen-binding molecules that block the negative regulatory activity of PD-L1 on PD1+ T cells and enhance T cell activation by transmitting positive signals via ICOS offer therapeutic potential in the treatment of cancer and other conditions where it is desirable to upregulate T cell immune responses. The fate of T cells in the tumor microenvironment and tumor-draining lymph nodes is influenced by the balance between inhibitory and activating receptors, and molecules that bind to and inhibit PD-L1 while acting as ICOS agonists effectively block negative signals (inhibitory PD-L1). 1 receptor) into a positive signal (from the ICOS co-activator receptor). The immune synapse between a T cell and an antigen-presenting cell (APC) or tumor cell can be envisioned as a receptor-dense space in which the balance of receptor occupancy determines signal transduction within the T cell, and this receptor occupancy is determined by the identity and concentration of receptors displayed on the surface of the APC / tumor cell involved. This is determined by the binding sites for ICOS and PD-L1. Multispecific molecules bearing binding sites for ICOS and PD-L1 can act directly at this immune synapse to alter the balance of signals received by T cells, shifting the balance toward TEff activation. Rather than separate antigen-binding molecules, the combination of anti-PD-L1 and anti-ICOS in one multispecific antigen-binding molecule provides a single agent that can act as a molecular switch. Multispecific molecules can cross-link ICOS and PD-L1 on different cells (Figure 1).

[0017] In addition to binding its two cognate antigens, the multispecific antigen-binding molecule may incorporate other moieties, such as antibody effector regions, to recruit cell-killing functions, which may further include, for example, depletion of Tregs that highly express ICOS on the cell surface and / or PD-L. Depletion of cancer cells expressing ICOS-1 tips the immune balance towards T cell activation and cancer cell killing. Bispecific antibodies that bind to ICOS and PD-L1 can induce ADCC against PD-L1+ immunosuppressive cells (e.g., MDSCs, tumor cells) and / or ADCC against ICOS+ immunosuppressive cells (e.g., Tregs).

[0018] The multispecific antigen-binding molecule according to the present invention comprises an antibody ( For example, it may be a bispecific antibody or a biconjugate antibody. Many different combinations of antibodies are possible, and many examples are provided herein. The antibody may be bivalent for both target antigens. For example, the antibody may be a FIT-Ig that contains two ICOS-binding Fab domains and two PD-L1-binding domains (e.g., as shown in Figure 2). Alternatively, the antibody may be a mAb that contains two ICOS-binding Fab domains and an Fc region that contains two binding sites for PD-L1 (i.e., a PD-L1-binding Fcab), as shown in Figure 3. 2 Examples of ICOS antibody and PD-L1 antibody sequences, including VH and VL domain sequences, are described herein and may be included in multispecific antibodies.

[0019] Multispecific antigen-binding molecules that bind ICOS and PD-L1 may increase the response rate of tumors that have already responded to PD-L1 or ICOS monotherapy, increasing the proportion of patients in whom an anti-tumor response is observed, potentially improving the level of response, reducing tumor growth, and prolonging survival compared to monotherapy. Some tumors do not respond to either anti-ICOS or anti-PD-L1 antibodies, but may respond to multispecific antibodies that bind ICOS and PD-L1. Anti-ICOS / anti-PD-L1 bispecific binding molecules may also be used to target tumors against antigens, e.g., tumor antigens. Multispecific antigen-binding molecules can be used to induce long-term memory, thereby providing protection against tumor regrowth. Thus, the multispecific approach described herein offers advantages in improving response rates, duration of response, and patient survival in the context of cancer treatment. Furthermore, multispecific antigen-binding molecules can be administered to patients using simpler treatment regimens compared to multiple separate formulations of different therapeutic agents. [Brief explanation of the drawings]

[0020] [Figure 1] Redirection of immune checkpoint regulation. Multispecific antigen-binding molecules result in simultaneous blockade of PD-L1 receptors on antigen-presenting cells (APCs) or tumor cells and agonism of ICOS receptors on T effector cells, switching negative regulatory signals to positive regulatory signals at the T cell immune synapse. [Figure 2] FIT-Ig format of a bispecific antibody that binds to ICOS and PD-L1. (i) Assembled FIT-Ig antibody (ii) Polypeptide chains contained in the FIT-Ig antibody. Construct #1 is a polypeptide comprising, in N to C orientation, the light chain variable region (VL) and light chain constant region (CL) of antibody "A," fused to the heavy chain variable region (VH) and heavy chain constant region (CH1, CH2, CH3) of antibody "B." Preferably, no linker is included between the CL domain and the VHB domain. Construct #2 is a polypeptide fusion of the heavy chain variable (VH) region and CH1 of antibody "A." Construct #3 is a polypeptide fusion of the light chain variable (VL) region and light chain constant (CL) region of antibody "B." FIT-Ig can be constructed where antibody "A" is anti-ICOS and antibody "B" is anti-PD-L1, or where antibody "A" is anti-PD-L1 and antibody "B" is anti-ICOS. [Figure 3] Example of a mAb2 IgG format of a bispecific antibody that binds to ICOS and PD-L1. mAb2 is a homodimeric IgG containing two anti-ICOS Fabs and two CH3 domains (anti-PD-L1 Fcab regions), each with three binding loops that form the PD-L1 binding site. [Figure 4-1](A) STIM001 and STIM003 mAb2 bind to recombinant human ICOS protein. Data are representative of three experiments. (B) STIM001 and STIM003 mAb2 bind to recombinant mouse ICOS protein. Data are representative of three experiments. [Figure 4-2] (C) Human STIM001 and STIM003 mAb2 binding to recombinant human PD-L1 protein. Data are representative of three experiments. (D) Mouse STIM001 and STIM003 mAb2 binding to recombinant mouse PD-L1 protein. Data are representative of three experiments. [Figure 5] Results of the ICOS FACS binding assay described in Example 4. A) mAb2 binding to human ICOS expressed on CHO cells. Data are representative of three experiments. B) mAb2 binding to mouse ICOS expressed on CHO cells. Data are representative of three experiments. [Figure 6] Results of human PD-L1 FACS binding assay as described in Example 4. A) Human PD-L1 binding FACS using anti-human IgG detection. Binding profiles of STIM001_289, STIM003_289, and IgG1_289 anti-PD-L1 mAb2 and their respective mAb controls. B) Human PD-L1 binding FACS using conjugated human ICOS-labeled AlexaFluor647 detection. Binding profiles of STIM001_289, STIM003_289, and IgG1_289 anti-PD-L1 mAb2 and their respective mAb controls. [Figure 7] Results of mouse PD-L1 FACS binding assay as described in Example 4. A) Mouse PD-L1 binding FACS using anti-human IgG detection. Binding profiles of STIM001_457, STIM003_457, and IgG1_438 anti-PD-L1 mAb2 and their respective monospecific mAb controls. B) Mouse PD-L1 binding FACS using conjugated human ICOS-labeled AlexaFluor647 detection. Binding profiles of STIM001_457, STIM003_457, and IgG1_438 anti-PD-L1 mAb2 and their respective monospecific mAb controls. [Figure 8] (A) Engagement of human STIM001 and STIM003 mAb2 Fc to human FcγRIIIa on effector cells as described in Example 5b. Data are representative of three experiments. (B) Engagement of murine STIM001 and STIM003 mAb2 Fc to FcγRIIIa on effector cells as described in Example 5b. Data are representative of three experiments. [Figure 9-1] Concentration-dependent study of STIM001_289- and STIM003_289-mediated ADCC on ICOS-transfected CCRF-CEM cells using freshly isolated NK cells as effector cells for three independent donors (Panels A, B, and C) as described in Example 5c. Effector and target cells (effector:target ratio 5:1) were incubated with antibody for 4 hours. Dye release from lysed target cells was measured as described in the kit manufacturer's instructions. 100% release was determined using lysis buffer. Basal killing (no Ab) is indicated by the dotted line at the bottom of each graph. [Figure 9-2] Concentration-dependent study of STIM001_289- and STIM003_289-mediated ADCC on ICOS-transfected CCRF-CEM cells using freshly isolated NK cells as effector cells for three independent donors (Panels A, B, and C) as described in Example 5c. Effector and target cells (effector:target ratio 5:1) were incubated with antibody for 4 hours. Dye release from lysed target cells was measured as described in the kit manufacturer's instructions. 100% release was determined using lysis buffer. Basal killing (no Ab) is indicated by the dotted line at the bottom of each graph. [Figure 10]A) Results of a mouse ICOS ligand neutralization HTRF assay using the mouse ICOS receptor as described in Example 6. Neutralization profiles of STIM001_289, STIM001_457, STIM003_289, and STIM003_457 mAb2. Data are representative of three experiments. B) Results of a human ICOS ligand neutralization HTRF assay using the human ICOS receptor as described in Example 6. Neutralization profiles of STIM001_289, STIM001_457, STIM003_289, and STIM003_457 mAb2. Data are representative of three experiments. [Figure 11] Results of the PD-L1 neutralization assay described in Example 7. A) Human PD-L1 neutralization FACS to human PD1. Binding profiles of STIM001_289, STIM003_289, and IgG1_289 anti-PD-L1 mAb2 and their respective mAb controls. B) Human PD-L1 neutralization FACS to human CD80. Binding profiles of STIM001_289, STIM003_289, and IgG1_289 anti-PD-L1 mAb2 and their respective mAb controls. [Figure 12] A) Data from a mouse PD-L1 neutralization assay (FACS) against mouse PD1 as described in Example 7. Binding profiles of STIM001_457, STIM003_457, and IgG1_438 anti-PD-L1 mAb2 and their respective controls. B) Data from a mouse PD-L1 neutralization assay (FACS) against mouse CD80. Binding profiles of STIM001_457, STIM003_457, and IgG1_438 anti-PD-L1 mAb2 and their respective controls. [Figure 13-1]Concentration-dependent study of the agonistic effects of STIM001_289 and STIM003_289 versus STIM001 and STIM003 on isolated human T cells costimulated with CD3 / CD28 Dynabeads for 3 days. IFN-γ production was used as a readout for ICOS agonism. All antibodies were plate-bound and compared to their isotype controls. Mean ± SD values ​​of technical replicates and a nonlinear regression curve (variable slope, 4 parameters) are shown for one donor (278) in panels A and B. Mean levels of IFN-γ induced by a single given dose (3.3 μM) for all four donors are shown. Each point represents an independent donor, identifiable by its number, and the median value for the four donors is indicated by the line. Significance was assessed using the Friedman statistical test, and p values ​​are shown on the graphs. [Figure 13-2] Concentration-dependent study of the agonistic effects of STIM001_289 and STIM003_289 versus STIM001 and STIM003 on isolated human T cells costimulated with CD3 / CD28 Dynabeads for 3 days. IFN-γ production was used as a readout for ICOS agonism. All antibodies were plate-bound and compared to their isotype controls. Mean ± SD values ​​of technical replicates and a nonlinear regression curve (variable slope, 4 parameters) are shown for one donor (278) in panels A and B. Mean levels of IFN-γ induced by a single given dose (3.3 μM) for all four donors are shown. Each point represents an independent donor, identifiable by its number, and the median value for the four donors is indicated by the line. Significance was assessed using the Friedman statistical test, and p values ​​are shown on the graphs. [Figure 14]Concentration-dependent study of the effects of STIM001_289, STIM003_289, and IgG1_289 versus PD-L1 AbV on cytokine production by CD45RO+ T cell cocultures (TCR activation) with autologous monocytes in the presence of CD3 antibodies. This assay, as described in Example 9, uses IFN-γ production as a readout of neutralization of the PD-1 / PD-L1 interaction by the test antibodies. All antibodies were compared to an isotype control (IgG1). Raw data from one independent donor (288) are shown in the top panel. Values ​​were normalized using basal IFN-γ levels (single / T cell dotted line) to calculate fold increases in IFN-γ. Example data comparing the increases in IFN-γ induced by a single given dose (10 nM) for all seven donors are shown in the bottom panel. Each point represents an independent donor, identified by its number, and the median is indicated by the line. Significance was assessed using the Friedman statistical test (*<0.05 and **<0.01). [Figure 15] Effect of STIM001_457 and STIM003_457 bispecific antibodies in the J558 syngeneic tumor study described in Example 10. Each treatment group is represented by a "spider plot" showing tumor size for individual animals (n=10 or n=8 per group). Both bispecific antibodies demonstrated significant antitumor efficacy, with 5 of 8 animals treated with STIM001_457 (B) and 4 of 8 animals treated with STIM003_457 (C) cured of their disease by day 37. The number of animals cured of disease is indicated at the bottom right of each graph. Planned dosing days are indicated by dotted lines (days 11, 15, 18, 22, 25, and 29). [Figure 16] Kaplan-Meier survival curves for % mouse survival / study time after different treatments as described in Example 10. Median survival times for animals in saline (open squares) and STIM003_457 (triangles) were 18 and 27.5 days, respectively. Median survival times were not available for STIM001_457 (filled circles). [Figure 17-1]Data from the CT26 in vivo efficacy study described in Example 11a. Each treatment group is represented by a "spider plot" showing tumor size for individual animals (n=10 per group). For each group, the number of animals cured of disease is indicated at the bottom left of each graph. Dosing occurred on days 6, 8, 10, 13, 15, and 17, with dosing times indicated by shaded areas. (A) Saline; (B) IgG1_457 LAGA control; (C) STIM003_457; (D) STIM001_457. [Figure 17-2] Data from the CT26 in vivo efficacy study described in Example 11a. Each treatment group is represented by a "spider plot" showing tumor size for individual animals (n=10 per group). For each group, the number of animals cured of disease is indicated at the bottom left of each graph. Dosing occurred on days 6, 8, 10, 13, 15, and 17, with dosing times indicated by shaded areas. (A) Saline; (B) IgG1_457 LAGA control; (C) STIM003_457; (D) STIM001_457. [Figure 18] Kaplan-Meier plot for the CT26 study described in Example 11a. Circles: saline control. Squares: IgG1_487 control. Upward triangles: STIM003_457. Downward triangles: STIM001_457. [Figure 19] Treatment with ICOS / PD-L1 antibodies results in long-term antitumor memory responses in animals previously cured of CT26 tumors. Mice cured of CT26 colon cancer, as described in Example 11b, were rechallenged subcutaneously in the left flank with either 2.5 x 10 EMT-6 cells (n = 4 mice per group) or 1 x 10 CT26 cells (n = 5 mice per group). Spider plots show tumor growth over 20 days after inoculation of EMT-6 or CT26 cells. [Figure 20-1]Results of the A20 in vivo efficacy study described in Example 12. Each treatment group is represented by a "spider plot" showing tumor size for individual animals (n=10 per group). For each group, the number of animals cured of disease is indicated at the bottom left of each graph. Dosing occurred on days 8, 11, 15, 18, 22, and 25. [Figure 20-2] Results of the A20 in vivo efficacy study described in Example 12. Each treatment group is represented by a "spider plot" showing tumor size for individual animals (n=10 per group). For each group, the number of animals cured of disease is indicated at the bottom left of each graph. Dosing occurred on days 8, 11, 15, 18, 22, and 25. [Figure 21] Results of the A20 in vivo efficacy study described in Example 12. Humane endpoint survival statistics were calculated from Kaplan-Meier curves using GraphPad Prism V7.0. This approach was used to determine whether a particular treatment is associated with improved survival. [Figure 22-1] Data from the EMT6 in vivo efficacy study described in Example 13. Each treatment group is represented by a "spider plot" showing tumor size for individual animals (n=10 per group). A) Saline, B) Anti-PD-L1 mAb2 control antibody, C) STIM003_457, D) STIM001_457. For each group, the number of animals cured of disease is indicated at the bottom left of each graph. Dosing occurred on days 6, 9, 13, 16, 20, and 23. [Figure 22-2] Data from the EMT6 in vivo efficacy study described in Example 13. Each treatment group is represented by a "spider plot" showing tumor size for individual animals (n=10 per group). A) Saline, B) Anti-PD-L1 mAb2 control antibody, C) STIM003_457, D) STIM001_457. For each group, the number of animals cured of disease is indicated at the bottom left of each graph. Dosing occurred on days 6, 9, 13, 16, 20, and 23. [Figure 23]Survival times (study duration) of animals treated with saline (closed circles), anti-PD-L1 mAb2 control antibody (squares), STIM003_457 (upward triangles), or STIM001_457 (downward triangles) as described in Example 13. Both ICOS / PD-L1 bispecific antibodies significantly improved the overall survival of animals compared to those treated with saline. [Figure 24-1] Bispecific efficacy in the EMT6 model as described in Example 13. A) IgG1 LAGA hybrid control mAb2 antibody with anti-PD-L1 457 Fcab, B) STIM003 in combination with anti-PD-L1 antibody (mouse IgG2a format), C) STIM001_457 bispecific antibody, D) STIM003_457 bispecific antibody. [Figure 24-2] Bispecific efficacy in the EMT6 model as described in Example 13. A) IgG1 LAGA hybrid control mAb2 antibody with anti-PD-L1 457 Fcab, B) STIM003 in combination with anti-PD-L1 antibody (mouse IgG2a format), C) STIM001_457 bispecific antibody, D) STIM003_457 bispecific antibody. [Figure 25] Kaplan-Meier (compassionate endpoint) showing superior efficacy of treatment with PD-L1 / ICOS bispecific antibodies (downward triangles for STIM001_457 and upward triangles for STIM003_457) compared to combined administration of anti-PD-L1 and anti-ICOS monospecific antibodies (filled diamonds) in the EMT6 model described in Example 13. Data from saline control treatments are shown as filled circles. Data for the IgG1 LAGA hybrid control mAb2 antibody with anti-PD-L1 457 Fcab are shown as open squares. [Figure 26-1] Representative examples from two independent experiments in the PD-L1-dependent ICOS agonism assay reported in Example 14. (A) BSA. (B) B7-H1-Fc. (C) Goat anti-human IgG Fcg fragment-specific F(ab')2. [Figure 26-2]Representative examples from two independent experiments in the PD-L1-dependent ICOS agonism assay reported in Example 14. (A) BSA. (B) B7-H1-Fc. (C) Goat anti-human IgG Fcg fragment-specific F(ab')2. [Figure 26-3] Representative examples from two independent experiments in the PD-L1-dependent ICOS agonism assay reported in Example 14. (A) BSA. (B) B7-H1-Fc. (C) Goat anti-human IgG Fcg fragment-specific F(ab')2. [Figure 27] Identification of four distinct quadrants on a dot plot graph of mAb2 antibody in a flow cytometric PD-L1 / ICOS cell recruitment assay. [Figure 28] Titration of mAb2 and monospecific antibodies in a PD-L1 / ICOS cell recruitment assay by flow cytometry. CHO human PD-L1 and CHO human ICOS were stained with CellTrace™ Far Red and CellTrace™ Violet, respectively, and co-incubated in the presence of antibody before fluorescent detection and identification of double-positive populations. Data shown are representative of two independent experiments. [Figure 29-1] FACS analysis revealed that STIM003_457 and STIM001_457 significantly depleted regulatory T cells (TRegs) in tumors and increased the effector cell:TReg ratio. BALB / c mice were treated with saline, STIM003_457, or STIM001_457 (n=8 per group) on days 13 and 15 after subcutaneous implantation of CT-26.WT tumor cells. The percentage of TRegs among total live tumor cells (A) and total CD4+ cells (B) was significantly reduced in response to both antibodies compared to saline. The ratio of CD4+ effector cells to TRegs (C) and the ratio of CD8+ cells to TRegs (D) were significantly increased compared to control. Kruskal-Wallis tests were performed, followed by post-hoc Dunn's test. *p<0.05; **p<0.01; ***p<0.001; ****p<0.0001. [Figure 29-2]FACS analysis revealed that STIM003_457 and STIM001_457 significantly depleted regulatory T cells (TRegs) in tumors and increased the effector cell:TReg ratio. BALB / c mice were treated with saline, STIM003_457, or STIM001_457 (n=8 per group) on days 13 and 15 after subcutaneous implantation of CT-26.WT tumor cells. The percentage of TRegs among total live tumor cells (A) and total CD4+ cells (B) was significantly reduced in response to both antibodies compared to saline. The ratio of CD4+ effector cells to TRegs (C) and the ratio of CD8+ cells to TRegs (D) were significantly increased compared to control. Kruskal-Wallis tests were performed, followed by post-hoc Dunn's test. *p<0.05; **p<0.01; ***p<0.001; ****p<0.0001. [Figure 30-1] FACS analysis shows that STIM003_457 and STIM001_457 have little effect on regulatory T cell (TReg) levels in the spleens of CT-26.WT tumor-bearing mice. STIM003_457 shows marginal TReg depletion as a percentage of total viable cells, while STIM001_457 has no effect (A). No clear changes were observed in the effect of the bispecific on TRegs as a percentage of total CD4+ cells (B). No significant changes were observed in the ratios of CD4+ effector cells to TRegs (C) and CD8+ cells to TRegs (D). Kruskal-Wallis tests were performed, followed by post-hoc Dunn's tests. *p<0.05. [Figure 30-2]FACS analysis shows that STIM003_457 and STIM001_457 have little effect on regulatory T cell (TReg) levels in the spleens of CT-26.WT tumor-bearing mice. STIM003_457 shows marginal TReg depletion as a percentage of total viable cells, while STIM001_457 has no effect (A). No clear changes were observed in the effect of the bispecific on TRegs as a percentage of total CD4+ cells (B). No significant changes were observed in the ratios of CD4+ effector cells to TRegs (C) and CD8+ cells to TRegs (D). Kruskal-Wallis tests were performed, followed by post-hoc Dunn's tests. *p<0.05. [Figure 31] FACS analysis shows increased ICOS ligand (ICOS-L) expression on B cells in the spleens of CT26-WT tumor-bearing mice treated with STIM003_457 and STIM001_457. Compared to saline, both bispecific antibodies caused a significant increase in the percentage of B cells expressing ICOS-L in the spleen (A). A significant increase in the mean fluorescence intensity (relative expression) of ICOS-L on B cells was also seen in both bispecific groups compared to the saline group (B). Kruskal-Wallis test was performed, followed by a post-hoc Dunn's test. *p<0.05. ****p<0.0001. [Figure 32] Graph showing the average body weight of mice over 46 days in different treatment groups. Vertical lines indicate the days animals were intraperitoneally dosed. Note that the slight decrease in average body weight observed from day 35 for Group 3 (aCTLA-4 monotherapy) and Group 7 (STIM003 / aPDL1 combination) is due to some animals dropping out of the tumor size study. [Figure 33] Spider plot graphs A-G show CT26 tumor size over time for individual animals with different treatment responses. "Triple combinations" of antibodies (against ICOS, PD-L1 and PD1, or CTLA-4) were associated with the most pronounced anti-tumor responses in the CT26 model. Vertical lines indicate the day animals were intraperitoneally administered. For combinations, antibodies were injected simultaneously. Numbers at the bottom right corner of each graph indicate the number of animals still on study at day 46 (40 days after initiation of treatment). DETAILED DESCRIPTION OF THE INVENTION

[0021] definition Unless otherwise defined herein, scientific and technical terms shall have the meanings commonly understood by those skilled in the art. Further, unless otherwise required by context, singular terms shall include pluralities and plural terms shall include the singular.

[0022] The singular terms "a," "an," and "the" include plural referents unless the context clearly indicates otherwise. Similarly, the word "or" is intended to include "and" unless the context clearly indicates otherwise. Although methods and materials similar or equivalent to those described herein can be used in the practice or testing of the present disclosure, suitable methods and materials are described below. The abbreviation "eg" is derived from the Latin "exempli gratia," and is used herein to indicate a non-limiting example. Thus, the abbreviation "eg" is synonymous with the term "for example."

[0023] In the specification and claims, the term "about" is used to modify, for example, the amounts, concentrations, volumes, processing temperatures, processing times, yields, flow rates, pressures, and similar values ​​and ranges of components in compositions used in describing embodiments of the present disclosure. The term "about" refers to variations in quantity that may occur, for example, through typical measuring and handling procedures for making a compound, composition, concentrate, or use formulation, through accidental errors in these procedures, through differences in purity in the manufacture, source, or starting materials or components used to carry out the method, and similar straightforward considerations. The term "about" also refers to amounts that vary with aging of a formulation having a particular initial concentration or mixture, and amounts that vary with the aging of a formulation having a particular initial concentration or mixture, and amounts that vary with the aging of a formulation having a particular initial concentration or mixture. It also encompasses amounts that vary by mixing or processing the formulation with concentrations or mixtures. When modified by the term "about," the appended claims include equivalents of these amounts.

[0024] As used herein, "administering" or "administration" refers to mucosal delivery, intradermal delivery, intravenous delivery, intramuscular delivery, and / or other administration methods described herein or known in the art. By any other method, a substance present outside the body (e.g., an antibody provided herein) can be detected. This refers to the act of injecting or otherwise physically delivering a drug (pD-L1 antibody) into a patient. When a condition is being treated, administration of the substance typically occurs after the onset of the disease or its symptoms. When a disease or its symptoms is being prevented, administration of the substance typically occurs before the onset of the disease or its symptoms.

[0025] The terms "antibody," "immunoglobulin," or "Ig" may be used interchangeably herein and refer to an immunoglobulin molecule that recognizes a target, such as a protein, polypeptide, peptide, carbohydrate, polynucleotide, lipid, or a combination of the foregoing, and specifically binds to the target at at least one antigen recognition site within the variable region of the immunoglobulin molecule. As used herein, the term "antibody" includes intact polyclonal antibodies, intact monoclonal antibodies, antibody fragments (e.g., Fab, Fab', F(ab')2, and Fµ fragments), single-chain Fµ (scFv) variants, bispecific antibodies (bispecific antibodies), and the like. Multispecific antibodies, chimeric antibodies, humanized antibodies, human antibodies, antigen-determining portions of antibodies, etc. The term "antibody" also encompasses fusion proteins containing the antigen-recognition site, as well as any other modified immunoglobulin molecule containing an antigen recognition site, so long as it exhibits the desired biological activity. The term "antibody" can also refer to a Y-shaped glycoprotein with a molecular weight of approximately 150 kDa, consisting of four polypeptide chains (two light (L) chains and two heavy (H) chains). Five mammalian Ig heavy chain isotypes exist, designated by the Greek letters alpha (α), delta (δ), epsilon (ε), gamma (γ), and mu (μ). The type of heavy chain defines the antibody class: IgA, IgD, IgE, IgG, and IgM, respectively. The γ and α classes are further divided into subclasses, e.g., IgG1, hIgG2, mIgG2A, mIgG2B, IgG3, IgG4, IgA1, and IgA2, based on differences in constant domain sequence and function. In mammals, two types of immunoglobulin light chains exist: λ and κ. The "variable region" or "variable domain" of an antibody refers to the amino-terminal domain of the heavy or light chain of the antibody. The variable domains of the heavy and light chains can be designated "VH" and "VL," respectively. These domains are generally the most variable parts of an antibody (relative to other antibodies of the same class) and contain the antigen-binding site.

[0026] The antibodies described herein may be oligoclonal, polyclonal, monoclonal (including full-length monoclonal), camelized, chimeric, CDR-grafted, multispecific, bispecific (including biconjugated), catalytic, chimeric, humanized, fully human, or anti-idiotypic antibodies, which may include antibodies that may be labeled in soluble or conjugated form, as well as fragments, variants, or derivatives thereof, either alone or in combination with other amino acid sequences provided by known techniques. Antibodies may be from any species. The antibodies described herein may be naked or conjugated to other molecules, such as toxins, radioisotopes, etc.

[0027] The terms "antigen-binding domain," "antigen-binding region," "antigen-binding fragment," and similar terms refer to the portion of an antibody (e.g., the complementarity-determining region (CDR)) that contains the amino acid residues that interact with an antigen and confer to the binder its specificity and affinity for the antigen. The antigen-binding region can be derived from any animal species, including rodents (e.g., rabbits, rats, or hamsters) and humans. Preferably, the antigen-binding region is of human origin. The antibodies described herein Antibody binding fragments include single chain Fv (scFv), single chain antibodies, domain antibodies, Fv fragments, Fab fragments, F(ab') fragments, F(ab')2 fragments, antibody fragments exhibiting desired biological activity, disulfide stabilized variable regions (dsFv), dimeric variable regions (diabodies), anti-idiotypes (antibodies), and the like. Id) antibodies (e.g., anti-Id antibodies against antibodies), intrabodies, linear antibodies, single-chain antibody molecules Antibodies and antibody fragments described herein may include antibodies, as well as multispecific antibodies formed from any of the antibody fragments and epitope-binding fragments described above. In particular, the antibodies and antibody fragments described herein may include immunoglobulin molecules and immunologically active fragments of immunoglobulin molecules, i.e., molecules containing antigen-binding sites. Digestion of antibodies with the enzyme papain results in two identical antigen-binding fragments, also known as "Fab" fragments, which lack antigen-binding activity but have the ability to crystallize. "Fab," as used herein, refers to an antibody fragment containing one constant domain and one variable domain from each of the heavy and light chains. The term "Fc region" herein defines the C-terminal region of an immunoglobulin heavy chain and includes the native-sequence Fc region and the variable Fc region. "Fc fragment" refers to the carboxy-terminal portions of both H chains held together by disulfides. The effector functions of an antibody are determined by the sequences in the Fc region, which is also recognized by Fc receptors (FcRs) found on certain types of cells. Digestion of antibodies with the enzyme pepsin results in an F(ab')2 fragment in which the two arms of the antibody molecule remain linked and contain two antigen-binding sites. The F(ab')2 fragment has the ability to cross-link antigen. "Fv," as used herein, refers to the minimum fragment of an antibody that retains both the antigen-recognition and antigen-binding sites. This region consists of a dimer of one heavy-chain variable domain and one light-chain variable domain associated by tight non-covalent or covalent bonds. In this configuration, the three CDRs of each variable domain interact to define an antigen-binding site on the surface of the VH-VL dimer. Collectively, the six CDRs confer antigen-binding specificity to the antibody. However, a single variable domain (or three CDRs specific for an antigen) can be bound to the antibody. Even half of an Fv containing only the CDRs) may bind to the entire binding site, albeit with lower affinity. , has the ability to recognize and bind to antigens.

[0028] As used herein, the term "monoclonal antibody" refers to an antibody obtained from a population of substantially homogeneous antibodies, i.e., the individual antibodies comprising the population differ from one another except for possible naturally occurring mutations and / or post-translational modifications (e.g., isomerization, amidation) that may be present in minor amounts. are identical. Monoclonal antibodies are highly specific and are directed against a single antigenic determinant or epitope. In contrast, polyclonal antibody preparations typically contain different antibodies directed against different antigenic determinants (or epitopes). As used herein, the term "monoclonal antibody" encompasses both intact and full-length monoclonal antibodies, as well as antibody fragments (e.g., Fab, Fab', F(ab')2, Fv), single-chain (scFv) variants, fusion proteins containing an antibody portion, and any other modified immunoglobulin molecule containing an antigen recognition site. Furthermore, "monoclonal antibody" refers to an antibody produced in any number of ways, including, but not limited to, hybridoma, phage selection, recombinant expression, and transgenic animals. As used herein, monoclonal antibodies include antibodies in which a portion of the heavy and / or light chains are derived from a particular species and belong to a particular antibody class or subclass. These may include "chimeric" antibodies (immunoglobulins) in which one chain is identical or homologous to a corresponding sequence in an antibody from another species belonging to a different antibody class or subclass, while the remaining chains are identical or homologous to corresponding sequences in an antibody from another species belonging to a different antibody class or subclass, as well as fragments of antibodies which exhibit a desired biological activity.

[0029] The term "humanized antibody" refers to a subset of chimeric antibodies in which a "hypervariable region" from a non-human immunoglobulin (donor antibody) replaces residues from a hypervariable region in a human immunoglobulin (recipient antibody). Generally, a humanized antibody will comprise substantially all of at least one, and typically two, variable regions, in which all or substantially all of the hypervariable loops correspond to those of a non-human immunoglobulin sequence and all or substantially all of the framework regions are those of a human immunoglobulin sequence, although the framework regions may contain one or more substitutions that improve antibody performance, e.g., binding affinity, isomerization, immunogenicity, etc. .

[0030] The term "bispecific antibody" refers to an antibody that contains specificity for two target molecules, and includes bispecific IgG (optionally IgG with a common light chain), DVD-Ig (DiGi Ammarino et al., "Design and generation of DVD-Ig™ molecules for dual-specific targeting," Meth. Mo. Biol., 2012, 889, 145-156), mAb 2 (See WO2008 / 003103, mAb 2 Fo the description of which is incorporated herein by reference), FIT-Ig (WO2015 / 1 03072, the description of the FIT-Ig backbone is incorporated herein by reference), mAb- dAb, Dock-Lock, Fab-arm exchange, SEEDbody, Triomab, LUZ-Y, Fcab, κλ-body, orthogonal Fab, scDiabody-Fc, diabody-Fc, tandem scFv-Fc, Fab-scFv-Fc, Fab-scFv, intracellular antibody, BiTE, diabody, DART, TandemAb, scDiabody, scDiabody-CH3, Diabody-CH3, triple body (Tr single body), miniantibody, minibody, TriBi minibody, scFv-CH Formats include 3KIH, scFv-CH-CL-scFv, F(ab')2-scFv, scFv-KIH, Fab-scFv-Fc, tetravalent HCab, ImmTAC, knob-in-hole, knob-in-hole with a common light chain, knob-in-hole with a common light chain and charge pair, charge pair, charge pair with a common light chain, DT-IgG, DutaMab, IgG(H)-scFv, scFv-(H)IgG, IgG(L)-scFv, scFv-(L)IgG, IgG(L,H)-Fv, IgG(H)-V, V(H)-IgG, IgG(L)-V, V(L)-IgG, KIH IgG-scFab, 2scFv-IgG, IgG-2scFv, scFv4-Ig, and zybody. For a review of bispecific formats, see Spiess, C., et al., Mol. Immunol (2015). In another embodiment, the bispecific molecule comprises another non-Ig format, such as a T-cell receptor binding domain, an immunoglobulin superfamily domain, a jawless variable lymphocyte receptor, a fibronectin domain (e.g., Adnectin™), an antibody constant domain (e.g., a CH3 domain, e.g., CH2 and / or CH3 of Fcab™) (wherein the constant domain is not a functional CH1 domain), an scFv, an (scFv)2, sc-diabody, scFab, centyrin and epitope binding domains derived from scaffolds selected from CTLA-4 (Evibody™), lipocalin domains, protein A (e.g., Affibody™ or SpA), such as the Z domain of protein A, A domains (e.g., Avimer™ or Maxibody™), epitope binding domains derived from heat shock proteins (GroEI and GroES), etc.), transferrin domains (e.g., transbodies), ankyrin repeat proteins proteins (e.g., DARPins™), peptide aptamers, C-type lectin domains (e.g., For example, Tetranectin™, human gamma-crystallin or human ubiquitin (affilin), PDZ domains, scorpion toxins, and Kunitz-type domains of human protease inhibitors. Contains an antibody fused to the main

[0031] In one embodiment, the bispecific antibody is a mAb 2 mAb 2 is a V from an intact antibody fused to a modified constant region. H and V L domains, which have been engineered to form antigen-binding sites known as "Fcabs." Fcab / mAb 2 The technology behind the format is described in more detail in WO2008 / 003103, mAb 2 The format description is incorporated herein by reference.

[0032] In one embodiment, a "bispecific antibody" does not include the FIT-Ig format. In one embodiment, a "bispecific antibody" is a mAb 2 In one embodiment, the bispecific antibody does not include the FIT-Ig format or the mAb 2 Format Not at all.

