Methods of using anti-IL-2 antibodies

Abstract

Described herein are methods of treatment, including dosage and administration regimens, using engineered anti-IL-2 antibodies. The engineered antibody can endow binding specificity to an IL-2-anti-IL-2 antibody compound modified receptor and inhibit binding of IL-2 and CD25. Engineered anti-IL-2 antibodies can promote expansion of effector immune cell subpopulations and reduce undesirable effects caused by IL-2. Methods of treatment using the engineered anti-IL-2 antibodies include the treatment of cancer, such as cancer manifested as solid tumors and metastases thereof.

Description

[0001] Sequence Listing Statement

[0002] This application contains a sequence listing, which has been submitted electronically in XML format and is hereby incorporated by reference in its entirety. The XML formatted sequence listing was created on November 10, 2022, is named P-621284-USP_10NOV22.XML, and is 97.7 kilobytes in size. Field of the Invention

[0003] The present disclosure relates generally to the field of antibodies. In one embodiment, the present disclosure describes the preparation and use of engineered anti-IL-2 antibodies that confer modified receptor binding specificity to IL-2. In certain embodiments, the present disclosure describes methods for treating cancer, including cancers that manifest as solid tumors.

[0004] background

[0005] Interleukin-2 (IL-2) is a 15.4 kDa type I cytokine with a four-helix bundle structure. Since its discovery over 30 years ago, the importance of IL-2 in regulating the immune system has been repeatedly demonstrated. IL-2 is primarily produced and secreted by antigen-activated CD4+ T cells. To a lesser extent, IL-2 is also produced by CD8+ T cells, natural killer (NK) cells, dendritic cells, and mast cells.

[0006] IL-2 signaling has two opposite effects. IL-2 can enhance the immune response and induce its proliferation by activating effector cells. In addition, IL-2 can downregulate the immune response by activating and proliferating CD4+ regulatory T (Treg) cells. In order to promote these functions, IL-2 mediates its effects by binding to the following two forms of IL-2 receptors: i) a trimeric receptor composed of the IL-2Rα (CD25) chain, the IL-2Rβ (CD122) chain, and the common IL-2Rγ (γc, CD132) chain, or ii) a dimeric receptor composed only of the IL-2Rβ subunit and the IL-2Rγ subunit. Both the dimeric and trimeric receptors are capable of transmitting IL-2 binding signaling via the STAT5 pathway. However, IL-2 binds to the αβγ receptor trimer 100 times more tightly than the βγ receptor dimer. It has been demonstrated that hIL-2 binds to the αβγ trimer with an affinity of approximately 10 pM, whereas hIL-2 binds to the βγ dimer with an affinity of 1 nM.

[0007] The difference in affinity with the dimer form and the trimer form of IL-2 receptor is one of the key mechanisms responsible for maintaining immune homeostasis in vivo. The activation of the trimeric receptor is related to the transcription mediated by FoxP3 in Treg, and Treg expresses more αβγ trimer IL-2 receptors on its membrane. In contrast, the combination of IL-2 and βγ dimers is related to the activation of NK cells and memory phenotype (MP) CD8+ cells, and the NK cells and memory phenotype (MP) CD8+ cells express relatively high levels of βγ dimers and very low levels of αβγ trimer IL-2 receptors. Due to the fact that under normal physiological conditions, the natural level of IL-2 is relatively low, the main function of IL-2 appears to be to promote immune tolerance by acting as Treg activation and proliferation factor. On the other hand, after the immune system is activated, IL-2 levels increase, and subsequently IL-2 can bind βγ dimers and promote activation and proliferation of memory phenotype effector T cells (MP) CD8+ and NK cells.

[0008] Since the early 1990s, high-dose IL-2 therapy has been used to treat melanoma and metastatic renal cell carcinoma, with a response rate of 10%-15%. Although effective, this approach has not been used for other cancers because IL-2-dependent adverse effects such as the potentially fatal vascular leak syndrome (VLS) make it impossible for many patients to consider receiving this therapy. The short half-life of the administered IL-2 requires very frequent administration, resulting in repeated surges in the levels of circulating IL-2, thereby exacerbating adverse effects. Finally, since wild-type IL-2 is not selective, it may also enhance the undesirable activation of Treg cells.

[0009] It has been found that certain antibodies can bind to IL-2 and regulate binding to the βγ dimer IL-2 receptor or the αβγ trimer IL-2 receptor. IL-2 complexed with these antibodies will have a relatively long half-life, and these IL-2 complexes will activate specific subsets of effector cells or immune cells. For example, the antibody S4B6-mouse IL-2 complex preferentially activates mouse effector cells in vivo, while the antibody JES6.1-mouse IL-2 complex preferentially activates mouse T regulatory cells in vivo. The regulatory mechanism of the JES-6.1 antibody has been elucidated. It has been shown that the JES6.1-mIL-2 complex binds to CD25 in vitro but does not bind to CD122.

[0010] Exogenous IL-2 therapy, even “non-α” therapies that do not bind to the CD25α subunit of the IL-2 trimeric receptor, leads to endogenous IL-2 production. Newly secreted endogenous IL-2 preferentially binds to the trimeric IL-2 receptor on Tregs, leading to the expansion of immunosuppressive Treg cells via a negative feedback loop.

[0011] Increased IL-2 has been suggested to play a role in viral infection. SARS-CoV-2 is a positive-strand RNA virus of the respiratory coronavirus family (coronaviridae). The virus enters the host by binding to angiotensin-converting enzyme 2 (ACE2) on lung and gastrointestinal tissues. The infectious process is characterized by an incubation period of ~7-14 days, followed by symptoms of dry cough, fever, and shortness of breath. Up to 20% of symptomatic individuals develop severe symptoms, and an average of 3% of cases are fatal due to lung failure. Earlier studies on members of the coronavirus family demonstrated that coronavirus infection leads to an increase in regulatory T lymphocytes, which may lead to delayed viral clearance. More recent studies of COVID-19 patients have shown that patients in the ICU have higher levels of IL-2, IL-7, IL-10, GSCF, IP10, MCP1, MIP1A, and TNF-α than non-ICU patients, suggesting a role for immunopathology in severe disease. Direct evidence for altered leukocyte homeostasis, derived from immunological profiles of peripheral blood leukocytes from patients infected with SARS-CoV2, suggests that, similar to some chronic infections, in COVID-19, impairment of CD4+ T cell function promotes overactivation and potentially subsequent exhaustion of CD8+ T cells. These perturbations in T cell subsets may ultimately attenuate host antiviral immunity. Therefore, therapeutic approaches that slow viral growth or enhance immune responses to eliminate viral load while reducing associated immunopathology would be highly beneficial.

[0012] The immune response to viruses is composed of the innate arm and the acquired arm of the immune system. The innate system uses toll-like receptors (TLRs) and retinoic acid-induced gene 1 (RIG-I) proteins to sense viral RNA / DNA and induce an initial response. This response includes the production of antiviral cytokines (such as interferon α), chemokines that bring the immune system to the site of infection, and mobilization of macrophage / dendritic cell arms. Natural killer (NK) cells (innate lymphoid) directly kill virus-infected cells in the absence of MHC class I expression. This occurs even when the virus has interfered with the MHC class I presentation system.

[0013] In addition, during the infection response, NK cells produce interferon-γ (IFN-γ), thereby increasing the expression of MHC class I on cells and enhancing the responsiveness of the acquired immune system. The acquired immune system is composed of T cells (CD4 and CD8) and B cells. CD4+ T cells recognize viral antigens in the context of MHC-II on antigen presenting cells, to amplify immune response (through cytokines) and induce B cell class switching and subsequently produce antiviral antibodies. CD4 cells, particularly the activation of Th1 cells, also release IFN-γ, thereby enhancing the presentation of viral antigens. CD8+ T cells show direct lysis to virus-infected cells presenting viral peptides in the context of MHC-I. The initial induction phase of the immune response typically requires 7-10 days to amplify T cell colonies and produce cells required for the removal of viruses.

[0014] IL-2 is a key mediator of the expansion and activation of T cells and NK cells. IL-2 is generally considered to play a major role in the secondary signals required for T cell activation. The expression of the dimeric (βγ) IL-2 receptor and the trimeric (αβγ) IL-2 receptor complex exhibits lineage selectivity, as the trimeric receptor containing CD25 (α subunit) is found to be highly expressed on regulatory T cells and activated short-lived cytotoxic effector T cell subsets, while the dimeric receptor is found to be highly expressed on naive T cells, memory T cells, and NK cells. Therefore, naive T cells, memory T cells, and NK cells can receive signaling via IL-2 binding to the dimeric receptor. Regulatory T cells rely on the high affinity trimeric receptor complex to enhance their function, which involves sequestering IL-2 from binding to memory T cells and naive T cells and thereby reducing the function of these cell populations. The mechanism of action of IL-2 is Figure 1 described in .

[0015] Effector T cell subsets also express trimeric IL-2 receptor complexes. Although these cells are highly active, the binding of IL-2 to effector T cell subsets induces activation-induced cell death (AICD). In addition, it has been shown that CD25 is expressed on the lung endothelium and vascular endothelium. This expression is correlated with pulmonary edema and vascular leakage in a mouse model using high-dose IL-2. It is shown that the expression of CD25 on lung cells is the cause of the pulmonary toxicity of high-dose IL-2 therapy. In addition, although lung endothelial cells express CD25 under steady-state conditions, the expression level of CD25 on these cells increases in vivo after mice are injected with IL-2. It has been shown that knocking out CD25 on non-immune cells or interfering with the CD25 binding epitope of IL-2 by using an immune complex of IL-2 and anti-IL-2 antibodies (IL-2 / mAb) can prevent IL-2-induced pulmonary edema and vascular leakage syndrome. It was also shown that in mice genetically modified to lack T and B cells and sublethally irradiated to remove remaining immune cells (NK, monocytes, DCs, and granulocytes), the addition of high-dose IL-2 resulted in significant pulmonary edema, indicative of a non-immune component.

[0016] Numerous studies have examined the dual role of IL-2 in clearing viral infections from the lungs. IL-2 has been shown to be essential for CD8+ T cell expansion for viral clearance. It has also been shown that IL-2 can mediate pulmonary edema. For example, in a mouse influenza model of influenza virus lung infection, memory CD4+ T cells have been shown to produce high levels of IL-2, and the presence of this IL-2 exacerbates the disease. Regulatory T cells are important for reducing the pathological damage to lung tissue caused by viral infection. It has been hypothesized and demonstrated that one mechanism by which Tregs control CD8+ effector cells is through high-affinity depletion of IL-2 via the CD25 trimeric receptor on Tregs. This effectively removes IL-2 from the expanded effector cells, subsequently limiting their availability and potentially reducing viral clearance. Tregs may also limit the effects of IL-2 on the lung endothelium by sequestering it from CD25+ endothelial cells. The outcome may depend on the Teff / Treg ratio. High levels of Teff (effector T cells) can lead to viral clearance, but can also result in excessive levels of IL-2 secreted by immune-activated cells, thus leading to pulmonary edema. In contrast, high Treg expansion can reduce the pathology of pulmonary edema, but also reduce viral clearance and lead to prolonged viral infection.

[0017] Recent data from COVID-19 patients suggest that higher viral loads lead to poorer outcomes; therefore, reduced viral clearance will be associated with worse outcomes. In a mouse model, an influenza A virus (IAV) infection model was used to demonstrate the role of Treg in reducing viral clearance, wherein mice infected with IAV showed higher levels of Treg in the lungs, spleen, and lymph nodes, as well as higher levels of viral load in lung tissue. This was observed even 6 weeks after the start of infection. This suggests that influenza A induces Treg expansion to avoid being cleared by the immune response. To evaluate whether enhancing the immune response will increase the clearance of IAV infection in the lungs, the researchers used mice previously infected with IAV and subsequently infected the mice with lymphocytic choriomeningitis virus (LCMV), which triggers a strong cytotoxic T lymphocyte response. The extensive immune response in the IAV-infected lungs also led to pulmonary edema and extensive lung tissue damage. Protection from severe pulmonary edema was achieved by treating mice carrying IAV with anti-CD25 blocking antibodies before administering LCMV. These data suggest that enhancing the immune response can induce viral clearance when Treg has already slowed viral clearance. Furthermore, it was shown that blocking IL-2 binding to CD25+ cells reduces the risk of immune-mediated pulmonary edema during viral clearance.

[0018] It has been shown that IL-2 given as a single-dose therapy enhances antiviral immune responses. The effect of IL-2 therapy during the expansion phase, contraction phase, and memory phase of T cells in mice infected with LCMV showed that IL-2 treatment during the expansion phase is not conducive to the survival of rapidly dividing effector T cells, which have transiently upregulated the expression of CD25. These effector T cells are then directed to AICD. In contrast, IL-2 therapy is highly beneficial during the contraction phase and leads to the survival and activation of virus-specific T cells. It has been observed that IL-2 treatment enhances the activation and proliferation of resting memory T cells. However, IL-2 therapy has its shortcomings. The half-life of IL-2 is short, so multiple administrations are required, such as daily loading doses followed by weekly administration, resulting in the risk of other related adverse events and increased immunogenicity. In addition, it is expected that the administration of exogenous high-dose IL-2 will bind it to CD25-positive endothelial cells. In fact, pulmonary edema and vascular leakage syndrome are the main serious adverse events of high-dose IL-2 therapy in oncology. Developing technologies that overcome these limitations is crucial for the use of IL-2 as a therapy.

[0019] One of ordinary skill in the art will recognize that the principles discussed above with respect to IL-2 and the treatment of viral infections will apply equally to IL-2 and the treatment of bacterial infections or the treatment of cancer.

[0020] Advances in biomolecular engineering have provided researchers with unprecedented opportunities to apply molecular design strategies to modify naturally occurring proteins and generate novel molecules for targeted disease therapies. On the one hand, evolving technologies and understanding of protein engineering have enabled the development of immunotherapeutics such as cytokine-based or antibody-based drugs. Consequently, there is a need for the development of engineered anti-IL-2 antibodies that can be used to modulate the function of IL-2 in certain disease states, such as, but not limited to, viral or bacterial infections and cancer.