[0033] In another embodiment, the bispecific antibody is a "dual binding antibody." As used herein, a "dual binding antibody" refers to an antibody in which both antigen-binding domains are V H / V L formed by pairs Bispecific antibodies that have been used include FIT-Ig (incorporated herein by reference). (See WO2015 / 103072, which is incorporated herein by reference), mAb-dAb, Dock-Lock, Fab-arm exchange, SEEDbody, Triomab, LUZ-Y, Fcab, κλ-body, orthogonal Fab, scDiabody-Fc, diabody-Fc, tandem scFv-Fc, Fab-scFv-Fc, Fab-scFv, intracellular antibody, BiTE, diabody, DART, TandAb, scDiabody, scDiabody-CH3, Diabody-CH3, triple body, miniantibody, minibody, scFv-CH3 KIH, scFv-CH-CL-scFv, F( ab')2-scFv, scFv-KIH, Fab-scFv-Fc, tetravalent HCab, These include ImmTAC, knobs-in-hole, knobs-in-hole with a common light chain, knobs-in-hole with a common light chain and charge pair, charge pair, charge pair with a common light chain, DT-IgG, DutaMab, IgG(H)-scFv, scFv-(H)IgG, IgG(L)-scFv, scFv-(L)IgG, IgG(L,H)-Fv, IgG(H)-V, V(H)-IgG, IgG(L)-V, V(L)-IgG, KIH IgG-scFab, 2scFv-IgG, IgG-2scFv, and scFv4-Ig.

[0034] The terms "hypervariable region," "CDR region," or "CDR" refer to the regions of an antibody variable domain that are hypervariable in sequence and / or form structurally defined loops. Thus, the antigen-binding site of an antibody contains six hypervariable regions: three in the VH (CDRH1, CDRH2, CDRH3) and three in the VL (CDRL1, CDRL2, CDRL3). These regions of the antibody heavy and light chains confer antigen-binding specificity to the antibody. CDRs can be defined according to the Kabat system (Kabat, EA et al., 1999). (See, for example, "Sequences of Proteins of Immunological Interest," 5th edit., NIH Publication no. 91-3242, USDapartment of Health and Human Services). Another system is described by Chothia et al. The system was devised by Chothia, C. & Lesk, A.M., 1987, "Ca nonical structures for the hypervariable The IMGT system (see Lefranc, MP, 1997, "Unique database numbering system for immunogenetic analysis", Immunol. Today, 18, 50) can be used to define CDRs. A human antibody typically contains three heavy chain CDRs and three light chain CDRs. The term CDR is used herein to refer to one or several of these regions. Those skilled in the art can easily compare different nomenclature systems to determine whether a particular sequence can be defined as a CDR.

[0035] A "human antibody" is an antibody that possesses an amino acid sequence that corresponds to that of an antibody produced by a human and / or that has been produced using any of the techniques for producing human antibodies. The term "specifically binds" refers to a measurable and reproducible interaction, such as binding between a target and an antibody, that determines the presence of the target in the presence of a homogeneous population of molecules, including biological molecules. For example, the binding of a target (epitope) to an antibody is not specifically binding to a target (epitope). An antibody that specifically binds to a target (which may be a target of interest) has a greater affinity, binding affinity, than it binds to other targets. The net result is an antibody that binds to its target more readily and / or for longer. is an antibody against an unrelated target, as measured, for example, by radioimmunoassay (RIA). The extent of binding is less than about 10% of the binding of the antibody to the target.

[0036] Antibodies or fragments thereof that specifically bind to the hPD-L1 antigen may be cross-reactive with related antigens. Preferably, antibodies or fragments thereof that specifically bind to the hPD-L1 antigen do not cross-react with other antigens (but optionally, with antigens from different species, e.g., rhesus monkeys, or mice). (It may cross-react with mouse PD-L1.) Fragments of may be identified, for example, by immunoassays, BIAcore™, or other techniques known in the art. An antibody or fragment thereof specifically binds to the hPD-L1 antigen when it binds to the hPD-L1 antigen with greater affinity than it does to any cross-reactive antigens, as determined using laboratory techniques such as radioimmunoassay (RIA) and enzyme-linked immunosorbent assay (ELISA). Typically, a specific or selective response will be at least 2-fold above background signal or noise, more typically more than 10-fold (e.g., more than 15-fold, 20-fold, more than, 50-fold, or 100-fold greater than background. For example, For a discussion of this, see pages 332-336 of Paul, ed., 1989, Fundamental Immunology Second Edition, Raven Press, New York.

[0037] The term "aliphatic amino acid" means that the amino acid R group is non-polar and hydrophobic. Hydrophobicity increases with the number of C atoms in the hydrocarbon chain. Glycine, alanine, valine, leucine, and isoleucine are aliphatic amino acids.

[0038] The term "aromatic amino acid" means that the amino acid R group contains an aromatic ring system. Phenylalanine, tyrosine, and tryptophan are aromatic amino acids. The term "hydroxyl-containing amino acid" means that the amino acid R group contains a hydroxyl group and is hydrophilic. Serine, cysteine, threonine, and methionine are hydroxyl-containing amino acids.

[0039] The term "basic amino acid" means that the amino acid R group contains nitrogen and is basic at neutral pH. Histidine, lysine, and arginine are basic amino acids.

[0040] The term "cyclic amino acid" means that the amino acid R group has an aliphatic ring structure. Proline is the only cyclic aliphatic amino acid.

[0041] The term "acidic amino acid" means that the amino acid R group is polar and negatively charged at physiological pH. Aspartic acid and glutamic acid are acidic amino acids.

[0042] The term "amide amino acid" means that the amino acid R group contains an amide group. Asparagine and glutamine are amide amino acids.

[0043] As used herein, "approval number" and "marketing authorization number" refer to a number issued by a regulatory authority when that authority determines that a particular medical product and / or composition may be sold and / or released in its area of ​​jurisdiction. As used herein, a "regulatory authority" refers to, for example, a regulatory authority that is responsible for evaluating the safety and effectiveness of a medical product and / or composition, and one of the authorities responsible for controlling the release / sale of such products and / or compositions in a given area. The Food and Drug Administration (FDA) in the United States and the European Medicines Agency (EPA) in Europe are just two examples of such regulatory authorities. Other non-limiting examples may include the SDA, MPA, MHPRA, IMA, ANMAT, Hong Kong Department of Health-Drug Office, CDSCO, Medsafe, and KFDA.

[0044] As used herein, the term "biomarker" refers to a gene that is differentially expressed in individuals with a disease of interest, e.g., a gene that is differentially expressed in individuals with cancer. In one embodiment, PD-L1 is a biomarker whose expression in a tumor can indicate whether a patient will respond to a particular type of treatment, specifically whether a patient will respond to a treatment that targets PD-L1, e.g., immunotherapy using an anti-PD-L1 antibody. In one embodiment, PD-L1 is a biomarker whose expression in a tumor can indicate whether a patient will respond to a particular type of treatment, specifically whether a patient will respond to a treatment that targets PD-1, e.g., immunotherapy using an anti-PD-1 antibody. In another embodiment, PD-L1 can be free or membrane-bound. In another embodiment, PD-L1 can be immobilized or non-immobilized.

[0045] As used herein, "buffer" refers to a chemical agent that can absorb a certain amount of acid or base without undergoing a strong shift in pH.

[0046] As used herein, the term "carrier" refers to a diluent, adjuvant (e.g., Freund's adjuvant (complete and incomplete)), excipient, or vehicle with which a therapeutic is administered. Such pharmaceutical carriers can be sterile liquids, such as water and oils, such as peanut oil, soybean oil, mineral oil, sesame oil, and the like, including those of petroleum, animal, vegetable, or synthetic origin. Water is a preferred carrier when the pharmaceutical composition is administered intravenously. Saline solutions and aqueous dextrose and glycerol solutions can also be used as liquid carriers, particularly for injectable solutions.

[0047] The terms "chemotherapeutic agent" or "chemotherapy" typically refer to therapeutic agents whose primary purpose is to destroy cancer cells by interfering with the ability of tumor cells to grow or multiply. There are many different types of chemotherapy agents, with over 50 approved chemotherapy agents available. Chemotherapy drugs can be classified based on how they work. Alkylating agents kill cancer cells by directly attacking DNA, the genetic material of genes. Cyclophosphamide is an alkylating agent. Antimetabolites interfere with the production of DNA, inhibiting cell growth and proliferation. An example of an antimetabolite is 5-fluorouracil (5-F Antitumor antibiotics are produced from natural sources such as fungi in the soil. They interfere with important cellular functions, including the production of DNA and cellular proteins. Doxorubicin and bleomycin belong to this group of chemotherapy drugs. Plant alkaloids prevent cells from dividing normally. Vinblastine and vincristine are plant alkaloids obtained from the periwinkle plant. Steroid hormones slow the growth of some hormone-dependent cancers. For example, tamoxifen is used to treat breast cancer, which depends on the hormone estrogen for growth. DNA damage response (DDR) inhibitors, such as PARP inhibitors, block DNA repair mechanisms after single- or double-strand breaks.

[0048] Examples of chemotherapeutic agents include adriamycin, doxorubicin, 5-fluorouracil, cytosine arabinoside (Ara-C), cyclophosphamide, thiotepa, and taxotere ( Docetaxel), Busulfan, Cytoxin, Taxol, Methotrexate, Syspro Suitable toxins and chemotherapeutic agents include platin, melphalan, vinblastine, bleomycin, etoposide, ifosfamide, mitomycin C, mitoxantrone, vincrestine, vinorelbine, carboplatin, teniposide, daunomycin, carminomycin, aminopterin, dactinomycin, mitomycin, esperamicin (see U.S. Pat. No. 4,675,187), melphalan, and other related nitrogen mustards. Suitable toxins and chemotherapeutic agents are described in Remington's Pharmaceutical Sciences, 19th Ed. (Mack Publishing Co. 1995) and Goodman and Gilman's The Pharmacological Basis of Therapeutics, 7th Ed. (MacMillan Publishing Co. 1995). Another example of a chemotherapeutic agent is an antibody. classes of toxins conjugated to, including but not limited to, pyrrolobenzodiazepines, maytansinoids, calicheamicins, etc. Other suitable toxins and / or Chemotherapeutic agents are known to those skilled in the art.

[0049] As used herein, the term "composition" is intended to encompass a product containing specified ingredients (e.g., an antibody of the invention), optionally in the specified amounts, and any product that results directly or indirectly from the combination of specified ingredients, optionally in the specified amounts.

[0050] As used herein, "comprising" or "comprises" The term "antibodies, fragments, uses, compositions, methods, and their corresponding components" refers to antibodies, fragments, uses, compositions, methods, and their corresponding components. Used in reference to an element, essential to a method or component, but open to the inclusion of unspecified elements, whether essential or not.

[0051] The term "consisting of" refers to the antibodies, fragments, uses, compositions, methods, and their respective components described herein, excluding any element not recited in the description of that embodiment.

[0052] As used herein, the term "consisting essentially of" refers to elements required for a given embodiment. The term permits the presence of elements that do not materially affect the basic and novel or functional characteristics of that embodiment.

[0053] As used herein, in the context of a polypeptide, the term "derivative" refers to a polypeptide comprising the amino acid sequence of an hPD-L1 polypeptide, a fragment of an hPD-L1 polypeptide, or an antibody that specifically binds to an hPD-L1 polypeptide that has been altered by the introduction of amino acid substitutions, deletions, or additions. As used herein, the term "derivative" also refers to an hPD-L1 polypeptide, a fragment of an hPD-L1 polypeptide, or an antibody that specifically binds to an hPD-L1 polypeptide that has been chemically modified, for example, by the covalent attachment of any type of molecule to the polypeptide. For example, and without limitation, an hPD-L1 polypeptide, a fragment of an hPD-L1 polypeptide, or an hPD-L1 antibody may be modified by, for example, glycosylation, acetylation, pegylation, phosphorylation, amidation, known protecting / blocking groups, or the like. Derivatives may be chemically modified by derivatization with linking groups, proteolytic cleavage, conjugation to cellular ligands or other proteins, etc. Derivatives are modified in such a way that either the type or location of the attached molecule is different from the native or starting peptide or polypeptide. Derivatives further include deletion of one or more chemical groups naturally present on the peptide or polypeptide. Derivatives of hPD-L1 polypeptides, hPD-L1 polypeptide fragments, or hPD-L1 antibodies may be chemically modified using techniques known to those skilled in the art, including, but not limited to, specific chemical cleavage, acetylation, formulation, metabolic synthesis with tunicamycin, etc. Furthermore, derivatives of hPD-L1 polypeptides, hPD-L1 polypeptide fragments, or hPD-L1 antibodies may contain one or more non-classical amino acids. Polypeptide derivatives retain similar or identical functions as the hPD-L1 polypeptides, hPD-L1 polypeptide fragments, or hPD-L1 antibodies described herein.

[0054] As used herein, the term "effector function" is intended to refer to one or more of antibody-dependent cellular cytotoxicity (ADCC), complement-dependent cytotoxicity (CDC)-mediated responses, Fc-mediated phagocytosis or antibody-dependent cellular phagocytosis (ADCP), and antibody recycling via the FcRn receptor.

[0055] "Effective amount" refers to an amount effective, at dosages and for periods of time necessary, to achieve the desired effect, including a therapeutic or prophylactic effect. A "therapeutically effective amount" is an amount that results in a measurable improvement or prevention of a particular disorder. "Therapeutically effective amount" refers to the minimum concentration required to achieve prophylactic effect. The therapeutically effective amount herein may vary depending on factors such as the stage of disease, age, sex, and weight of the patient, and the ability of the antibody to elicit a desired response in an individual. A therapeutically effective amount is also one in which the beneficial effects of treatment outweigh any toxic or harmful effects of the antibody. A "prophylactically effective amount" refers to an amount effective at the dosage and for the period necessary to achieve the desired prophylactic effect. In some embodiments, an effective amount of an antibody of the present invention is from about 0.1 mg / kg (mg of antibody per kg of subject body weight) to about 100 mg / kg. In certain embodiments, an effective amount of an antibody provided herein is from about 0.1 mg / kg, about 0 .5mg / kg, about 1mg / kg, 3mg / kg, 5mg / kg, about 10mg / kg, about 15 mg / kg, about 20mg / kg, about 25mg / kg, about 30mg / kg, about 35mg / kg, Approx. 40mg / kg, approx. 45mg / kg, approx. 50mg / kg, approx. 60mg / kg, approx. 70mg / kg, about 80 mg / kg, about 90 mg / kg, or about 100 mg / kg (or a range therein) In some embodiments, "effective amount," as used herein, also refers to the amount of an antibody of the invention to achieve a specified result (e.g., inhibition of hPD-L1 biological activity in a cell).

[0056] As used herein, the term "epitope" refers to a localized region on the surface of an antigen, such as an hPD-L1 polypeptide or hPD-L1 polypeptide fragment, that can bind to one or more antigen-binding regions of an antibody, has antigenic or immunogenic activity, and is capable of eliciting an immune response in an animal, preferably a mammal, and most preferably a human. An epitope with immunogenic activity is a portion of a polypeptide that elicits an antibody response in an animal. An epitope with antigenic activity is a portion of a polypeptide to which an antibody specifically binds, as determined by any method known in the art, such as the immunoassays described herein. An antigenic epitope need not necessarily be immunogenic. Epitopes usually consist of chemically active surface groupings of molecules, such as amino acids or sugar side chains, and have specific three-dimensional structural characteristics, as well as specific charge characteristics. The region of a polypeptide that contributes to an epitope may be contiguous amino acids of the polypeptide, or an epitope may be assembled from two or more non-contiguous regions of the polypeptide. An epitope may or may not be a three-dimensional surface feature of an antigen. In certain embodiments, an hPD-L1 epitope is a three-dimensional surface feature of an hPD-L1 polypeptide (e.g., a trimer). In another embodiment, the hPD-L1 epitope The fragment is a linear characteristic of the hPD-L1 polypeptide (e.g., a trimeric or monomeric form). The antibodies provided herein may be in a monomeric (denatured) form. An antibody may specifically bind to an epitope on hPD-L1 in its trimeric (native) form, an epitope on hPD-L1 in its trimeric (native) form, or an epitope on hPD-L1 in both its monomeric (denatured) and trimeric (native) forms. In certain embodiments, the antibodies provided herein specifically bind to an epitope on hPD-L1 in its trimeric form, but do not specifically bind to hPD-L1 in its monomeric form.

[0057] As used herein, the term "excipient" refers to an inert substance commonly used as a diluent, vehicle, preservative, binder, or stabilizer for drugs, and includes proteins (e.g., serum albumin, etc.), amino acids (e.g., aspartic acid, glutamic acid, Examples of suitable surfactants include amino acids, lysine, arginine, glycine, histidine, etc., fatty acids and phospholipids (e.g., alkyl sulfonates, caprylic acid, etc.), surfactants (e.g., SDS, polysorbates, nonionic surfactants, etc.), saccharides (e.g., sucrose, maltose, trehalose, etc.), and polyols (e.g., mannitol, sorbitol, etc.). Also included are, but not limited to, Remington's Pharmaceutical Sciences (1999), which is incorporated herein by reference in its entirety. 90) See also Mack Publishing Co., Easton, Pa. .

[0058] As used herein, the terms "fixed" or "fixation" refer to a chemical process by which biological tissue is preserved to prevent autolysis or decay. Generally, fixation involves exposing the tissue to a chemical compound, e.g., an aldehyde such as alcohol or formaldehyde, to halt ongoing biochemical reactions. In some instances, fixation can also increase the mechanical strength and stability of the treated tissue. The term "unfixed" refers to tissue that has not been subjected to a chemical process to prevent tissue decay. As used herein, "surface-expressed" means that the protein is embedded in or spans the cell membrane, or is associated with a protein that is embedded in or spans the cell membrane (i.e., a membrane-associated protein). In one embodiment, a surface-expressed protein contains one or more transmembrane domains. In another embodiment, the protein is indirectly associated with the outer or inner surface of the cell membrane through association with another transmembrane protein (i.e., the surface-expressed protein does not span the cell membrane itself). In general, a surface-expressed protein integrated into a cell membrane or expressed endogenously within a cell is more likely to fold in the correct structure than a recombinantly produced free form of the same protein. As used herein, in the context of a peptide or polypeptide, the term "fragment" refers to a peptide or polypeptide that comprises less than the full-length amino acid sequence. Such fragments may include, for example, truncations at the amino terminus, truncations at the carboxy terminus, and / or or may result from internal deletion of residues from the amino acid sequence. Fragments may result, for example, from alternative RNA splicing or from in vivo protease activity. In certain embodiments, a PD-L1 Fragment comprises a polypeptide comprising an amino acid sequence of at least 5 contiguous amino acid residues, at least 10 contiguous amino acid residues, at least 15 contiguous amino acid residues, at least 20 contiguous amino acid residues, at least 25 contiguous amino acid residues, at least 40 contiguous amino acid residues, at least 50 contiguous amino acid residues, at least 60 contiguous amino acid residues, at least 70 contiguous amino acid residues, at least 80 contiguous amino acid residues, at least 90 contiguous amino acid residues, at least 100 contiguous amino acid residues, at least 125 contiguous amino acid residues, at least 150 contiguous amino acid residues, at least 175 contiguous amino acid residues, at least 200 contiguous amino acid residues, or at least 250 contiguous amino acid residues of an hPD-L1 polypeptide or an antibody that specifically binds to an hPD-L1 polypeptide. In certain embodiments, a fragment of an hPD-L1 polypeptide or an antibody that specifically binds to an hPD-L1 antigen retains at least one, at least two, or at least three functions of the polypeptide or antibody.

[0059] The term "free" refers to a polypeptide, e.g., PD-L1, or fragments and variants thereof, in combination with a buffer, where the polypeptide is not bound to a cell surface or cell membrane. Thus, the term "free" refers to a polypeptide capable of surface expression (i.e., associated with one or more membranes). The nucleotide sequence of the nucleotide sequence (containing the transmembrane domain or membrane-binding domain) is expressed on the surface of the cell in its current state. Free polypeptides can refer to polypeptides that are not bound to proteins expressed on the surface of a cell or that are not encapsulated in a solution or that are not bound to proteins expressed on the surface of a cell. Free polypeptides can also refer to free recombinant or natural or unbound polypeptides. In the context of phage display, free antigens can be selected in solution (referred to herein as "soluble selection") or can be adsorbed to a surface, for example, the surface of a 96-well plate (referred to herein as "biopanning selection"). ").

[0060] The term "fusion protein", as used herein, refers to a polypeptide comprising the amino acid sequence of an antibody and the amino acid sequence of a heterologous polypeptide or protein (i.e., a polypeptide or protein that is not normally part of an antibody (e.g., a non-anti-hPD-L1 antigen antibody)). The term "fusion", when used in reference to hPD-L1 or anti-hPD-L1 antibodies, refers to a peptide or polypeptide, or fragment, variant, and / or derivative thereof. Fusion proteins refer to the conjugation of a heterologous peptide or polypeptide to a heterologous peptide or polypeptide. Preferably, the fusion protein retains the biological activity of hPD-L1 or an anti-hPD-L1 antibody. In certain embodiments, the fusion protein comprises the VH domain, VL domain, VH CDRs (one, two, or three VH CDRs), and / or VL CDRs (one, two, or three VL CDRs) of an hPD-L1 antibody, and the fusion protein specifically binds to an hPD-L1 epitope. do.

[0061] The term "heavy chain," when used in reference to antibodies, can be alpha (α), delta (δ), epsilon (ε), gamma (γ), and mu (μ), based on the amino acid sequence of the heavy chain constant domain. These different types of heavy chains are well known and This gives rise to five classes of antibodies, IgA, IgD, IgE, IgG, and IgM, respectively, with four subclasses of IgG, namely, IgG1, IgG1, IgG3, and IgG4. Preferably, the heavy chain is a human heavy chain. Multiple heavy chain constant region alleles of each immunoglobulin or immunoglobulin subclass exist in the human population. The nucleotide and amino acid sequences of these allelic variants are accessible on publicly available databases such as IMGT, ENSEMBL Swiss-Prot, and Uniprot. Allelic variants can also be identified in various genome sequencing projects. In one embodiment, the antibodies and antibody fragments disclosed herein are derived from human IGHG1*01 (SEQ ID NOs: 340, 341, and 537), IGHG1*02 (SEQ ID NOs: 340, 341, and 537), or IGHG1*03 (SEQ ID NOs: 340, 341, and 537). , and 537), IGHG1*03 (SEQ ID NOs: 523 and 524), IGHG1*04 (SEQ ID NOs: 525 and 526), ​​and IGHG1*05 (SEQ ID NOs: 340, 341, and 537). In one embodiment, the antibodies and antibody fragments disclosed herein comprise human IGHG2*01 (SEQ ID NOs: 527 and 528), IGHG2*02 (SEQ ID NOs: 529 and 530), IGHG2*03 (SEQ ID NOs: 527 and 528), IGHG2*04 (SEQ ID NOs: 531 and 532), IGHG2*05 (SEQ ID NOs: 533 and 534), IGHG2*06 (SEQ ID NOs: 535 and 536), IGHG2*07 (SEQ ID NOs: 537 and 538), IGHG2*08 (SEQ ID NOs: 539 and 540), IGHG2*09 (SEQ ID NOs: 541 and 542), IGHG2*10 (SEQ ID NOs: 543 and 544), 32), IGHG2*05 (SEQ ID NOs: 527 and 528), and IGHG2*06 (SEQ ID NOs: 533 and 534). In one embodiment, the antibodies and antibody fragments disclosed herein comprise proteins encoded by IgG3 constant region alleles, including, but not limited to, human IGHG3*01, IGHG3*02, IGHG3*03, IGHG3*04, IGHG3*05, IGHG3*06, IGHG3*07, IGHG3*08, IGHG3*09, IGHG3*10, IGHG3*11, IGHG3*12, IGHG3*13, IGHG3*14, IGHG3*15, IGHG3*16, IGHG3*17, IGHG3*18, and IGHG3*19. In one embodiment, the antibodies and antibody fragments disclosed herein comprise proteins encoded by IgG4 constant region alleles, including, but not limited to, human IGHG4*01 (SEQ ID NOs: 192 and 193), IGHG4*02 (SEQ ID NOs: 194 and 195), IGHG4*03 (SEQ ID NOs: 196 and 197), and IGHG4*04 (SEQ ID NOs: 192 and 193). In another example, the heavy chain is a truncated IgG isotype, e.g., a truncated IgG4. In certain embodiments, antibodies of the invention comprise a human gamma 4 constant region. In another embodiment, the heavy chain constant region does not bind to an Fc-gamma receptor and comprises, for example, a Leu235Glu mutation. In another embodiment, the heavy chain constant region comprises a Ser228Pro mutation to increase stability. In another embodiment, the heavy chain constant region is IgG4-PE (SEQ ID NO: 199). In another embodiment, the antibodies and antibody fragments disclosed herein comprise a heavy chain constant region encoded by a mouse IgG1 constant region allele, including but not limited to, mouse IGHG1*01 or IGHG1*02. In one embodiment, the antibodies and antibody fragments disclosed herein comprise a heavy chain constant region encoded by a mouse IgG2 constant region allele, including but not limited to, mouse IGHG2A*01, IGHG2A*02, IGHG2B*01, IGHG2B*02, IGHG2C*01, IGHG2C*02, or IGHG2C*03. In one embodiment, the antibodies or antibody fragments disclosed herein comprise a heavy chain constant region encoded by a mouse IgG2 constant region allele, including but not limited to, mouse IGHG2A*01, IGHG2A*02, IGHG2B*01, IGHG2B*02, IGHG2C*01, IGHG2C*02, or IGHG2C*03. Includes proteins encoded by mouse IgG3 constant region alleles, including but not limited to GHG3*01.

[0062] As used herein, the term "host" refers to an animal, preferably a mammal, and most preferably a human.

[0063] As used herein, the term "host cell" refers to a particular subject cell transfected with a nucleic acid molecule and to the progeny or potential progeny of such a cell. The progeny of such a cell may not be identical to the parent cell transfected with the nucleic acid molecule due to mutations or environmental influences that may occur in subsequent generations, or due to integration of the nucleic acid molecule into the host cell genome.

[0064] As used herein, the term "IL-2 cytokine" refers to a cytokine-like molecule that has activity similar to wild-type IL-2. It may have activity at the high (αβγ) affinity IL-2 receptor and / or the intermediate affinity (αβ) IL-2 receptor. A cytokine is a molecule that binds to IL-2. The mutant IL-2 cytokine may have one or more amino acid deletions, substitutions, or additions.

[0065] As used herein, "immunomodulatory agent," and variations thereof, including but not limited to immunomodulator, refer to agents that modulate the host's immune system. In certain embodiments, an immunomodulatory agent is an immunosuppressant. In certain other embodiments, an immunomodulatory agent is an immunostimulatory agent. In accordance with the present invention, immunomodulatory agents used in the combination therapies of the invention do not include anti-hPD-L1 antibodies or antigen-binding fragments. Immunomodulatory agents include, but are not limited to, small molecules, peptides, polypeptides, proteins, fusion proteins, antibodies, inorganic molecules, mimetics, and organic molecules.

[0066] The term "in combination" in the context of administration of other therapeutic agents refers to the use of two or more therapeutic agents. The use of the term "in combination" does not restrict the order in which the therapeutic agents are administered to a subject with a disease. A first therapy may be administered (e.g., 1 minute, 45 minutes, 30 minutes, 4 The dose may be administered 5 minutes, 1 hour, 2 hours, 4 hours, 6 hours, 12 hours, 24 hours, 48 ​​hours, 72 hours, 96 hours, 1 week, 2 weeks, 3 weeks, 4 weeks, 5 weeks, 6 weeks, 8 weeks, or 12 weeks before, simultaneously with, or subsequently (e.g., 1 minute, 45 minutes, 30 minutes, 45 minutes, 1 hour, 2 hours, 4 hours, 6 hours, 12 hours, 24 hours, 48 ​​hours, 72 hours, 96 hours, 1 week, 2 weeks, 3 weeks, 4 weeks, 5 weeks, 6 weeks, 8 weeks, or 12 weeks after). Any additional therapeutic agent may be administered in any order with the other additional therapeutic agents. In certain embodiments, the antibodies of the invention are administered in combination with one or more therapeutic agents (e.g., those intended to treat an hPD-L1-mediated disease). Therapies other than the antibodies of the present invention currently administered to prevent, treat, manage, and / or ameliorate Therapeutic agents that can be administered in combination with the antibodies of the present invention include: Non-limiting examples include analgesics, anesthetics, antibiotics, or immunomodulators, or any other medication listed in the United States Pharmacopeia and / or Physician's Guide.

[0067] As used herein, the term "immunocytokine" refers to an antibody format fused to a cytokine molecule. The antibody format can be any of those described herein, and the cytokine can be fused directly or via a linker or chemical conjugation to the N-terminus or C-terminus of the heavy or light chain of the antibody format.

[0068] As used herein, "injection device" refers to a device designed to perform injections, which is the step of temporarily fluidly connecting the injection device to human tissue, typically subcutaneous tissue. Injection further includes administering a quantity of liquid medication into the tissue and separating or removing the injection device from the tissue. In some embodiments, the injection device can be an intravenous or IV device, which is a type of injection device used when the target tissue is blood in the circulatory system, e.g., blood in a blood vessel. A common, but non-limiting, example of an injection device is a needle and syringe.

[0069] As used herein, "instructions" refers to written, printed, or graphic material on the immediate container of an article, e.g., written material displayed on a vial containing a pharmaceutically active agent, or a display detailing the composition and use of a product of interest included in a kit containing the composition of interest. The instructions describe the method of treatment that is intended to be administered or performed.

[0070] An "isolated" or "purified" antibody or protein is one that is free from components of its production environment (e.g., For example, an antibody or protein is substantially free of cellular material or other contaminating proteins from the cell or tissue source from which the antibody is derived, or, if chemically synthesized, substantially free of chemical precursors or other chemicals. The term "substantially free of cellular material" includes preparations of an antibody in which the antibody is separated from cellular components of the cells from which it is isolated or recombinantly produced. Thus, an antibody that is substantially free of cellular material has less than about 30%, 20%, 10%, or 5% (by dry weight) of heterologous protein (referred to herein as "contaminating protein"). If the antibody is recombinantly produced, the antibody may be cultured in a medium that also substantially contains the medium. Preferably, the antibody is free of culture medium, i.e., culture medium comprises about 20%, 10%, or 5% of the volume of the protein preparation. If the antibody is produced by chemical synthesis, the antibody is preferably substantially free of chemical precursors or other chemicals, i.e., the antibody is separated from chemical precursors or other chemicals involved in the synthesis of the protein. Thus, such antibody preparations have less than about 30%, 20%, 10%, or 5% (by dry weight) of chemical precursors or compounds other than the antibody of interest. In a preferred embodiment, the antibodies of the invention are isolated or purified.

[0071] The term "Kabat numbering" and similar terms are art-recognized and refer to amino acid residues that are more variable (i.e., , hypervariable) (Kabat et al. (197 1) Ann.NY Acad.Sci.190:382-391 and Kabat et. (1991) Sequences of Proteins of Immunological Interest, Fifth Edition, US Department of Health and Human Services, NIH Publication No. 91-3242). In the case of the heavy chain variable region, the hypervariable region is Typically, the range is amino acid positions 31 to 35 for CDR1, amino acid positions 50 to 65 for CDR2, and amino acid positions 95 to 102 for CDR3.

[0072] As used herein, "label" or "labeled" refers to the addition of a detectable moiety to a polypeptide, such as a radiolabel, a fluorescent label, an enzyme label, a chemiluminescent label, or a biotinyl group or gold. Radioisotopes or radionuclides include: 3 H, 14 C. 15 N, 35 S, 90 Y, 99 Tc, 115 In, 125 I, 131 Fluorescent labels may include 1, fluorescent labels may include rhodamine, lanthanide fluorophores, or FITC, and fluorescent labels may include horseradish peroxidase, β-galactosidase, luciferase, alkaline phosphatase. Additional labels include, by way of example and not limitation, enzymes such as glucose-6-phosphate dehydratase ("G6PDH"), alpha-D-galactosidase, glucose oxidase, glucose amylase, carbonic anhydrase, acetylcholinesterase, lysozyme, malate dehydrogenase, and peroxidase; dyes (e.g., cyanine dyes, For example, Cy5™, Cy5.5™, or Cy7™; fluorescein and the like; Derivatives of fluorescent dyes, GFP (for "green fluorescent protein"), other fluorescent proteins ( For example, mCherry, mTomato), dansyl, umbelliferone, phycoerythroyltransferase Phycocyanin, allophycocyanin, o-phthalaldehyde hyde), and additional fluorescent labels or fluorophores such as fluorescamine; lanthanide cryptates fluorophores such as phosphates and chelates, e.g., europium (PerkinElmer and Cisbio assay); isoluminol, luminol, and dioxetanes, etc. chemiluminescent labels or chemiluminescent fluorescers; sensitizers; coenzymes; enzyme substrates; particles such as latex or carbon particles; metal sols; microcrystals; liposomes; cells, etc., which contain dyes, catalysts, or other detectable groups; molecules such as biotin, digoxigenin, or 5-bromodeoxyuridine; Diptheria toxin or its cytotoxic fragment or variant, botulinum toxin A , B, C, D, E, or F, ricin or a cytotoxic fragment thereof, e.g., ricin A, abrin or a cytotoxic fragment thereof, saporin or a cytotoxic fragment thereof, pokeweed antiviral toxin or a cytotoxic fragment thereof, and bryodin 1 or a cytotoxic fragment thereof.

[0073] The term "light chain," when used in reference to an antibody, refers to an immunoglobulin light chain, of which there are two types in mammals: lambda (λ) and kappa (κ). Preferably, the light chain is a human light chain. Preferably, the light chain constant region is a human constant region. Multiple light chain constant region alleles exist in the human population. The nucleotide and amino acid sequences of these allelic variants are accessible on publicly available databases such as IMGT, ENSEMBL Swiss-Prot, and Uniprot. In one embodiment, the antibodies and antibody fragments disclosed herein are derived from IGKC*01 (SEQ ID NO: 206 and and 207), IGKC*02 (SEQ ID NOs: 208 and 209), IGKC*03 (SEQ ID NOs: 210 and 211), IGKC*04 (SEQ ID NOs: 212 and 213), and IGKC*05 (SEQ ID NOs: 214 and 215). In one embodiment, the antibodies and antibody fragments disclosed herein include proteins encoded by IGLC1*01 (SEQ ID NOs: 216 and 217), IGLC1*02 (SEQ ID NOs: 219 and 220), IGLC1*03 (SEQ ID NOs: 221 and 222), IGLC1*04 (SEQ ID NOs: 223 and 224), IGLC1*05 (SEQ ID NOs: 225 and 226), IGLC1*06 (SEQ ID NOs: 227 and 228), IGLC1*07 (SEQ ID NOs: 229 and 230), IGLC1*08 (SEQ ID NOs: 231 and 232), IGLC1*09 (SEQ ID NOs: 233 and 234), IGLC1*100 (SEQ ID NOs: 235 and 236), IGLC1*110 (SEQ ID NOs: 237 and 238), IGLC1*120 (SEQ ID NOs: 239 and 239), IGLC1*130 (SEQ ID NOs: 239 and 239), IGLC1*140 (SEQ ID NOs: 239 and 239), IGLC1*150 (SEQ ID NOs: 239 and 239), IGLC1*160 (SEQ ID NOs: 239 and 239), IGLC1*170 (SEQ ID NOs: 239 and 239), IGLC1*180 (SEQ ID NOs: 239 and 239), IGLC1*190 (SEQ ID NOs: 239 and 239), IGLC1*191 (SEQ ID NOs: 239 and 239), IGLC1 Nos. 218, 219, and 220), IGLC2*01 (SEQ ID NOs. 221, 222, and 538), IGLC2*02 (SEQ ID NOs. 224 and 225), IGLC2*03 (SEQ ID NOs. 224 and 225), IGLC3*01 (SEQ ID NOs. 226 and 227), IGLC3*02 (SEQ ID NOs. 228 and 229), IGLC3*03 (SEQ ID NOs. 230 and 231), IGLC3*0 In another embodiment, the antibodies and antibody fragments disclosed herein comprise proteins encoded by human lambda constant region alleles, including, but not limited to, IGLC1*01, IGLC2*01, or IGLC3*01. In another embodiment, the antibodies and antibody fragments disclosed herein comprise proteins encoded by human lambda constant region alleles, including, but not limited to, IGLC4 (SEQ ID NOs: 232 and 233), IGLC6*01 (SEQ ID NOs: 234 and 235), IGLC7*01 (SEQ ID NOs: 236 and 237), IGLC7*02 (SEQ ID NOs: 236 and 237), and IGLC7*03 (SEQ ID NOs: 535 and 536). In another embodiment, the antibodies and antibody fragments disclosed herein comprise a light chain constant region encoded by a mouse kappa constant region allele, including, but not limited to, IGKC*01, IGKC*03, or IGKC*03. In another embodiment, the antibodies and antibody fragments disclosed herein comprise a light chain constant region encoded by a mouse lambda constant region allele, including, but not limited to, IGLC1*01, IGLC2*01, or IGLC3*01.