[0021] Overview

[0022] In one aspect, disclosed herein is a method of treating cancer in a subject, comprising the step of administering to the subject a composition comprising an anti-IL-2 antibody comprising a heavy chain variable region (VH) and a light chain variable region (VL), wherein the VH comprises heavy chain complementarity determining regions (HCDRs) HCDR1, HCDR2, and HCDR3, and the VL comprises light chain complementarity determining regions (LCDRs) LCDR1, LCDR2, and LCDR3, wherein the CDRs have the following amino acid sequences:

[0023] (a) HCDR1 comprises the amino acid sequence of SEQ ID NO: 62, HCDR2 comprises the amino acid sequence of SEQ ID NO: 63, HCDR3 comprises the amino acid sequence of SEQ ID NO: 64, LCDR1 comprises the amino acid sequence of SEQ ID NO: 65, LCDR2 comprises the amino acid sequence of DAS, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 67;

[0024] (b) HCDR1 comprises the amino acid sequence of SEQ ID NO: 38, HCDR2 comprises the amino acid sequence of SEQ ID NO: 39, HCDR3 comprises the amino acid sequence of SEQ ID NO: 40, LCDR1 comprises the amino acid sequence of SEQ ID NO: 41, LCDR2 comprises the amino acid sequence of YAS, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 43;

[0025] (c) HCDR1 comprises the amino acid sequence of SEQ ID NO: 44, HCDR2 comprises the amino acid sequence of SEQ ID NO: 45, HCDR3 comprises the amino acid sequence of SEQ ID NO: 46, LCDR1 comprises the amino acid sequence of SEQ ID NO: 47, LCDR2 comprises the amino acid sequence of YAS, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 49;

[0026] (d) HCDR1 comprises the amino acid sequence of SEQ ID NO: 50, HCDR2 comprises the amino acid sequence of SEQ ID NO: 51, HCDR3 comprises the amino acid sequence of SEQ ID NO: 52, LCDR1 comprises the amino acid sequence of SEQ ID NO: 53, LCDR2 comprises the amino acid sequence of YAS, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 55; or

[0027] (e) HCDR1 comprises the amino acid sequence of SEQ ID NO: 56, HCDR2 comprises the amino acid sequence of SEQ ID NO: 57, HCDR3 comprises the amino acid sequence of SEQ ID NO: 58, LCDR1 comprises the amino acid sequence of SEQ ID NO: 59, LCDR2 comprises the amino acid sequence of YAS, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 61,

[0028] wherein the dosage of the anti-IL-2 antibody is between about 0.5 mg / kg and 12 mg / kg of the subject, and wherein the antibody promotes the differential growth of immune cell subsets and reduces undesirable effects caused by IL-2, thereby treating the cancer in the subject.

[0029] In another aspect, disclosed herein is a method of treating cancer in a subject, comprising administering to the subject an anti-IL-2 antibody and IL-2, wherein the IL-2 antibody comprises a heavy chain variable region (VH) and a light chain variable region (VL), wherein the VH comprises heavy chain complementarity determining regions (HCDRs) HCDR1, HCDR2, and HCDR3, and the VL comprises light chain complementarity determining regions (LCDRs) LCDR1, LCDR2, and LCDR3, wherein the CDRs have the following amino acid sequences:

[0030] (a) HCDR1 comprises the amino acid sequence of SEQ ID NO: 62, HCDR2 comprises the amino acid sequence of SEQ ID NO: 63, HCDR3 comprises the amino acid sequence of SEQ ID NO: 64, LCDR1 comprises the amino acid sequence of SEQ ID NO: 65, LCDR2 comprises the amino acid sequence of DAS, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 67;

[0031] (b) HCDR1 comprises the amino acid sequence of SEQ ID NO: 38, HCDR2 comprises the amino acid sequence of SEQ ID NO: 39, HCDR3 comprises the amino acid sequence of SEQ ID NO: 40, LCDR1 comprises the amino acid sequence of SEQ ID NO: 41, LCDR2 comprises the amino acid sequence of YAS, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 43;

[0032] (c) HCDR1 comprises the amino acid sequence of SEQ ID NO: 44, HCDR2 comprises the amino acid sequence of SEQ ID NO: 45, HCDR3 comprises the amino acid sequence of SEQ ID NO: 46, LCDR1 comprises the amino acid sequence of SEQ ID NO: 47, LCDR2 comprises the amino acid sequence of YAS, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 49;

[0033] (d) HCDR1 comprises the amino acid sequence of SEQ ID NO: 50, HCDR2 comprises the amino acid sequence of SEQ ID NO: 51, HCDR3 comprises the amino acid sequence of SEQ ID NO: 52, LCDR1 comprises the amino acid sequence of SEQ ID NO: 53, LCDR2 comprises the amino acid sequence of YAS, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 55; or

[0034] (e) HCDR1 comprises the amino acid sequence of SEQ ID NO: 56, HCDR2 comprises the amino acid sequence of SEQ ID NO: 57, HCDR3 comprises the amino acid sequence of SEQ ID NO: 58, LCDR1 comprises the amino acid sequence of SEQ ID NO: 59, LCDR2 comprises the amino acid sequence of YAS, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 61,

[0035] wherein the IL-2 is administered by subcutaneous injection, wherein the dose of the anti-IL-2 antibody is between about 0.5 mg / kg-12 mg / kg of the subject, and wherein the antibody promotes differential growth of immune cell subsets and reduces undesirable effects caused by IL-2, thereby treating the cancer in the subject.

[0036] In another aspect, disclosed herein is a method of treating cancer in a subject, comprising administering to the subject an anti-IL-2 antibody, IL-2, and a checkpoint inhibitor, wherein the IL-2 antibody comprises a heavy chain variable region (VH) and a light chain variable region (VL), wherein the VH comprises heavy chain complementarity determining regions (HCDRs) HCDR1, HCDR2, and HCDR3, and the VL comprises light chain complementarity determining regions (LCDRs) LCDR1, LCDR2, and LCDR3, wherein the CDRs have the following amino acid sequences:

[0037] (a) HCDR1 comprises the amino acid sequence of SEQ ID NO: 62, HCDR2 comprises the amino acid sequence of SEQ ID NO: 63, HCDR3 comprises the amino acid sequence of SEQ ID NO: 64, LCDR1 comprises the amino acid sequence of SEQ ID NO: 65, LCDR2 comprises the amino acid sequence of DAS, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 67;

[0038] (b) HCDR1 comprises the amino acid sequence of SEQ ID NO: 38, HCDR2 comprises the amino acid sequence of SEQ ID NO: 39, HCDR3 comprises the amino acid sequence of SEQ ID NO: 40, LCDR1 comprises the amino acid sequence of SEQ ID NO: 41, LCDR2 comprises the amino acid sequence of YAS, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 43;

[0039] (c) HCDR1 comprises the amino acid sequence of SEQ ID NO: 44, HCDR2 comprises the amino acid sequence of SEQ ID NO: 45, HCDR3 comprises the amino acid sequence of SEQ ID NO: 46, LCDR1 comprises the amino acid sequence of SEQ ID NO: 47, LCDR2 comprises the amino acid sequence of YAS, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 49;

[0040] (d) HCDR1 comprises the amino acid sequence of SEQ ID NO: 50, HCDR2 comprises the amino acid sequence of SEQ ID NO: 51, HCDR3 comprises the amino acid sequence of SEQ ID NO: 52, LCDR1 comprises the amino acid sequence of SEQ ID NO: 53, LCDR2 comprises the amino acid sequence of YAS, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 55; or

[0041] (e) HCDR1 comprises the amino acid sequence of SEQ ID NO: 56, HCDR2 comprises the amino acid sequence of SEQ ID NO: 57, HCDR3 comprises the amino acid sequence of SEQ ID NO: 58, LCDR1 comprises the amino acid sequence of SEQ ID NO: 59, LCDR2 comprises the amino acid sequence of YAS, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 61,

[0042] wherein the IL-2 is administered by subcutaneous injection, wherein the dose of the anti-IL-2 antibody is between about 0.5 mg / kg-12 mg / kg of the subject, wherein the antibody promotes differential growth of immune cell subsets and reduces undesirable effects caused by IL-2, and wherein the checkpoint inhibitor comprises PD-1, PD-L1, CTLA-4, TIGIT, TIM-3, B7-H3, CD73, LAG3, CD27, CD70, 4-1BB, GITR, OX40, SIRP-α (CD47), CD39, ILDR2, VISTA, BTLA or VTCN-1, thereby treating the cancer in the subject.

[0043] In another aspect, disclosed herein is a method of treating a solid cancer in a subject, comprising administering to the subject a composition comprising an anti-IL-2 antibody comprising a heavy chain variable region (VH) and a light chain variable region (VL), wherein the VH comprises heavy chain complementarity determining regions (HCDRs) HCDR1, HCDR2, and HCDR3, and the VL comprises light chain complementarity determining regions (LCDRs) LCDR1, LCDR2, and LCDR3, wherein the CDRs have the following amino acid sequences:

[0044] (a) HCDR1 comprises the amino acid sequence of SEQ ID NO: 62, HCDR2 comprises the amino acid sequence of SEQ ID NO: 63, HCDR3 comprises the amino acid sequence of SEQ ID NO: 64, LCDR1 comprises the amino acid sequence of SEQ ID NO: 65, LCDR2 comprises the amino acid sequence of DAS, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 67;

[0045] (b) HCDR1 comprises the amino acid sequence of SEQ ID NO: 38, HCDR2 comprises the amino acid sequence of SEQ ID NO: 39, HCDR3 comprises the amino acid sequence of SEQ ID NO: 40, LCDR1 comprises the amino acid sequence of SEQ ID NO: 41, LCDR2 comprises the amino acid sequence of YAS, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 43;

[0046] (c) HCDR1 comprises the amino acid sequence of SEQ ID NO: 44, HCDR2 comprises the amino acid sequence of SEQ ID NO: 45, HCDR3 comprises the amino acid sequence of SEQ ID NO: 46, LCDR1 comprises the amino acid sequence of SEQ ID NO: 47, LCDR2 comprises the amino acid sequence of YAS, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 49;

[0047] (d) HCDR1 comprises the amino acid sequence of SEQ ID NO: 50, HCDR2 comprises the amino acid sequence of SEQ ID NO: 51, HCDR3 comprises the amino acid sequence of SEQ ID NO: 52, LCDR1 comprises the amino acid sequence of SEQ ID NO: 53, LCDR2 comprises the amino acid sequence of YAS, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 55; or

[0048] (e) HCDR1 comprises the amino acid sequence of SEQ ID NO: 56, HCDR2 comprises the amino acid sequence of SEQ ID NO: 57, HCDR3 comprises the amino acid sequence of SEQ ID NO: 58, LCDR1 comprises the amino acid sequence of SEQ ID NO: 59, LCDR2 comprises the amino acid sequence of YAS, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 61,

[0049] wherein the dosage of the anti-IL-2 antibody is between about 0.5 mg / kg and 12 mg / kg of the subject, and wherein the antibody promotes the differential growth of immune cell subsets and reduces undesirable effects caused by IL-2, thereby treating the cancer in the subject.

[0050] In related aspects of the methods disclosed herein, multiple doses of a composition comprising an anti-IL2 antibody are administered.

[0051] In another related aspect of the methods disclosed herein, the methods further comprise administering at least a single low dose of IL-2, wherein the low dose of IL-2 comprises about 15×10 3 IU / Kg-500×10 3 In another related aspect, the administration of IL-2 comprises subcutaneous administration. In another related aspect, the IL-2 is administered before, simultaneously with, or after the administration of the anti-IL-2 antibody. In another further related aspect, the IL-2 is administered in multiple doses. In another further related aspect, multiple doses of IL-2 are administered before, simultaneously with, or after the administration of the anti-IL-2 antibody, or any combination thereof.

[0052] In another related aspect of the method disclosed herein, the method further comprises administering a checkpoint inhibitor. In another related aspect, the checkpoint inhibitor includes PD-L1, PD-1, CTLA-4, TIGIT, TIM-3, B7-H3, CD73, LAG3, CD27, CD70, 4-1BB, GITR, OX40, SIRP-α (CD47), CD39, ILDR2, VISTA, BTLA or VTCN-1.

[0053] In another related aspect of the methods disclosed herein, solid cancers include melanoma, metastatic melanoma, a primary melanoma and metastatic melanoma, renal cell carcinoma (RCC), non-small cell lung cancer (NSCLC), head and neck cancer, head and neck squamous cell carcinoma (HNSCC), pancreatic cancer, lung cancer, thyroid cancer, bladder cancer, nasopharyngeal cancer, colorectal cancer (CRC), cholangiocarcinoma (bile duct cancer), uterine cancer, cervical cancer, gallbladder cancer, squamous cell carcinoma of the skin, etc.

[0054] In yet another related aspect of the methods disclosed herein, the method comprises second-line therapy or third-line therapy, or a combination thereof.

[0055] In another related aspect of the methods disclosed herein, treating the subject reduces the size of the tumor, inhibits or reduces the growth of the tumor, or inhibits or reduces metastasis of the tumor, or any combination thereof.

[0056] In yet another related aspect of the methods disclosed herein, VH and VL have the amino acid sequences

[0057] (a) VH comprises the amino acid sequence of SEQ ID NO: 26, and VL comprises the amino acid sequence of SEQ ID NO: 27;

[0058] (b) VH comprises the amino acid sequence of SEQ ID NO: 20, and VL comprises the amino acid sequence of SEQ ID NO: 21;

[0059] (c) VH comprises the amino acid sequence of SEQ ID NO: 22, and VL comprises the amino acid sequence of SEQ ID NO: 23;

[0060] (d) VH comprises the amino acid sequence of SEQ ID NO: 24, and VL comprises the amino acid sequence of SEQ ID NO: 25; or

[0061] (e) VH comprises the amino acid sequence of SEQ ID NO: 36, and VL comprises the amino acid sequence of SEQ ID NO: 37.

[0062] In another related aspect, the antibody comprises an IgG, IgA, IgM, IgE, IgD, Fv, scFv, Fab, F(ab')2, minibody, diabody, or triabody. In yet another related aspect, the antibody comprises a heavy chain comprising a mutation that reduces binding to an Fcγ receptor. In yet another related aspect, the mutation comprises an L234A, L235A mutation. In yet another related aspect,

[0063] (a) when HCDR1 comprises the amino acid sequence of SEQ ID NO: 62, HCDR2 comprises the amino acid sequence of SEQ ID NO: 63, HCDR3 comprises the amino acid sequence of SEQ ID NO: 64, LCDR1 comprises the amino acid sequence of SEQ ID NO: 65, LCDR2 comprises the amino acid sequence of DAS, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 67, the amino acid sequence of the full-length heavy chain is set forth in SEQ ID NO: 72, and the amino acid sequence of the full-length light chain is set forth in SEQ ID NO: 73;

[0064] (b) when HCDR1 comprises the amino acid sequence of SEQ ID NO:44, HCDR2 comprises the amino acid sequence of SEQ ID NO:45, HCDR3 comprises the amino acid sequence of SEQ ID NO:46, LCDR1 comprises the amino acid sequence of SEQ ID NO:47, LCDR2 comprises the amino acid sequence of YAS, and LCDR3 comprises the amino acid sequence of SEQ ID NO:49, the amino acid sequence of the full-length heavy chain is set forth in SEQ ID NO:68 and the amino acid sequence of the full-length light chain is set forth in SEQ ID NO:69; and

[0065] (c) when HCDR1 comprises the amino acid sequence of SEQ ID NO: 50, HCDR2 comprises the amino acid sequence of SEQ ID NO: 51, HCDR3 comprises the amino acid sequence of SEQ ID NO: 52, LCDR1 comprises the amino acid sequence of SEQ ID NO: 53, LCDR2 comprises the amino acid sequence of YAS, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 55, the amino acid sequence of the full-length heavy chain is set forth in SEQ ID NO: 70, and the amino acid sequence of the full-length light chain is set forth in SEQ ID NO: 71.