[0074] "Percent Amino Acid Sequence Identity (%)" related to peptide, polypeptide, or antibody sequences %)" and "homology" align sequences to achieve maximum percent sequence identity; It is defined as the percentage of amino acid residues in a candidate sequence that are identical to a particular peptide or polypeptide sequence, after introducing gaps, if necessary, and without considering any conservative substitutions as part of the sequence identity. Alignment for purposes of determining percent amino acid sequence identity can be accomplished in a variety of ways that are within the skill of the art, for example, using publicly available computer software such as BLAST, BLAST-2, ALIGN, or MEG ALIGN™ (DNASTAR) software. In one embodiment, The % homology is about 70%. In one embodiment, the % homology is about 75%. In one embodiment, the % homology is about 80%. In one embodiment, the % homology is about 85%. In one embodiment, the % homology is about 90%. In one embodiment, the % homology is about 92%. In one embodiment, the % homology is about 95%. In one embodiment, the % homology is about 97%. In one embodiment, the % homology is about 98%. In one embodiment, the % homology is about 99%. In one embodiment, the % homology is 100%.

[0075] The terms "native" or "natural" when used in reference to biological materials, e.g., nucleic acid molecules, polypeptides, host cells, refer to those found in nature and that have not been manipulated by human beings.

[0076] As used herein, "packaging" refers to a container in which components are suitable for distribution and / or use. Packaging refers to the way in which something is packed and / or contained within a sealed unit. Bags, syringes, ampoules, vials, tubes, clamshell packaging, barriers, and / or Containers for maintaining sterility, labeling, etc. may be included.

[0077] As used herein, the term "pharmaceutically acceptable" means approved by a federal or state regulatory agency or listed in the United States Pharmacopoeia, the European Pharmacopoeia, or other generally recognized pharmacopoeias, for use in animals, and more particularly in humans.

[0078] As used herein, the terms "polynucleotide," "nucleotide," "nucleic acid," "nucleic acid molecule," and other similar terms are used interchangeably and include DNA, RNA, mRNA, etc.

[0079] As used herein, "prevent," "preventing," and "prevention" refer to the use of a therapy or combination of therapies provided herein (e.g., a prophylactic or therapeutic agent, such as an antibody of the invention). "Inhibition of hPD-L1-mediated disease" refers to the complete or partial inhibition of the onset, recurrence, development, or metastasis of an hPD-L1-mediated disease and / or symptoms thereof resulting from the administration of a combination of

[0080] The term "soluble" refers to polypeptides, such as PD-L1 and variants or fragments thereof, that lack one or more transmembrane or cytoplasmic domains found in native or membrane-bound forms. In one embodiment, a "soluble" form of PD-L1 lacks both the transmembrane and cytoplasmic domains.

[0081] The term "subject" or "patient" refers to any animal, including, but not limited to, a mammal. As used herein, the term "mammal" refers to any animal that either nurses its young and gives birth to live young (eutherian or placental mammals), or produces eggs (metatherian or non-placental mammals). "mammalian" refers to any vertebrate animal, either a mammal or a mammalian species. Examples of mammalian species include humans and other mammals. primates (including non-human primates such as chimpanzees and other ape and monkey species); livestock animals such as cattle, sheep, pigs, goats, and horses; domestic mammals such as dogs and cats; laboratory animals including rodents such as mice, rats (including cotton rats), and guinea pigs; poultry, wild fowl, and game birds, including chickens, turkeys and other pheasants, ducks, geese, and the like.

[0082] As used herein, "substantially all" refers to at least about 60%, at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 98%, at least about 99%, or about 100%.

[0083] As used herein, the term “substantially free of surfactants” refers to a preparation of an antibody that specifically binds to the hPD-L1 antigen that contains less than 0.0005%, less than 0.0003%, or less than 0.0001% surfactant, and / or less than 0.0005%, 0 Refers to formulations containing less than 0.0003% or less than 0.0001% surfactant.

[0084] As used herein, the term "substantially free of salts" refers to a preparation of an antibody that specifically binds to the hPD-L1 antigen that contains less than 0.0005%, less than 0.0003%, or less than 0.0001% inorganic salts.

[0085] As used herein, the term "surfactant" refers to organic substances with amphiphilic structures, i.e., they are composed of groups with opposite solubility tendencies, typically oil-soluble hydrocarbon chains and water-soluble ionic groups. Depending on the charge of the surface-active moiety, surfactants can be classified into anionic, cationic, and nonionic surfactants. Surfactants are often used as wetting agents, emulsifiers, solubilizers, and dispersants for the preparation of various pharmaceutical compositions and biological materials.

[0086] As used herein, the term "tag" refers to a polypeptide and / or polynucleotide that encodes, for example, hPD-L1 or an hPD-L1 antibody, or an antigen-binding fragment thereof. The term "detectable" or "detection" as used herein refers to any type of moiety attached to a polypeptide. For example, a polynucleotide encoding hPD-L1, an hPD-L1 antibody, or an antigen-binding fragment thereof may include one or more additional tag-encoding nucleotide sequences that encode, for example, a detectable moiety or a moiety that assists in affinity purification. When translated, the tag and antibody may be in the form of a fusion protein. The terms "detectable" or "detection" as used herein with respect to a tag refer to any tag that allows the presence of the tag to be visualized or otherwise determined and / or measured (e.g., by quantification). A non-limiting example of a detectable tag is a fluorescent tag.

[0087] As used herein, the term "therapeutic agent" refers to a therapeutic agent for treating an hPD-L1 mediated disease and / or or symptoms associated therewith. In certain embodiments, the term "therapeutic agent" refers to an antibody of the invention. In certain other embodiments, the term "therapeutic agent" refers to an agent other than an antibody of the invention. Preferably, a therapeutic agent is used to treat, manage, or ameliorate an hPD-L1-mediated disease and / or symptoms associated therewith. Therapeutic Agents are agents that are known to be useful for, or have been used or are currently being used for, hi certain embodiments, the Therapeutic Agent is a fully human anti-hPD-L1 antibody, for example a fully human anti-hPD-L1 monoclonal antibody.

[0088] As used herein, the term "therapy" refers to a treatment for a hPD-L1 mediated disease, such as In certain embodiments, the term "therapy" refers to any protocol, method, and / or agent that can be used by a medical professional or other appropriate professional in the prevention, management, treatment, and / or amelioration of a disease (e.g., cancer). Any of the compounds known to those skilled in the art to be useful in the prevention, management, treatment, and / or amelioration of hPD-L1 mediated diseases. This refers to biologic, supportive, and / or other therapies used in the treatment of cancer.

[0089] The terms "treat," "treatment," and "treating" refer to the progression, severity, and / or stage of an hPD-L1 mediated disease (e.g., cancer) resulting from the administration of one or more therapeutic agents (including, but not limited to, the administration of one or more prophylactic or therapeutic agents, such as an antibody of the invention). In certain embodiments, these terms refer to a reduction or inhibition of hPD-L1 binding to PD-1, a reduction or inhibition of hPD-L1 binding to CD80, and / or an inhibition or amelioration of one or more symptoms associated with an hPD-L1 mediated disease, such as cancer. In certain embodiments, these terms refer to a reduction in PD-1 and / or CD80. Reducing or inhibiting binding of hPD-L1 to and / or hPD-L1-mediated diseases such as cancer In one example, the cells are human cells. In a specific embodiment, the prophylactic agent is a fully human anti-hPD-L1 antibody, for example a fully human anti-hPD-L1 monoclonal antibody.

[0090] The terms "variable region" or "variable domain" refer to the portions of the light and heavy chains, typically the amino-terminal 120-130 amino acids in the heavy chain and approximately 100-110 amino acids in the light chain, which vary widely in sequence among antibodies and are used in the binding and specificity of each particular antibody to its particular antigen. Antibody variability is concentrated in regions called complementarity-determining regions (CDRs), while the more highly conserved regions in the variable domains are called framework regions (FRs). The CDRs of PD-L1 and the heavy chain are primarily responsible for the interaction of the antibody with the antigen. Unless otherwise specified, the numbering of amino acid positions used herein for PD-L1 antibody sequences is based on Kabat et al. (1991) Sequences of proteins of immunological interest. (US Department of Health and Human Services Ices, Washington, DC) 5th ed. ("Kabat et al.") according to the EU index. In a preferred embodiment, the variable region is It is a variable region.

[0091] Definitions of common terms in cell biology and molecular biology can be found in "The Merck Manual of Diagnosis and Therapy", 19th Edition (published by Merck Research Laboratories), 2006 (ISBN 0-911910-19-0), Robert S. Porter et al. (eds.), The Encyclopedia of Molecular Biology (published by Blackwell Science Ltd.), 1994 (I SBN 0-632-02182-9), Benjamin Lewin,Genes X (published by Jones & Bartlett Publishing), 2009 (IS BN-10:0763766321), Kendrew et al. (Eds.), Mo lecular Biology and Biotechnology:a Comprehensive Desk Reference(VCH Publishers, Inc.), 1995 (ISBN 1-56081-569-8), and Cur The details can be found in Routine Protocols in Protein Sciences 2009, Wiley Intersciences, Coligan et al., eds.

[0092] Unless otherwise stated, the present invention is not limited to the methods described in, for example, Sambrook et al., Molecular Cloning: A Laboratory Manual (4th ed.), Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY, USA (2012), Davis et al., Basic Methods in Molecular Biology, Elsevier Science Publishing, Inc., New York, USA (1995), or Methods in Enzymology: Guide to Molecular Cloning Techniques Vol. 152, S.L. Berger and A.R. Kimmel Eds., Academic Press Inc., San Diego, USA (1987), Current Protocols in Protein Science (CPPS) (John E. Coli John Wiley and Sons, Inc.) ,Current Protocols in Cell Biology(CPCB)( Juan S. Bonifacino et al. ed., John Wiley and Sons, Inc.), and Culture by R. Ian Freshney of Animal Cells: A Manual of Basic Technique (Publisher: Wiley-Liss); 5th edition (2005), Ani mal Cell Culture Methods(Methods in Cell These were performed using standard procedures as described in Biology, Vol. 57, Jennie P. Mather and David Barnes editors, Academic Press, 1st edition, 1998. all of which are incorporated herein by reference in their entirety.

[0093] Other terms are defined herein in the description of various aspects of the invention.

[0094] Antibodies against PD-L1 The antigen-binding site of any anti-PD-L1 antibody may be used in the multispecific antibodies of the invention. Numerous examples of anti-PD-L1 antibodies are disclosed herein, and others are known in the art. Characterization data for many of the anti-PD-L1 antibodies mentioned herein are published in US 9,567,399 and US 9,617,338, both of which are incorporated herein by reference.

[0095] 1D05 is a heavy chain variable region (V) of SEQ ID NO: 33, which comprises the CDRH1 amino acid sequence of SEQ ID NO: 27 (IMGT) or SEQ ID NO: 30 (Kabat), the CDRH2 amino acid sequence of SEQ ID NO: 28 (IMGT) or SEQ ID NO: 31 (Kabat), and the CDRH3 amino acid sequence of SEQ ID NO: 29 (IMGT) or SEQ ID NO: 32 (Kabat). H ) amino acid sequence. H The heavy chain nucleic acid sequence of the domain is SEQ ID NO: 34. 1D05 contains a light chain variable region (V) of SEQ ID NO: 43, which contains a CDRL1 amino acid sequence of SEQ ID NO: 37 (IMGT) or SEQ ID NO: 40 (Kabat), a CDRL2 amino acid sequence of SEQ ID NO: 38 (IMGT) or SEQ ID NO: 41 (Kabat), and a CDRL3 amino acid sequence of SEQ ID NO: 39 (IMGT) or SEQ ID NO: 42 (Kabat). L ) amino acid sequence. L The light chain nucleic acid sequence of domain V is SEQ ID NO: 44. H The domain may be combined with any of the heavy chain constant region sequences described herein, for example, SEQ ID NO:193, SEQ ID NO:195, SEQ ID NO:197, SEQ ID NO:199, SEQ ID NO:201, SEQ ID NO:203, SEQ ID NO:205, SEQ ID NO:340, SEQ ID NO:524, SEQ ID NO:526, SEQ ID NO:528, SEQ ID NO:530, SEQ ID NO:532, or SEQ ID NO:534. LThe domain may be combined with any of the light chain constant region sequences described herein, for example, SEQ ID NOs: 207, 209, 211, 213, 215, 217, 219, 221, 223, 225, 227, 229, 231, 233, 235, 237, 536, and 538. The full-length heavy chain amino acid sequence is SEQ ID NO: 35 (SEQ ID NO: 36 for the heavy chain nucleic acid sequence). The full-length light chain amino acid sequence is SEQ ID NO: 45 (SEQ ID NO: 46 for the light chain nucleic acid sequence).

[0096] 84G09 is a heavy chain variable (V) nucleotide sequence of SEQ ID NO: 13, which comprises a CDRH1 amino acid sequence of SEQ ID NO: 7 (IMGT) or SEQ ID NO: 10 (Kabat), a CDRH2 amino acid sequence of SEQ ID NO: 8 (IMGT) or SEQ ID NO: 11 (Kabat), and a CDRH3 amino acid sequence of SEQ ID NO: 9 (IMGT) or SEQ ID NO: 12 (Kabat). H ) region amino acid sequence. H The heavy chain nucleic acid sequence of the domain is SEQ ID NO: 14. 84G09 contains a light chain variable region (V) of SEQ ID NO: 23, which contains a CDRL1 amino acid sequence of SEQ ID NO: 17 (IMGT) or SEQ ID NO: 20 (Kabat), a CDRL2 amino acid sequence of SEQ ID NO: 18 (IMGT) or SEQ ID NO: 21 (Kabat), and a CDRL3 amino acid sequence of SEQ ID NO: 19 (IMGT) or SEQ ID NO: 22 (Kabat). L ) amino acid sequence. L The light chain nucleic acid sequence of domain V is SEQ ID NO: 24. H The domain may be combined with any of the heavy chain constant region sequences described herein, for example, SEQ ID NO:193, SEQ ID NO:195, SEQ ID NO:197, SEQ ID NO:199, SEQ ID NO:201, SEQ ID NO:203, SEQ ID NO:205, SEQ ID NO:340, SEQ ID NO:524, SEQ ID NO:526, SEQ ID NO:528, SEQ ID NO:530, SEQ ID NO:532, or SEQ ID NO:534. L The domain may be combined with any of the light chain constant region sequences described herein, for example, SEQ ID NOs: 207, 209, 211, 213, 215, 217, 219, 221, 223, 225, 227, 229, 231, 233, 235, 237, 536, and 538. The full-length heavy chain amino acid sequence is SEQ ID NO: 15 (heavy chain nucleic acid sequence SEQ ID NO: 16). The full-length light chain amino acid sequence is SEQ ID NO: 25 (light chain nucleic acid sequence SEQ ID NO: 26).

[0097] 1D05 HC variant 1 comprises a heavy chain variable (V) variant of SEQ ID NO: 47, which comprises a CDRH1 amino acid sequence of SEQ ID NO: 27 (IMGT) or SEQ ID NO: 30 (Kabat), a CDRH2 amino acid sequence of SEQ ID NO: 28 (IMGT) or SEQ ID NO: 31 (Kabat), and a CDRH3 amino acid sequence of SEQ ID NO: 29 (IMGT) or SEQ ID NO: 32 (Kabat). H ) region amino acid sequence of SEQ ID NO: 43. 1D05 HC variant 1 has a light chain variable region (V) of SEQ ID NO: 43, which contains a CDRL1 amino acid sequence of SEQ ID NO: 37 (IMGT) or SEQ ID NO: 40 (Kabat), a CDRL2 amino acid sequence of SEQ ID NO: 38 (IMGT) or SEQ ID NO: 41 (Kabat), and a CDRL3 amino acid sequence of SEQ ID NO: 39 (IMGT) or SEQ ID NO: 42 (Kabat). L ) amino acid sequence. L The light chain nucleic acid sequence of domain V is SEQ ID NO: 44. H The domain may be combined with any of the heavy chain constant region sequences described herein, for example, SEQ ID NO:193, SEQ ID NO:195, SEQ ID NO:197, SEQ ID NO:199, SEQ ID NO:201, SEQ ID NO:203, SEQ ID NO:205, SEQ ID NO:340, SEQ ID NO:524, SEQ ID NO:526, SEQ ID NO:528, SEQ ID NO:530, SEQ ID NO:532, or SEQ ID NO:534. L The domain may be combined with any of the light chain constant region sequences described herein, for example, SEQ ID NOs: 207, 209, 211, 213, 215, 217, 219, 221, 223, 225, 227, 229, 231, 233, 235, 237, 536, and 538. The full-length light chain amino acid sequence is SEQ ID NO: 45 (light chain nucleic acid sequence SEQ ID NO: 46).

[0098] 1D05 HC variant 2 comprises a heavy chain variable (V) variant of SEQ ID NO: 48, which comprises a CDRH1 amino acid sequence of SEQ ID NO: 27 (IMGT) or SEQ ID NO: 30 (Kabat), a CDRH2 amino acid sequence of SEQ ID NO: 28 (IMGT) or SEQ ID NO: 31 (Kabat), and a CDRH3 amino acid sequence of SEQ ID NO: 29 (IMGT) or SEQ ID NO: 32 (Kabat). H ) region amino acid sequence of SEQ ID NO: 43. 1D05 HC Variant 2 has a light chain variable region (V) of SEQ ID NO: 43, which contains a CDRL1 amino acid sequence of SEQ ID NO: 37 (IMGT) or SEQ ID NO: 40 (Kabat), a CDRL2 amino acid sequence of SEQ ID NO: 38 (IMGT) or SEQ ID NO: 41 (Kabat), and a CDRL3 amino acid sequence of SEQ ID NO: 39 (IMGT) or SEQ ID NO: 42 (Kabat). L ) amino acid sequence. L The light chain nucleic acid sequence of domain V is SEQ ID NO: 44. H The domain may be combined with any of the heavy chain constant region sequences described herein, for example, SEQ ID NO:193, SEQ ID NO:195, SEQ ID NO:197, SEQ ID NO:199, SEQ ID NO:201, SEQ ID NO:203, SEQ ID NO:205, SEQ ID NO:340, SEQ ID NO:524, SEQ ID NO:526, SEQ ID NO:528, SEQ ID NO:530, SEQ ID NO:532, or SEQ ID NO:534. L The domain may be combined with any of the light chain constant region sequences described herein, for example, SEQ ID NOs: 207, 209, 211, 213, 215, 217, 219, 221, 223, 225, 227, 229, 231, 233, 235, 237, 536, and 538. The full-length light chain amino acid sequence is SEQ ID NO: 45 (light chain nucleic acid sequence SEQ ID NO: 46).

[0099] 1D05 HC variant 3 comprises a heavy chain variable (V) variant of SEQ ID NO: 49, which comprises a CDRH1 amino acid sequence of SEQ ID NO: 27 (IMGT) or SEQ ID NO: 30 (Kabat), a CDRH2 amino acid sequence of SEQ ID NO: 28 (IMGT) or SEQ ID NO: 31 (Kabat), and a CDRH3 amino acid sequence of SEQ ID NO: 29 (IMGT) or SEQ ID NO: 32 (Kabat). H) region amino acid sequence of SEQ ID NO: 43. 1D05 HC Variant 3 has a light chain variable region (V) of SEQ ID NO: 43, which contains a CDRL1 amino acid sequence of SEQ ID NO: 37 (IMGT) or SEQ ID NO: 40 (Kabat), a CDRL2 amino acid sequence of SEQ ID NO: 38 (IMGT) or SEQ ID NO: 41 (Kabat), and a CDRL3 amino acid sequence of SEQ ID NO: 39 (IMGT) or SEQ ID NO: 42 (Kabat). L ) amino acid sequence. L The light chain nucleic acid sequence of domain V is SEQ ID NO: 44. H The domain may be any of the heavy chain constant region sequences described herein, e.g., SEQ ID NO:193, SEQ ID NO:195, SEQ ID NO:197, SEQ ID NO:199, SEQ ID NO:201, SEQ ID NO:203, SEQ ID NO:205, SEQ ID NO:206, SEQ ID NO:207, SEQ ID NO:208, SEQ ID NO:209, SEQ ID NO:210, SEQ ID NO:211, SEQ ID NO:212, SEQ ID NO:213, SEQ ID NO:214, SEQ ID NO:215, SEQ ID NO:216, SEQ ID NO:217, SEQ ID NO:218, SEQ ID NO:219, SEQ ID NO:220, SEQ ID NO:221, SEQ ID NO:222, SEQ ID NO:223, SEQ ID NO:224, S It may be combined with sequence number 340, SEQ ID NO: 524, SEQ ID NO: 526, SEQ ID NO: 528, SEQ ID NO: 530, SEQ ID NO: 532, or SEQ ID NO: 534. L The domain may be combined with any of the light chain constant region sequences described herein, for example, SEQ ID NOs: 207, 209, 211, 213, 215, 217, 219, 221, 223, 225, 227, 229, 231, 233, 235, 237, 536, and 538. The full-length light chain amino acid sequence is SEQ ID NO: 45 (light chain nucleic acid sequence SEQ ID NO: 46).

[0100] 1D05 HC variant 4 comprises a heavy chain variable (V) variant of SEQ ID NO: 342, which comprises a CDRH1 amino acid sequence of SEQ ID NO: 27 (IMGT) or SEQ ID NO: 30 (Kabat), a CDRH2 amino acid sequence of SEQ ID NO: 28 (IMGT) or SEQ ID NO: 31 (Kabat), and a CDRH3 amino acid sequence of SEQ ID NO: 29 (IMGT) or SEQ ID NO: 32 (Kabat). H ) region amino acid sequence of SEQ ID NO: 43. 1D05 HC variant 4 has a light chain variable region (V) of SEQ ID NO: 43, which contains a CDRL1 amino acid sequence of SEQ ID NO: 37 (IMGT) or SEQ ID NO: 40 (Kabat), a CDRL2 amino acid sequence of SEQ ID NO: 38 (IMGT) or SEQ ID NO: 41 (Kabat), and a CDRL3 amino acid sequence of SEQ ID NO: 39 (IMGT) or SEQ ID NO: 42 (Kabat). L ) amino acid sequence. LThe light chain nucleic acid sequence of domain V is SEQ ID NO: 44. H The domain may be combined with any of the heavy chain constant region sequences described herein, for example, SEQ ID NO:193, SEQ ID NO:195, SEQ ID NO:197, SEQ ID NO:199, SEQ ID NO:201, SEQ ID NO:203, SEQ ID NO:205, SEQ ID NO:340, SEQ ID NO:524, SEQ ID NO:526, SEQ ID NO:528, SEQ ID NO:530, SEQ ID NO:532, or SEQ ID NO:534. L The domain may be combined with any of the light chain constant region sequences described herein, for example, SEQ ID NOs: 207, 209, 211, 213, 215, 217, 219, 221, 223, 225, 227, 229, 231, 233, 235, 237, 536, and 538. The full-length light chain amino acid sequence is SEQ ID NO: 45 (light chain nucleic acid sequence SEQ ID NO: 46).

[0101] 1D05 LC variant 1 comprises a heavy chain variable (V) nucleotide sequence of SEQ ID NO: 33, which comprises a CDRH1 amino acid sequence of SEQ ID NO: 27 (IMGT) or SEQ ID NO: 30 (Kabat), a CDRH2 amino acid sequence of SEQ ID NO: 28 (IMGT) or SEQ ID NO: 31 (Kabat), and a CDRH3 amino acid sequence of SEQ ID NO: 29 (IMGT) or SEQ ID NO: 32 (Kabat). H ) region amino acid sequence. H The heavy chain nucleic acid sequence of the domain is SEQ ID NO: 34. 1D05 LC variant 1 comprises a light chain variable region (V) of SEQ ID NO: 50, which comprises a CDRL1 amino acid sequence of SEQ ID NO: 37 (IMGT) or SEQ ID NO: 40 (Kabat), and a CDRL3 amino acid sequence of SEQ ID NO: 39 (IMGT) or SEQ ID NO: 42 (Kabat). L The CDRL2 sequence of 1D05 LC variant 1 has the amino acid sequence V of SEQ ID NO: 50. L As defined by the Kabat or IMGT system from the sequence. HThe domain may be combined with any of the heavy chain constant region sequences described herein, for example, SEQ ID NO:193, SEQ ID NO:195, SEQ ID NO:197, SEQ ID NO:199, SEQ ID NO:201, SEQ ID NO:203, SEQ ID NO:205, or SEQ ID NO:340, SEQ ID NO:524, SEQ ID NO:526, SEQ ID NO:528, SEQ ID NO:530, SEQ ID NO:532, or SEQ ID NO:534. L The domain may be combined with any of the light chain constant region sequences described herein, for example, SEQ ID NOs: 207, 209, 211, 213, 215, 217, 219, 221, 223, 225, 227, 229, 231, 233, 235, 237, 536, and 538. The full-length heavy chain amino acid sequence is set forth in SEQ ID NO: 35 ( The heavy chain nucleic acid sequence is SEQ ID NO: 36).

[0102] 1D05 LC variant 2 comprises a heavy chain variable (V) nucleotide sequence of SEQ ID NO: 33, which comprises a CDRH1 amino acid sequence of SEQ ID NO: 27 (IMGT) or SEQ ID NO: 30 (Kabat), a CDRH2 amino acid sequence of SEQ ID NO: 28 (IMGT) or SEQ ID NO: 31 (Kabat), and a CDRH3 amino acid sequence of SEQ ID NO: 29 (IMGT) or SEQ ID NO: 32 (Kabat). H ) region amino acid sequence. H The heavy chain nucleic acid sequence of domain 1D05 LC variant 2 is SEQ ID NO: 34. The CDRL1 amino acid sequence of SEQ ID NO: 37 (IMGT) or SEQ ID NO: 40 (Kabat), SEQ ID NO: a light chain variable region (V) of SEQ ID NO: 51, comprising a CDRL2 amino acid sequence of SEQ ID NO: 38 (IMGT) or SEQ ID NO: 41 (Kabat), and a CDRL3 amino acid sequence of SEQ ID NO: 39 (IMGT) or SEQ ID NO: 42 (Kabat); L ) amino acid sequence. H The domain may be combined with any of the heavy chain constant region sequences described herein, for example, SEQ ID NO:193, SEQ ID NO:195, SEQ ID NO:197, SEQ ID NO:199, SEQ ID NO:201, SEQ ID NO:203, SEQ ID NO:205, SEQ ID NO:340, SEQ ID NO:524, SEQ ID NO:526, SEQ ID NO:528, SEQ ID NO:530, SEQ ID NO:532, or SEQ ID NO:534. LThe domain may be combined with any of the light chain constant region sequences described herein, for example, SEQ ID NOs: 207, 209, 211, 213, 215, 217, 219, 221, 223, 225, 227, 229, 231, 233, 235, 237, 536, and 538. The full-length heavy chain amino acid sequence is SEQ ID NO: 35 (heavy chain nucleic acid sequence SEQ ID NO: 36).

[0103] 1D05 LC variant 3 comprises a heavy chain variable (V) nucleotide sequence of SEQ ID NO: 33, which comprises a CDRH1 amino acid sequence of SEQ ID NO: 27 (IMGT) or SEQ ID NO: 30 (Kabat), a CDRH2 amino acid sequence of SEQ ID NO: 28 (IMGT) or SEQ ID NO: 31 (Kabat), and a CDRH3 amino acid sequence of SEQ ID NO: 29 (IMGT) or SEQ ID NO: 32 (Kabat). H ) region amino acid sequence. H The heavy chain nucleic acid sequence of the domain is SEQ ID NO: 34. 1D05 LC variant 3 comprises a light chain variable region (V) of SEQ ID NO: 298, which contains a CDRL1 amino acid sequence of SEQ ID NO: 37 (IMGT) or SEQ ID NO: 40 (Kabat), and a CDRL3 amino acid sequence of SEQ ID NO: 39 (IMGT) or SEQ ID NO: 42 (Kabat). L The CDRL2 sequence of 1D05 LC variant 3 has the amino acid sequence V L As defined by the Kabat or IMGT system from the sequence. L The light chain nucleic acid sequence of domain V is SEQ ID NO: 44. H The domain may be combined with any of the heavy chain constant region sequences described herein, for example, SEQ ID NO:193, SEQ ID NO:195, SEQ ID NO:197, SEQ ID NO:199, SEQ ID NO:201, SEQ ID NO:203, SEQ ID NO:205, or SEQ ID NO:340, SEQ ID NO:524, SEQ ID NO:526, SEQ ID NO:528, SEQ ID NO:530, SEQ ID NO:532, or SEQ ID NO:534. LThe domain may be combined with any of the light chain constant region sequences described herein, for example, SEQ ID NOs: 207, 209, 211, 213, 215, 217, 219, 221, 223, 225, 227, 229, 231, 233, 235, 237, 536, and 538. The full-length heavy chain amino acid sequence is SEQ ID NO: 35 (SEQ ID NO: 36 for the heavy chain nucleic acid sequence). The full-length light chain amino acid sequence is SEQ ID NO: 45 (SEQ ID NO: 46 for the light chain nucleic acid sequence).

[0104] 411B08 is a heavy chain variable (V) nucleotide sequence of SEQ ID NO: 58, which comprises a CDRH1 amino acid sequence of SEQ ID NO: 52 (IMGT) or SEQ ID NO: 55 (Kabat), a CDRH2 amino acid sequence of SEQ ID NO: 53 (IMGT) or SEQ ID NO: 56 (Kabat), and a CDRH3 amino acid sequence of SEQ ID NO: 54 (IMGT) or SEQ ID NO: 57 (Kabat). H ) region amino acid sequence. H The heavy chain nucleic acid sequence of the domain is SEQ ID NO: 59. 411B08 contains a light chain variable region (V) of SEQ ID NO: 68, which contains a CDRL1 amino acid sequence of SEQ ID NO: 62 (IMGT) or SEQ ID NO: 65 (Kabat), a CDRL2 amino acid sequence of SEQ ID NO: 63 (IMGT) or SEQ ID NO: 66 (Kabat), and a CDRL3 amino acid sequence of SEQ ID NO: 64 (IMGT) or SEQ ID NO: 67 (Kabat). L ) amino acid sequence. L The light chain nucleic acid sequence of domain V is SEQ ID NO: 69. H The domain may be combined with any of the heavy chain constant region sequences described herein, for example, SEQ ID NO:193, SEQ ID NO:195, SEQ ID NO:197, SEQ ID NO:199, SEQ ID NO:201, SEQ ID NO:203, SEQ ID NO:205, SEQ ID NO:340, SEQ ID NO:524, SEQ ID NO:526, SEQ ID NO:528, SEQ ID NO:530, SEQ ID NO:532, or SEQ ID NO:534. LThe domain may be combined with any of the light chain constant region sequences described herein, for example, SEQ ID NOs: 207, 209, 211, 213, 215, 217, 219, 221, 223, 225, 227, 229, 231, 233, 235, 237, 536, and 538. The full-length heavy chain amino acid sequence is SEQ ID NO: 60 (SEQ ID NO: 61 for the heavy chain nucleic acid sequence). The full-length light chain amino acid sequence is SEQ ID NO: 70 (SEQ ID NO: 71 for the light chain nucleic acid sequence).

[0105] 411C04 contains the heavy chain variable (V) sequence of SEQ ID NO: 78, which comprises the CDRH1 amino acid sequence of SEQ ID NO: 72 (IMGT) or SEQ ID NO: 75 (Kabat), the CDRH2 amino acid sequence of SEQ ID NO: 73 (IMGT) or SEQ ID NO: 76 (Kabat), and the CDRH3 amino acid sequence of SEQ ID NO: 74 (IMGT) or SEQ ID NO: 77 (Kabat). H ) region amino acid sequence. H The heavy chain nucleic acid sequence of the domain is SEQ ID NO: 79. 411C04 contains a light chain variable region (V) of SEQ ID NO: 88, which contains a CDRL1 amino acid sequence of SEQ ID NO: 82 (IMGT) or SEQ ID NO: 85 (Kabat), a CDRL2 amino acid sequence of SEQ ID NO: 83 (IMGT) or SEQ ID NO: 86 (Kabat), and a CDRL3 amino acid sequence of SEQ ID NO: 84 (IMGT) or SEQ ID NO: 87 (Kabat). L ) amino acid sequence. L The light chain nucleic acid sequence of domain V is SEQ ID NO: 89. H The domain may be combined with any of the heavy chain constant region sequences described herein, for example, SEQ ID NO:193, SEQ ID NO:195, SEQ ID NO:197, SEQ ID NO:199, SEQ ID NO:201, SEQ ID NO:203, SEQ ID NO:205, SEQ ID NO:340, SEQ ID NO:524, SEQ ID NO:526, SEQ ID NO:528, SEQ ID NO:530, SEQ ID NO:532, or SEQ ID NO:534. LThe domain may be combined with any of the light chain constant region sequences described herein, for example, SEQ ID NOs: 207, 209, 211, 213, 215, 217, 219, 221, 223, 225, 227, 229, 231, 233, 235, 237, 536, and 538. The full-length heavy chain amino acid sequence is SEQ ID NO: 80 (SEQ ID NO: 81 for the heavy chain nucleic acid sequence). The full-length light chain amino acid sequence is SEQ ID NO: 90 (SEQ ID NO: 91 for the light chain nucleic acid sequence).

[0106] 411D07 is a heavy chain variable (V) antibody having the amino acid sequence of SEQ ID NO: 98, which comprises the CDRH1 amino acid sequence of SEQ ID NO: 92 (IMGT) or SEQ ID NO: 95 (Kabat), the CDRH2 amino acid sequence of SEQ ID NO: 93 (IMGT) or SEQ ID NO: 96 (Kabat), and the CDRH3 amino acid sequence of SEQ ID NO: 94 (IMGT) or SEQ ID NO: 97 (Kabat). H ) region amino acid sequence. H The heavy chain nucleic acid sequence of the domain is SEQ ID NO: 99. 411D07 has a CDRL1 amino acid sequence of SEQ ID NO: 102 (IMGT) or SEQ ID NO: 105 (Kabat), a CDRL2 amino acid sequence of SEQ ID NO: 103 (IMGT) or SEQ ID NO: 106 (Kabat), and a CDRL3 amino acid sequence of SEQ ID NO: 104 (IMGT). SEQ ID NO: 10, which contains the CDRL3 amino acid sequence of SEQ ID NO: 107 (Kabat) 8 light chain variable region (V L ) amino acid sequence. L The light chain nucleic acid sequence of domain V is SEQ ID NO: 109. H The domain may be combined with any of the heavy chain constant region sequences described herein, for example, SEQ ID NO:193, SEQ ID NO:195, SEQ ID NO:197, SEQ ID NO:199, SEQ ID NO:201, SEQ ID NO:203, SEQ ID NO:205, SEQ ID NO:340, SEQ ID NO:524, SEQ ID NO:526, SEQ ID NO:528, SEQ ID NO:530, SEQ ID NO:532, or SEQ ID NO:534. LThe domain can be combined with any of the light chain constant region sequences described herein, for example, SEQ ID NOs: 207, 209, 211, 213, 215, 217, 219, 221, 223, 225, 227, 229, 231, 233, 235, 237, 536, and 538. The full-length heavy chain amino acid sequence can be SEQ ID NO: 100 (heavy chain nucleic acid sequence). The full length light chain amino acid sequence is SEQ ID NO: 110 (light chain nucleic acid sequence SEQ ID NO: 111).