[0066] In another aspect, disclosed herein is a method of treating cancer in a subject, comprising administering to the subject an anti-IL-2 antibody and IL-2, wherein the IL-2 antibody comprises a heavy chain variable region (VH) and a light chain variable region (VL), wherein the VH comprises heavy chain complementarity determining regions (HCDRs) HCDR1, HCDR2, and HCDR3, and the VL comprises light chain complementarity determining regions (LCDRs) LCDR1, LCDR2, and LCDR3, wherein the CDRs have the following amino acid sequences:

[0067] (a) HCDR1 comprises the amino acid sequence of SEQ ID NO: 62, HCDR2 comprises the amino acid sequence of SEQ ID NO: 63, HCDR3 comprises the amino acid sequence of SEQ ID NO: 64, LCDR1 comprises the amino acid sequence of SEQ ID NO: 65, LCDR2 comprises the amino acid sequence of DAS, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 67;

[0068] (b) HCDR1 comprises the amino acid sequence of SEQ ID NO: 38, HCDR2 comprises the amino acid sequence of SEQ ID NO: 39, HCDR3 comprises the amino acid sequence of SEQ ID NO: 40, LCDR1 comprises the amino acid sequence of SEQ ID NO: 41, LCDR2 comprises the amino acid sequence of YAS, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 43;

[0069] (c) HCDR1 comprises the amino acid sequence of SEQ ID NO: 44, HCDR2 comprises the amino acid sequence of SEQ ID NO: 45, HCDR3 comprises the amino acid sequence of SEQ ID NO: 46, LCDR1 comprises the amino acid sequence of SEQ ID NO: 47, LCDR2 comprises the amino acid sequence of YAS, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 49;

[0070] (d) HCDR1 comprises the amino acid sequence of SEQ ID NO: 50, HCDR2 comprises the amino acid sequence of SEQ ID NO: 51, HCDR3 comprises the amino acid sequence of SEQ ID NO: 52, LCDR1 comprises the amino acid sequence of SEQ ID NO: 53, LCDR2 comprises the amino acid sequence of YAS, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 55; or

[0071] (e) HCDR1 comprises the amino acid sequence of SEQ ID NO: 56, HCDR2 comprises the amino acid sequence of SEQ ID NO: 57, HCDR3 comprises the amino acid sequence of SEQ ID NO: 58, LCDR1 comprises the amino acid sequence of SEQ ID NO: 59, LCDR2 comprises the amino acid sequence of YAS, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 61,

[0072] wherein the IL-2 is administered by subcutaneous injection, wherein the dose of the anti-IL-2 antibody is between about 0.5 mg / kg-12 mg / kg of the subject, and wherein the antibody promotes differential growth of immune cell subsets and reduces undesirable effects caused by IL-2, thereby treating the cancer in the subject.

[0073] In yet another aspect, disclosed herein is a method of treating a solid cancer in a subject, comprising administering to the subject an anti-IL-2 antibody, IL-2, and a checkpoint inhibitor, wherein the IL-2 antibody comprises a heavy chain variable region (VH) and a light chain variable region (VL), wherein the VH comprises heavy chain complementarity determining regions (HCDRs) HCDR1, HCDR2, and HCDR3, and the VL comprises light chain complementarity determining regions (LCDRs) LCDR1, LCDR2, and LCDR3, wherein the CDRs have the following amino acid sequences:

[0074] (a) HCDR1 comprises the amino acid sequence of SEQ ID NO: 62, HCDR2 comprises the amino acid sequence of SEQ ID NO: 63, HCDR3 comprises the amino acid sequence of SEQ ID NO: 64, LCDR1 comprises the amino acid sequence of SEQ ID NO: 65, LCDR2 comprises the amino acid sequence of DAS, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 67;

[0075] (b) HCDR1 comprises the amino acid sequence of SEQ ID NO: 38, HCDR2 comprises the amino acid sequence of SEQ ID NO: 39, HCDR3 comprises the amino acid sequence of SEQ ID NO: 40, LCDR1 comprises the amino acid sequence of SEQ ID NO: 41, LCDR2 comprises the amino acid sequence of YAS, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 43;

[0076] (c) HCDR1 comprises the amino acid sequence of SEQ ID NO: 44, HCDR2 comprises the amino acid sequence of SEQ ID NO: 45, HCDR3 comprises the amino acid sequence of SEQ ID NO: 46, LCDR1 comprises the amino acid sequence of SEQ ID NO: 47, LCDR2 comprises the amino acid sequence of YAS, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 49;

[0077] (d) HCDR1 comprises the amino acid sequence of SEQ ID NO: 50, HCDR2 comprises the amino acid sequence of SEQ ID NO: 51, HCDR3 comprises the amino acid sequence of SEQ ID NO: 52, LCDR1 comprises the amino acid sequence of SEQ ID NO: 53, LCDR2 comprises the amino acid sequence of YAS, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 55; or

[0078] (e) HCDR1 comprises the amino acid sequence of SEQ ID NO: 56, HCDR2 comprises the amino acid sequence of SEQ ID NO: 57, HCDR3 comprises the amino acid sequence of SEQ ID NO: 58, LCDR1 comprises the amino acid sequence of SEQ ID NO: 59, LCDR2 comprises the amino acid sequence of YAS, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 61,

[0079] Wherein the IL-2 is administered by subcutaneous injection, wherein the dose of the anti-IL-2 antibody is between about 0.5 mg / kg-12 mg / kg of the subject, wherein the antibody promotes the differential growth of immune cell subsets and reduces the undesirable effects caused by IL-2, and wherein the checkpoint inhibitor includes PD-L1, PD-1, CTLA-4, TIGIT, TIM-3, B7-H3, CD73, LAG3, CD27, CD70, 4-1BB, GITR, OX40, SIRP-α (CD47), CD39, ILDR2, VISTA, BTLA or VTCN-1, thereby treating the cancer in the subject. In related aspects, the checkpoint inhibitor includes a PD-L1 checkpoint inhibitor.

[0080] In another aspect, disclosed herein is a method of treating non-small cell lung cancer (NSCLC) in a subject, comprising administering to the subject a composition comprising an anti-IL-2 antibody comprising a heavy chain variable region (VH) and a light chain variable region (VL), wherein the VH comprises heavy chain complementarity determining regions (HCDRs) HCDR1, HCDR2, and HCDR3, and the VL comprises light chain complementarity determining regions (LCDRs) LCDR1, LCDR2, and LCDR3, wherein the CDRs have the following amino acid sequences:

[0081] (a) HCDR1 comprises the amino acid sequence of SEQ ID NO: 62, HCDR2 comprises the amino acid sequence of SEQ ID NO: 63, HCDR3 comprises the amino acid sequence of SEQ ID NO: 64, LCDR1 comprises the amino acid sequence of SEQ ID NO: 65, LCDR2 comprises the amino acid sequence of DAS, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 67;

[0082] (b) HCDR1 comprises the amino acid sequence of SEQ ID NO: 38, HCDR2 comprises the amino acid sequence of SEQ ID NO: 39, HCDR3 comprises the amino acid sequence of SEQ ID NO: 40, LCDR1 comprises the amino acid sequence of SEQ ID NO: 41, LCDR2 comprises the amino acid sequence of YAS, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 43;

[0083] (c) HCDR1 comprises the amino acid sequence of SEQ ID NO: 44, HCDR2 comprises the amino acid sequence of SEQ ID NO: 45, HCDR3 comprises the amino acid sequence of SEQ ID NO: 46, LCDR1 comprises the amino acid sequence of SEQ ID NO: 47, LCDR2 comprises the amino acid sequence of YAS, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 49;

[0084] (d) HCDR1 comprises the amino acid sequence of SEQ ID NO: 50, HCDR2 comprises the amino acid sequence of SEQ ID NO: 51, HCDR3 comprises the amino acid sequence of SEQ ID NO: 52, LCDR1 comprises the amino acid sequence of SEQ ID NO: 53, LCDR2 comprises the amino acid sequence of YAS, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 55; or

[0085] (e) HCDR1 comprises the amino acid sequence of SEQ ID NO: 56, HCDR2 comprises the amino acid sequence of SEQ ID NO: 57, HCDR3 comprises the amino acid sequence of SEQ ID NO: 58, LCDR1 comprises the amino acid sequence of SEQ ID NO: 59, LCDR2 comprises the amino acid sequence of YAS, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 61,

[0086] wherein the dosage of the anti-IL-2 antibody is between about 0.5 mg / kg and 12 mg / kg of the subject, and wherein the antibody promotes the differential growth of immune cell subsets and reduces undesirable effects caused by IL-2, thereby treating the NSCLC in the subject.

[0087] In another aspect, disclosed herein is a method of treating non-small cell lung cancer (NSCLC) in a subject, comprising administering to the subject an anti-IL-2 antibody and IL-2, wherein the IL-2 antibody comprises a heavy chain variable region (VH) and a light chain variable region (VL), wherein the VH comprises heavy chain complementarity determining regions (HCDRs) HCDR1, HCDR2, and HCDR3, and the VL comprises light chain complementarity determining regions (LCDRs) LCDR1, LCDR2, and LCDR3, wherein the CDRs have the following amino acid sequences:

[0088] (a) HCDR1 comprises the amino acid sequence of SEQ ID NO: 62, HCDR2 comprises the amino acid sequence of SEQ ID NO: 63, HCDR3 comprises the amino acid sequence of SEQ ID NO: 64, LCDR1 comprises the amino acid sequence of SEQ ID NO: 65, LCDR2 comprises the amino acid sequence of DAS, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 67;

[0089] (b) HCDR1 comprises the amino acid sequence of SEQ ID NO: 38, HCDR2 comprises the amino acid sequence of SEQ ID NO: 39, HCDR3 comprises the amino acid sequence of SEQ ID NO: 40, LCDR1 comprises the amino acid sequence of SEQ ID NO: 41, LCDR2 comprises the amino acid sequence of YAS, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 43;

[0090] (c) HCDR1 comprises the amino acid sequence of SEQ ID NO: 44, HCDR2 comprises the amino acid sequence of SEQ ID NO: 45, HCDR3 comprises the amino acid sequence of SEQ ID NO: 46, LCDR1 comprises the amino acid sequence of SEQ ID NO: 47, LCDR2 comprises the amino acid sequence of YAS, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 49;

[0091] (d) HCDR1 comprises the amino acid sequence of SEQ ID NO: 50, HCDR2 comprises the amino acid sequence of SEQ ID NO: 51, HCDR3 comprises the amino acid sequence of SEQ ID NO: 52, LCDR1 comprises the amino acid sequence of SEQ ID NO: 53, LCDR2 comprises the amino acid sequence of YAS, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 55; or

[0092] (e) HCDR1 comprises the amino acid sequence of SEQ ID NO: 56, HCDR2 comprises the amino acid sequence of SEQ ID NO: 57, HCDR3 comprises the amino acid sequence of SEQ ID NO: 58, LCDR1 comprises the amino acid sequence of SEQ ID NO: 59, LCDR2 comprises the amino acid sequence of YAS, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 61,

[0093] wherein the dose of the anti-IL-2 antibody is between about 0.5 mg / kg-12 mg / kg of the subject, and wherein the antibody promotes differential growth of immune cell subsets and reduces undesirable effects caused by IL-2, and wherein the IL-2 is administered subcutaneously, thereby treating the NSCLC in the subject.

[0094] In another aspect, disclosed herein is a method of treating non-small cell lung cancer (NSCLC) in a subject, comprising administering to the subject an anti-IL-2 antibody, IL-2, and a checkpoint inhibitor, wherein the IL-2 antibody comprises a heavy chain variable region (VH) and a light chain variable region (VL), wherein the VH comprises heavy chain complementarity determining regions (HCDRs) HCDR1, HCDR2, and HCDR3, and the VL comprises light chain complementarity determining regions (LCDRs) LCDR1, LCDR2, and LCDR3, wherein the CDRs have the following amino acid sequences:

[0095] (a) HCDR1 comprises the amino acid sequence of SEQ ID NO: 62, HCDR2 comprises the amino acid sequence of SEQ ID NO: 63, HCDR3 comprises the amino acid sequence of SEQ ID NO: 64, LCDR1 comprises the amino acid sequence of SEQ ID NO: 65, LCDR2 comprises the amino acid sequence of DAS, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 67;

[0096] (b) HCDR1 comprises the amino acid sequence of SEQ ID NO: 38, HCDR2 comprises the amino acid sequence of SEQ ID NO: 39, HCDR3 comprises the amino acid sequence of SEQ ID NO: 40, LCDR1 comprises the amino acid sequence of SEQ ID NO: 41, LCDR2 comprises the amino acid sequence of YAS, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 43;

[0097] (c) HCDR1 comprises the amino acid sequence of SEQ ID NO: 44, HCDR2 comprises the amino acid sequence of SEQ ID NO: 45, HCDR3 comprises the amino acid sequence of SEQ ID NO: 46, LCDR1 comprises the amino acid sequence of SEQ ID NO: 47, LCDR2 comprises the amino acid sequence of YAS, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 49;

[0098] (d) HCDR1 comprises the amino acid sequence of SEQ ID NO: 50, HCDR2 comprises the amino acid sequence of SEQ ID NO: 51, HCDR3 comprises the amino acid sequence of SEQ ID NO: 52, LCDR1 comprises the amino acid sequence of SEQ ID NO: 53, LCDR2 comprises the amino acid sequence of YAS, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 55; or

[0099] (e) HCDR1 comprises the amino acid sequence of SEQ ID NO: 56, HCDR2 comprises the amino acid sequence of SEQ ID NO: 57, HCDR3 comprises the amino acid sequence of SEQ ID NO: 58, LCDR1 comprises the amino acid sequence of SEQ ID NO: 59, LCDR2 comprises the amino acid sequence of YAS, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 61,

[0100] wherein the dose of the anti-IL-2 antibody is between about 0.5 mg / kg-12 mg / kg of the subject, and wherein the antibody promotes differential growth of immune cell subsets and reduces undesirable effects caused by IL-2, wherein the IL-2 is administered subcutaneously, and wherein the checkpoint inhibitor comprises PD-1, PD-L1, CTLA-4, TIGIT, TIM-3, B7-H3, CD73, LAG3, CD27, CD70, 4-1BB, GITR, OX40, SIRP-α (CD47), CD39, ILDR2, VISTA, BTLA or VTCN-1, thereby treating the NSCLC in the subject.

[0101] In another aspect, disclosed herein is a method of treating renal cell carcinoma (RCC) in a subject, comprising administering to the subject an anti-IL-2 antibody comprising a heavy chain variable region (VH) and a light chain variable region (VL), wherein the VH comprises heavy chain complementarity determining regions (HCDRs) HCDR1, HCDR2, and HCDR3, and the VL comprises light chain complementarity determining regions (LCDRs) LCDR1, LCDR2, and LCDR3, wherein the CDRs have the following amino acid sequences:

[0102] (a) HCDR1 comprises the amino acid sequence of SEQ ID NO: 62, HCDR2 comprises the amino acid sequence of SEQ ID NO: 63, HCDR3 comprises the amino acid sequence of SEQ ID NO: 64, LCDR1 comprises the amino acid sequence of SEQ ID NO: 65, LCDR2 comprises the amino acid sequence of DAS, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 67;

[0103] (b) HCDR1 comprises the amino acid sequence of SEQ ID NO: 38, HCDR2 comprises the amino acid sequence of SEQ ID NO: 39, HCDR3 comprises the amino acid sequence of SEQ ID NO: 40, LCDR1 comprises the amino acid sequence of SEQ ID NO: 41, LCDR2 comprises the amino acid sequence of YAS, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 43;