[0107] 385F01 contains the heavy chain variable (V) sequence of SEQ ID NO: 118, which comprises the CDRH1 amino acid sequence of SEQ ID NO: 112 (IMGT) or SEQ ID NO: 115 (Kabat), the CDRH2 amino acid sequence of SEQ ID NO: 113 (IMGT) or SEQ ID NO: 116 (Kabat), and the CDRH3 amino acid sequence of SEQ ID NO: 114 (IMGT) or SEQ ID NO: 117 (Kabat). H ) region amino acid sequence. H The heavy chain nucleic acid sequence of domain 385F01 is SEQ ID NO: 119. 385F01 contains a light chain variable region (V) of SEQ ID NO: 128, which contains a CDRL1 amino acid sequence of SEQ ID NO: 122 (IMGT) or SEQ ID NO: 125 (Kabat), a CDRL2 amino acid sequence of SEQ ID NO: 123 (IMGT) or SEQ ID NO: 126 (Kabat), and a CDRL3 amino acid sequence of SEQ ID NO: 124 (IMGT) or SEQ ID NO: 127 (Kabat). L ) amino acid sequence. L The light chain nucleic acid sequence of domain V is SEQ ID NO: 129. H The domains are the heavy chain constant region sequences described herein. It may be combined with any of the sequences, for example, SEQ ID NO:193, SEQ ID NO:195, SEQ ID NO:197, SEQ ID NO:199, SEQ ID NO:201, SEQ ID NO:203, SEQ ID NO:205, SEQ ID NO:340, SEQ ID NO:524, SEQ ID NO:526, SEQ ID NO:528, SEQ ID NO:530, SEQ ID NO:532, or SEQ ID NO:534. LThe domain may be combined with any of the light chain constant region sequences described herein, for example, SEQ ID NOs: 207, 209, 211, 213, 215, 217, 219, 221, 223, 225, 227, 229, 231, 233, 235, 237, 536, and 538. The full-length heavy chain amino acid sequence is SEQ ID NO: 120 (heavy chain nucleic acid sequence SEQ ID NO: 121). The full-length light chain amino acid sequence is SEQ ID NO: 130 (light chain nucleic acid sequence SEQ ID NO: 131).

[0108] 386H03 is a heavy chain variable (V) nucleotide sequence of SEQ ID NO: 158, which comprises the CDRH1 amino acid sequence of SEQ ID NO: 152 (IMGT) or SEQ ID NO: 155 (Kabat), the CDRH2 amino acid sequence of SEQ ID NO: 153 (IMGT) or SEQ ID NO: 156 (Kabat), and the CDRH3 amino acid sequence of SEQ ID NO: 154 (IMGT) or SEQ ID NO: 157 (Kabat). H ) region amino acid sequence. H The heavy chain nucleic acid sequence of the domain is SEQ ID NO: 159. 386H03 contains a light chain variable region (V) of SEQ ID NO: 168, which contains a CDRL1 amino acid sequence of SEQ ID NO: 162 (IMGT) or SEQ ID NO: 165 (Kabat), a CDRL2 amino acid sequence of SEQ ID NO: 163 (IMGT) or SEQ ID NO: 166 (Kabat), and a CDRL3 amino acid sequence of SEQ ID NO: 164 (IMGT) or SEQ ID NO: 167 (Kabat). L ) amino acid sequence. L The light chain nucleic acid sequence of domain V is SEQ ID NO: 169. H The domain may be combined with any of the heavy chain constant region sequences described herein, for example, SEQ ID NO:193, SEQ ID NO:195, SEQ ID NO:197, SEQ ID NO:199, SEQ ID NO:201, SEQ ID NO:203, SEQ ID NO:205, SEQ ID NO:340, SEQ ID NO:524, SEQ ID NO:526, SEQ ID NO:528, SEQ ID NO:530, SEQ ID NO:532, or SEQ ID NO:534. LThe domain may be combined with any of the light chain constant region sequences described herein, for example, SEQ ID NOs: 207, 209, 211, 213, 215, 217, 219, 221, 223, 225, 227, 229, 231, 233, 235, 237, 536, and 538. The full-length heavy chain amino acid sequence is SEQ ID NO: 160 (SEQ ID NO: 161 for the heavy chain nucleic acid sequence). The full-length light chain amino acid sequence is SEQ ID NO: 170 (SEQ ID NO: 171 for the light chain nucleic acid sequence).

[0109] 389A03 is a heavy chain variable (V) nucleotide sequence of SEQ ID NO: 178, which comprises a CDRH1 amino acid sequence of SEQ ID NO: 172 (IMGT) or SEQ ID NO: 175 (Kabat), a CDRH2 amino acid sequence of SEQ ID NO: 173 (IMGT) or SEQ ID NO: 176 (Kabat), and a CDRH3 amino acid sequence of SEQ ID NO: 174 (IMGT) or SEQ ID NO: 177 (Kabat). H ) region amino acid sequence. H The heavy chain nucleic acid sequence of the domain is SEQ ID NO: 179. 389A03 contains a light chain variable region (V) of SEQ ID NO: 188, which contains a CDRL1 amino acid sequence of SEQ ID NO: 182 (IMGT) or SEQ ID NO: 185 (Kabat), a CDRL2 amino acid sequence of SEQ ID NO: 183 (IMGT) or SEQ ID NO: 186 (Kabat), and a CDRL3 amino acid sequence of SEQ ID NO: 184 (IMGT) or SEQ ID NO: 187 (Kabat). L ) amino acid sequence. L The light chain nucleic acid sequence of domain V is SEQ ID NO: 189. H The domain may be combined with any of the heavy chain constant region sequences described herein, for example, SEQ ID NO:193, SEQ ID NO:195, SEQ ID NO:197, SEQ ID NO:199, SEQ ID NO:201, SEQ ID NO:203, SEQ ID NO:205, SEQ ID NO:340, SEQ ID NO:524, SEQ ID NO:526, SEQ ID NO:528, SEQ ID NO:530, SEQ ID NO:532, or SEQ ID NO:534. LThe domain may be combined with any of the light chain constant region sequences described herein, for example, SEQ ID NOs: 207, 209, 211, 213, 215, 217, 219, 221, 223, 225, 227, 229, 231, 233, 235, 237, 536, and 538. The full-length heavy chain amino acid sequence is SEQ ID NO: 180 (SEQ ID NO: 181 for the heavy chain nucleic acid sequence). The full-length light chain amino acid sequence is SEQ ID NO: 190 (SEQ ID NO: 191 for the light chain nucleic acid sequence).

[0110] 413D08 is a heavy chain variable (V) nucleotide sequence of SEQ ID NO: 138, which comprises a CDRH1 amino acid sequence of SEQ ID NO: 132 (IMGT) or SEQ ID NO: 135 (Kabat), a CDRH2 amino acid sequence of SEQ ID NO: 133 (IMGT) or SEQ ID NO: 136 (Kabat), and a CDRH3 amino acid sequence of SEQ ID NO: 134 (IMGT) or SEQ ID NO: 137 (Kabat). H ) region amino acid sequence. H The heavy chain nucleic acid sequence of the domain is SEQ ID NO: 139. 413D08 contains a light chain variable region (V) of SEQ ID NO: 148, which contains a CDRL1 amino acid sequence of SEQ ID NO: 142 (IMGT) or SEQ ID NO: 145 (Kabat), a CDRL2 amino acid sequence of SEQ ID NO: 143 (IMGT) or SEQ ID NO: 146 (Kabat), and a CDRL3 amino acid sequence of SEQ ID NO: 144 (IMGT) or SEQ ID NO: 147 (Kabat). L ) amino acid sequence. L The light chain nucleic acid sequence of domain V is SEQ ID NO: 149. H The domain may be combined with any of the heavy chain constant region sequences described herein, for example, SEQ ID NO:193, SEQ ID NO:195, SEQ ID NO:197, SEQ ID NO:199, SEQ ID NO:201, SEQ ID NO:203, SEQ ID NO:205, SEQ ID NO:340, SEQ ID NO:524, SEQ ID NO:526, SEQ ID NO:528, SEQ ID NO:530, SEQ ID NO:532, or SEQ ID NO:534. LThe domain may be combined with any of the light chain constant region sequences described herein, for example, SEQ ID NOs: 207, 209, 211, 213, 215, 217, 219, 221, 223, 225, 227, 229, 231, 233, 235, 237, 536, and 538. The full-length heavy chain amino acid sequence is SEQ ID NO: 140 (heavy chain nucleic acid sequence SEQ ID NO: 141). The full-length light chain amino acid sequence is SEQ ID NO: 150 (light chain nucleic acid sequence SEQ ID NO: 151).

[0111] 413G05 is a heavy chain variable (V) nucleotide sequence of SEQ ID NO: 244, which comprises a CDRH1 amino acid sequence of SEQ ID NO: 238 (IMGT) or SEQ ID NO: 241 (Kabat), a CDRH2 amino acid sequence of SEQ ID NO: 239 (IMGT) or SEQ ID NO: 242 (Kabat), and a CDRH3 amino acid sequence of SEQ ID NO: 240 (IMGT) or SEQ ID NO: 243 (Kabat). H ) region amino acid sequence. H The heavy chain nucleic acid sequence of the domain is SEQ ID NO: 245. 413G05 contains a light chain variable region (V) of SEQ ID NO: 254, which contains a CDRL1 amino acid sequence of SEQ ID NO: 248 (IMGT) or SEQ ID NO: 251 (Kabat), a CDRL2 amino acid sequence of SEQ ID NO: 249 (IMGT) or SEQ ID NO: 252 (Kabat), and a CDRL3 amino acid sequence of SEQ ID NO: 250 (IMGT) or SEQ ID NO: 253 (Kabat). L ) amino acid sequence. L The light chain nucleic acid sequence of domain V is SEQ ID NO: 255. H The domain may be combined with any of the heavy chain constant region sequences described herein, for example, SEQ ID NO:193, SEQ ID NO:195, SEQ ID NO:197, SEQ ID NO:199, SEQ ID NO:201, SEQ ID NO:203, SEQ ID NO:205, SEQ ID NO:340, SEQ ID NO:524, SEQ ID NO:526, SEQ ID NO:528, SEQ ID NO:530, SEQ ID NO:532, or SEQ ID NO:534. LThe domain may be combined with any of the light chain constant region sequences described herein, for example, SEQ ID NOs: 207, 209, 211, 213, 215, 217, 219, 221, 223, 225, 227, 229, 231, 233, 235, 237, 536, and 538. The full-length heavy chain amino acid sequence is SEQ ID NO: 246 (heavy chain nucleic acid sequence SEQ ID NO: 247). The full-length light chain amino acid sequence is SEQ ID NO: 256 (light chain nucleic acid sequence SEQ ID NO: 257).

[0112] 413F09 contains the heavy chain variable (V) sequence of SEQ ID NO: 264, which comprises the CDRH1 amino acid sequence of SEQ ID NO: 258 (IMGT) or SEQ ID NO: 261 (Kabat), the CDRH2 amino acid sequence of SEQ ID NO: 259 (IMGT) or SEQ ID NO: 262 (Kabat), and the CDRH3 amino acid sequence of SEQ ID NO: 260 (IMGT) or SEQ ID NO: 263 (Kabat). H ) region amino acid sequence. H The heavy chain nucleic acid sequence of the domain is SEQ ID NO: 265. 413F09 contains a light chain variable region (V) of SEQ ID NO: 274, which contains a CDRL1 amino acid sequence of SEQ ID NO: 268 (IMGT) or SEQ ID NO: 271 (Kabat), a CDRL2 amino acid sequence of SEQ ID NO: 269 (IMGT) or SEQ ID NO: 272 (Kabat), and a CDRL3 amino acid sequence of SEQ ID NO: 270 (IMGT) or SEQ ID NO: 273 (Kabat). L ) amino acid sequence. L The light chain nucleic acid sequence of domain V is SEQ ID NO: 275. H The domains are the heavy chain constant region sequences described herein. It may be combined with any of the sequences, for example, SEQ ID NO:193, SEQ ID NO:195, SEQ ID NO:197, SEQ ID NO:199, SEQ ID NO:201, SEQ ID NO:203, SEQ ID NO:205, SEQ ID NO:340, SEQ ID NO:524, SEQ ID NO:526, SEQ ID NO:528, SEQ ID NO:530, SEQ ID NO:532, or SEQ ID NO:534. LThe domain may be combined with any of the light chain constant region sequences described herein, for example, SEQ ID NOs: 207, 209, 211, 213, 215, 217, 219, 221, 223, 225, 227, 229, 231, 233, 235, 237, 536, and 538. The full-length heavy chain amino acid sequence is SEQ ID NO: 266 (SEQ ID NO: 267 for the heavy chain nucleic acid sequence). The full-length light chain amino acid sequence is SEQ ID NO: 276 (SEQ ID NO: 277 for the light chain nucleic acid sequence).

[0113] 414B06 is a heavy chain variable (V) nucleotide sequence of SEQ ID NO: 284, which comprises a CDRH1 amino acid sequence of SEQ ID NO: 278 (IMGT) or SEQ ID NO: 281 (Kabat), a CDRH2 amino acid sequence of SEQ ID NO: 279 (IMGT) or SEQ ID NO: 282 (Kabat), and a CDRH3 amino acid sequence of SEQ ID NO: 280 (IMGT) or SEQ ID NO: 283 (Kabat). H ) region amino acid sequence. H The heavy chain nucleic acid sequence of the domain is SEQ ID NO: 285. 414B06 contains the light chain variable region (V) of SEQ ID NO: 294, which comprises the CDRL1 amino acid sequence of SEQ ID NO: 288 (IMGT) or SEQ ID NO: 291 (Kabat), the CDRL2 amino acid sequence of SEQ ID NO: 289 (IMGT) or SEQ ID NO: 292 (Kabat), and the CDRL3 amino acid sequence of SEQ ID NO: 290 (IMGT) or SEQ ID NO: 293 (Kabat). L ) amino acid sequence. L The light chain nucleic acid sequence of domain V is SEQ ID NO: 295. H The domain may be combined with any of the heavy chain constant region sequences described herein, for example, SEQ ID NO:193, SEQ ID NO:195, SEQ ID NO:197, SEQ ID NO:199, SEQ ID NO:201, SEQ ID NO:203, SEQ ID NO:205, SEQ ID NO:340, SEQ ID NO:524, SEQ ID NO:526, SEQ ID NO:528, SEQ ID NO:530, SEQ ID NO:532, or SEQ ID NO:534. LThe domain may be combined with any of the light chain constant region sequences described herein, for example, SEQ ID NOs: 207, 209, 211, 213, 215, 217, 219, 221, 223, 225, 227, 229, 231, 233, 235, 237, 536, and 538. The full-length heavy chain amino acid sequence is SEQ ID NO: 286 (heavy chain nucleic acid sequence SEQ ID NO: 287). The full-length light chain amino acid sequence is SEQ ID NO: 296 (light chain nucleic acid sequence SEQ ID NO: 297).

[0114] 416E01 contains a heavy chain variable region (V) of SEQ ID NO: 349, which contains the CDRH1 amino acid sequence of SEQ ID NO: 343 (IMGT) or SEQ ID NO: 346 (Kabat), the CDRH2 amino acid sequence of SEQ ID NO: 344 (IMGT) or SEQ ID NO: 347 (Kabat), and the CDRH3 amino acid sequence of SEQ ID NO: 345 (IMGT) or SEQ ID NO: 348 (Kabat). H ) amino acid sequence. H The heavy chain nucleic acid sequence of the domain is SEQ ID NO: 350. 416E01 contains a light chain variable region (V) of SEQ ID NO: 359, which contains a CDRL1 amino acid sequence of SEQ ID NO: 353 (IMGT) or SEQ ID NO: 356 (Kabat), a CDRL2 amino acid sequence of SEQ ID NO: 354 (IMGT) or SEQ ID NO: 357 (Kabat), and a CDRL3 amino acid sequence of SEQ ID NO: 355 (IMGT) or SEQ ID NO: 358 (Kabat). L ) amino acid sequence. L The light chain nucleic acid sequence of domain V is SEQ ID NO: 360. H The domain may be combined with any of the heavy chain constant region sequences described herein, for example, SEQ ID NO:193, SEQ ID NO:195, SEQ ID NO:197, SEQ ID NO:199, SEQ ID NO:201, SEQ ID NO:203, SEQ ID NO:205, SEQ ID NO:340, SEQ ID NO:524, SEQ ID NO:526, SEQ ID NO:528, SEQ ID NO:530, SEQ ID NO:532, or SEQ ID NO:534. LThe domain may be combined with any of the light chain constant region sequences described herein, for example, SEQ ID NOs: 207, 209, 211, 213, 215, 217, 219, 221, 223, 225, 227, 229, 231, 233, 235, 237, 536, and 538. The full-length heavy chain amino acid sequence is SEQ ID NO: 351 (SEQ ID NO: 352 for the heavy chain nucleic acid sequence). The full-length light chain amino acid sequence is SEQ ID NO: 361 (SEQ ID NO: 362 for the light chain nucleic acid sequence).

[0115] Concepts regarding PD-L1 antibodies Concept 1. An antibody or fragment thereof that specifically binds to hPD-L1 as defined by SEQ ID NO: 1 and competes with antibody 1D05 for binding to said hPD-L1, wherein the antibody or fragment comprises a V H X1, X2, and X3 are independently any amino acid, and X4 is either present or absent, and if present, can be any amino acid.

[0116] In these concepts, the antibody or fragment may or may not include a bispecific antibody. In one embodiment, in these concepts, the antibody or fragment includes a bispecific antibody. In one embodiment, the bispecific antibody does not include the FIT-Ig format. In one embodiment, a "bispecific antibody" is a mAb 2 In one embodiment, the bispecific antibody does not include the FIT-Ig format or the mAb 2 In one embodiment, the antibodies or fragments in these concepts include bispecific antibodies, but do not include bispecific antibodies having the FIT-Ig format. In one embodiment, the antibodies or fragments in these concepts include bispecific antibodies, but do not include bispecific antibodies having the mAb format. 2 In one embodiment, the antibody or fragment in this context includes a bispecific antibody, but does not include a bispecific antibody having a FIT-Ig format or a mAb format. 2In another embodiment, the antibody or fragment in this context does not include a bispecific antibody having a bispecific format.

[0117] Preferably, antibodies or fragments thereof that specifically bind to the hPD-L1 antigen do not cross-react with other antigens (but optionally, with antigens from different species, such as rhesus monkeys, cynomolgus monkeys, or (It may cross-react with mouse PD-L1.) The fragments may be identified, for example, by immunoassays, BIAcore™, or other techniques known in the art. An antibody or fragment thereof specifically binds to the hPD-L1 antigen when it binds to the hPD-L1 antigen with greater affinity than it binds to any cross-reactive antigens, as determined using laboratory techniques such as radioimmunoassay (RIA) and enzyme-linked immunosorbent assay (ELISA). Typically, a specific or selective reaction is at least 2-fold background signal or noise, more typically more than 10-fold background. See, for example, Paul, ed., 1989, Fundamental Immunology Second Edition, Raven, 1999, for a discussion of antibody specificity. See pages 332-336 of the New York Press.

[0118] In one embodiment, the antibody or fragment is a human antibody. In one embodiment, the antibody or fragment is a human antibody or fragment. In one embodiment, the antibody or fragment is a fully human antibody or fragment. In one embodiment, the antibody or fragment is a fully human monoclonal antibody or fragment.

[0119] Concept 1a: An antibody or fragment thereof that specifically binds to hPD-L1 as defined by SEQ ID NO: 1 and competes with antibody 411B08 for binding to said hPD-L1, wherein the antibody or fragment comprises a V HAlso provided are antibodies or fragments thereof comprising the domain, wherein X1, X2, X3, X4, and X5 are independently any amino acid.

[0120] Concept 1b: An antibody or fragment thereof that specifically binds to hPD-L1 as defined by SEQ ID NO: 1 and competes with antibody 411B08 for binding to said hPD-L1, wherein the antibody or fragment comprises a V H Also provided are antibodies or fragments thereof comprising the domain, wherein X1 is any amino acid.

[0121] As provided in the concepts or embodiments herein, an anti-PD-L1 antibody or immunocytokinin The kine was determined by surface plasmon resonance to have a K of less than 50 nM, less than 40 nM, or less than 30 nM. D In another embodiment, the anti-PD-L1 antibody or immunocytokine may bind to PD-L1, e.g., human PD-L1, having a K of less than 20 nM, less than 15 nM, or less than 10 nM as determined by surface plasmon resonance. D The anti-PD-L1 or immunocytokine may bind to PD-L1, e.g., human PD-L1, having a K of less than 8 nM, less than 5 nM, less than 4 nM, less than 3 nM, less than 2 nM, or less than 1 nM as determined by surface plasmon resonance. D It can bind to PD-L1 having the formula K, e.g., human PD-L1. D may be 0.9 nM or less, 0.8 nM or less, 0.7 nM or less, 0.6 nM or less, 0.5 nM or less, 0.4 nM or less, 0.3 nM or less, 0.2 nM or less, or 0.1 nM or less.

[0122] In another embodiment, K D is in the range of 0.01 to 1 nM, or in the range of 0.05 to 2 nM, or in the range of 0.05 to 1 nM. D may relate to hPD-L1, cynomolgus monkey PD-L1, and / or mouse PD-L1.

[0123] In another embodiment, the anti-PD-L1 antibodies described herein have a K of about 0.5-10 μM, for example, about 1-8 μM or about 1-7 μM. ON speed (e.g., SP at 25°C or 37°C) In another embodiment, K ON The rate is about 1-5 μM, e.g. For example, about 1 μM, about 1.5 μM, about 2 μM, about 2.5 μM, or about 3 μM. ON The rate is about 3.5 μM, about 4 μM, about 4.5, about 5 μM, or about 5.5 μM.

[0124] In another embodiment, the anti-PD-L1 antibodies described herein have a K OFF speed (e.g., 25°C or 3 7°C as measured by SPR. OFF The speed is about 0.5 In another embodiment, the K OFF The rate is about 0.6 mM, about 0.7 mM, about 0.8 mM, or about 0.9 mM.

[0125] In another embodiment, the anti-PD-L1 antibodies (and and immunocytokines) were improved over other anti-PD-L1 antibodies and immunocytokines Thus, in one embodiment, an anti-PD-L1 antibody (or immunoglobulin) is administered to a subject in need of treatment with PD-L1. The immunocytokine is expressed at a concentration of approximately 100 μg / mL or within the range of approximately 100 to 350 μg / mL. At present levels, it is expressed in HEK293 cells, e.g., HEK293T cells. In another embodiment, the expression level is greater than about 350 μg / mL.

[0126] In another embodiment, the anti-PD-L1 antibody (or immunocytokine) is at about 100 μg / mL or at an expression level in the range of about 100 to 350 μg / mL, in CHO cells, e.g., Ex In another embodiment, the expression level is about 350 μg / m Exceeds L.

[0127] In another embodiment, the anti-PD-L1 antibody (or immunocytokine) is at about 100 μg / mL or at an expression level in the range of about 100 to 350 μg / mL, in CHO cells, e.g., Ex In another embodiment, the expression level is greater than about 350 μg / mL. Human IgG1 (formulated as SEQ ID NO: 340) was cultured in O-E7 EBNA cells in a volume of 2 L. The antibody produced has an expression level of approximately 115 μg / mL. Human IgG1 (SEQ ID NO: 1) was cultured in CHO-E7 EBNA cells in a volume of 2 L as described herein. The antibody described herein as 1414B06, formatted as human IgG1 (SEQ ID NO: 340) in CHO-E7 EBNA cells in a 2 L volume, has an expression level of approximately 783 μg / mL. The antibody described herein as 413G05, formatted as a human IgG1 (SEQ ID NO: 340) in CHO-E7 EBNA cells at a volume of 2 L, has an expression level of approximately 383 μg / mL.

[0128] In any of these expression systems, expression is carried out in a volume of about 0.5 mL to 3 mL, for example, about 0.5 mL to 2 mL. In any of these expression systems, the anti-PD-L1 antibody (or immunocytokine) may be expressed from a pTT5 vector. In any of these expression systems, the anti-PD-L1 antibody (or immunocytokine) may be expressed in conjunction with a lipid transfection reagent, and may optionally be expressed in CHO cells, for example Expi-CHO cells. In any of these expression systems, the anti-PD-L1 antibody (or immunocytokine) may be expressed in combination with a lipid transfection reagent, and may optionally be expressed in CHO cells, for example Expi-CHO cells. The IL-16 receptor agonist (IL-16 receptor agonist) can be expressed in conjunction with a PEI transfection reagent, optionally with CH In either of these expression systems, the anti-PD-L1 antibody (or immunocytokine) can be expressed in a helper plasmid (e.g., CHO-E7 EBNA cells). , AKT helper plasmid), and optionally in CHO cells, e.g. It can be expressed in HO-E7 EBNA cells. In all of these expression systems, the expression levels range from about 100 μg / mL to about 1500 μg / mL. μg / mL, for example, about 100 μg / mL to about 1000 μg / mL, about 200 μg / mL to about 1000 μg / mL, or about 350 μg / mL to about 1000 μg / mL. In any of the expression systems, the lower limit of expression can be about 100 μg / mL, about 200 μg / mL, about 300 μg / mL, or about 400 μg / mL. In another embodiment, the lower limit of expression can be about 500 μg / mL, about 600 μg / mL, about 700 μg / mL, or about 800 μg / mL. In any of these expression systems, the upper limit of expression can be about 2000 μg / mL. , about 1800 μg / mL, about 1600 μg / mL, or about 1500 μg / mL. In another embodiment, the upper limit of expression can be about 1250 μg / mL, about 1000 μg / mL, about 900 μg / mL, or about 800 μg / mL.

[0129] In another embodiment, the expression system is a Lonza expression system, e.g., Lonza X-Ceed ( In the Lonza expression system, expression is carried out in a volume of approximately 30 mL to 2 L, e.g., 5 mL. This can be carried out in volumes of about 0 mL to 1 L or 1 L to 2 L. In the Lonza expression system, the anti-PD-L1 antibody (or immunocytokine) can be expressed in conjunction with electroporation, and optionally without any helper plasmid. In the Lonza expression system, the anti-PD-L1 antibody (or immunocytokine) can be expressed at about 1 g / L, or about 900 mg / L, or about 800 mg / L. or about 700 mg / L. In another embodiment, the Lonza expression system In this case, the anti-PD-L1 antibody (or immunocytokine) is administered at a concentration of about 600 mg / L, or about 50 mg / L. In the Lonza expression system, the anti-PD-L1 antibody (or immunocytokine) may be expressed at a level of about 400 mg / L to about 2 g / L, for example, about 500 mg / L to about 1.5 g / L, or about 500 mg / L to about 1 g / L. In another embodiment, the expression level is greater than about 1 g / L.

[0130] Concept 2. The antibody or fragment of Concept 1, wherein X1 is a hydroxyl-containing amino acid, optionally T. In one embodiment, the hydroxyl-containing amino acid is serine. In one embodiment, the hydroxyl-containing amino acid is cysteine. In one embodiment, the hydroxyl-containing amino acid is threonine. In one embodiment, the hydroxyl-containing amino acid is methionine. In one embodiment, the hydroxyl-containing amino acid is serine or cysteine. In one embodiment, the hydroxyl-containing amino acid is serine or threonine. In one embodiment, the hydroxyl-containing amino acid is serine or methionine. In one embodiment, the hydroxyl-containing amino acid is cysteine ​​or threonine. In one embodiment, the hydroxyl-containing amino acid is cysteine ​​or methionine. In one embodiment, the hydroxyl-containing amino acid is threonine or methionine. In one embodiment, the hydroxyl-containing amino acid is selected from serine, cysteine, threonine, and methionine. can be.

[0131] Concept 2a. The antibody or fragment according to Concept 1a, wherein X1 is an aliphatic amino acid or an amide amino acid. In one embodiment, X1 is selected from asparagine (N) and valine (V). In one embodiment, X1 is valine. In one embodiment, X1 is asparagine.

[0132] Concept 2b. The antibody or fragment according to Concept 1b, wherein X1 is an aliphatic amino acid. In one embodiment, X1 is selected from alanine (A) or valine (V). In one embodiment, X1 is valine. In one embodiment, X1 is alanine.

[0133] Concept 3. The antibody or fragment of Concept 1 or Concept 2, wherein X2 is a basic amino acid, optionally K. In one embodiment, the hydroxyl-containing amino acid is histidine. In one embodiment, the hydroxyl-containing amino acid is lysine. In one embodiment, the hydroxyl-containing amino acid is arginine. In one embodiment, the hydroxyl-containing amino acid is histidine or lysine. In one embodiment, the hydroxyl-containing amino acid is histidine or arginine. In one embodiment, the hydroxyl-containing amino acid is lysine or arginine. In one embodiment, the hydroxyl-containing amino acid is selected from histidine, lysine, and arginine.

[0134] Concept 3a. The antibody or fragment according to Concept 1a or Concept 2a, wherein X2 is an aliphatic amino acid or an amide amino acid. In one embodiment, X2 is selected from leucine (L), isoleucine (I), valine (V), asparagine (N), and glutamine (Q). In one embodiment, X2 is selected from leucine (L), isoleucine (I), and valine (V). In one embodiment, X2 is selected from asparagine (N) and glutamine (Q). In one embodiment, X2 is selected from leucine (L) and glutamine (Q). In one embodiment, X2 is leucine (L). In one embodiment, X2 is glutamine (Q).

[0135] Concept 4. The antibody or fragment of any one of Concepts 1-3, wherein X2 is a hydroxyl-containing amino acid, optionally S or T. In one embodiment, the hydroxyl-containing amino acid is serine. In one embodiment, the hydroxyl-containing amino acid is cysteine. In one embodiment, the hydroxyl-containing amino acid is threonine. In one embodiment, the hydroxyl-containing amino acid is methionine. In one embodiment, the hydroxyl-containing amino acid is serine or cysteine. In one embodiment, the hydroxyl-containing amino acid is serine or threonine. In one embodiment, the hydroxyl-containing amino acid is serine or methionine. In one embodiment, the hydroxyl-containing amino acid is cysteine ​​or threonine. In one embodiment, the hydroxyl-containing amino acid is cysteine ​​or methionine. In one embodiment, the hydroxyl-containing amino acid is threonine or methionine. In one embodiment, the hydroxyl-containing amino acid is selected from serine, cysteine, threonine, and methionine.

[0136] Concept 4a. The antibody or fragment of any one of Concepts 1a, 2a, or 3a, wherein X3 is an aromatic amino acid. In one embodiment, X3 is selected from phenylalanine (F), tyrosine (Y), and tryptophan (W). In one embodiment, X3 is selected from tyrosine (Y) and tryptophan (W). In one embodiment, X3 is tyrosine (Y). In one embodiment, X3 is tryptophan (W).

[0137] Concept 5. The antibody or fragment of any one of Concepts 1-4, wherein X3 is an aromatic amino acid, optionally W. In one embodiment, the hydroxyl-containing amino acid is phenylalanine. In one embodiment, the hydroxyl-containing amino acid is tyrosine. In one embodiment, The hydroxyl-containing amino acid is tryptophan. In one embodiment, the hydroxyl-containing amino acid is phenylalanine or tyrosine. In one embodiment, the hydroxyl-containing amino acid is phenylalanine or tryptophan. In one embodiment, the hydroxyl-containing amino acid is tyrosine or tryptophan. In one embodiment, the hydroxyl-containing amino acid is selected from phenylalanine, tyrosine, and tryptophan.

[0138] Concept 5a. The antibody or fragment of any one of Concepts 1a, 2a, 3a, or 4a, wherein X4 is an aromatic amino acid. In one embodiment, X4 is selected from phenylalanine (F), tyrosine (Y), and tryptophan (W). In one embodiment, X4 is tyrosine ( In one embodiment, X4 is selected from tyrosine (Y) and phenylalanine (F). In one embodiment, X4 is phenylalanine (F).

[0139] Concept 6. The antibody or fragment of any one of Concepts 1 to 5, wherein X4 is absent.

[0140] Concept 6a. The antibody or fragment of Concept 1a, 2a, 3a, 4a, or 5a, wherein X5 is an aliphatic amino acid or a hydroxyl-containing amino acid. In one embodiment, X5 is selected from leucine (L), isoleucine (I), valine (V), serine (S), cysteine ​​(C), and threonine (T). In one embodiment, X5 is selected from leucine (L), isoleucine (I), and valine (V). In one embodiment, X5 is selected from serine (S), cysteine ​​(C), and threonine (T). In one embodiment, X5 is selected from leucine (L) and serine (S). In one embodiment, X5 is selected from serine ( In one embodiment, X5 is leucine (L).

[0141] Concept 7. The antibody or fragment of any one of Concepts 1 to 5, wherein X4 is present.

[0142] Concept 8. The antibody or fragment of Concept 7, wherein X4 is an aliphatic amino acid, optionally G. In one embodiment, the hydroxyl-containing amino acid is selected from glycine, alanine, valine, leucine, and isoleucine. In one embodiment, the hydroxyl-containing amino acid is selected from glycine and alanine. In one embodiment, the hydroxyl-containing amino acid is selected from glycine and valine. In one embodiment, the hydroxyl-containing amino acid is selected from glycine and leucine. In one embodiment, the hydroxyl-containing amino acid is selected from glycine and isoleucine. In one embodiment, the hydroxyl-containing amino acid is selected from alanine and valine. In one embodiment, the hydroxyl-containing amino acid is selected from alanine and leucine. In one embodiment, the hydroxyl-containing amino acid is selected from alanine and isoleucine. In one embodiment, the hydroxyl-containing amino acid is selected from valine and leucine. In one embodiment, the hydroxyl-containing amino acid is selected from valine and isoleucine. In one embodiment, the hydroxyl-containing amino acid is selected from leucine and isoleucine. In one embodiment, the hydroxyl-containing amino acids are selected from three of each of glycine, alanine, valine, leucine, and isoleucine, hi one embodiment, the hydroxyl-containing amino acids are selected from four of each of glycine, alanine, valine, leucine, and isoleucine.

[0143] Concept 9. The antibody or fragment thereof, optionally according to any one of Concepts 1-8, that specifically binds to hPD-L1 and competes with antibody 1D05 for binding to hPD-L1, wherein the antibody or fragment comprises a CDRH3 sequence of SEQ ID NO: 29 or 32, or a CDRH3 sequence of SEQ ID NO: 29 or 32 with 6 or fewer amino acid substitutions. H An antibody or fragment thereof, comprising a domain.

[0144] Concept 9a: A compound that specifically binds to hPD-L1 and is anti-hPD-L1 9. The antibody or fragment thereof of any one of claims 1 to 8, which competes with antibody 84G09, wherein the antibody or fragment comprises a CDRH3 sequence of SEQ ID NO: 9 or 12, or a CDRH3 sequence of SEQ ID NO: 9 or 12 containing six or fewer amino acid substitutions. H An antibody or fragment thereof, comprising a domain.

[0145] Concept 9b: The antibody or fragment thereof, optionally according to any one of Concepts 1-8, that specifically binds to hPD-L1 and competes with antibody 411B08 for binding to hPD-L1, wherein the antibody or fragment comprises a CDRH3 sequence of SEQ ID NO: 54 or 57, or a CDRH3 sequence of SEQ ID NO: 54 or 57 with 6 or fewer amino acid substitutions. H An antibody or fragment thereof, comprising a domain.

[0146] Concept 9c: The antibody or fragment thereof, optionally according to any one of Concepts 1-8, that specifically binds to hPD-L1 and competes with antibody 411C04 for binding to hPD-L1, wherein the antibody or fragment comprises a CDRH3 sequence of SEQ ID NO: 74 or 77, or a CDRH3 sequence of SEQ ID NO: 74 or 77 with 6 or fewer amino acid substitutions. H An antibody or fragment thereof, comprising a domain.

[0147] Concept 9d: The antibody or fragment thereof, optionally according to any one of Concepts 1-8, that specifically binds to hPD-L1 and competes with antibody 411D07 for binding to hPD-L1, wherein the antibody or fragment comprises a CDRH3 sequence of SEQ ID NO: 94 or 97, or a CDRH3 sequence of SEQ ID NO: 94 or 97 with 3 or fewer amino acid substitutions. H An antibody or fragment thereof, comprising a domain.

[0148] Concept 9e: The antibody or fragment thereof, optionally according to any one of Concepts 1-8, that specifically binds to hPD-L1 and competes with antibody 385F01 for binding to hPD-L1, wherein the antibody or fragment comprises a CDRH3 sequence of SEQ ID NO: 114 or 117, or a CDRH3 sequence of SEQ ID NO: 114 or 117 with 6 or fewer amino acid substitutions. H An antibody or fragment thereof, comprising a domain.

[0149] Concept 9f: The antibody or fragment thereof, optionally according to any one of Concepts 1-8, that specifically binds to hPD-L1 and competes with antibody 386H03 for binding to hPD-L1, wherein the antibody or fragment comprises a CDRH3 sequence of SEQ ID NO: 144 or 147, or a CDRH3 sequence of SEQ ID NO: 144 or 147 with 3 or fewer amino acid substitutions. H An antibody or fragment thereof, comprising a domain.

[0150] Concept 9g: The antibody or fragment thereof, optionally according to any one of Concepts 1-8, that specifically binds to hPD-L1 and competes with antibody 389A03 for binding to hPD-L1, wherein the antibody or fragment comprises a CDRH3 sequence of SEQ ID NO: 174 or 177, or a CDRH3 sequence of SEQ ID NO: 174 or 177 with 6 or fewer amino acid substitutions. H An antibody or fragment thereof, comprising a domain.

[0151] Concept 9h: The antibody or fragment thereof, optionally according to any one of Concepts 1-8, that specifically binds to hPD-L1 and competes with antibody 413D08 for binding to hPD-L1, wherein the antibody or fragment comprises a CDRH3 sequence of SEQ ID NO: 134 or 137, or a CDRH3 sequence of SEQ ID NO: 134 or 137 with 5 or fewer amino acid substitutions. H An antibody or fragment thereof, comprising a domain.

[0152] Concept 9i: The antibody or fragment thereof, optionally according to any one of Concepts 1-8, that specifically binds to hPD-L1 and competes with antibody 413G05 for binding to hPD-L1, wherein the antibody or fragment has a CDRH3 sequence of SEQ ID NO: 240 or 243; or a V comprising the CDRH3 sequence of SEQ ID NO: 240 or 243 containing six or fewer amino acid substitutions H An antibody or fragment thereof, comprising a domain.

[0153] Concept 9j: The antibody or fragment thereof, optionally according to any one of Concepts 1-8, that specifically binds to hPD-L1 and competes with antibody 413F09 for binding to hPD-L1, wherein the antibody or fragment comprises a CDRH3 sequence of SEQ ID NO: 260 or 263, or a CDRH3 sequence of SEQ ID NO: 260 or 263 with 6 or fewer amino acid substitutions. H An antibody or fragment thereof, comprising a domain.

[0154] Concept 9k: The antibody or fragment thereof, optionally according to any one of Concepts 1-8, that specifically binds to hPD-L1 and competes with antibody 414B06 for binding to hPD-L1, wherein the antibody or fragment comprises a CDRH3 sequence of SEQ ID NO: 280 or 283, or a CDRH3 sequence of SEQ ID NO: 280 or 283 with 6 or fewer amino acid substitutions. H An antibody or fragment thereof, comprising a domain.

[0155] Concept 91: The antibody or fragment thereof, optionally according to any one of Concepts 1-8, that specifically binds to hPD-L1 and competes with antibody 416E01 for binding to hPD-L1, wherein the antibody or fragment comprises a CDRH3 sequence of SEQ ID NO: 345 or 348, or a CDRH3 sequence of SEQ ID NO: 345 or 348 with 6 or fewer amino acid substitutions. H An antibody or fragment thereof, comprising a domain.

[0156] In all of concepts 9, 9a-l, 17, 17a-l, 18, 18a-l, 19, 19a-l, 22, 22a-l, 23, 23a-l, 24, and 24a-l, in one embodiment the CDRs comprise one amino acid substitution, which may be a conservative amino acid substitution. In all of concepts 9, 9a-l, 17, 17a-l, 18, 18a-l, 19, 19a-l, 22, 22a-l, 23, 23a-l, 24, and 24a-l, in one embodiment the CDRs comprise two amino acid substitutions, which may be conservative amino acid substitutions. In all of concepts 9, 9a-l, 17, 17a-l, 18, 18a-l, 19, 19a-l, 22, 22a, 22b, 22d, 22f, 22g, 24, and 24a-l, in one embodiment the CDRs comprise three amino acid substitutions, which may be conservative amino acid substitutions. In all of concepts 9, 9a-c, 9e, 9g-k, 17, 17a-c, 17e, 17g-l, 19, 19a, 22, 22d, 22f, 22g, 24, and 24a-l, in one embodiment the CDRs comprise four amino acid substitutions, which may be conservative amino acid substitutions. In all of Concepts 9, 9a-c, 9e, 9g-l, 17, 17a-c, 17e, 17g-l, 22d, 22f, and 22g, in one embodiment, the CDRs comprise five amino acid substitutions, which may be conservative amino acid substitutions. In all of Concepts 9, 9a-c, 9e, 9g, 9i-l, 17, 17a-c, 17e, 17g, and 17i-l, in one embodiment, the CDRs comprise six amino acid substitutions, which may be conservative amino acid substitutions. Amino acid substitutions include modifications in which an amino acid is replaced with a different naturally occurring amino acid residue. Such substitutions may be classified as "conservative," in which an amino acid residue contained in a polypeptide is replaced with another naturally occurring amino acid having similar characteristics in terms of either polarity, side chain functionality, or size. This conservative substitution is well known in the art. Substitutions encompassed by the present invention may also be "non-conservative," in which an amino acid residue present in the peptide is substituted with an amino acid having different properties, such as a naturally occurring amino acid from a different group (e.g., substituting alanine for an anionic or hydrophobic amino acid), or alternatively, a naturally occurring amino acid is substituted with an unconventional amino acid.

[0157] In one embodiment, conservative amino acid substitutions are as described herein, for example, substitutions include substitution of Y with F, substitution of T with S or K, substitution of P with A, substitution of E with D or Q, substitution of N with D or G, substitution of R with K, substitution of G with N or A, substitution of S or substitution of T with K, substitution of D with N or E, substitution of I with L or V, substitution of F with Y, substitution of S with T or A, substitution of R with K, substitution of G with N or A, substitution of K with R, substitution of A with S, K, or P. In another embodiment, conservative amino acid substitutions can be substitution of Y with F, T with A or S, I with L or V, W with Y, M with L, N with D, G with A, T with A or S, D with N, I with L or V, F with Y or L, S with A or T, and A with S, G, T, or V.

[0158] Concept 10. V that specifically binds to hPD-L1 and contains a CDRH3 of 12-20 amino acids H domain, human V H gene segment, a human D gene segment, and a human J H Derived from the recombination of gene segments, human J H An antibody or fragment wherein the gene segment is IGHJ5 (e.g., IGHJ5*02).

[0159] In one embodiment, CDRH3 is 14-17 amino acids and is a human J H The gene segment is IGHJ5 (e.g., IGHJ5*02).

[0160] Concept 10a: A V molecule that specifically binds to hPD-L1 and contains a CDRH3 of 8 to 16 amino acids. H domain, human V H gene segment, a human D gene segment, and a human J HDerived from the recombination of gene segments, human J H Also provided are antibodies or fragments wherein the gene segment is selected from IGHJ4 (e.g., IGHJ4*02), IGHJ5 (e.g., IGHJ5*02), and IGHJ6 (e.g., IGHJ6*02). H In another embodiment, the gene segment is IGHJ6 (e.g., IGHJ6*02). In another embodiment, the CDRH3 is 10 to 17 amino acids and is a human J H In another embodiment, the gene segment is IGHJ6 (e.g., IGHJ6*02). H In another embodiment, the gene segment is IGHJ4 (e.g., IGHJ4*02). In another embodiment, the CDRH3 is 7 to 17 amino acids and is a human J H The gene segment is IGHJ4 (e.g., IGHJ4*02).

[0161] Optionally, the antibody of Concept 10 or 10a has any of the properties of Concepts 1-9, including binding affinity, K on and K off rates, expression level, half-life, etc.

[0162] Concept 11. Human V H The gene segment is IGHV3 (e.g., IGHV3-9*01 The antibody or fragment of Concept 10 or 10a, wherein the antibody or fragment is IGHV3-9, such as IGHV4-9.

[0163] Concept 11a, Human V H The gene segment is an IGHV3 (e.g., IGHV3- or IGHV3-9, such as IGHV3-9*01, or IGHV3-7, such as IGHV3-7*01, or IGHV3-33, such as IGHV3-33*01, or IGHV3-11, such as IGHV3-11*01, or IGHV3-23, such as IGHV3-23*04) or IGHV4 (e.g., IGHV4-4, such as IGHV4-4*02, or IGHV4-39, such as IGHV4-39*01). Also provided is an antibody or fragment according to Concept 10 or 10a. In one embodiment, human V H The gene segment may be IGHV3 (e.g., IGHV3-7* In one embodiment, the human IGHV is an IGHV3-7, such as IGHV 01. H The gene segment is In one embodiment, the human V is an IGHV3 (e.g., an IGHV3-33, such as IGHV3-33*01). H The gene segment may be an IGHV3 (e.g., IGHV3-11*01 or an IGHV3-11*02 gene). In one embodiment, the human V H The gene segment is IGHV3 (e.g., IGHV3-23, such as IGHV3-23*04). H The gene segment can be an IGHV4 (e.g., an IGHV such as IGHV4-4*02). 4-4). In one embodiment, human V H The gene segments are selected from IGHV4 (e.g., I IGHV4-39, such as GHV4-39*01).

[0164] As concept 11b, the human D gene segment is IGHD1 (e.g., IGHD1-2 IGHD1-20 (e.g., IGHD3-10, such as IGHD3-10*01), IGHD4 (e.g., IGHD4-11, such as IGHD4-11*01), IGHD5 (e.g., IGHD5-7, such as IGHD5-18*01), and IGHD6 (e.g., For example, IGHD6-13 such as IGHD6-13*01) Also provided are antibodies or fragments of any of the above. In one embodiment, the human D gene segment is selected from the group consisting of IGHD1 (e.g., IGHD1-2, such as IGHD1-20*01). In one embodiment, the human D gene segment is IGHD3 (e.g., IGHD3-10, such as IGHD3-10*01). In one embodiment, the human D gene segment is IGHD4 (e.g., IGHD4-11, such as IGHD4-11*01). In one embodiment, the human D gene segment is IGHD5 (e.g., IGHD5-19*01). In one embodiment, the human D gene segment is IGHD 6 (e.g., IGHD6-13, such as IGHD6-13*01).

[0165] In concepts 10, 11, and 11a, V H , D H , and J H The gene segments are as described in the antibody combinations in Table 5 herein below. In one embodiment, the heavy chain of the antibody is an IGHV3 (e.g., IGHV3-7*01, such as IGHV3-7*01). 7), IGHD4 (e.g., IGHD4-11, such as IGHD4-11*01), and IGH In one embodiment, the heavy chain of the antibody is derived from a combination of IGHV4 (e.g., IGHV4-4, such as IGHV4-4*02), IGHD3 (e.g., In one embodiment, the heavy chain of the antibody is derived from a combination of IGHV4 (e.g., IGHV4-39, such as IGHV4-39*01), IGHD6 (e.g., IGHD6-13, such as IGHD6-13*01), and IGHJ1 (e.g., IGHJ1*01). In one embodiment, the heavy chain of the antibody is a polypeptide of IGHV3 (e.g., IGHV3-33*01, etc.). In one embodiment, the IgG1 gene is derived from a combination of IGHD5 (e.g., IGHD5-18, such as IGHD5-18*01), IGHDV3-33), IGHD5 (e.g., IGHD5-18, such as IGHD5-18*01), and IGHJ6 (e.g., IGHJ6*02). The heavy chain of the antibody can be selected from IGHV3 (e.g., IGHV3-11, such as IGHV3-11*01), IGHD1 (e.g., IGHD1-20, such as IGHD1-20*01), and IGHJ6 (e.g., In one embodiment, the heavy chain of the antibody is derived from the combination IGHJ6*02. V3 (e.g., IGHV3-23 such as IGHV3-23*04), IGHD5 (e.g., I In one embodiment, the heavy chain of the antibody is derived from a combination of IGHV3 (e.g., IGHV3-7, such as IGHV3-7*01), IGHD5 (e.g., IGHD5-24, such as IGHD5-24*01), and IGHJ4 (e.g., IGHJ4*02). In embodiments, the heavy chain of the antibody is an IGHV3 (e.g., IGHV3-23*04, etc.) V3-23), IGHD6 (e.g., IGHD6-13 such as IGHD6-13*01), and and IGHJ4 (e.g., IGHJ4*02).

[0166] Concept 12. The antibody or fragment is a Vκ gene derived from the recombination of a human Vκ gene segment and a Jκ gene segment. L domain, and the human Vκ gene segment is IGκV1D( For example, IGκV1D-39 such as IGκV1D-39*01) 11, 11a, or 11b.

[0167] As concept 12a, the human Vκ gene segment is IGκV1 (e.g., IGκV1- IGκV1-17 (e.g., IGκV1-17*01), or IGκV1-9 (e.g., IGκV1-9*d01), or IGκV1D-12 (e.g., IGκV1D-12*02), or IGκV1D-39 (e.g., IGκV1D-39*01) and IGκV4 (e.g., IGκV4- 1*01) (IGκV4-1, such as IGκV4-1) Also provided is an antibody or fragment according to any one of 11b. In one embodiment, the human Vκ gene segment is IGκV1 (e.g., IGκV1-17, such as IGκV1-17*01). In one embodiment, the human Vκ gene segment is IGκV1 (e.g., IGκV In one embodiment, the human Vκ gene segment is IGκV1-9, such as IGκV1-9*d01. In one embodiment, the human Vκ gene segment is IGκV1 (e.g., IGκV1D-12, such as IGκV1D-12*02). In one embodiment, the human Vκ gene segment is IGκV1D-39, such as 9*01. is IGκV1 IGκV4 (e.g., IGκV4-1 such as IGκV4-1*01).

[0168] As concept 12b, the human Jκ gene segment is IGκJ1 (e.g., IGκJ1* Also provided is an antibody or fragment of Concept 10, 10a, 11, or 11a, wherein the antibody or fragment is selected from IGκJ2 (e.g., IGκJ2*01), IGκJ2 (e.g., IGκJ2*04), IGκJ3 (e.g., IGκJ3*01), IGκJ4 (e.g., IGκJ4*01), or IGκJ5 (e.g., IGκJ5*01). In one embodiment, the human Jκ gene segment is IGκJ1 (e.g., IGκ In one embodiment, the human Jκ gene segment is IGκJ2 (e.g., IGκJ2*04). In one embodiment, the human Jκ gene segment is IGκJ In one embodiment, the human Jκ gene segment is IGκJ3 (e.g., IGκJ4*01). In one embodiment, the human Jκ gene segment is IGκJ5 (e.g., IGκJ5*01).

[0169] In both Concepts 12 and 12a, the Vκ and Jκ gene segments are as set forth in the antibody combinations in Table 5 herein below. In one embodiment, the light chain of the antibody is derived from a combination of IGKV1D (e.g., IGKV1D-12, such as IGKV1D-12*02) and IGKJ3 (e.g., IGKJ3*01). In one embodiment, the light chain of the antibody is derived from a combination of IGKV4 (e.g., IGKV4-1, such as IGKV14-1*01) and IGKJ2 (e.g., IGKJ2*04). In one embodiment, the light chain of the antibody is derived from a combination of IGKV1 (e.g., IGKV1-17, such as IGKV1-17*01) and IGKJ1 (e.g., In one embodiment, the light chain of the antibody is derived from the combination IGKV1 D (e.g., IGKV1D-12, such as IGKV1D-12*02) and IGKJ4 (e.g., In one embodiment, the light chain of the antibody is derived from a combination of IGKV1 (e.g., IGKV1-9 such as IGKV1-9*d01) and IGKJ5 (e.g., IGKJ5*01). In one embodiment, the light chain of the antibody is derived from a combination of IGKV1D (e.g., IGKV1D-12 such as IGKV1D-12*02) and IGKJ5 (e.g., IGKJ5*01).

[0170] Concept 13. An antibody or fragment thereof that specifically binds to the same epitope as that specifically bound by antibody 1D05.

[0171] Concept 13a. An antibody or fragment thereof that specifically binds to the same epitope as the antibody 84G09 specifically binds.

[0172] Concept 13b. An antibody or fragment thereof that specifically binds to the same epitope as antibody 411B08 specifically binds.

[0173] Concept 13c. An antibody or fragment thereof that specifically binds to the same epitope as antibody 411C04.

[0174] Concept 13d. An epitope identical to the epitope specifically bound by antibody 411D07 An antibody or fragment thereof that specifically binds to

[0175] Concept 13e. An antibody or fragment thereof that specifically binds to the same epitope as antibody 385F01 specifically binds.

[0176] Concept 13f. An antibody or fragment thereof that specifically binds to the same epitope as the antibody 386H03 specifically binds.

[0177] Concept 13g. An antibody or fragment thereof that specifically binds to the same epitope as the antibody 389A03 specifically binds.

[0178] Concept 13h. An antibody or fragment thereof that specifically binds to the same epitope as the antibody 413D08 specifically binds.

[0179] Concept 13i. An antibody or fragment thereof that specifically binds to the same epitope as the antibody 413G05 specifically binds.

[0180] Concept 13j. An antibody or fragment thereof that specifically binds to the same epitope as antibody 413F09.

[0181] Concept 13k. An antibody or fragment thereof that specifically binds to the same epitope as antibody 414B06.

[0182] Concept 131. An antibody or fragment thereof that specifically binds to the same epitope as the antibody 416E01 specifically binds.

[0183] The antibodies described in these concepts have sequences as described herein above.

[0184] In one embodiment, an antibody is provided that specifically binds to an epitope that is substantially similar to the epitope bound by the antibody in Concepts 13, 13a-13l.

[0185] Contact amino acid residues involved in the interaction of an antibody with an antigen can be determined by various methods known in the art. In one embodiment, sequential substitution of amino acids in the antigen sequence (e.g., using standard molecular biology techniques to mutate the DNA in the coding sequence), in this case Substitution of hPD-L1 with alanine (also known as alanine scanning) or another unrelated amino acid may provide a residue that reduces or eliminates the ability of the antibody to recognize the antigen of interest. Binding may be assessed using standard techniques, including but not limited to, SPR, HTRF, and ELISA (described elsewhere herein). Altering the charge of the side chain of an amino acid in the antigen sequence (e.g., changing a lysine to a glutamic acid) may also be used. exchanging polar and non-polar residues (e.g., serine for leucine), etc. Other substitutions may be made to enhance disruption of binding. Alanine scanning or other amino substitution methods can be performed either with recombinant soluble antigen, or, if the target is a cell membrane target, directly on cells using transient or stable expression of the mutated version. In one embodiment, contact residues between the antibody and the antigen are determined (i.e., Protein crystallography can be used to determine the epitope to which an antibody binds. Crystallography allows direct visualization of contact residues involved in antibody-antigen interactions. Alongside standard X-ray crystallography, cryo-electron microscopy has been used to determine contact residues between antibodies and HIV capsid proteins (Lee, Jeong Hyun, et al. “Antibodies to a conformational ep. (See, "Antope on gp41 neutralizes HIV-1 by destabilizing the Env spike." Nature communications, 6, (2015)). In one embodiment, the antibody binds to a linear epitope. If the antibody recognizes the antigen sequence, short peptides can be generated based on the antigen sequence, and the binding of the antibody to these peptides can be detected by a variety of methods including, but not limited to, SPR, HTRF, ELISA ( can be assessed using standard techniques such as those described elsewhere herein. Further investigation of the pitope can be provided by performing an alanine scan on any peptides that show binding. Pepscan technology (http: / / www.pepscan.com / ) is used, which uses chemical attachment of peptides onto a backbone instead of linear peptides. Conformational scanning can be performed using a nucleotide sequence, which has been used to identify discontinuous epitopes on CD20-targeting antibodies (Niederfellner, Gerhard, et al. al. “Epitope characterization and crystal structure of GA101 provide insights into the molecular basis for type I / II dist In one embodiment, to identify the binding epitope, For example, limited protein digestion and mass spectrophotometric measurements can be used. The antibody-antigen complex is digested with a protease, such as, but not limited to, trypsin. The digested complex peptides are compared with digestion mass spectrophotometric measurements of the antibody alone and the antigen alone to determine whether specific epitopes are protected by complex formation. Competitive binding, a further step involving amino acid substitution, can be used to narrow down the individual amino acid residues involved in the interaction (see, e.g., Suckau, Detlev, et al., "Molecular epitopes"). (See, "Epitope identification by limited proteolysis of an immobilized antigen-antibody complex and mass spectrometric peptide mapping." Proceedings of the National Academy of Sciences, 87.24, (1990), 9848-9852.) Thus, in one embodiment, the contact residues of an epitope are identified by unrelated amino acid sequences. In another embodiment, the unrelated amino acid scan (e.g., alanine scan) is performed using a technique selected from SPR, HTRF, ELISA, X-ray crystallography, cryo-electron microscopy, and limited protein digestion in combination with mass spectrometry. In one embodiment, the unrelated amino acid scan (e.g., alanine scan) is performed using a technique selected from SPR, HTRF, ELISA, X-ray crystallography, cryo-electron microscopy, and limited protein digestion in combination with mass spectrometry. For example, alanine scanning) is performed using HTRF. An amino acid scan (e.g., an alanine scan) is performed using ELISA. When an alanine scan is performed using either ELISA or HTRF, an amino acid residue is identified as contributing to the epitope if there is at least a 25% reduction in signal. In one embodiment, the signal reduction is at least 30%. In one embodiment, the signal reduction is at least 35%. In one embodiment, the signal reduction is at least 40%. In one embodiment, the signal reduction is at least 45%. In one embodiment, the signal reduction is at least 50%. In one embodiment, the signal reduction is at least 55%. In one embodiment, the signal reduction is at least 60%. In one embodiment, the signal reduction is at least 70%. In one embodiment, the signal reduction is at least 75%. In one embodiment, the signal reduction is at least 80%. In one embodiment, the signal reduction is at least 85%. In one embodiment, the signal reduction is at least 90%. When an alanine scan is performed using SPR, an amino acid residue is identified as contributing to the epitope if there is at least a 10-fold reduction in affinity. In one embodiment, the affinity reduction is at least 15-fold. In one embodiment, the affinity reduction is at least 20-fold. In one embodiment, the affinity reduction is at least 30-fold. In one embodiment, the affinity reduction is at least 40-fold. In one embodiment, the affinity reduction is at least 10-fold. The reduction is at least 50-fold. In one embodiment, the reduction in affinity is at least 100-fold. Thus, in one embodiment, the contact residues of the epitope are identified by X-ray crystallography. Thus, in one embodiment, the contact residues of the epitope are identified by cryo-electron microscopy. Thus, in one embodiment, the contact residues of the epitope are identified by a combination of limited protein digestion and mass spectrometry.

[0186] Concept 14. The antibody or fragment of Concept 13, wherein the epitope is identified by an independent amino acid scan or X-ray crystallography.

[0187] Concept 15. The antibody or fragment of Concept 14, wherein the contact residues of the epitope are defined by at least a 10-fold reduction in affinity in an unrelated amino acid scan, e.g., an alanine scan, as determined by SPR. In one embodiment, the reduction in affinity is at least 15-fold. In one embodiment, the reduction in affinity is at least 20-fold. In one embodiment, the reduction in affinity is at least 30-fold. In one embodiment, the reduction in affinity is at least 40-fold. In one embodiment, the reduction in affinity is at least 50-fold. In one embodiment, the reduction in affinity is at least 100-fold. SPR can be performed as described herein above.

[0188] Concept 16. An antibody or fragment thereof that competes with antibody 1D05 for binding to hPD-L1.

[0189] Competition may be determined by surface plasmon resonance (SPR), a technique that will be readily apparent to one of ordinary skill in the art. SPR may be performed using Biacore™, Proteon™, or another standard SPR technique. The competition may be due, for example, to antibodies or fragments that bind to the same or overlapping epitopes of hPD-L1. In one embodiment, competition may be determined by ELISA, a technique that will be readily apparent to one of ordinary skill in the art. In one embodiment, competition may be determined by homogeneous time-resolved fluorescence (HTRF), a technique that will be readily apparent to one of ordinary skill in the art. In one embodiment, competition may be determined by fluorescence-activated cell sorting (FAC). S), techniques which will be readily apparent to those skilled in the art. Competition is determined by ForteBio Octet® Bio-Layer Interferometry (BLI), a technique readily apparent to those skilled in the art.

[0190] In one embodiment, the antibody or fragment competes with hPD-1 (or a fusion protein thereof) for binding to cell surface-expressed hPD-L1 (e.g., in a dose-dependent manner). In one embodiment, the antibody or fragment competes with hPD-1 (or a fusion protein thereof) for binding to soluble hPDL-1. In one embodiment, the antibody or fragment competes with the target protein (e.g., in a dose-dependent manner). Partially inhibits the binding of PD-1 and / or CD80 to cell surface-expressed PD-L1, etc. In another embodiment, the antibody or fragment partially or completely inhibits the binding of hPD-1 and / or CD80 to soluble hPD-L1. The antibody or fragment partially or completely increases secretion of IFNγ, CD25, and IL-2 from cells with cell surface-expressed PD-1. In one embodiment, the antibody or fragment partially or completely inhibits binding of CD80 to soluble hPD-L1, but does not show any detectable inhibition of PD-1 binding to cell surface-expressed PD-L1. In one embodiment, the antibody or fragment partially or completely inhibits binding of CD80 to soluble hPD-L1, but does not show any detectable inhibition of PD-1 binding to soluble PD-L1.

[0191] As used herein, "inhibit," "inhibition," "inhibiting," and the like, as used herein, refer to the inhibition of an antagonist (e.g., an antibody or fragment thereof) by a ligand (e.g., partially or completely inhibits the binding of receptors (e.g., CD80 or PD-1) to the target antigen (PD-L1). Antagonists are capable of binding to epitopes that interfere with either the function or the target. If the peptide completely blocks the binding site of the ligand, the ligand binding is completely prevented (this Blockage can be physical, in the case of overlapping epitopes, or steric, if the antagonist is large enough to prevent ligand binding to its unique epitope), and the ligand is released from the circulation. Therefore, the concentration of circulating ligand may appear to be increased. If the epitope to which the antagonist binds partially blocks the binding site of the ligand, the ligand may be able to bind, but only weakly (partial inhibition). In this case, the ligand binds in an orientation different from the natural binding interaction. Some of these may be removed from the circulation, but not as many as would be removed if the ligand-binding site were completely free and available for binding. Thus, inhibition refers to the physical interaction between the ligand and the receptor. Inhibition can be measured by HTRF, which is described in more detail elsewhere herein and in Mathis (1995) Clinical Chemistry 41(9), 1391-1397. Inhibition can also be measured by flow cytometry, where the receptor is expressed on cells, or by ELISA, where the receptor is adsorbed to a plate.

[0192] Concept 16a. An antibody or fragment thereof that competes with antibody 84G09 for binding to hPD-L1.

[0193] Concept 16b. An antibody or fragment thereof that competes with antibody 411B08 for binding to hPD-L1.

[0194] Concept 16c. An antibody or fragment thereof that competes with antibody 411C04 for binding to hPD-L1.

[0195] Concept 16d. An antibody or fragment thereof that competes with antibody 411D07 for binding to hPD-L1.

[0196] Concept 16e. An antibody or fragment thereof that competes with antibody 385F01 for binding to hPD-L1.

[0197] Concept 16f. An antibody or fragment thereof that competes with antibody 386H03 for binding to hPD-L1.

[0198] Concept 16g. An antibody or fragment thereof that competes with antibody 389A03 for binding to hPD-L1.

[0199] Concept 16h. An antibody or fragment thereof that competes with antibody 413D08 for binding to hPD-L1.

[0200] Concept 16i. An antibody or fragment thereof that competes with antibody 413G05 for binding to hPD-L1.

[0201] Concept 16j. An antibody or fragment thereof that competes with antibody 413F09 for binding to hPD-L1.

[0202] Concept 16k. An antibody or fragment thereof that competes with antibody 414B06 for binding to hPD-L1.

[0203] Concept 161. An antibody or fragment thereof that competes with antibody 416E01 for binding to hPD-L1.

[0204] The antibody has a sequence as described herein above.

[0205] Concept 17.V H 17. The antibody or fragment of any one of Concepts 10 to 16, wherein the domain comprises the CDRH3 sequence of SEQ ID NO: 29 or 32, or the CDRH3 sequence of SEQ ID NO: 29 or 32 with six or fewer amino acid substitutions.

[0206] Concept 17a:V H Any one of concepts 10 to 16 (provided that if dependent on concept 13, it is dependent on concept 13a and if dependent on concept 16), wherein the domain comprises a CDRH3 sequence of SEQ ID NO: 9 or 12, or a CDRH3 sequence of SEQ ID NO: 9 or 12 containing six or fewer amino acid substitutions. (If dependent, subject to concept 16a) or a fragment thereof.

[0207] Concept 17b:V H Any one of concepts 10 to 16 (provided that if subject to concept 13, then subject to concept 13b, and if subject to concept 16, then subject to concept 13b), wherein the domain comprises a CDRH3 sequence of SEQ ID NO: 54 or 57, or a CDRH3 sequence of SEQ ID NO: 54 or 57 containing six or fewer amino acid substitutions. 6, then subject to concept 16b).

[0208] Concept 17c:V HAny one of concepts 10 to 16 (provided that if subject to concept 13, then subject to concept 13c, and if subject to concept 16, then subject to concept 13c), wherein the domain comprises a CDRH3 sequence of SEQ ID NO: 74 or 77, or a CDRH3 sequence of SEQ ID NO: 74 or 77 containing six or fewer amino acid substitutions. 6, or if dependent on concept 16c, the antibody or fragment thereof.

[0209] Concept 17d:V H Any one of concepts 10 to 16 (provided that if subject to concept 13, then subject to concept 13d, and if subject to concept 14, then subject to concept 15), wherein the domain comprises a CDRH3 sequence of SEQ ID NO: 94 or 97, or a CDRH3 sequence of SEQ ID NO: 94 or 97 containing no more than three amino acid substitutions. 6, when dependent on concept 16d), or a fragment thereof.

[0210] Concept 17e:V H Any one of concepts 10 to 16 (provided that if subject to concept 13, then subject to concept 13e) wherein the domain comprises the CDRH3 sequence of SEQ ID NO: 114 or 117, or the CDRH3 sequence of SEQ ID NO: 114 or 117 containing six or fewer amino acid substitutions. , and if dependent on Concept 16, dependent on Concept 16e).

[0211] Concept 17f:V H Any one of concepts 10 to 16 (provided that if subject to concept 13, then subject to concept 13f) wherein the domain comprises the CDRH3 sequence of SEQ ID NO: 144 or 147, or the CDRH3 sequence of SEQ ID NO: 144 or 147 containing no more than three amino acid substitutions. (or, if dependent on Concept 16, dependent on Concept 16f).

[0212] Concept 17g:V H Any one of Concepts 10 to 16 (provided that if subject to Concept 13, then subject to Concept 13g) wherein the domain comprises the CDRH3 sequence of SEQ ID NO: 174 or 177, or the CDRH3 sequence of SEQ ID NO: 174 or 177 containing six or fewer amino acid substitutions. (or, if dependent on Concept 16, dependent on Concept 16g).

[0213] Concept 17h:V H Any one of concepts 10 to 16 (provided that if subject to concept 13, then subject to concept 13h) wherein the domain comprises the CDRH3 sequence of SEQ ID NO: 134 or 137, or the CDRH3 sequence of SEQ ID NO: 134 or 137 containing 5 or fewer amino acid substitutions. , and if dependent on Concept 16, dependent on Concept 16h).

[0214] Concept 17i:V H Any one of concepts 10 to 16 (provided that if subject to concept 13, then subject to concept 13i) wherein the domain comprises a CDRH3 sequence of SEQ ID NO: 240 or 243, or a CDRH3 sequence of SEQ ID NO: 240 or 243 containing six or fewer amino acid substitutions. (or, if dependent on Concept 16, dependent on Concept 16i).

[0215] Concept 17j:V H Any one of concepts 10 to 16 (provided that if subject to concept 13, then subject to concept 13j) wherein the domain comprises a CDRH3 sequence of SEQ ID NO: 260 or 263, or a CDRH3 sequence of SEQ ID NO: 260 or 263 containing six or fewer amino acid substitutions. (or, if dependent on Concept 16, dependent on Concept 16j).

[0216] Concept 17k:V H Any one of Concepts 10 to 16 (provided that if subject to Concept 13, then subject to Concept 13k) wherein the domain comprises a CDRH3 sequence of SEQ ID NO: 280 or 283, or a CDRH3 sequence of SEQ ID NO: 280 or 283 containing six or fewer amino acid substitutions. , and if dependent on Concept 16, dependent on Concept 16k).

[0217] Concept 17l:V HAny one of concepts 10 to 16 (provided that if subject to concept 13, then subject to concept 131) wherein the domain comprises the CDRH3 sequence of SEQ ID NO: 345 or 348, or the CDRH3 sequence of SEQ ID NO: 345 or 348 containing six or fewer amino acid substitutions. , and if dependent on Concept 16, dependent on Concept 16l).

[0218] Concept 18.V H 18. The antibody or fragment of any of concepts 1 to 17, wherein the domain comprises the CDRH1 sequence of SEQ ID NO: 27 or 30, or the CDRH1 sequence of SEQ ID NO: 27 or 30 comprising 3, 2, or 1 amino acid substitutions.

[0219] Concept 18a:V H Any of Concepts 1 to 17 (provided that if subject to Concept 9, it is subject to Concept 9a, and if subject to Concept 13, it is subject to Concept 9b), in which the domain comprises the CDRH1 sequence of SEQ ID NO: 7 or 10, or the CDRH1 sequence of SEQ ID NO: 7 or 10 containing 3, 2, or 1 amino acid substitutions. If it is subject to concept 13a, if it is subject to concept 16, it is subject to concept 16a, and if it is subject to concept 17, it is subject to concept 17a).

[0220] Concept 18b:V H Any of concepts 1 to 17 (provided that if it is subject to concept 9, it is subject to concept 9b, and if it is subject to concept 13), wherein the domain comprises the CDRH1 sequence of SEQ ID NO: 52 or 55, or the CDRH1 sequence of SEQ ID NO: 52 or 55 containing 3, 2, or 1 amino acid substitutions. (If dependent on concept 16, it is dependent on concept 16b; if dependent on concept 17, it is dependent on concept 17b).

[0221] Concept 18c:V HAny of concepts 1 to 17 (provided that if subject to concept 9, then subject to concept 9c, and if subject to concept 13), wherein the domain comprises the CDRH1 sequence of SEQ ID NO: 72 or 75, or the CDRH1 sequence of SEQ ID NO: 72 or 75 containing 3, 2, or 1 amino acid substitutions. (If dependent on concept 13c, then dependent on concept 13; if dependent on concept 16, then dependent on concept 16c; if dependent on concept 17, then dependent on concept 17c).

[0222] Concept 18d:V H Any of concepts 1 to 17 (provided that if it is subject to concept 9, it is subject to concept 9d, and if it is subject to concept 13), wherein the domain comprises the CDRH1 sequence of SEQ ID NO: 92 or 95, or the CDRH1 sequence of SEQ ID NO: 92 or 95 containing 3, 2, or 1 amino acid substitutions. (If dependent on concept 13d, then dependent on concept 13d; if dependent on concept 16, then dependent on concept 16d; if dependent on concept 17, then dependent on concept 17d).

[0223] Concept 18e:V H Any of concepts 1 to 17 (provided that when subject to concept 9, it is subject to concept 9e), wherein the domain comprises the CDRH1 sequence of SEQ ID NO: 112 or 115, or the CDRH1 sequence of SEQ ID NO: 112 or 115 containing 3, 2, or 1 amino acid substitutions; If subject to Concept 13, it is subject to Concept 13e; if subject to Concept 16, it is subject to Concept 16e; if subject to Concept 17, it is subject to Concept 17e) piece.