[0104] (c) HCDR1 comprises the amino acid sequence of SEQ ID NO: 44, HCDR2 comprises the amino acid sequence of SEQ ID NO: 45, HCDR3 comprises the amino acid sequence of SEQ ID NO: 46, LCDR1 comprises the amino acid sequence of SEQ ID NO: 47, LCDR2 comprises the amino acid sequence of YAS, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 49;

[0105] (d) HCDR1 comprises the amino acid sequence of SEQ ID NO: 50, HCDR2 comprises the amino acid sequence of SEQ ID NO: 51, HCDR3 comprises the amino acid sequence of SEQ ID NO: 52, LCDR1 comprises the amino acid sequence of SEQ ID NO: 53, LCDR2 comprises the amino acid sequence of YAS, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 55; or

[0106] (e) HCDR1 comprises the amino acid sequence of SEQ ID NO: 56, HCDR2 comprises the amino acid sequence of SEQ ID NO: 57, HCDR3 comprises the amino acid sequence of SEQ ID NO: 58, LCDR1 comprises the amino acid sequence of SEQ ID NO: 59, LCDR2 comprises the amino acid sequence of YAS, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 61,

[0107] wherein the dose of the anti-IL-2 antibody is between about 0.5 mg / kg and 12 mg / kg of the subject, and wherein the antibody promotes the differential growth of immune cell subsets and reduces undesirable effects caused by IL-2, thereby treating the RCC in the subject.

[0108] In another aspect, disclosed herein is a method of treating melanoma in a subject, comprising administering to the subject a composition comprising an anti-IL-2 antibody comprising a heavy chain variable region (VH) and a light chain variable region (VL), wherein the VH comprises heavy chain complementarity determining regions (HCDRs) HCDR1, HCDR2, and HCDR3, and the VL comprises light chain complementarity determining regions (LCDRs) LCDR1, LCDR2, and LCDR3, wherein the CDRs have the following amino acid sequences:

[0109] (a) HCDR1 comprises the amino acid sequence of SEQ ID NO: 62, HCDR2 comprises the amino acid sequence of SEQ ID NO: 63, HCDR3 comprises the amino acid sequence of SEQ ID NO: 64, LCDR1 comprises the amino acid sequence of SEQ ID NO: 65, LCDR2 comprises the amino acid sequence of DAS, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 67;

[0110] (b) HCDR1 comprises the amino acid sequence of SEQ ID NO: 38, HCDR2 comprises the amino acid sequence of SEQ ID NO: 39, HCDR3 comprises the amino acid sequence of SEQ ID NO: 40, LCDR1 comprises the amino acid sequence of SEQ ID NO: 41, LCDR2 comprises the amino acid sequence of YAS, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 43;

[0111] (c) HCDR1 comprises the amino acid sequence of SEQ ID NO: 44, HCDR2 comprises the amino acid sequence of SEQ ID NO: 45, HCDR3 comprises the amino acid sequence of SEQ ID NO: 46, LCDR1 comprises the amino acid sequence of SEQ ID NO: 47, LCDR2 comprises the amino acid sequence of YAS, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 49;

[0112] (d) HCDR1 comprises the amino acid sequence of SEQ ID NO: 50, HCDR2 comprises the amino acid sequence of SEQ ID NO: 51, HCDR3 comprises the amino acid sequence of SEQ ID NO: 52, LCDR1 comprises the amino acid sequence of SEQ ID NO: 53, LCDR2 comprises the amino acid sequence of YAS, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 55; or

[0113] (e) HCDR1 comprises the amino acid sequence of SEQ ID NO: 56, HCDR2 comprises the amino acid sequence of SEQ ID NO: 57, HCDR3 comprises the amino acid sequence of SEQ ID NO: 58, LCDR1 comprises the amino acid sequence of SEQ ID NO: 59, LCDR2 comprises the amino acid sequence of YAS, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 61,

[0114] wherein the melanoma comprises a primary tumor or a metastatic melanoma, or a combination thereof, wherein the dose of the anti-IL-2 antibody is between about 0.5 mg / kg and 12 mg / kg of the subject, and wherein the antibody promotes differential growth of immune cell subsets and reduces undesirable effects caused by IL-2, thereby treating the melanoma in the subject.

[0115] In another aspect, disclosed herein is a method of treating cancer in a subject, comprising administering to the subject an anti-IL-2 antibody, IL-2, and a checkpoint inhibitor, wherein the IL-2 antibody comprises a heavy chain variable region (VH) and a light chain variable region (VL), wherein the VH comprises heavy chain complementarity determining regions (HCDRs) HCDR1, HCDR2, and HCDR3, and the VL comprises light chain complementarity determining regions (LCDRs) LCDR1, LCDR2, and LCDR3, wherein the CDRs have the following amino acid sequences:

[0116] (a) HCDR1 comprises the amino acid sequence of SEQ ID NO: 62, HCDR2 comprises the amino acid sequence of SEQ ID NO: 63, HCDR3 comprises the amino acid sequence of SEQ ID NO: 64, LCDR1 comprises the amino acid sequence of SEQ ID NO: 65, LCDR2 comprises the amino acid sequence of DAS, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 67;

[0117] (b) HCDR1 comprises the amino acid sequence of SEQ ID NO: 38, HCDR2 comprises the amino acid sequence of SEQ ID NO: 39, HCDR3 comprises the amino acid sequence of SEQ ID NO: 40, LCDR1 comprises the amino acid sequence of SEQ ID NO: 41, LCDR2 comprises the amino acid sequence of YAS, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 43;

[0118] (c) HCDR1 comprises the amino acid sequence of SEQ ID NO: 44, HCDR2 comprises the amino acid sequence of SEQ ID NO: 45, HCDR3 comprises the amino acid sequence of SEQ ID NO: 46, LCDR1 comprises the amino acid sequence of SEQ ID NO: 47, LCDR2 comprises the amino acid sequence of YAS, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 49;

[0119] (d) HCDR1 comprises the amino acid sequence of SEQ ID NO: 50, HCDR2 comprises the amino acid sequence of SEQ ID NO: 51, HCDR3 comprises the amino acid sequence of SEQ ID NO: 52, LCDR1 comprises the amino acid sequence of SEQ ID NO: 53, LCDR2 comprises the amino acid sequence of YAS, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 55; or

[0120] (e) HCDR1 comprises the amino acid sequence of SEQ ID NO: 56, HCDR2 comprises the amino acid sequence of SEQ ID NO: 57, HCDR3 comprises the amino acid sequence of SEQ ID NO: 58, LCDR1 comprises the amino acid sequence of SEQ ID NO: 59, LCDR2 comprises the amino acid sequence of YAS, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 61,

[0121] wherein the dose of the anti-IL-2 antibody is between about 0.5 mg / kg-12 mg / kg of the subject, wherein the dose of IL-2 is a low dose, wherein the checkpoint inhibitor is a PD-L1 checkpoint inhibitor, and wherein the antibody promotes differential growth of immune cell subsets and reduces undesirable effects caused by IL-2, thereby treating the cancer in the subject.

[0122] In another related aspect, the method further comprises administering IL-2. In another related aspect, the administration of IL-2 comprises subcutaneous administration. In another related aspect, IL-2 is administered in a single dose. In yet another related aspect, IL-2 is administered before, simultaneously with, or after the administration of the anti-IL-2 antibody. In yet another related aspect, IL-2 is administered in multiple doses. In yet another related aspect, multiple doses of IL-2 are administered before, simultaneously with, or after the administration of the anti-IL-2 antibody, or any combination thereof. In yet another related aspect, the dose of IL-2 is about 15×10 3 IU / Kg to 270×10 3 IU / Kg between the subjects.

[0123] In another related aspect, the method further comprises administering a checkpoint inhibitor. In another related aspect, the method further comprises administering IL-2 and a checkpoint inhibitor.