[0224] Concept 18f:V H Any of concepts 1 to 17 (provided that when subject to concept 9, it is subject to concept 9f), wherein the domain comprises the CDRH1 sequence of SEQ ID NO: 142 or 145, or the CDRH1 sequence of SEQ ID NO: 142 or 145 containing 3, 2, or 1 amino acid substitutions; If subject to Concept 13, it is subject to Concept 13f; if subject to Concept 16, it is subject to Concept 16f; if subject to Concept 17, it is subject to Concept 17f) piece.

[0225] Concept 18g:V H Any of concepts 1 to 17 (provided that when subject to concept 9, it is subject to concept 9g), wherein the domain comprises the CDRH1 sequence of SEQ ID NO: 172 or 175, or the CDRH1 sequence of SEQ ID NO: 172 or 175 containing 3, 2, or 1 amino acid substitutions; If subject to Concept 13, it is subject to Concept 13g; if subject to Concept 16, it is subject to Concept 16g; if subject to Concept 17, it is subject to Concept 17g) piece.

[0226] Concept 18h:V H Any of concepts 1 to 17 (provided that when subject to concept 9, it is subject to concept 9h), wherein the domain comprises the CDRH1 sequence of SEQ ID NO: 132 or 135, or the CDRH1 sequence of SEQ ID NO: 132 or 135 containing 3, 2, or 1 amino acid substitutions; If subject to Concept 13, it is subject to Concept 13h; if subject to Concept 16, it is subject to Concept 16h; if subject to Concept 17, it is subject to Concept 17h) piece.

[0227] Concept 18i:V H Any of concepts 1 to 17 (provided that when subject to concept 9, it is subject to concept 9i), wherein the domain comprises the CDRH1 sequence of SEQ ID NO: 238 or 241, or the CDRH1 sequence of SEQ ID NO: 238 or 241 containing 3, 2, or 1 amino acid substitutions; If subject to Concept 13, it is subject to Concept 13i; if subject to Concept 16, it is subject to Concept 16i; if subject to Concept 17, it is subject to Concept 17i) piece.

[0228] Concept 18j:V HAny of concepts 1 to 17 (provided that when subject to concept 9, it is subject to concept 9j), wherein the domain comprises the CDRH1 sequence of SEQ ID NO: 258 or 261, or the CDRH1 sequence of SEQ ID NO: 258 or 261 containing 3, 2, or 1 amino acid substitutions; If subject to Concept 13, it is subject to Concept 13j; if subject to Concept 16, it is subject to Concept 16j; if subject to Concept 17, it is subject to Concept 17j) piece.

[0229] Concept 18k:V H Any of concepts 1 to 17 (provided that when subject to concept 9, it is subject to concept 9k), wherein the domain comprises the CDRH1 sequence of SEQ ID NO: 278 or 281, or the CDRH1 sequence of SEQ ID NO: 278 or 281 containing 3, 2, or 1 amino acid substitutions; If subject to Concept 13, it is subject to Concept 13k; if subject to Concept 16, it is subject to Concept 16k; if subject to Concept 17, it is subject to Concept 17k) piece.

[0230] Concept 18l:V H Any of concepts 1 to 17 (provided that when subject to concept 9, it is subject to concept 91), wherein the domain comprises the CDRH1 sequence of SEQ ID NO: 343 or 346, or the CDRH1 sequence of SEQ ID NO: 343 or 346 containing 3, 2, or 1 amino acid substitutions; If subject to Concept 13, it is subject to Concept 13l; if subject to Concept 16, it is subject to Concept 16l; if subject to Concept 17, it is subject to Concept 17l) piece.

[0231] Concept 19.V H 19. The antibody or fragment of any of concepts 1 to 18, wherein the domain comprises the CDRH2 sequence of SEQ ID NO: 28 or 31, or the CDRH2 sequence of SEQ ID NO: 28 or 31 with no more than four amino acid substitutions.

[0232] Concept 19a:V HAny of Concepts 1 to 18 (provided that if subject to Concept 9, it is subject to Concept 9a, and if subject to Concept 13, it is subject to Concept 14), in which the domain comprises the CDRH2 sequence of SEQ ID NO: 8 or 11, or the CDRH2 sequence of SEQ ID NO: 8 or 11 containing no more than four amino acid substitutions. if subject to Concept 13a, if subject to Concept 16, subject to Concept 16a, if subject to Concept 17, subject to Concept 17a, and if subject to Concept 18, subject to Concept 18a).

[0233] Concept 19b:V H Any of concepts 1 to 18 (provided that if it is subject to concept 9, it is subject to concept 9b, and if it is subject to concept 13), wherein the domain comprises the CDRH2 sequence of SEQ ID NO: 53 or 56, or the CDRH2 sequence of SEQ ID NO: 53 or 56 containing 3, 2, or 1 amino acid substitutions. (If dependent on Concept 13b, then dependent on Concept 13; if dependent on Concept 16, then dependent on Concept 16b; if dependent on Concept 17, then dependent on Concept 17b; if dependent on Concept 18, then dependent on Concept 18b).

[0234] Concept 19c:V H Any of concepts 1 to 18 (provided that if subject to concept 9, then subject to concept 9c, and if subject to concept 13), wherein the domain comprises the CDRH2 sequence of SEQ ID NO: 73 or 76, or the CDRH2 sequence of SEQ ID NO: 73 or 76 containing 3, 2, or 1 amino acid substitutions. (If dependent on Concept 13c, then dependent on Concept 13; if dependent on Concept 16, then dependent on Concept 16c; if dependent on Concept 17, then dependent on Concept 17c; if dependent on Concept 18, then dependent on Concept 18c).

[0235] Concept 19d:V H Any of concepts 1 to 18 (provided that if it is subject to concept 9, it is subject to concept 9d, and if it is subject to concept 13), wherein the domain comprises the CDRH2 sequence of SEQ ID NO: 93 or 96, or the CDRH2 sequence of SEQ ID NO: 93 or 96 containing 3, 2, or 1 amino acid substitutions. (If dependent on concept 13d, then dependent on concept 13; if dependent on concept 16, then dependent on concept 16d; if dependent on concept 17, then dependent on concept 17d; if dependent on concept 18, then dependent on concept 18d).

[0236] Concept 19e:V H Any of concepts 1 to 18 (provided that when subject to concept 9, it is subject to concept 9e), wherein the domain comprises the CDRH2 sequence of SEQ ID NO: 113 or 116, or the CDRH2 sequence of SEQ ID NO: 113 or 116 containing 3, 2, or 1 amino acid substitutions; If subject to concept 13, it is subject to concept 13e; if subject to concept 16, it is subject to concept 16e; if subject to concept 17, it is subject to concept 17e; and if subject to concept 18, it is subject to concept 18e).

[0237] Concept 19f:V H Any of concepts 1 to 18 (provided that when subject to concept 9, it is subject to concept 9f), wherein the domain comprises the CDRH2 sequence of SEQ ID NO: 143 or 146, or the CDRH2 sequence of SEQ ID NO: 143 or 146 containing 3, 2, or 1 amino acid substitutions; If subject to Concept 13, it is subject to Concept 13f; if subject to Concept 16, it is subject to Concept 16f; if subject to Concept 17, it is subject to Concept 17f; and if subject to Concept 18, it is subject to Concept 18f).

[0238] Concept 19g:V H Any of concepts 1 to 18 (provided that when subject to concept 9, it is subject to concept 9g), wherein the domain comprises the CDRH2 sequence of SEQ ID NO: 173 or 176, or the CDRH2 sequence of SEQ ID NO: 173 or 176 containing 3, 2, or 1 amino acid substitutions; If subject to Concept 13, it is subject to Concept 13g; if subject to Concept 16, it is subject to Concept 16g; if subject to Concept 17, it is subject to Concept 17g; and if subject to Concept 18, it is subject to Concept 18g).

[0239] Concept 19h:V H Any of concepts 1 to 18 (provided that if subject to concept 9, then subject to concept 9h), wherein the domain comprises the CDRH2 sequence of SEQ ID NO: 133 or 136, or the CDRH2 sequence of SEQ ID NO: 133 or 136 containing 3, 2, or 1 amino acid substitutions; If subject to Concept 13, it is subject to Concept 13h; if subject to Concept 16, it is subject to Concept 16h; if subject to Concept 17, it is subject to Concept 17h; and if subject to Concept 18, it is subject to Concept 18h).

[0240] Concept 19i:V H Any of concepts 1 to 18 (provided that if subject to concept 9, then subject to concept 9i), wherein the domain comprises the CDRH2 sequence of SEQ ID NO: 239 or 242, or the CDRH2 sequence of SEQ ID NO: 239 or 242 containing 3, 2, or 1 amino acid substitutions; If subject to Concept 13, it is subject to Concept 13i; if subject to Concept 16, it is subject to Concept 16i; if subject to Concept 17, it is subject to Concept 17i; and if subject to Concept 18, it is subject to Concept 18i).

[0241] Concept 19j:V H Any of concepts 1 to 18 (provided that when subject to concept 9, it is subject to concept 9j), wherein the domain comprises the CDRH2 sequence of SEQ ID NO: 259 or 262, or the CDRH2 sequence of SEQ ID NO: 259 or 262 containing 3, 2, or 1 amino acid substitutions; If subject to Concept 13, it is subject to Concept 13j; if subject to Concept 16, it is subject to Concept 16j; if subject to Concept 17, it is subject to Concept 17j; and if subject to Concept 18, it is subject to Concept 18j).

[0242] Concept 19k:V HAny of concepts 1 to 18 (provided that when subject to concept 9, it is subject to concept 9k), wherein the domain comprises the CDRH2 sequence of SEQ ID NO: 279 or 282, or the CDRH2 sequence of SEQ ID NO: 279 or 282 containing 3, 2, or 1 amino acid substitutions; An antibody or fragment thereof according to any one of the preceding claims (if dependent on Concept 13, then dependent on Concept 13k, if dependent on Concept 16, then dependent on Concept 16k, if dependent on Concept 17, then dependent on Concept 17k, and if dependent on Concept 18, then dependent on Concept 18k).

[0243] Concept 19l:V H Any of concepts 1 to 18 (provided that when subject to concept 9, it is subject to concept 91), wherein the domain comprises the CDRH2 sequence of SEQ ID NO: 344 or 347, or the CDRH2 sequence of SEQ ID NO: 344 or 347 containing 3, 2, or 1 amino acid substitutions; If subject to Concept 13, it is subject to Concept 13l; if subject to Concept 16, it is subject to Concept 16l; if subject to Concept 17, it is subject to Concept 17l; and if subject to Concept 18, it is subject to Concept 18l).

[0244] Concept 20.V H 20. The antibody or fragment of any of Concepts 1 to 19, wherein the domain comprises the amino acid sequence of SEQ ID NO: 33, or a heavy chain variable domain amino acid sequence that is at least 80% (e.g., at least 85% or at least 90%) identical to SEQ ID NO: 33.

[0245] Concept 20a:V H Any of concepts 1 to 19 (provided that when subordinate to concept 9, approximately 1 to 19) in which the domain comprises the amino acid sequence of SEQ ID NO: 13 or a heavy chain variable domain amino acid sequence that is at least 80% (e.g., at least 85% or at least 90%) identical to SEQ ID NO: 13. If subject to concept 13, then subject to concept 13a; if subject to concept 16, then subject to concept 16a; if subject to concept 17, then subject to concept 17a; if subject to concept 18, then subject to concept 18a; and if subject to concept 19, then subject to concept 19a).

[0246] Concept 20b:V H Any of concepts 1 to 19 (provided that when subordinate to concept 9, approximately 1 to 19) wherein the domain comprises the amino acid sequence of SEQ ID NO: 58 or a heavy chain variable domain amino acid sequence that is at least 80% (e.g., at least 85% or at least 90%) identical to SEQ ID NO: 58. If subject to concept 13, then subject to concept 13b; if subject to concept 16, then subject to concept 16b; if subject to concept 17, then subject to concept 17b; if subject to concept 18, then subject to concept 18b; and if subject to concept 19, then subject to concept 19b).

[0247] Concept 20c:V H Any of concepts 1 to 19 (provided that when subordinate to concept 9, approximately 1 to 19) wherein the domain comprises the amino acid sequence of SEQ ID NO: 78 or a heavy chain variable domain amino acid sequence that is at least 80% (e.g., at least 85% or at least 90%) identical to SEQ ID NO: 78. if subject to concept 13, then subject to concept 13c; if subject to concept 16, then subject to concept 16c; if subject to concept 17, then subject to concept 17c; if subject to concept 18, then subject to concept 18c; and if subject to concept 19, then subject to concept 19c).

[0248] Concept 20d:V H Any of concepts 1 to 19 (provided that when subordinate to concept 9, approximately 1 to 19) wherein the domain comprises the amino acid sequence of SEQ ID NO: 98 or a heavy chain variable domain amino acid sequence that is at least 80% (e.g., at least 85% or at least 90%) identical to SEQ ID NO: 98. if subject to concept 13, then subject to concept 13d; if subject to concept 16, then subject to concept 16d; if subject to concept 17, then subject to concept 17d; if subject to concept 18, then subject to concept 18d; and if subject to concept 19, then subject to concept 19d).

[0249] Concept 20e:V H Any of concepts 1 to 19 (provided that, when subordinate to concept 9), wherein the domain comprises the amino acid sequence of SEQ ID NO: 118 or a heavy chain variable domain amino acid sequence that is at least 80% (e.g., at least 85% or at least 90%) identical to SEQ ID NO: 118. if subject to concept 9e, if subject to concept 13, then subject to concept 13e, if subject to concept 16, then subject to concept 16e, if subject to concept 17, then subject to concept 17e, if subject to concept 18, then subject to concept 18e, and if subject to concept 19, then subject to concept 19e).

[0250] Concept 20f:V H Any of concepts 1 to 19 (provided that, when subordinate to concept 9), wherein the domain comprises the amino acid sequence of SEQ ID NO: 158 or a heavy chain variable domain amino acid sequence that is at least 80% (e.g., at least 85% or at least 90%) identical to SEQ ID NO: 158. if subject to concept 9f, if subject to concept 13, then subject to concept 13f, if subject to concept 16, then subject to concept 16f, if subject to concept 17, then subject to concept 17f, if subject to concept 18, then subject to concept 18f, and if subject to concept 19, then subject to concept 19f).

[0251] Concept 20g:V H Any of concepts 1 to 19 (provided that, when subordinate to concept 9), wherein the domain comprises the amino acid sequence of SEQ ID NO: 178 or a heavy chain variable domain amino acid sequence that is at least 80% (e.g., at least 85% or at least 90%) identical to SEQ ID NO: 178. if subject to Concept 9g, if subject to Concept 13, then subject to Concept 13g, if subject to Concept 16, then subject to Concept 16g, if subject to Concept 17, then subject to Concept 17g, if subject to Concept 18, then subject to Concept 18g, and if subject to Concept 19, then subject to Concept 19g).

[0252] Concept 20h:V H The domain has the amino acid sequence of SEQ ID NO: 138 or SEQ ID NO: 138 and Any of concepts 1 to 19 (provided that it is dependent on concept 9) comprising heavy chain variable domain amino acid sequences that are at least 80% (e.g., at least 85% or at least 90%) identical. if subject to concept 9h, if subject to concept 13, then subject to concept 13h, if subject to concept 16, then subject to concept 16h, if subject to concept 17, then subject to concept 17h, if subject to concept 18, then subject to concept 18h, and if subject to concept 19, then subject to concept 19h).

[0253] Concept 20i:V H Any of concepts 1 to 19 (provided that, when subordinate to concept 9), wherein the domain comprises the amino acid sequence of SEQ ID NO: 244 or a heavy chain variable domain amino acid sequence that is at least 80% (e.g., at least 85% or at least 90%) identical to SEQ ID NO: 244. if subject to concept 9i, if subject to concept 13, then subject to concept 13i, if subject to concept 16, then subject to concept 16i, if subject to concept 17, then subject to concept 17i, if subject to concept 18, then subject to concept 18i, and if subject to concept 19, then subject to concept 19i).

[0254] Concept 20j:V H Any of concepts 1 to 19 (provided that, when subordinate to concept 9), wherein the domain comprises the amino acid sequence of SEQ ID NO: 264 or a heavy chain variable domain amino acid sequence that is at least 80% (e.g., at least 85% or at least 90%) identical to SEQ ID NO: 264. if subject to Concept 13, then subject to Concept 13j; if subject to Concept 16, then subject to Concept 16j; if subject to Concept 17, then subject to Concept 17j; if subject to Concept 18, then subject to Concept 18j; and if subject to Concept 19, then subject to Concept 19j).

[0255] Concept 20k:V H Any of concepts 1 to 19 (provided that, when subordinate to concept 9), wherein the domain comprises the amino acid sequence of SEQ ID NO: 284 or a heavy chain variable domain amino acid sequence that is at least 80% (e.g., at least 85% or at least 90%) identical to SEQ ID NO: 284. (if dependent on concept 9k, if dependent on concept 13, then dependent on concept 13k, if dependent on concept 16, then dependent on concept 16k, if dependent on concept 17, then dependent on concept 17k, if dependent on concept 18, then dependent on concept 18k, and if dependent on concept 19, then dependent on concept 19k).

[0256] Concept 20l:V H Any of concepts 1 to 19 (provided that, when subordinate to concept 9), wherein the domain comprises the amino acid sequence of SEQ ID NO: 349 or a heavy chain variable domain amino acid sequence that is at least 80% (e.g., at least 85% or at least 90%) identical to SEQ ID NO: 349. if subject to Concept 9L, if subject to Concept 13, then subject to Concept 13L, if subject to Concept 16, then subject to Concept 16L, if subject to Concept 17, then subject to Concept 17L, if subject to Concept 18, then subject to Concept 18L, and if subject to Concept 19, then subject to Concept 19L).

[0257] In one embodiment, the amino acid sequence is at least 70% identical to the designated sequence. In one embodiment, the amino acid sequence is at least 75% identical to the designated sequence. In one embodiment, the amino acid sequence is at least 95% identical to the designated sequence. In one embodiment, the amino acid sequence is at least 96% identical to the designated sequence. In one embodiment, the amino acid sequence is at least 97% identical to the designated sequence. In one embodiment, the amino acid sequence is at least 98% identical to the designated sequence. In one embodiment, the amino acid sequence is at least 99% identical to the designated sequence. In one embodiment, the amino acid sequence is at least 99.5% identical to the designated sequence.

[0258] Concept 21. V H 21. The antibody or fragment of any of concepts 1 to 20, comprising a first and a second copy of the domain.

[0259] Concept 22. A V comprising a CDRL1 sequence of SEQ ID NO: 37 or 40, or a CDRL1 sequence of SEQ ID NO: 37 or 40 containing no more than three amino acid substitutions. L 22. The antibody or fragment of any one of concepts 1 to 21, comprising a domain.

[0260] Concept 22a: V comprising the CDRL1 sequence of SEQ ID NO: 17 or 20, or the CDRL1 sequence of SEQ ID NO: 17 or 20 containing no more than three amino acid substitutions L Any of concepts 1 to 21, including the domain (however, if it is subordinate to concept 9, it is subordinate to concept 9a, and if it is subordinate to concept 13, it is subordinate to concept 1). (wherein the antibody or fragment thereof is subject to Concept 13a if subject to Concept 16, subject to Concept 16a if subject to Concept 17, subject to Concept 17a if subject to Concept 18, subject to Concept 18a if subject to Concept 19, subject to Concept 19a if subject to Concept 20, and subject to Concept 20a if subject to Concept 20).

[0261] Concept 22b: V comprising the CDRL1 sequence of SEQ ID NO: 62 or 65, or the CDRL1 sequence of SEQ ID NO: 62 or 65 containing no more than three amino acid substitutions LAny of concepts 1 to 21, including the domain (however, if it is subordinate to concept 9, it is subordinate to concept 9b, and if it is subordinate to concept 13, it is subordinate to concept 1). (wherein the antibody or fragment thereof is subject to Concept 13b if subject to Concept 16, subject to Concept 16b if subject to Concept 17, subject to Concept 17b if subject to Concept 18, subject to Concept 18b if subject to Concept 19, subject to Concept 19b if subject to Concept 20, and subject to Concept 20b if subject to Concept 20).

[0262] Concept 22c: V comprising the CDRL1 sequence of SEQ ID NO: 82 or 85, or the CDRL1 sequence of SEQ ID NO: 82 or 85 containing two or one amino acid substitutions L Any of concepts 1 to 21, including the domain (however, if it is subordinate to concept 9, it is subordinate to concept 9c, and concept 13 (wherein, if dependent on concept 13c, then dependent on concept 13; where dependent on concept 16, then dependent on concept 16c; where dependent on concept 17, then dependent on concept 17c; where dependent on concept 18, then dependent on concept 18c; where dependent on concept 19, then dependent on concept 19c; and where dependent on concept 20, then dependent on concept 20c).

[0263] Concept 22d: V comprising the CDRL1 sequence of SEQ ID NO: 102 or 105, or the CDRL1 sequence of SEQ ID NO: 102 or 105 containing 5 or fewer amino acid substitutions L Any of concepts 1 to 21, including the domain (however, if it is subordinate to concept 9, it is subordinate to concept 9d, and concept 1 3, then subject to concept 13d; if subject to concept 16, then subject to concept 16d; if subject to concept 17, then subject to concept 17d; if subject to concept 18, then subject to concept 18d; if subject to concept 19, then subject to concept 19d; and if subject to concept 20, then subject to concept 20d).

[0264] Concept 22e: V comprising the CDRL1 sequence of SEQ ID NO: 122 or 125, or the CDRL1 sequence of SEQ ID NO: 122 or 125 containing two or one amino acid substitutions LAny of concepts 1 to 21, including the domain (however, if it is subordinate to concept 9, it is subordinate to concept 9e, If subject to concept 13, it is subject to concept 13e; if subject to concept 16, it is subject to concept 16e; if subject to concept 17, it is subject to concept 17e; if subject to concept 18, it is subject to concept 18e; if subject to concept 19, it is subject to concept 19e; and if subject to concept 20, it is subject to concept 20e).

[0265] Concept 22f: V comprising the CDRL1 sequence of SEQ ID NO: 162 or 165, or the CDRL1 sequence of SEQ ID NO: 162 or 165 containing 5 or fewer amino acid substitutions L Any of concepts 1 to 21, including the domain (however, if it is subordinate to concept 9, it is subordinate to concept 9f, and concept 1 If it is subordinate to concept 3, it is subordinate to concept 13f; if it is subordinate to concept 16, it is subordinate to concept 16f; if it is subordinate to concept 17, it is subordinate to concept 17f; if it is subordinate to concept 18, it is subordinate to concept 19f; If subject to concept 18f, if subject to concept 19, then subject to concept 19f, and if subject to concept 20, then subject to concept 20f).

[0266] Concept 22g: A V comprising the CDRL1 sequence of SEQ ID NO: 182 or 185, or the CDRL1 sequence of SEQ ID NO: 182 or 185 containing 5 or fewer amino acid substitutions. L Any of concepts 1 to 21, including the domain (however, if it is subordinate to concept 9, it is subordinate to concept 9g, and concept 1 3, then subject to concept 13g; if subject to concept 16, then subject to concept 16g; if subject to concept 17, then subject to concept 17g; if subject to concept 18, then subject to concept 18g; if subject to concept 19, then subject to concept 19g; and if subject to concept 20, then subject to concept 20g).

[0267] Concept 22h:V LAny of concepts 1 to 21 (provided that if it is dependent on concept 9, it is dependent on concept 9h, and concept 1 is dependent on concept 9h), wherein the domain comprises the CDRL1 sequence of SEQ ID NO: 142 or 145, or the CDRL1 sequence of SEQ ID NO: 142 or 145 containing two or one amino acid substitutions. 3, then subject to concept 13h; if subject to concept 16, then subject to concept 16h; if subject to concept 17, then subject to concept 17h; if subject to concept 18, then subject to concept 18h; if subject to concept 19, then subject to concept 19h; and if subject to concept 20, then subject to concept 20h).

[0268] Concept 22i: V comprising the CDRL1 sequence of SEQ ID NO: 248 or 251, or the CDRL1 sequence of SEQ ID NO: 248 or 251 containing two or one amino acid substitutions L Any of concepts 1 to 21, including the domain (however, if it is subordinate to concept 9, it is subordinate to concept 9i, If subject to concept 13, it is subject to concept 13i; if subject to concept 16, it is subject to concept 16i; if subject to concept 17, it is subject to concept 17i; if subject to concept 18, it is subject to concept 18i; if subject to concept 19, it is subject to concept 19i; and if subject to concept 20, it is subject to concept 20i).

[0269] Concept 22j: A V comprising the CDRL1 sequence of SEQ ID NO: 268 or 271, or the CDRL1 sequence of SEQ ID NO: 268 or 271 containing two or one amino acid substitutions L Any of concepts 1 to 21, including the domain (however, if it is subordinate to concept 9, it is subordinate to concept 9j, If subject to Concept 13, it is subject to Concept 13j; if subject to Concept 16, it is subject to Concept 16j; if subject to Concept 17, it is subject to Concept 17j; if subject to Concept 18, it is subject to Concept 18j; if subject to Concept 19, it is subject to Concept 19j; and if subject to Concept 20, it is subject to Concept 20j).

[0270] Concept 22k: A V comprising the CDRL1 sequence of SEQ ID NO: 288 or 291, or the CDRL1 sequence of SEQ ID NO: 288 or 291 containing two or one amino acid substitutions L Any of concepts 1 to 21, including the domain (however, if it is subordinate to concept 9, it is subordinate to concept 9k, An antibody or fragment thereof according to the invention (where it is dependent on Concept 13, it is dependent on Concept 13k; where it is dependent on Concept 16, it is dependent on Concept 16k; where it is dependent on Concept 17, it is dependent on Concept 17k; where it is dependent on Concept 18, it is dependent on Concept 18k; where it is dependent on Concept 19, it is dependent on Concept 19k; and where it is dependent on Concept 20, it is dependent on Concept 20k).

[0271] Concept 221: A V comprising the CDRL1 sequence of SEQ ID NO: 353 or 356, or the CDRL1 sequence of SEQ ID NO: 353 or 356 containing two or one amino acid substitutions L Any of concepts 1 to 21, including the domain (however, if it is subordinate to concept 9, it is subordinate to concept 9l, (If subject to Concept 13, it is subject to Concept 13l; if subject to Concept 16, it is subject to Concept 16l; if subject to Concept 17, it is subject to Concept 17l; if subject to Concept 18, it is subject to Concept 18l; if subject to Concept 19, it is subject to Concept 19l; and if subject to Concept 20, it is subject to Concept 20l).

[0272] Concept 23. A V comprising a CDRL2 sequence of SEQ ID NO: 38 or 41, or a CDRL2 sequence of SEQ ID NO: 38 or 41 containing two or one amino acid substitutions, such as a CDRL2 sequence of SEQ ID NO: 50. L Domain or V L 23. The antibody or fragment of any one of concepts 1 to 22, comprising a domain.

[0273] Concept 23a: V comprising the CDRL2 sequence of SEQ ID NO: 18 or 21, or the CDRL2 sequence of SEQ ID NO: 18 or 21 containing two or one amino acid substitutions L Domain or V L Any of concepts 1 to 22, including the domain (however, if it is subordinate to concept 9, concept 9 a, if dependent on concept 13, then dependent on concept 13a, if dependent on concept 16, then dependent on concept 16a, if dependent on concept 17, then dependent on concept 17a, if dependent on concept 18, then dependent on concept 18a, if dependent on concept 19, then dependent on concept 19a, if dependent on concept 20, then dependent on concept 20a, and if dependent on concept 22, then dependent on concept 22a).

[0274] Concept 23b: V comprising the CDRL2 sequence of SEQ ID NO: 63 or 66, or the CDRL2 sequence of SEQ ID NO: 63 or 66 containing one amino acid substitution L Domain or V L Any of concepts 1 to 22, including the domain (however, if it is subordinate to concept 9, it is subordinate to concept 9b) , if subject to concept 13, then subject to concept 13b; if subject to concept 16, then subject to concept 16b; if subject to concept 17, then subject to concept 17b; if subject to concept 18, then subject to concept 18b; if subject to concept 19, then subject to concept 19b; if subject to concept 20, then subject to concept 20b; and if subject to concept 22, then subject to concept 22b).

[0275] Concept 23c: A V comprising the CDRL2 sequence of SEQ ID NO: 83 or 86, or the CDRL2 sequence of SEQ ID NO: 83 or 86 containing one amino acid substitution L Domain or V L Any of concepts 1 to 22, including the domain (however, if it is subordinate to concept 9, it is subordinate to concept 9c) , if subject to concept 13, then subject to concept 13c; if subject to concept 16, then subject to concept 16c; if subject to concept 17, then subject to concept 17c; if subject to concept 18, then subject to concept 18c; if subject to concept 19, then subject to concept 19c; if subject to concept 20, then subject to concept 20c; and if subject to concept 22, then subject to concept 22c).

[0276] Concept 23d: A V comprising the CDRL2 sequence of SEQ ID NO: 103 or 106, or the CDRL2 sequence of SEQ ID NO: 103 or 106 containing one amino acid substitution L Domain or V L Any of concepts 1 to 22, including the domain (however, if subordinate to concept 9, concept 9d) if subject to concept 13, then subject to concept 13d; if subject to concept 16, then subject to concept 16d; if subject to concept 17, then subject to concept 17d; if subject to concept 18, then subject to concept 18d; if subject to concept 19, then subject to concept 19d; if subject to concept 20, then subject to concept 20d; if subject to concept 22, then subject to concept 22d).

[0277] Concept 23e: A V comprising the CDRL2 sequence of SEQ ID NO: 123 or 126, or the CDRL2 sequence of SEQ ID NO: 123 or 126 containing one amino acid substitution. L Domain or V L Any of concepts 1 to 22, including the domain (however, if subordinate to concept 9, concept 9e) if subject to concept 13, then subject to concept 13e; if subject to concept 16, then subject to concept 16e; if subject to concept 17, then subject to concept 17e; if subject to concept 18, then subject to concept 18e; if subject to concept 19, then subject to concept 19e; if subject to concept 20, then subject to concept 20e; if subject to concept 22, then subject to concept 22e).

[0278] Concept 23f: A V comprising the CDRL2 sequence of SEQ ID NO: 153 or 156, or the CDRL2 sequence of SEQ ID NO: 153 or 156 containing one amino acid substitution L Domain or V L Any of concepts 1 to 22, including the domain (however, if subordinate to concept 9, concept 9f) (wherein the antibody or fragment thereof is dependent on concept 13, then dependent on concept 13f, where dependent on concept 16, then dependent on concept 16f, where dependent on concept 17, then dependent on concept 17f, where dependent on concept 18, then dependent on concept 18f, where dependent on concept 19, then dependent on concept 19f, where dependent on concept 20, then dependent on concept 20f, and where dependent on concept 22, then dependent on concept 22f).

[0279] Concept 23g: A V comprising the CDRL2 sequence of SEQ ID NO: 183 or 186, or the CDRL2 sequence of SEQ ID NO: 183 or 186 containing one amino acid substitution. L Domain or V L Any of concepts 1 to 22, including the domain (however, if subordinate to concept 9, concept 9g) if subject to concept 13, then subject to concept 13g; if subject to concept 16, then subject to concept 16g; if subject to concept 17, then subject to concept 17g; if subject to concept 18, then subject to concept 18g; if subject to concept 19, then subject to concept 19g; if subject to concept 20, then subject to concept 20g; if subject to concept 22, then subject to concept 22g).

[0280] Concept 23h: A V comprising the CDRL2 sequence of SEQ ID NO: 143 or 146, or the CDRL2 sequence of SEQ ID NO: 143 or 146 containing one amino acid substitution L Domain or V L Any of concepts 1 to 22, including the domain (however, if subordinate to concept 9, concept 9h) if subject to concept 13, then subject to concept 13h; if subject to concept 16, then subject to concept 16h; if subject to concept 17, then subject to concept 17h; if subject to concept 18, then subject to concept 18h; if subject to concept 19, then subject to concept 19h; if subject to concept 20, then subject to concept 20h; and if subject to concept 22, then subject to concept 22h).

[0281] Concept 23i: A V comprising the CDRL2 sequence of SEQ ID NO: 249 or 252, or the CDRL2 sequence of SEQ ID NO: 249 or 252 containing one amino acid substitution L Domain or V L Any of Concepts 1 to 22, including the domain (however, if subordinate to Concept 9, Concept 9i) if subject to concept 13, then subject to concept 13i; if subject to concept 16, then subject to concept 16i; if subject to concept 17, then subject to concept 17i; if subject to concept 18, then subject to concept 18i; if subject to concept 19, then subject to concept 19i; if subject to concept 20, then subject to concept 20i; if subject to concept 22, then subject to concept 22i).

[0282] Concept 23j: A V comprising the CDRL2 sequence of SEQ ID NO: 269 or 272, or the CDRL2 sequence of SEQ ID NO: 269 or 272 containing one amino acid substitution L Domain or V L Any of concepts 1 to 22, including the domain (however, if subordinate to concept 9, concept 9j) if subject to concept 13, then subject to concept 13j; if subject to concept 16, then subject to concept 16j; if subject to concept 17, then subject to concept 17j; if subject to concept 18, then subject to concept 18j; if subject to concept 19, then subject to concept 19j; if subject to concept 20, then subject to concept 20j; if subject to concept 22, then subject to concept 22j).

[0283] Concept 23k: A V comprising the CDRL2 sequence of SEQ ID NO: 289 or 292, or the CDRL2 sequence of SEQ ID NO: 289 or 292 containing one amino acid substitution. L Domain or V L Any of concepts 1 to 22, including the domain (however, if subordinate to concept 9, concept 9k) If it is subordinate to concept 13, it is subordinate to concept 13k, and if it is subordinate to concept 16, it is subordinate to concept 17. (if dependent on concept 16k, if dependent on concept 17, then dependent on concept 17k, if dependent on concept 18, then dependent on concept 18k, if dependent on concept 19, then dependent on concept 19k, if dependent on concept 20, then dependent on concept 20k, and if dependent on concept 22, then dependent on concept 22k).

[0284] Concept 231: A V comprising the CDRL2 sequence of SEQ ID NO: 354 or 357, or the CDRL2 sequence of SEQ ID NO: 354 or 357 containing one amino acid substitution. L Domain or V L Any of concepts 1 to 22, including the domain (however, if subordinate to concept 9, concept 91) if subject to concept 13, then subject to concept 13l; if subject to concept 16, then subject to concept 16l; if subject to concept 17, then subject to concept 17l; if subject to concept 18, then subject to concept 18l; if subject to concept 19, then subject to concept 19l; if subject to concept 20, then subject to concept 20l; if subject to concept 22, then subject to concept 22l).

[0285] Concept 24. A V comprising a CDRL3 sequence of SEQ ID NO: 39 or 42, or a CDRL3 sequence of SEQ ID NO: 39 or 42 containing no more than four amino acid substitutions. L Domain or V L 24. The antibody or fragment of any one of concepts 1 to 23, comprising a domain.

[0286] Concept 24a: A V comprising a CDRL3 sequence of SEQ ID NO: 19 or 22, or a CDRL3 sequence of SEQ ID NO: 19 or 22 containing no more than four amino acid substitutions. L Domain or V L Any of concepts 1 to 23, including the domain (however, if subordinate to concept 9, it is subordinate to concept 9a) and if subject to concept 13, then subject to concept 13a; if subject to concept 16, then subject to concept 16a; if subject to concept 17, then subject to concept 17a; if subject to concept 18, then subject to concept 18a; if subject to concept 19, then subject to concept 19a; if subject to concept 20, then subject to concept 20a; if subject to concept 22, then subject to concept 22a; and if subject to concept 23, then subject to concept 23a). .

[0287] Concept 24b: A V comprising a CDRL3 sequence of SEQ ID NO: 64 or 67, or a CDRL3 sequence of SEQ ID NO: 64 or 67 containing no more than four amino acid substitutions. L Domain or V L Any of concepts 1 to 23, including the domain (however, if subordinate to concept 9, it shall be subordinate to concept 9b) and if subject to concept 13, then subject to concept 13b; if subject to concept 16, then subject to concept 16b; if subject to concept 17, then subject to concept 17b; if subject to concept 18, then subject to concept 18b; if subject to concept 19, then subject to concept 19b; if subject to concept 20, then subject to concept 20b; if subject to concept 22, then subject to concept 22b; and if subject to concept 23, then subject to concept 23b). .

[0288] Concept 24c: A V comprising a CDRL3 sequence of SEQ ID NO: 84 or 87, or a CDRL3 sequence of SEQ ID NO: 84 or 87 containing no more than four amino acid substitutions. L Domain or V L Any of concepts 1 to 23, including the domain (however, if subordinate to concept 9, it shall be subordinate to concept 9c) and if subject to concept 13, then subject to concept 13c; if subject to concept 16, then subject to concept 16c; if subject to concept 17, then subject to concept 17c; if subject to concept 18, then subject to concept 18c; if subject to concept 19, then subject to concept 19c; if subject to concept 20, then subject to concept 20c; if subject to concept 22, then subject to concept 22c; and if subject to concept 23, then subject to concept 23c). .

[0289] Concept 24d: A V comprising a CDRL3 sequence of SEQ ID NO: 104 or 107, or a CDRL3 sequence of SEQ ID NO: 104 or 107 containing no more than four amino acid substitutions. L Domain or Applicable V L Any of concepts 1 to 23, including the domain (however, if it is subordinate to concept 9, concept if subject to concept 13, then subject to concept 13d; if subject to concept 16, then subject to concept 16d; if subject to concept 17, then subject to concept 17d; if subject to concept 18, then subject to concept 18d; if subject to concept 19, then subject to concept 19d; if subject to concept 20, then subject to concept 20d; if subject to concept 22, then subject to concept 22d; if subject to concept 23, then subject to concept 23d) The fragment.

[0290] Concept 24e: A V comprising the CDRL3 sequence of SEQ ID NO: 124 or 127, or the CDRL3 sequence of SEQ ID NO: 124 or 127 containing no more than four amino acid substitutions. L Domain or V L Any of concepts 1 to 23, including the domain (however, if it is subordinate to concept 9, concept if subject to concept 13, then subject to concept 13e; if subject to concept 16, then subject to concept 16e; if subject to concept 17, then subject to concept 17e; if subject to concept 18, then subject to concept 18e; if subject to concept 19, then subject to concept 19e; if subject to concept 20, then subject to concept 20e; if subject to concept 22, then subject to concept 22e; if subject to concept 23, then subject to concept 23e The fragment.

[0291] Concept 24f: A V comprising a CDRL3 sequence of SEQ ID NO: 164 or 167, or a CDRL3 sequence of SEQ ID NO: 164 or 167 containing no more than four amino acid substitutions. L Domain or V L Any of concepts 1 to 23, including the domain (however, if it is subordinate to concept 9, concept if subject to concept 13, then subject to concept 13f; if subject to concept 16, then subject to concept 16f; if subject to concept 17, then subject to concept 17f; if subject to concept 18, then subject to concept 18f; if subject to concept 19, then subject to concept 19f; if subject to concept 20, then subject to concept 20f; if subject to concept 22, then subject to concept 22f; if subject to concept 23, then subject to concept 23f The fragment.

[0292] Concept 24g: A V comprising the CDRL3 sequence of SEQ ID NO: 184 or 187, or the CDRL3 sequence of SEQ ID NO: 184 or 187 containing no more than four amino acid substitutions. L Domain or V L Any of concepts 1 to 23, including the domain (however, if it is subordinate to concept 9, concept if subject to concept 13, then subject to concept 13g; if subject to concept 16, then subject to concept 16g; if subject to concept 17, then subject to concept 17g; if subject to concept 18, then subject to concept 18g; if subject to concept 19, then subject to concept 19g; if subject to concept 20, then subject to concept 20g; if subject to concept 22, then subject to concept 22g; if subject to concept 23, then subject to concept 23g); The fragment.

[0293] Concept 24h: A ​​V comprising the CDRL3 sequence of SEQ ID NO: 144 or 147, or the CDRL3 sequence of SEQ ID NO: 144 or 147 containing no more than four amino acid substitutions. L Domain or V L Any of concepts 1 to 23, including the domain (however, if it is subordinate to concept 9, concept if subject to concept 13, then subject to concept 13h; if subject to concept 16, then subject to concept 16h; if subject to concept 17, then subject to concept 17h; if subject to concept 18, then subject to concept 18h; if subject to concept 19, then subject to concept 19h; if subject to concept 20, then subject to concept 20h; if subject to concept 22, then subject to concept 22h; if subject to concept 23, then subject to concept 23h The fragment.

[0294] Concept 24i: A V comprising a CDRL3 sequence of SEQ ID NO: 250 or 253, or a CDRL3 sequence of SEQ ID NO: 250 or 253 containing no more than four amino acid substitutions. L Domain or Applicable V L Any of concepts 1 to 23, including the domain (however, if it is subordinate to concept 9, concept if subject to concept 13, then subject to concept 13i; if subject to concept 16, then subject to concept 16i; if subject to concept 17, then subject to concept 17i; if subject to concept 18, then subject to concept 18i; if subject to concept 19, then subject to concept 19i; if subject to concept 20, then subject to concept 20i; if subject to concept 22, then subject to concept 22i; if subject to concept 23, then subject to concept 23i) The fragment.

[0295] Concept 24j: A V comprising a CDRL3 sequence of SEQ ID NO: 270 or 273, or a CDRL3 sequence of SEQ ID NO: 270 or 273 containing no more than four amino acid substitutions. L Domain or V L Any of concepts 1 to 23, including the domain (however, if it is subordinate to concept 9, concept if subject to concept 13, then subject to concept 13j; if subject to concept 16, then subject to concept 16j; if subject to concept 17, then subject to concept 17j; if subject to concept 18, then subject to concept 18j; if subject to concept 19, then subject to concept 19j; if subject to concept 20, then subject to concept 20j; if subject to concept 22, then subject to concept 22j; if subject to concept 23, then subject to concept 23j The fragment.

[0296] Concept 24k: A V comprising the CDRL3 sequence of SEQ ID NO: 290 or 293, or the CDRL3 sequence of SEQ ID NO: 290 or 293 containing no more than four amino acid substitutions. L Domain or V L Any of concepts 1 to 23, including the domain (however, if it is subordinate to concept 9, concept if subject to concept 13, then subject to concept 13k; if subject to concept 16, then subject to concept 16k; if subject to concept 17, then subject to concept 17k; if subject to concept 18, then subject to concept 18k; if subject to concept 19, then subject to concept 19k; if subject to concept 20, then subject to concept 20k; if subject to concept 22, then subject to concept 22k; if subject to concept 23, then subject to concept 23k The fragment.

[0297] Concept 241: A V comprising the CDRL3 sequence of SEQ ID NO: 355 or 358, or the CDRL3 sequence of SEQ ID NO: 355 or 358 containing no more than four amino acid substitutions. L Domain or V L Any of concepts 1 to 23, including the domain (however, if it is subordinate to concept 9, concept if subject to concept 13, then subject to concept 13l; if subject to concept 16, then subject to concept 16l; if subject to concept 17, then subject to concept 17l; if subject to concept 18, then subject to concept 18l; if subject to concept 19, then subject to concept 19l; if subject to concept 20, then subject to concept 20l; if subject to concept 22, then subject to concept 22l; if subject to concept 23, then subject to concept 23l The fragment.

[0298] Concept 25. The amino acid sequence of SEQ ID NO: 43, or a sequence at least 80% identical to SEQ ID NO: 43 (e.g., and VL domain sequences in the light chain sequences of SEQ ID NOs: 50, 51, or 298. L Domain or V L 25. The antibody or fragment of any one of concepts 1 to 24, comprising a domain.

[0299] Concept 25a:V LAny of concepts 1 to 24 (provided that when subordinate to concept 9, approximately 25%), wherein the domain comprises the amino acid sequence of SEQ ID NO: 23 or a light chain variable domain amino acid sequence that is at least 80% (e.g., at least 85% or at least 90%) identical to SEQ ID NO: 23. If it is subordinate to concept 9a, then it is subordinate to concept 13a; if it is subordinate to concept 16, then it is subordinate to concept 16a; if it is subordinate to concept 17, then it is subordinate to concept 17a; if it is subordinate to concept 18, then it is subordinate to concept 18a; if it is subordinate to concept 19, then it is subordinate to concept 19a; if it is subordinate to concept 20, then it is subordinate to concept 20a; if it is subordinate to concept 22, then it is subordinate to concept 23a; An antibody or fragment thereof according to claim 22a, if dependent on concept 23 then dependent on concept 23a, and if dependent on concept 24 then dependent on concept 24a).

[0300] Concept 25b:V L Any of concepts 1 to 24 (provided that when subordinate to concept 9, approximately 1 to 24) wherein the domain comprises the amino acid sequence of SEQ ID NO: 68 or a light chain variable domain amino acid sequence that is at least 80% (e.g., at least 85% or at least 90%) identical to SEQ ID NO: 68. if subject to concept 13, then subject to concept 13b; if subject to concept 16, then subject to concept 16b; if subject to concept 17, then subject to concept 17b; if subject to concept 18, then subject to concept 18b; if subject to concept 19, then subject to concept 19b; if subject to concept 20, then subject to concept 20b; if subject to concept 22, then subject to concept 22a; if subject to concept 23, then subject to concept 23b; if subject to concept 24, then subject to concept 24b).

[0301] Concept 25c:V L Any of concepts 1 to 24 (provided that when subordinate to concept 9, approximately 1 to 24) wherein the domain comprises the amino acid sequence of SEQ ID NO: 88 or a light chain variable domain amino acid sequence that is at least 80% (e.g., at least 85% or at least 90%) identical to SEQ ID NO: 88. if subject to concept 13, then subject to concept 13c; if subject to concept 16, then subject to concept 16c; if subject to concept 17, then subject to concept 17c; if subject to concept 18, then subject to concept 18c; if subject to concept 19, then subject to concept 19c; if subject to concept 20, then subject to concept 20c; if subject to concept 22, then subject to concept 22c; if subject to concept 23, then subject to concept 23c; if subject to concept 24, then subject to concept 24c).

[0302] Concept 25d:V L Any of concepts 1 to 24 (provided that, when subject to concept 9), wherein the domain comprises the amino acid sequence of SEQ ID NO: 108 or a light chain variable domain amino acid sequence that is at least 80% (e.g., at least 85% or at least 90%) identical to SEQ ID NO: 108. (if subject to concept 9d, if subject to concept 13, then subject to concept 13d, if subject to concept 16, then subject to concept 16d, if subject to concept 17, then subject to concept 17d, if subject to concept 18, then subject to concept 18d, if subject to concept 19, then subject to concept 19d, if subject to concept 20, then subject to concept 20d, if subject to concept 22, then subject to concept 22d, if subject to concept 23, then subject to concept 23d, and if subject to concept 24, then subject to concept 24d).

[0303] Concept 25e:V L Any of concepts 1 to 24 (provided that, when subject to concept 9), wherein the domain comprises the amino acid sequence of SEQ ID NO: 128 or a light chain variable domain amino acid sequence that is at least 80% (e.g., at least 85% or at least 90%) identical to SEQ ID NO: 128. (if dependent on concept 9e, then dependent on concept 13, then dependent on concept 13e, if dependent on concept 16, then dependent on concept 16e, if dependent on concept 17, then dependent on concept 17e, if dependent on concept 18, then dependent on concept 18e, if dependent on concept 19, then dependent on concept 19e, if dependent on concept 20, then dependent on concept 20e, if dependent on concept 22, then dependent on concept 22e, if dependent on concept 23, then dependent on concept 23e, and if dependent on concept 24, then dependent on concept 24e).

[0304] Concept 25f:V L Any of concepts 1 to 24 (provided that, when subject to concept 9), wherein the domain comprises the amino acid sequence of SEQ ID NO: 168 or a light chain variable domain amino acid sequence that is at least 80% (e.g., at least 85% or at least 90%) identical to SEQ ID NO: 168. is subordinate to concept 9f; if it is subordinate to concept 13, it is subordinate to concept 13f; if it is subordinate to concept 16, it is subordinate to concept 16f; if it is subordinate to concept 17, it is subordinate to concept 17f; if it is subordinate to concept 18, it is subordinate to concept 18f; if it is subordinate to concept 19, it is subordinate to concept 19f; if it is subordinate to concept 20, it is subordinate to concept 20f; if it is subordinate to concept 22, it is subordinate to concept 23f; If it is dependent on Concept 22f, if it is dependent on Concept 23, it is dependent on Concept 23f, and if it is dependent on Concept 24, it is dependent on Concept 24f).

[0305] Concept 25g:V L Any of concepts 1 to 24 (provided that, when subject to concept 9), wherein the domain comprises the amino acid sequence of SEQ ID NO: 188 or a light chain variable domain amino acid sequence that is at least 80% (e.g., at least 85% or at least 90%) identical to SEQ ID NO: 188. (if subject to Concept 9g, if subject to Concept 13, then subject to Concept 13g, if subject to Concept 16, then subject to Concept 16g, if subject to Concept 17, then subject to Concept 17g, if subject to Concept 18, then subject to Concept 18g, if subject to Concept 19, then subject to Concept 19g, if subject to Concept 20, then subject to Concept 20g, if subject to Concept 22, then subject to Concept 22g, if subject to Concept 23, then subject to Concept 23g, and if subject to Concept 24, then subject to Concept 24g).

[0306] Concept 25h:V L Any of concepts 1 to 24 (provided that, when subject to concept 9), wherein the domain comprises the amino acid sequence of SEQ ID NO: 148, or a light chain variable domain amino acid sequence that is at least 80% (e.g., at least 85% or at least 90%) identical to SEQ ID NO: 148. (if subject to Concept 9h, if subject to Concept 13, then subject to Concept 13h, if subject to Concept 16, then subject to Concept 16h, if subject to Concept 17, then subject to Concept 17h, if subject to Concept 18, then subject to Concept 18h, if subject to Concept 19, then subject to Concept 19h, if subject to Concept 20, then subject to Concept 20h, if subject to Concept 22, then subject to Concept 22h, if subject to Concept 23, then subject to Concept 23h, and if subject to Concept 24, then subject to Concept 24h).

[0307] Concept 25i:V L Any of concepts 1 to 24 (provided that, when subject to concept 9), wherein the domain comprises the amino acid sequence of SEQ ID NO: 254 or a light chain variable domain amino acid sequence that is at least 80% (e.g., at least 85% or at least 90%) identical to SEQ ID NO: 254. (if dependent on concept 9i, then dependent on concept 13, then dependent on concept 13i, if dependent on concept 16, then dependent on concept 16i, if dependent on concept 17, then dependent on concept 17i, if dependent on concept 18, then dependent on concept 18i, if dependent on concept 19, then dependent on concept 19i, if dependent on concept 20, then dependent on concept 20i, if dependent on concept 22, then dependent on concept 22i, if dependent on concept 23, then dependent on concept 23i, and if dependent on concept 24, then dependent on concept 24i).

[0308] Concept 25j:V L Any of concepts 1 to 24 (provided that, when subject to concept 9), wherein the domain comprises the amino acid sequence of SEQ ID NO: 274 or a light chain variable domain amino acid sequence that is at least 80% (e.g., at least 85% or at least 90%) identical to SEQ ID NO: 274. (if subject to Concept 9j, if subject to Concept 13, then subject to Concept 13j, if subject to Concept 16, then subject to Concept 16j, if subject to Concept 17, then subject to Concept 17j, if subject to Concept 18, then subject to Concept 18j, if subject to Concept 19, then subject to Concept 19j, if subject to Concept 20, then subject to Concept 20j, if subject to Concept 22, then subject to Concept 22j, if subject to Concept 23, then subject to Concept 23j, and if subject to Concept 24, then subject to Concept 24j).

[0309] Concept 25k:V L Any of concepts 1 to 24 (provided that, when subject to concept 9), wherein the domain comprises the amino acid sequence of SEQ ID NO: 294 or a light chain variable domain amino acid sequence that is at least 80% (e.g., at least 85% or at least 90%) identical to SEQ ID NO: 294. is subordinate to concept 9k, if subordinate to concept 13, it is subordinate to concept 13k, if subordinate to concept 16, it is subordinate to concept 16k, if subordinate to concept 17, it is subordinate to concept 17k, if subordinate to concept 18, it is subordinate to concept 18k, if subordinate to concept 19, it is subordinate to concept 19k, if subordinate to concept 20, it is subordinate to concept 20k, and if subordinate to concept 22, it is subordinate to concept 17k. If dependent on concept 22k, if dependent on concept 23, then dependent on concept 23k, and if dependent on concept 24, then dependent on concept 24k). The amino acid sequence is at least 70% identical to the designated sequence. In one embodiment, the amino acid sequence is at least 75% identical to the designated sequence. In one embodiment, the amino acid sequence is at least 95% identical to the designated sequence. In one embodiment, the amino acid sequence is at least 96% identical to the designated sequence. In one embodiment, the amino acid sequence is at least 97% identical to the designated sequence. In one embodiment, the amino acid sequence is at least 98% identical to the designated sequence. In one embodiment, the amino acid sequence is at least 99% identical to the designated sequence. In one embodiment, the amino acid sequence is at least 99.5% identical to the designated sequence.

[0310] Concept 25l:V L Any of concepts 1 to 24 (provided that, when subordinate to concept 9), wherein the domain comprises the amino acid sequence of SEQ ID NO: 359 or a light chain variable domain amino acid sequence that is at least 80% (e.g., at least 85% or at least 90%) identical to SEQ ID NO: 359. (if subject to Concept 9L, then subject to Concept 13L, if subject to Concept 13, then subject to Concept 13L, if subject to Concept 16, then subject to Concept 16L, if subject to Concept 17, then subject to Concept 17L, if subject to Concept 18, then subject to Concept 18L, if subject to Concept 19, then subject to Concept 19L, if subject to Concept 20, then subject to Concept 20L, if subject to Concept 22, then subject to Concept 22L, if subject to Concept 23, then subject to Concept 23L, and if subject to Concept 24, then subject to Concept 24L).

[0311] Concept 26.V L Domain or V L 22. The antibody or fragment of any one of concepts 12 to 21, comprising a first and a second copy of the domain.

[0312] Concept 27. The antibody or fragment of any of Concepts 1-26, which specifically binds to cynomolgus PD-L1 as defined by SEQ ID NO: 2. In one embodiment, the antibody or fragment has a potency of less than 1 nM (e.g., 1 nM to 0.01 pM, or 1 nM to 0.1 pM, or 1 nM In one embodiment, the antibody or fragment binds to cynomolgus PDL-1 with an affinity of 1 pM or less. The fragments are less than 10 nM (e.g., 10 nM to 0.01 pM, or 10 nM to 0.1 pM, or binds to cynomolgus monkey PDL-1 with an affinity of 10 nM to 1 pM. The antibody or fragment may have a concentration of less than 0.1 nM (e.g., 0.1 nM to 0.01 pM, or 0.1 nM Binds to cynomolgus monkey PDL-1 with an affinity of ~0.1 pM, or 0.1 nM to 1 pM In one embodiment, the antibody or fragment has a potency of less than 0.01 nM (e.g., 0.011 nM to 0. Binds to cynomolgus monkey PDL-1 with an affinity of 0.01 pM or 0.01 nM to 0.1 pM In one embodiment, the antibody or fragment binds to cynomolgus PD-L1 with an affinity that is within 2-fold of its affinity to hPD-L1. In one embodiment, the antibody or fragment binds to cynomolgus PD-L1 with an affinity that is within 4-fold of its affinity to hPD-L1. In one embodiment, the antibody or fragment binds to cynomolgus PD-L1 with an affinity that is within 5-fold of its affinity to hPD-L1. In one embodiment, the antibody or fragment binds to cynomolgus PD-L1 with an affinity that is within 6-fold of its affinity to hPD-L1. In one embodiment, the antibody or fragment binds to cynomolgus PD-L1 with an affinity that is within 8-fold of its affinity to hPD-L1. In one embodiment, the antibody or fragment binds to cynomolgus PD-L1 with an affinity that is within 10-fold of its affinity to hPD-L1. In one embodiment, the antibody or fragment does not detectably bind to cynomolgus monkey PD-L1. In one embodiment, the antibody or fragment does not detectably bind to murine PD-L1. In one embodiment, the antibody or fragment has a concentration of less than 1 nM (e.g., 1 nM to 0.01 pM, or 1 It binds to mouse PDL-1 with an affinity of 0.1 nM to 0.1 pM, or 1 nM to 1 pM. In embodiments, the antibody or fragment has a concentration of less than 10 nM (e.g., 10 nM to 0.01 pM, or 1 It binds to mouse PDL-1 with an affinity of 0 nM to 1 pM, or 10 nM to 1 pM. In embodiments, the antibody or fragment has a concentration of less than 0.1 nM (e.g., 0.1 nM to 0.01 pM, or binds to mouse PDL-1 with an affinity of 0.1 nM to 0.1 pM, or 0.1 nM to 1 pM. In one embodiment, the antibody or fragment has a binding activity of less than 0.01 nM (e.g., 0.011 nM Binds to mouse PDL-1 with an affinity of ~0.01 pM or 0.01 nM to 0.1 pM .

[0313] Concept 28. The antibody or fragment of any of Concepts 1-27, wherein the antibody or fragment comprises a kappa light chain. Kappa light chain constant region amino acid and nucleotide sequences can be found in SEQ ID NOs: 206 and 215. In one embodiment, the light chain can be a lambda light chain. Lambda light chain constant region amino acid and nucleotide sequences can be found in SEQ ID NOs: 216-237, as well as SEQ ID NOs: 535, 536, and 538.

[0314] Concept 29. The amino acid substitution is a conservative amino acid substitution, and optionally the conservative substitution is: 1) Alanine (A), serine (S), threonine (T), 2) Aspartic acid (D), glutamic acid (E), 3) Asparagine (N), Glutamine (Q), 4) Arginine (R), Lysine (K), 5) Isoleucine (I), Leucine (L), Methionine (M), Valine (V), and 6) Six amino acids selected from phenylalanine (F), tyrosine (Y), and tryptophan (W) 29. The antibody or fragment of any one of concepts 9-28, wherein the antibody or fragment is from one of groups (each group containing amino acids that are conservative substitutions for one another).

[0315] Conservative substitutions may be as described above in Concept 9.

[0316] Concept 30. The antibody or fragment of any of Concepts 1-29, wherein the antibody or fragment comprises a constant region, e.g., a human constant region, e.g., an effector-null human constant region, e.g., an IgG4 constant region or an IgG1 constant region, optionally wherein the constant region is IgG4-PE (SEQ ID NO: 199), or a deleted IgG1 as defined in SEQ ID NO: 205. In other embodiments, the antibody or fragment is any of the isotypes or constant regions defined herein above. In one embodiment, the constant region is wild-type human IgG1 (SEQ ID NO: 340). For example, the constant region comprises an effector-null human constant region, optionally having ADCC and / or CDC activity. In one embodiment, the constant region is an IgG1 constant region that is ADCC and / or C In another embodiment, the constant region is engineered to enhance DC and / or ADCP. , which have been engineered for enhanced effector function.

[0317] The IgG4 constant region may be any of the IgG4 constant region amino acid sequences or may be encoded by any of the nucleic acid sequences of SEQ ID NOs: 192 to 203. The heavy chain constant region may be an IgG4 containing both the Leu235Glu mutation and the Ser228Pro mutation. This "IgG4-PE" heavy chain constant region (SEQ ID NOs: 198, 199, 200) may be encoded by any of the nucleic acid sequences of SEQ ID NOs: 192 to 203. 00, and 201) is effector null.

[0318] An alternative effector-null human constant region contains the L235A and / or G237A mutations. IgG1*01 alleles (e.g., LAGA, SEQ ID NO: 204, SEQ ID NO: 205) In one embodiment, the IgG1 disclosed herein is a truncated IgG1. The antibody or antibody fragment to be treated comprises an IgG1 heavy chain constant region, the sequence of which is at positions 235 and / or It is ranked 237th (EU index numbering) and contains alanine.

[0319] The antibody-dependent cellular phagocytosis (ADCP) mechanism is described in Gul et al., "Antibody-Dependent Phagocytosis of Tumor Cells by Macrophages: A Potent Effector Mechanism of Monoclonal Antibody Therapy of Cancer." er”, Cancer Res., 75(23), December 1, 2015.

[0320] The potency of Fc-mediated effects can be enhanced by engineering the Fc domain by a variety of established techniques to increase affinity for certain Fc receptors, thereby creating a diverse profile of potential enhanced activation. This can be achieved by modification of one or several amino acid residues (see, e.g., Lazar et al.). (e.g., as described in J. Am. Chem. Soc. Soc. 2006, 14:103(11):4005-10; the modifications disclosed therein are incorporated herein by reference). IgG1 constant regions containing additions (e.g., N297Q, EU index numbering) have been shown to enhance binding to Fc receptors. In one embodiment, such mutations are at positions 239, 332, and 330 for the human IgG1 constant region (or other IgG isotypes). In one embodiment, the residues are one or more of the residues selected from: The antibody or fragment may contain the amino acids N297Q, S239D, I332E, and A330L (EU index). and a human IgG1 constant region having one or more mutations independently selected from the following: . In another embodiment, increased affinity for Fc receptors is achieved by modifying the native glycosylation profile of the Fc domain, for example, by generating underfucosylated or defucosylated variants (Natsume et al., 2009, Drug Des.Devel.Ther.,3:7-16 or Zhou Q, Biotechnol.Bioeng,2008,Feb 15,99(3):652-65 (The modifications described therein are incorporated herein by reference.) A nonfucosylated antibody is one in which the Fc complex N-glycan fragments lack fucose residues. These glycoengineered antibodies, which lack the core fucose residue from the Fc N-glycan, may exhibit stronger ADCC than their fucosylated counterparts due to enhanced FcγRIIIa binding ability. For example, to increase ADCC, residues in the hinge region can be modified to increase binding to FcγRIII (e.g., Sh (See, e.g., ields et al., 2001, J. Biol. Chem., Mar 2;276(9):6591-604; the modifications described therein are incorporated herein by reference.) Thus, in one embodiment, the antibody or fragment is a wild-type human IgG The antibody or fragment comprises a human IgG heavy chain constant region that is a variant of the heavy chain constant region, wherein the variant human IgG heavy chain constant region binds to a human Fcγ receptor selected from the group consisting of FcyRIIB and FcyRIIA with higher affinity than the wild-type human IgG heavy chain constant region binds to the human Fcγ receptor. In one embodiment, the antibody or fragment comprises a human IgG heavy chain constant region that is a variant of the wild-type human IgG heavy chain constant region, wherein the variant human IgG heavy chain constant region binds to human FcγRIIB with higher affinity than the wild-type human IgG heavy chain constant region binds to human FcγRIIB. In one embodiment, the variant human IgG heavy chain constant region is a variant human IgG1, variant human IgG2, or variant human IgG4 heavy chain constant region. In one embodiment, the variant human IgG heavy chain constant region comprises one or more amino acid mutations selected from G236D, P238D, S239D, S267E, L328F, and L328E (EU index numbering system). In another embodiment, the variant human IgG heavy chain constant region has one or more substitutions selected from the group consisting of: S267E and L328F; P238D and L328E; P238D and one or more substitutions selected from the group consisting of E233D, G237D, H268D, P271G, and A330R; P238D, E233D, G237D, H268D, P271G, and A330R; G236D and S267E; S239D and S267E; V262E, S267E, and L328F; and V264E, S267E, and L328F (EU index numbering system). In another embodiment, the mutations The human IgG heavy chain constant region further comprises one or more amino acid mutations that reduce the affinity of the IgG for human FcγRIIIA, human FcγRIIA, or human FcγRI. In one embodiment, FcγRIIB is expressed on cells selected from the group consisting of macrophages, monocytes, B cells, dendritic cells, endothelial cells, and activated T cells. In one embodiment, the variant human IgG heavy chain constant region comprises the amino acid mutations G236A, S239D, F243L, T256A, K290A, R292P, S298A, Y300L, V305I, A330L, I332E, E333A, K334A, A339T, and P396L (EU index). In one embodiment, the variant human IgG heavy chain constant region comprises one or more of the following: are S239D; T256A; K290A; S298A; I332E; E333A; K334A; A339T; S239D and I332E; S239D, A330L, and I332E; S298A, E333A, and K334A; G236A, S239D, and I332E; and F243L, R292P, Y300L, V305I, and P396L (E U-indexed numbering system). In one embodiment, the variant human IgG heavy chain constant region comprises S239D, A330L, or I332E amino acid mutations (EU index numbering system). In one embodiment, the variant human IgG heavy chain constant region comprises S239D and and I332E amino acid mutations (EU index numbering system). In one embodiment, the antibody or fragment thereof comprises an afucosylated Fc region. In another embodiment, the antibody or fragment thereof is defucosylated. In another embodiment, the antibody or fragment thereof is fucosylated.

[0321] In another embodiment, the antibodies and fragments disclosed herein may contain a triple mutation (M252Y / S254T / T256E) that enhances binding to FcRn. For a discussion of the mutations in Table 2 affecting FcRn binding, see Dall et al., Immunol 2002;169:5171-5180 (the mutations described therein are incorporated herein by reference). incorporated into the book).

[0322] Similarly, enhanced CDC can be achieved by amino acid changes that increase affinity for C1q, the first component of the classical complement activation cascade (Idusogie (See, e.g., et al., J. Immunol., 2001, 166:2571-2575; the modifications described are incorporated herein by reference). Another approach is to The goal is to create a chimeric Fc domain made from human IgG1 and human IgG3 segments that takes advantage of the higher affinity of IgG3 for C1q (Natsume et al., 2011). (T al., 2008, Cancer Res., 68:3863-3872; modifications are incorporated herein by reference). In another embodiment, the antibodies or The antibody fragments may be used to modify C1q binding and / or reduce or eliminate CDC activity. Alternatively, the present invention may include mutated amino acids at residues 329, 331, and / or 322. In embodiments, the antibodies or antibody fragments disclosed herein may comprise an Fc region with modifications at residues 231 and 239, whereby amino acids are substituted to alter the ability of the antibody to fix complement. In one embodiment, the antibody or fragment comprises E345K, E430G, R344D, and D356R, particularly R344D and D356R (EU index no. The constant region comprises one or more mutations selected from the double mutations comprising the nucleotide sequence of ...

[0323] The antibody may have a heavy chain constant region that binds to one or more types of Fc receptor but does not induce cellular effector function, i.e., does not mediate ADCC, CDC, or ADCP activity. Such a constant region may not be able to bind to a particular Fc receptor responsible for triggering ADCC, CDC, or ADCP activity. The antibody may have a heavy chain constant region that does not bind to Fcγ receptors. Thus, in one embodiment, the constant region may contain a Leu235Glu mutation (EU index numbering system).

[0324] In another embodiment, the antibodies and fragments disclosed herein have increased or decreased serum half-life. In one embodiment, one or more of the mutations T252L, T254S, or T256F are introduced to increase the biological half-life of the antibody. Biological half-life can also be increased by modifying the heavy chain constant region CH1 domain or CL region to contain salvage receptor binding epitopes taken from two loops of the CH2 domain of the Fc region of IgG, as described in U.S. Patent Nos. 5,869,046 and 6,121,022, the modifications described therein being incorporated herein by reference. In another embodiment, the Fc hinge region of an antibody or antigen-binding fragment of the invention is mutated to decrease the biological half-life of the antibody or fragment. Introducing one or more amino acid mutations into the CH2-CH3 domain interface region of the Fc hinge fragment results in impaired Staphylococcus protein A (SpA) binding of the antibody or fragment compared to native Fc hinge domain SpA binding. Other methods for increasing serum half-life are known to those skilled in the art. Thus, in one embodiment, the antibody or fragment is PEGylated. In another embodiment, the antibody or fragment is fused to an albumin-binding domain, e.g., an albumin-binding single domain antibody (dAb). In another embodiment, the antibody or fragment is PASylated (i.e., a large PAS (XL-Protein) forms a non-charged random coil structure with hydrodynamic volume. in GmbH)). In another embodiment, the antibody or fragment is XTENylated® / rPEGylated (i.e., genetic fusion of a non-exact repeat peptide sequence (Amunix, Versartis) with a therapeutic peptide). In another embodiment, the antibody or fragment is ELPylated (i.e., genetic fusion of a polypeptide sequence consisting of an ELP repeat sequence (Ph These various half-life extending fusions are described in detail by Strohl, BioDrugs (2015) 29:215-239, e.g., those fusions in Tables 2 and 6, which are incorporated herein by reference.

[0325] The antibody may have a modified constant region that increases stability. Thus, in one embodiment, the heavy chain constant region contains a Ser228Pro mutation. In another embodiment, the antibodies and fragments disclosed herein contain a heavy chain hinge region modified to alter the number of cysteine ​​residues. This modification can be used to facilitate assembly of the light and heavy chains or to increase or decrease the stability of the antibody.

[0326] Concept 31. The antibody or fragment of Concept 30, wherein the constant region is a murine constant region. In another embodiment, the constant region can be of any non-human mammalian origin, such as rat, mouse, hamster, guinea pig, dog, cat, horse, chicken, llama, dromedary, etc. In one embodiment, the constant region is a rat constant region. In another embodiment, the constant region is a llama constant region. The murine constant region can be any of the isotypes or alleles described herein above.

[0327] Concept 32. The antibody of Concept 30 or Concept 3, wherein the constant region has CDC and / or ADCC activity. 1. The antibody or fragment described in 1.

[0328] Concept 33. a)V H The domain comprises the amino acid sequence of SEQ ID NO: 33, and V L Domain is sequence number No. 43, b)V H The domain comprises an amino acid sequence at least 85% identical to SEQ ID NO: 33. , and V L the domain comprises an amino acid sequence at least 85% identical to SEQ ID NO: 43; c)V H Domain V of SEQ ID NO: 47 H domain, and V L Do The main amino acid sequence of SEQ ID NO: 43 is contained, d)V H Domain V of SEQ ID NO: 48H domain, and V L Do The main amino acid sequence of SEQ ID NO: 43 is contained, e)V H Domain V of SEQ ID NO: 49 H domain, and V L Do The main amino acid sequence of SEQ ID NO: 43 is contained, f)V H Domain V of SEQ ID NO: 342 H domain, and V L the domain comprises the amino acid sequence of SEQ ID NO: 43; g)V H The domain comprises the amino acid sequence of SEQ ID NO: 33, and V L Domain is sequence number No. 50 V L comprising the amino acid sequence of the domain, h)V H Domain V of SEQ ID NO: 47 H domain, and V L Do The main sequence is V of sequence number 50 L comprising the amino acid sequence of the domain, i)V H Domain V of SEQ ID NO: 48 H domain, and V L Do The main sequence is V of sequence number 50 L comprising the amino acid sequence of the domain, j)V H Domain V of SEQ ID NO: 49 H domain, and V L Do The main sequence is V of sequence number 50 L comprising the amino acid sequence of the domain, k)V H Domain V of SEQ ID NO: 342 H domain, and V L Domain V of SEQ ID NO: 50 Lcomprising the amino acid sequence of the domain, l)V H The domain comprises the amino acid sequence of SEQ ID NO: 33, and V L Domain is sequence number No. 51 V L comprising the amino acid sequence of the domain, m)V H Domain V of SEQ ID NO: 47 H domain, and V L Do The main V is sequence number 51 L comprising the amino acid sequence of the domain, n)V H Domain V of SEQ ID NO: 48 H domain, and V L Do The main V is sequence number 51 L comprising the amino acid sequence of the domain, o)V H Domain V of SEQ ID NO: 49 H domain, and V L Do The main V is sequence number 51 L comprising the amino acid sequence of the domain, p)V H Domain V of SEQ ID NO: 342 H domain, and V L Domain V of SEQ ID NO: 51 L comprising the amino acid sequence of the domain, q)V H The domain comprises the amino acid sequence of SEQ ID NO: 33, and V L Domain is sequence number No. 298 V L comprising the amino acid sequence of the domain, r)V H Domain V of SEQ ID NO: 47 H domain, and V L Do The main sequence is V of sequence number 298 L comprising the amino acid sequence of the domain, s)V HDomain V of SEQ ID NO: 48 H domain, and V L Do The main sequence is V of sequence number 298 L comprising the amino acid sequence of the domain, t)V H Domain V of SEQ ID NO: 49 H domain, and V L Do The main sequence is V of sequence number 298 L comprising the amino acid sequence of the domain, u)V H Domain V of SEQ ID NO: 342 H domain, and V L Domain V of SEQ ID NO: 298 L comprising the amino acid sequence of the domain, v)V H The domain comprises the amino acid sequence of SEQ ID NO: 58, and V L Domain is sequence number No. 68, w)V H The domain comprises an amino acid sequence at least 85% identical to SEQ ID NO: 58. , and V L the domain comprises an amino acid sequence at least 85% identical to SEQ ID NO: 68, x)V H The domain comprises the amino acid sequence of SEQ ID NO: 78, and V L Domain is sequence number No. 88, y)V H The domain comprises an amino acid sequence at least 85% identical to SEQ ID NO: 78. , and V L the domain comprises an amino acid sequence at least 85% identical to SEQ ID NO: 88; z)V H The domain comprises the amino acid sequence of SEQ ID NO: 98, and V L Domain is sequence number No. 108, aa)V HThe domain contains an amino acid sequence that is at least 85% identical to SEQ ID NO:98. Mikatsu V L a domain comprising an amino acid sequence at least 85% identical to SEQ ID NO: 108; bb)V H The domain comprises the amino acid sequence of SEQ ID NO: 118, and V L Domain is distributed containing the amino acid sequence of sequence number 128, cc)V H An amino acid sequence in which the domain is at least 85% identical to SEQ ID NO: 118 Contains and V L a domain comprising an amino acid sequence at least 85% identical to SEQ ID NO: 128; dd)V H The domain comprises the amino acid sequence of SEQ ID NO: 158, and V L Domain is distributed containing the amino acid sequence of sequence number 168, ee)V H An amino acid sequence whose domain is at least 85% identical to SEQ ID NO: 158 Contains and V L a domain comprising an amino acid sequence at least 85% identical to SEQ ID NO: 168; ff)V H The domain comprises the amino acid sequence of SEQ ID NO: 178, and V L Domain is distributed containing the amino acid sequence of sequence number 188, gg)V H An amino acid sequence in which the domain is at least 85% identical to SEQ ID NO: 178 Contains and V L a domain comprising an amino acid sequence at least 85% identical to SEQ ID NO: 188; hh)V H The domain comprises the amino acid sequence of SEQ ID NO: 138, and V L Domain is distributed containing the amino acid sequence of sequence number 148, ii) V H An amino acid sequence whose domain is at least 85% identical to SEQ ID NO: 138 Contains and VL a domain comprising an amino acid sequence at least 85% identical to SEQ ID NO: 148; jj)V H The domain comprises the amino acid sequence of SEQ ID NO: 244, and V L Domain is distributed containing the amino acid sequence of sequence number 254, kk)V H An amino acid sequence in which the domain is at least 85% identical to SEQ ID NO: 244 Contains and V L a domain comprising an amino acid sequence at least 85% identical to SEQ ID NO: 254; ll)V H The domain comprises the amino acid sequence of SEQ ID NO: 264, and V L Domain is distributed containing the amino acid sequence of sequence number 274, mm)V H An amino acid sequence in which the domain is at least 85% identical to SEQ ID NO: 264 Contains and V L a domain comprising an amino acid sequence at least 85% identical to SEQ ID NO: 274; nn)V H The domain comprises the amino acid sequence of SEQ ID NO: 284, and V L Domain is distributed containing the amino acid sequence of sequence number 294, oo)V H An amino acid sequence in which the domain is at least 85% identical to SEQ ID NO: 284 Contains and V L a domain comprising an amino acid sequence at least 85% identical to SEQ ID NO: 294; pp)V H The domain comprises the amino acid sequence of SEQ ID NO: 349, and V L Domain is distributed containing the amino acid sequence of sequence number 359, qq)V H An amino acid sequence in which the domain is at least 85% identical to SEQ ID NO: 349 Contains and V L33. The antibody of any one of concepts 1 to 32, wherein the domain comprises an amino acid sequence that is at least 85% identical to SEQ ID NO:359.

[0329] In one embodiment, the amino acid sequence is at least 70% identical to the designated sequence. In one embodiment, the amino acid sequence is at least 75% identical to the designated sequence. In one embodiment, the amino acid sequence is at least 95% identical to the designated sequence. In one embodiment, the amino acid sequence is at least 96% identical to the designated sequence. In one embodiment, the amino acid sequence is at least 97% identical to the designated sequence. In one embodiment, the amino acid sequence is at least 98% identical to the designated sequence. In one embodiment, the amino acid sequence is at least 99% identical to the designated sequence. In one embodiment, the amino acid sequence is at least 99.5% identical to the designated sequence.

[0330] Concept 34. An antibody comprises a heavy chain and a light chain; a) the heavy chain amino acid sequence comprises the amino acid sequence of SEQ ID NO: 35, and the light chain amino acid sequence comprises the amino acid sequence of SEQ ID NO: 45, b) an amino acid sequence whose heavy chain amino acid sequence is at least 85% identical to SEQ ID NO: 35; and the light chain amino acid sequence is at least 85% identical to SEQ ID NO: 45. Including, c) the heavy chain amino acid sequence comprises the amino acid sequence of SEQ ID NO: 47, and the light chain amino acid sequence comprises the amino acid sequence of SEQ ID NO: 45, d) the heavy chain amino acid sequence comprises the amino acid sequence of SEQ ID NO: 48, and the light chain amino acid sequence comprises the amino acid sequence of SEQ ID NO: 45, e) the heavy chain amino acid sequence comprises the amino acid sequence of SEQ ID NO: 49, and the light chain amino acid sequence comprises the amino acid sequence of SEQ ID NO: 45, f) the heavy chain amino acid sequence comprises the amino acid sequence of SEQ ID NO: 342, and the light chain amino acid sequence the string comprises the amino acid sequence of SEQ ID NO: 45, g) the heavy chain amino acid sequence comprises the amino acid sequence of SEQ ID NO: 35, and the light chain amino acid sequence comprises the amino acid sequence of SEQ ID NO: 50, h) the heavy chain amino acid sequence comprises the amino acid sequence of SEQ ID NO: 47, and the light chain amino acid sequence comprises the amino acid sequence of SEQ ID NO: 50, i) the heavy chain amino acid sequence comprises the amino acid sequence of SEQ ID NO: 48, and the light chain amino acid sequence comprises the amino acid sequence of SEQ ID NO: 50, j) the heavy chain amino acid sequence comprises the amino acid sequence of SEQ ID NO: 49, and the light chain amino acid sequence comprises the amino acid sequence of SEQ ID NO: 50, k) the heavy chain amino acid sequence comprises the amino acid sequence of SEQ ID NO: 342, and the light chain amino acid sequence the string comprises the amino acid sequence of SEQ ID NO: 50, l) the heavy chain amino acid sequence comprises the amino acid sequence of SEQ ID NO: 35, and the light chain amino acid sequence comprises the amino acid sequence of SEQ ID NO: 51, m) the heavy chain amino acid sequence comprises the amino acid sequence of SEQ ID NO: 47, and the light chain amino acid sequence comprises the amino acid sequence of SEQ ID NO: 51, n) the heavy chain amino acid sequence comprises the amino acid sequence of SEQ ID NO: 48, and the light chain amino acid sequence comprises the amino acid sequence of SEQ ID NO: 51, o) the heavy chain amino acid sequence comprises the amino acid sequence of SEQ ID NO: 49, and the light chain amino acid sequence comprises the amino acid sequence of SEQ ID NO: 51, p) the heavy chain amino acid sequence comprises the amino acid sequence of SEQ ID NO: 342, and the light chain amino acid sequence the string comprises the amino acid sequence of SEQ ID NO: 51, q) the heavy chain amino acid sequence comprises the amino acid sequence of SEQ ID NO: 35, and the light chain amino acid sequence comprises the amino acid sequence of SEQ ID NO: 298; r) the heavy chain amino acid sequence comprises the amino acid sequence of SEQ ID NO: 47, and the light chain amino acid sequence comprises the amino acid sequence of SEQ ID NO: 298; s) the heavy chain amino acid sequence comprises the amino acid sequence of SEQ ID NO: 48, and the light chain amino acid sequence comprises the amino acid sequence of SEQ ID NO: 298; t) the heavy chain amino acid sequence comprises the amino acid sequence of SEQ ID NO: 49, and the light chain amino acid sequence comprises the amino acid sequence of SEQ ID NO: 298; u) the heavy chain amino acid sequence comprises the amino acid sequence of SEQ ID NO: 342, and the light chain amino acid sequence the string comprises the amino acid sequence of SEQ ID NO: 298, v) the heavy chain amino acid sequence comprises the amino acid sequence of SEQ ID NO: 60, and the light chain amino acid sequence comprises the amino acid sequence of SEQ ID NO: 70, w) a heavy chain amino acid sequence having an amino acid sequence at least 85% identical to SEQ ID NO: 60; and the light chain amino acid sequence comprises an amino acid sequence at least 85% identical to SEQ ID NO: 70, x) the heavy chain amino acid sequence comprises the amino acid sequence of SEQ ID NO: 80, and the light chain amino acid sequence comprises the amino acid sequence of SEQ ID NO: 90, y) an amino acid sequence whose heavy chain amino acid sequence is at least 85% identical to SEQ ID NO: 80; and the light chain amino acid sequence comprises an amino acid sequence at least 85% identical to SEQ ID NO: 90; z) the heavy chain amino acid sequence comprises the amino acid sequence of SEQ ID NO: 100, and the light chain amino acid sequence the string comprises the amino acid sequence of SEQ ID NO: 110, aa) a heavy chain amino acid sequence that is at least 85% identical to SEQ ID NO: 100 and the light chain amino acid sequence comprises an amino acid sequence that is at least 85% identical to SEQ ID NO: 110, bb) the heavy chain amino acid sequence comprises the amino acid sequence of SEQ ID NO: 120, and the light chain amino acid sequence the sequence comprises the amino acid sequence of SEQ ID NO: 130, cc) a heavy chain amino acid sequence having at least 85% identity to SEQ ID NO: 120; and the light chain amino acid sequence comprises an amino acid sequence at least 85% identical to SEQ ID NO: 130, dd) the heavy chain amino acid sequence comprises the amino acid sequence of SEQ ID NO: 160, and the light chain amino acid sequence the sequence comprises the amino acid sequence of SEQ ID NO: 170, ee) an amino acid sequence whose heavy chain amino acid sequence is at least 85% identical to SEQ ID NO: 160 and the light chain amino acid sequence comprises an amino acid sequence that is at least 85% identical to SEQ ID NO: 170, ff) the heavy chain amino acid sequence comprises the amino acid sequence of SEQ ID NO: 180, and the light chain amino acid sequence the sequence comprises the amino acid sequence of SEQ ID NO: 190, gg) an amino acid sequence whose heavy chain amino acid sequence is at least 85% identical to SEQ ID NO: 180 and the light chain amino acid sequence comprises an amino acid sequence that is at least 85% identical to SEQ ID NO: 190, hh) the heavy chain amino acid sequence comprises the amino acid sequence of SEQ ID NO: 140, and the light chain amino acid sequence the sequence comprises the amino acid sequence of SEQ ID NO: 150, ii) an amino acid sequence whose heavy chain amino acid sequence is at least 85% identical to SEQ ID NO: 140; and the light chain amino acid sequence comprises an amino acid sequence that is at least 85% identical to SEQ ID NO: 150, jj) the heavy chain amino acid sequence comprises the amino acid sequence of SEQ ID NO: 246, and the light chain amino acid sequence the sequence comprises the amino acid sequence of SEQ ID NO: 256, kk) a heavy chain amino acid sequence having at least 85% identity to SEQ ID NO: 246 and the light chain amino acid sequence comprises an amino acid sequence at least 85% identical to SEQ ID NO: 256, ll) the heavy chain amino acid sequence comprises the amino acid sequence of SEQ ID NO: 266, and the light chain amino acid sequence the sequence comprising the amino acid sequence of SEQ ID NO: 276, mm) a heavy chain amino acid sequence having at least 85% identity to SEQ ID NO: 266 and the light chain amino acid sequence comprises an amino acid sequence at least 85% identical to SEQ ID NO: 276, nn) the heavy chain amino acid sequence comprises the amino acid sequence of SEQ ID NO: 286, and the light chain amino acid sequence the sequence comprising the amino acid sequence of SEQ ID NO: 296, oo) an amino acid sequence whose heavy chain amino acid sequence is at least 85% identical to SEQ ID NO: 286 and the light chain amino acid sequence comprises an amino acid sequence at least 85% identical to SEQ ID NO: 296, pp) the heavy chain amino acid sequence comprises the amino acid sequence of SEQ ID NO: 351, and the light chain amino acid sequence the sequence comprising the amino acid sequence of SEQ ID NO: 361, qq) an amino acid sequence whose heavy chain amino acid sequence is at least 85% identical to SEQ ID NO: 351 34. The antibody of any of Concepts 1 to 33, wherein the antibody comprises a light chain amino acid sequence comprising a sequence at least 85% identical to SEQ ID NO: 361.

[0331] In one embodiment, the amino acid sequence is at least 70% identical to the designated sequence. In one embodiment, the amino acid sequence is at least 75% identical to the designated sequence. In one embodiment, the amino acid sequence is at least 95% identical to the designated sequence. In one embodiment, the amino acid sequence is at least 96% identical to the designated sequence. In one embodiment, the amino acid sequence is at least 97% identical to the designated sequence. In one embodiment, the amino acid sequence is at least 98% identical to the designated sequence. In one embodiment, the amino acid sequence is at least 98% identical to the specified sequence. In one embodiment, the amino acid sequence is at least 99% identical to the specified sequence. In one embodiment, the amino acid sequence is at least 99.5% identical to the specified sequence.

[0332] Concept 35. The antibody or fragment of any of Concepts 1-34, which competes with antibody 1D05 for binding to hPDL-1, and optionally wherein the competition for binding to hPD-L1 is carried out using SPR. SPR can be carried out as described herein above or as described in Concept 16.

[0333] Concept 36. The antibody or fragment can inhibit PD-L1-mediated T cell suppression and, optionally, provide costimulation either by direct CD3 / CD28 stimulation or superantigen stimulation. The antibody or fragment of any of concepts 1-35, wherein T cell suppression is measured by an increase in one or more of IFNγ, IL-2, CD25, or T cell proliferation in an assay that provides costimulation by co-incubation with cells capable of inducing a T cell response, or by co-incubation with cells capable of inducing a T cell response. Measurements can be performed using any suitable technique. For example, measurements can be performed using ELISA, HTRF, BRDU uptake (proliferation), electrochemiluminescence (ECL), or flow cytometry (e.g., FACS). These techniques are well known to those of skill in the art and are described elsewhere herein. In one embodiment, the assay is flow cytometry. In one embodiment, the assay is ELISA. In one embodiment, the assay is HTRF. In one embodiment, T cell suppression is measured by an increase in IFNγ. In one embodiment, T cell suppression is measured by an increase in IL-2. In one embodiment, T cell suppression is measured by an increase in CD25. In one embodiment, T cell suppression is measured by an increase in IFNγ and IL-2. In one embodiment, T cell suppression is measured by an increase in IFNγ and CD25. In one embodiment, T cell suppression is measured by an increase in CD25 and IL-2. In one embodiment, T cell suppression is measured by an increase in IFNγ, IL-2, and CD25. In one embodiment, costimulation is provided by direct CD3 / CD28 stimulation. In some embodiments, costimulation is provided by a superantigen, such as staphylococcal enterotoxin B (SEB). In one embodiment, the assay provides costimulation by co-incubation with a cell capable of inducing a T cell response. The cell can be an antigen-presenting cell (APC), e.g., a monocyte, a B cell, or a dendritic cell. In one embodiment, the assay provides costimulation by co-incubation with an APC. In one embodiment, the assay provides costimulation by co-incubation with a monocyte. In one embodiment, the assay provides costimulation by co-incubation with a B cell. In one embodiment, the assay provides costimulation by co-incubation with a dendritic cell.

[0334] Concept 37. A bispecific antibody or fusion protein comprising the antibody or fragment thereof according to any one of Concepts 1 to 36.

[0335] Concept 37a: A dual-binding antibody or fusion protein comprising an antibody or fragment thereof according to any one of Concepts 1 to 36. Dual-binding antibody has the meaning described above.

[0336] Concept 38. Bispecific formats include DVD-Ig, mAb 2 , FIT-Ig, mAb-dAb, dock and lock, SEEDbody, scDiabody-Fc, diabody-Fc, tandem scFv-Fc, Fab-scFv-Fc, Fab-scFv, intrabody, BiTE, diabody, DART, TandAb, scDiabody, scDiabody-CH3, Diabody-CH3, minibody, knobs-in-holes, knobs-in-hole with common light chain, knobs-in-hole with common light chain and charge pair, charge pair , charge pairs with a common light chain, especially mAbs 2 mAb, e.g., knobs-in-hole, common light chain, and charge pair and FIT-Ig with knobs-in-hole, common light chain, and common light chain 2and knob-in-hole with FIT-Ig.

[0337] In one embodiment, the bispecific format is DVD-Ig, mAb 2 , FIT-Ig, mAb-dAb, Dock-Lock, Fab-Arm Exchange, SEEDbody, Triomab, LUZ-Y, Fcab, κλ-body, Orthogonal Fab, scDiabody-Fc, diabody-Fc, tandem scFv-Fc, Fab-scFv-Fc, Fab-scFv, intracellular antibody, BiTE, diabody, DART, TandemAb, scDiabody, scDiabody-CH3, Diabody-CH3, Triple body body), miniantibody, minibody, TriBi minibody, scFv-CH3KIH, scFv-CH-CL-scFv, F(ab')2-scFv, scFv-KIH, Fab-scFv-Fc, tetravalent HCab, ImmTAC, knob-in-hole, knob-in-hole with common light chain, knob-in-hole with common light chain and charge pair, charge pair, charge pair with common light chain, DT-IgG, DutaMab, IgG(H)-scFv, scFv-(H)IgG, IgG(L)-scFv, scFv-(L)IgG, IgG(L,H)-Fv, IgG(H)-V, V(H)-IgG, IgG(L)-V, V( L)-IgG, KIH IgG-scFab, 2scFv-IgG, IgG-2scFv , scFv4-Ig, and zybody.

[0338] In one embodiment, the bispecific format is DVD-Ig, FIT-Ig, mAb-dAb, Dock-Lock, Fab-arm exchange, SEEDbody, Triomab, LUZ-Y, Fcab, κλ-body, orthogonal Fab, scDiabody-Fc, diabody-Fc, tandem scFv-Fc, Fab-scFv-Fc, Fab-scFv, intrabody, BiTE, diabody, DART, TandAb, scDiabody, scDiabody-CH3, Diabody-CH3, triplebody, miniantibody, minibody, TriBi minibody, scFv-CH3KIH, scFv-CH-CL-scFv, F(ab')2-scFv, scFv-KIH, Fab-scFv-Fc, tetravalent HCa b, ImmTAC, knob-in-hole, knob-in-hole with common light chain, knob-in-hole with common light chain and charge pair, charge pair, charge pair with common light chain, DT-IgG, DutaMab, IgG(H)-scFv, scFv-(H)IgG, IgG(L)-scFv, scFv-(L)IgG, IgG(L,H)-Fv, IgG(H)-V, V(H)-IgG, IgG(L)-V, V(L)-IgG, KIH IgG-scFab, 2scFv-IgG, IgG-2scFv, scFv4-Ig, and zybodies, such as DVD-Ig, FIT-Ig, mAb-dAb, Dock-Lock, SEEDbody, scDiabody-Fc, diabody-Fc, tandem scFv-Fc, Fab-scFv-Fc, Fab-scFv, intrabody, BiTE, diabody, DART, TandAb, scDiabody, scDiabody-CH3, Diabody-CH3, minibody, knobs-in-hole, knobs-in-hole with a common light chain, knobs-in-hole with a common light chain and charge pair, charge pair, charge pair with a common light chain, especially knobs-in-hole, knobs-in-hole with a common light chain, knobs-in-hole with a common light chain and charge pair, and FIT-Ig, such as FIT-Ig.

[0339] In one embodiment, the bispecific format is DVD-Ig, mAb 2, mAb-dAb, Dock-Lock, Fab-arm exchange, SEEDbody, Triomab, LUZ-Y, Fcab, κλ-body, Orthogonal Fab, scDiabody-Fc, diabody-Fc, tandem scFv-Fc, Fab-scFv-Fc, Fab-scFv, intracellular antibody, BiTE, diabody, DART, TandAb, scDiabod y, scDiabody-CH3, Diabody-CH3, triple body, miniantibody, minibody, TriBi minibody, scFv-CH3KIH, scFv-CH-CL-scFv, F(ab')2-scFv, scFv-KIH, Fab-scFv-Fc, tetravalent HCab, ImmTAC, knob-in-hole, knob-in-hole with common light chain, knob-in-hole with common light chain and charge pair, charge pair, charge pair with common light chain, DT-IgG, DutaMab, IgG(H)-scFv, scFv-(H)IgG, IgG(L)-scFv, scFv-(L)IgG, IgG(L,H)-Fv, IgG(H)-V, V(H)-IgG, IgG(L)-V, V(L)-IgG, KIH IgG-scFab, 2scFv-IgG, IgG-2scFv, scFv4-Ig, as well as zybodies, e.g., DVD-Ig, mAb 2 , mAb-dAb, Dock-Lock, SEEDbody, scDiabody-Fc, diabody-Fc, tandem scFv-Fc, Fab-scFv-Fc, Fab-scFv, intrabody, BiTE, diabody, DART, TandAb, scDiabody, scDiabody-CH3, Diabody-CH3, minibody, knob-in-hole, knob-in-hole with common light chain, knob-in-hole with common light chain and charge pair, charge pair, charge pair with common light chain, mAb, among others 2 , knobs-in-hole, knobs-in-hole with a common light chain, knobs-in-hole with a common light chain and charge pair, and knobs-in-hole with a common light chain, e.g., mAb 2 is selected from.

[0340] In one embodiment, the bispecific format includes DVD-Ig, mAb-dAb, Dock-Lock, Fab-arm exchange, SEEDbody, Triomab, LUZ-Y, Fcab, κλ-body, orthogonal Fab, scDiabody-Fc, diabody-Fc, tandem scFv-Fc, Fab-scFv-Fc, Fab-scFv, intrabody, BiTE, diabody, DART, TandAb, scDiabody, scDiabody-CH3, Diabody-CH3, triplebody, miniantibody, minibody, TriBiminibody, scFv-CH3KIH, scFv-CH-CL-scFv, F(ab')2-scFv, scFv-KIH, Fab-scFv-Fc, tetravalent HCab, ImmTAC, knob-in-hole, knob-in-hole with common light chain, knob-in-hole with common light chain and charge pair, charge pair, charge pair with common light chain, DT-IgG, DutaMab, IgG(H)-scFv, scFv-(H)IgG, IgG(L)-scFv, scFv-(L)IgG, IgG(L,H)-Fv, IgG(H)-V, V( H)-IgG, IgG(L)-V, V(L)-IgG, KIH IgG-scFab, 2sc Fv-IgG, IgG-2 scFv, scFv4-Ig, and zybodies such as DVD-Ig, mAb-dAb, Dock-Lock, SEEDbody, scDiabody-Fc, diabody-Fc, tandem scFv-Fc, Fab-scFv-Fc, Fab-scFv, intrabody, BiTE, diabody, DART, TandAb, scDiabody, scDiabody-CH3, Diabody-CH3, minibody, knobs-in-hole, knobs-in-hole with a common light chain, knobs-in-hole with a common light chain and charge pair, charge pair, charge pair with a common light chain, especially selected from knobs-in-hole, knobs-in-hole with a common light chain, knobs-in-hole with a common light chain and charge pair, and knobs-in-hole with a common light chain.

[0341] Concept 39. Bispecific antibodies combine hPD-L1 with an immune checkpoint inhibitor (e.g., , PD-1, CTLA-4, TIGIT, TIM-3, LAG-3, and VISTA, e.g., TIGIT, TIM-3, and LAG-3), immunomodulatory agents (e.g., BTLA, hHVEM, CSF1R, CCR4, CD39, CD40, CD73, CD96, CXCR2, CXCR4, CD200, GARP, SIRPα, CXCL9, CXCL10, CXCL11, and CD155, e.g., GARP, SIRPα, CXCR4, BTLA, hVEM, and CSF1R), immunostimulatory agents (e.g., CD137, GITR, OX40, CD4 0, CXCR3 (e.g., an agonistic anti-CXCR3 antibody), CD27, CD3, ICOS (e.g., an agonistic anti-ICOS antibody), e.g., ICOS, CD137, GITR, and OX40. 39. The bispecific antibody of claim 38.

[0342] Concept 39a. A bispecific antibody is one or more of the CDRs (e.g., CDRH3 and CDRH4). DRL3) or any of the variable region sequences of the antibodies described herein below in aspect 1a. Contains V H , V L , or V H and V L hPD-L1 with a pair of and a dual antibody that binds to another target antigen selected from: a VISTA antibody, e.g., TIGIT, TIM-3, and LAG-3; an immunomodulatory agent (e.g., BTLA, hHVEM, CSF1R, CCR4, CD39, CD40, CD73, CD96, CXCR2, CXCR4, CD200, GARP, SIRPα, CXCL9, CXCL10, CXCL11, and CD155, e.g., GARP, SIRPα, CXCR4, BTLA, hVEM, and CSF1R); an immunostimulatory agent (e.g., CD137, GITR, OX40, CD40, CXCR3 (e.g., an agonistic anti-CXCR3 antibody), CD27, CD3, ICOS (e.g., an agonistic anti-ICOS antibody), e.g., ICOS, CD137, GITR, and OX40). Specific antibodies.

[0343] Concept 39b. Bispecific antibodies combine hPD-L1 with immune checkpoint inhibitors (e.g., for example, PD-1, CTLA-4, TIGIT, TIM-3, LAG-3, and VISTA, e.g., TIGIT, TIM-3, and LAG-3), immunomodulatory agents (e.g., BTLA, hHVEM, CSF1R, CCR4, CD39, CD40, CD73, CD96, CXCR2, CXCR4, CD200, GARP, SIRPα, CXCL9, CXCL10, CXCL11, and CD155, e.g., GARP, SIRPα, CXCR4, BTLA, hVEM, and CSF1R), immunostimulatory agents (e.g., CD137, GITR, OX40, CD40, CXCR3 (e.g., agonistic anti-CXCR3 antibodies), CD3, ICOS (e.g., For example, agonistic anti-ICOS antibodies), for example, ICOS, CD137, GITR, and OX40), and another target antigen selected from concept 37 or concept 38. 38. The bispecific antibody according to 38.

[0344] In one embodiment, the other target antigen is an immune checkpoint inhibitor such as PD-1, CTLA-4, TIGIT, TIM-3, LAG-3, and VISTA, e.g., TIGIT, CTLA-4, TIM-3, and LAG-3. In one embodiment, the other target antigen is an immunomodulatory agent such as BTLA, hHVEM, CSF1R, CCR4, CD39, CD40, CD73, CD96, CXCR2, CXCR4, CD200, GARP, SIRPα, CXCL9, CXCL10, CXCL11, and CD155, e.g., GARP, SIRPα, CXCR4, BTLA, hVEM, and CSF1R. In one embodiment, the other target antigen is CD137, GITR, OX40, CD40, CXCR3 (e.g., agonist and immunostimulants such as CD137, GITR, OX40, CD40, CXCR3 (e.g., agonistic anti-CXCR3 antibody), CD3, CD27, and ICOS (e.g., agonistic anti-ICOS antibody), or CD137, GITR, OX40, CD40, CXCR3 (e.g., agonistic anti-CXCR3 antibody), CD3 and ICOS (e.g., agonistic anti-ICOS antibody), e.g., ICOS, CD137, GITR, and OX40.

[0345] In one embodiment, the other target antigen is CTLA-4. In one embodiment, the other target antigen is TIGIT. In one embodiment, the other target antigen is TIM-3. In one embodiment, the other target antigen is LAG-3. In one embodiment, the other target antigen is GITR. In one embodiment, the other target antigen is VISTA. In one embodiment, the other target antigen is CD137. In one embodiment, the other target antigen is SIRPα. In one embodiment, the other target antigen is CXCL10. In one embodiment, the other target antigen is CD155. In one embodiment, the other target antigen is CD40.

[0346] In another embodiment, the bispecific antibody binds to another target antigen, which is PD-1, and the binding to PD-1 is achieved by using CDR sequences (e.g., CD4 sequences) as described herein below in Aspect 1A. an antigen-binding domain (e.g., VRH3 and / or CDRL3) or a sequence comprising a variable region sequence; H , V L , or V H and V L (pair of)

[0347] In another embodiment, the bispecific antibody binds to another target antigen, which is CTLA4, and the binding to CTLA4 is achieved by CDR sequences as described herein below in Aspect 1A (e.g., an antigen-binding domain (e.g., V) having either a sequence including a variable region sequence (e.g., V H , V L , or V H and V L (pair of)

[0348] In another embodiment, the bispecific antibody binds to another target antigen, which is TIGIT, and the binding to TIGIT is achieved by a CDR sequence as described herein below in Aspect 1A (e.g., an antigen-binding domain (e.g., V) having either a sequence including a variable region sequence (e.g., V H , V L , or V H and V L (pair of)

[0349] In another embodiment, the bispecific antibody binds to another target antigen, which is TIM-3, and the binding to TIM-3 is achieved by a CDR sequence as described herein below in Aspect 1A (e.g., an antigen-binding domain (e.g., V) having either a sequence including a variable region sequence (e.g., V H , V L , or V H and V L (pair of)

[0350] In another embodiment, the bispecific antibody binds to another target antigen which is LAG3, and the binding to LAG3 is achieved by using CDR sequences (e.g., CD4, CD5, CD6, CD7, CD8, CD9, CD10, CD11, CD12, CD13, CD14, CD15, CD16, CD17, CD18, CD19, CD20, CD21, CD22, CD23, CD19, CD24, CD25, CD30, CD19, CD26, CD27, CD28, CD31, CD12, CD29, CD32, CD13, CD29, CD29, CD33, CD14, CD29, CD34, CD15, CD29, CD29, CD35, CD46, CD16, CD29, CD17, CD29, CD29, CD35, CD18, CD29, CD29, CD36, CD19, CD an antigen-binding domain (e.g., VRH3 and / or CDRL3) or a sequence comprising a variable region sequence; H , V L , or V H and V L (pair of)

[0351] In another embodiment, the bispecific antibody binds to another target antigen, which is VISTA, and the binding to VISTA is achieved by CDR sequences as described herein below in Aspect 1A (e.g., an antigen-binding domain (e.g., V) having either a sequence including a variable region sequence (e.g., V H , V L , or V H and V L (pair of)

[0352] In another embodiment, the bispecific antibody binds to another target antigen that is BTLA, and the binding to BTLA is achieved by using CDR sequences (e.g., CD4s) as described herein below in Aspect 1A. an antigen-binding domain (e.g., VRH3 and / or CDRL3) or a sequence comprising a variable region sequence; H , V L , or V H and V L (pair of)

[0353] In another embodiment, the bispecific antibody binds to another target antigen, which is hHVEM, and the binding to hHVEM is achieved by CDR sequences as described herein below in Aspect 1A (e.g., an antigen-binding domain (e.g., V) having either a sequence including a variable region sequence (...

Claims

1. a multispecific antibody that binds to ICOS and PD-L1; a pharmaceutically acceptable excipient, diluent, or carrier; Anti-CTLA-4 antibody or anti-PD-1 antibody A composition comprising:

2. 1. A multispecific antibody that binds to ICOS and PD-L1 for use in a method of treating cancer in a human patient, the method comprising administering to the patient the multispecific antibody and an anti-CTLA-4 antibody or an anti-PD-1 antibody.

3. A method of treating cancer in a human patient comprising administering to the patient (i) a multispecific antibody that binds to ICOS and PD-L1, and (ii) an anti-CTLA-4 antibody or an anti-PD-1 antibody.

4. the cancer is Treg-associated and / or tests positive for ICOS and FOXP3 expression The antibody for use according to claim 2 or the method according to claim 3, wherein the antibody exhibits the following:

5. The composition, antibody or method of any of claims 1 to 4, wherein the anti-CTLA-4 antibody is ipilimumab or tremelimumab.

6. The composition, antibody or method of any of claims 1 to 5, wherein the anti-PD-1 antibody is pembrolizumab, nivolumab or genolimu- zumab.

7. 7. The composition, antibody or method of any of claims 1-6, wherein the antibody comprises an ICOS binding site provided by a VH domain amino acid sequence that is at least 90% identical to SEQ ID NO: 408 and a VL domain amino acid sequence that is at least 90% identical to SEQ ID NO:

415.

8. 8. The composition, antibody or method of any of claims 1 to 7, wherein the antibody comprises an ICOS binding site provided by a VH domain amino acid sequence that is at least 95% identical to SEQ ID NO:

408.

9. the antibody comprises an ICOS binding site provided by a VH domain having HCDR1, HCDR2, and HCDR3 sequences; HCDR1 is SEQ ID NO:405, which optionally contains a conservative substitution at residue 28; SEQ ID NO: 1 in which HCDR2 may contain substitutions at residue 59, residue 63 and / or residue 64 406, HCDR3 may contain substitutions at residue 108, residue 109 and / or residue 112. Column number 407 A composition, antibody or method according to any one of claims 1 to 8.

10. 10. The composition, antibody or method of claim 9, wherein the substitution at residue 28 of HCDR1 is a conservative substitution, e.g., V28F.

11. the substitution of residue 59 of HCDR2 is N59I; the substitution at residue 63 of HCDR2 is G63D, and / or the substitution of residue 64 of HCDR2 is D64N; 11. The composition, antibody or method of claim 9 or claim 10.

12. the substitution at residue 108 of HCDR3 is F108Y; the substitution at residue 109 of HCDR3 is Y109F; and / or the substitution at residue 112 of HCDR3 is H112N; A composition, antibody or method according to any one of claims 9 to 11.

13. 13. The composition, antibody or method of any of claims 9 to 12, comprising a VH domain having an HCDR1 sequence, an HCDR2 sequence and an HCDR3 sequence, wherein HCDR1 is SEQ ID NO: 405, HCDR2 is SEQ ID NO: 406 and HCDR3 is SEQ ID NO:

407.

14. 14. The composition, antibody or method of any preceding claim, wherein the antibody comprises the VH domain amino acid sequence of SEQ ID NO:

408.

15. 15. The composition, antibody or method of any of claims 1 to 14, wherein the antibody comprises a VL domain amino acid sequence that is at least 95% identical to SEQ ID NO:

415.

16. the antibody comprises a VL domain having an LCDR1 sequence, an LCDR2 sequence, and an LCDR3 sequence; LCDR1 is SEQ ID NO:412, optionally comprising a substitution at residue 36; LCDR2 is SEQ ID NO: 413; LCDR3 is SEQ ID NO: 414, which may contain a substitution at residue 108 or residue 109; A composition, antibody or method according to any one of claims 1 to 15.

17. 17. The composition, antibody or method of claim 16, wherein the substitution at residue 36 of LCDR1 is R36S.

18. the substitution at residue 108 of said LCDR3 is D108G and / or 18. The composition, antibody or method of claim 16 or claim 17, wherein the substitution at 109 is M109N.

19. 19. The composition, antibody or method of any of claims 15 to 18, comprising a VL domain having LCDR1, LCDR2 and LCDR3 sequences, wherein LCDR1 is SEQ ID NO: 412, LCDR2 is SEQ ID NO: 413 and LCDR3 is SEQ ID NO:

414.

20. 20. The composition, antibody or method of any preceding claim, comprising the VL domain amino acid sequence of SEQ ID NO:415.

Images