[0124] The method of claim 10, wherein the checkpoint inhibitor comprises PD-L1, PD-1, CTLA-4, TIGIT, TIM-3, B7-H3, CD73, LAG3, CD27, CD70, 4-1BB, GITR, OX40, SIRP-α (CD47), CD39, ILDR2, VISTA, BTLA or VTCN-1. In another related aspect, the checkpoint is PD-L1. In yet another related aspect, the PD-L1 checkpoint inhibitor is avelumab.

[0125] In another related aspect, cancer includes solid cancer.In another related aspect, solid cancer includes non-small cell lung cancer (NSCLC), head and neck cancer, head and neck squamous cell carcinoma (HNSCC), pancreatic cancer, lung cancer, thyroid cancer, bladder cancer, nasopharyngeal carcinoma, melanoma, metastatic melanoma, primary melanoma and metastatic melanoma, colorectal cancer (CRC), bladder cancer, bile duct cancer (bile duct cancer), uterine cancer, cervical cancer, gallbladder cancer or renal cell carcinoma (RCC).In another other related aspect, solid cancer includes non-small cell lung cancer (NSCLC), melanoma, metastatic melanoma, primary melanoma and metastatic melanoma or renal cell carcinoma (RCC).In another other related aspect, solid cancer includes non-small cell lung cancer (NSCLC).In another other related aspect, non-small cell lung cancer (NSCLC) includes unresectable advanced cancer or metastatic cancer.

[0126] In related aspects, the method comprises second-line treatment or third-line treatment, or a combination thereof.

[0127] In another related aspect, administration of the anti-IL2 antibody continues for between about 3 months and 1 year.

[0128] In another related aspect, the method of treating the subject reduces the size of a tumor, inhibits or reduces tumor growth, or inhibits or reduces metastasis of the tumor, or any combination thereof.

[0129] In another related aspect, VH and VL have the amino acid sequences

[0130] (a) VH comprises the amino acid sequence of SEQ ID NO: 26, and VL comprises the amino acid sequence of SEQ ID NO: 27;

[0131] (b) VH comprises the amino acid sequence of SEQ ID NO: 20, and VL comprises the amino acid sequence of SEQ ID NO: 21;

[0132] (c) VH comprises the amino acid sequence of SEQ ID NO: 22, and VL comprises the amino acid sequence of SEQ ID NO: 23;

[0133] (d) VH comprises the amino acid sequence of SEQ ID NO: 24, and VL comprises the amino acid sequence of SEQ ID NO: 25; or

[0134] (e) VH comprises the amino acid sequence of SEQ ID NO: 36, and VL comprises the amino acid sequence of SEQ ID NO: 37.

[0135] In another related aspect, the antibody comprises an IgG, IgA, IgM, IgE, IgD, Fv, scFv, Fab, F(ab')2, minibody, diabody, or triabody. In another related aspect, the antibody comprises a heavy chain comprising a mutation that reduces binding to Fcγ receptors. In yet another related aspect, the mutation comprises L234A or L235A mutations.

[0136] In another related aspect,

[0137] (a) when HCDR1 comprises the amino acid sequence of SEQ ID NO: 62, HCDR2 comprises the amino acid sequence of SEQ ID NO: 63, HCDR3 comprises the amino acid sequence of SEQ ID NO: 64, LCDR1 comprises the amino acid sequence of SEQ ID NO: 65, LCDR2 comprises the amino acid sequence of DAS, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 67, the amino acid sequence of the full-length heavy chain is set forth in SEQ ID NO: 72, and the amnioin sequence of the full-length light chain is set forth in SEQ ID NO: 73;

[0138] (b) when HCDR1 comprises the amino acid sequence of SEQ ID NO:44, HCDR2 comprises the amino acid sequence of SEQ ID NO:45, HCDR3 comprises the amino acid sequence of SEQ ID NO:46, LCDR1 comprises the amino acid sequence of SEQ ID NO:47, LCDR2 comprises the amino acid sequence of YAS, and LCDR3 comprises the amino acid sequence of SEQ ID NO:49, the amino acid sequence of the full-length heavy chain is set forth in SEQ ID NO:68 and the amino acid sequence of the full-length light chain is set forth in SEQ ID NO:69; and

[0139] (c) when HCDR1 comprises the amino acid sequence of SEQ ID NO: 50, HCDR2 comprises the amino acid sequence of SEQ ID NO: 51, HCDR3 comprises the amino acid sequence of SEQ ID NO: 52, LCDR1 comprises the amino acid sequence of SEQ ID NO: 53, LCDR2 comprises the amino acid sequence of YAS, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 55, the amino acid sequence of the full-length heavy chain is set forth in SEQ ID NO: 70, and the amino acid sequence of the full-length light chain is set forth in SEQ ID NO: 71.

[0140] In another related aspect, undesirable effects caused by IL-2 include activation of regulatory T cells, CD25 +One or more of apoptosis of T effector cells, IL-2-induced pulmonary edema, IL-2-induced pneumonia, or IL-2-induced vascular leakage. In further related aspects, the anti-IL2 antibody binds to IL-2, and wherein the binding reduces or eliminates the binding of IL-2 to CD25. In yet another further related aspect, the binding of IL-2 to CD132 / CD122 is not reduced or inhibited. BRIEF DESCRIPTION OF THE DRAWINGS

[0142] This patent or patent application file contains at least one drawing executed in color. Copies of this patent or patent application publication with color drawing(s) will be provided by the Office upon request and payment of the necessary fee.

[0143] The present disclosure of both the production and methods of using engineered anti-IL-2 antibodies, as well as objects, features, and advantages thereof, may be best understood by reference to the following detailed description when read in conjunction with the accompanying drawings, in which:

[0144] Figure 1 . A schematic diagram of the mechanism of action of IL-2 and its dual role in controlling the immune response is presented.

[0145] Figure 2 . A schematic diagram of anti-IL-2 antibody-directed immunotherapy is presented.

[0146] Figure 3A and Figure 3B A schematic diagram of the progression of COVID-19 infection and potential anti-IL-2 therapy as an adjunctive intervention is presented. Figure 3A Modified from Shi Y et al. (2020) COVID-19 infection: the perspectives on immune responses. Cell Death & Differentiation, Vol. 27, pp. 1451-1454 (doi:10.1038 / s41418-020-0530-3). Figure 1 .

[0147] Figures 4A-4D The JES6.1 antibody and human IL-2 ( Figure 4A ), mouse IL-2 ( Figure 4B ) binding, and JES6.1RMC antibody binding to human IL-2 ( Figure 4C ) and mouse IL-2 ( Figure 4D ) Representative SPR sensorgrams of binding.

[0148] Figures 5A-5CResults for IL-2 binding of the YSD clone expressing the JES6.1 antibody in scFv format are presented. The X-axis fluorescence level corresponds to the level of Jes6.1 scFv expression, and the Y-axis fluorescence level corresponds to the binding of human IL-2 or mouse IL-2. Figure 5A Negative controls without IL-2 are shown. Figure 5B Shown are the JES6.1 YSD clones together with 1000 nM human IL-2. Figure 5C Shown are YSD expressing mouse IL-2 incubated with 100 nM labeled JES6.1.

[0149] Figure 6A Binding of isolated yeast surface display clones to IL-2 (0.1 nM) is presented. Mean fluorescence intensity (Em655 nM) was normalized to yeast surface expression levels. Negative YSD clones were labeled with 500 nM hIL-2.

[0150] Figure 6B Nonspecific binding of YSD clones to the OX40 / PD-1 / TNFR2 mixture is shown. These clones were labeled with 500 nM of the mixture. TNFR2-binding yeast clones were used as positive controls.

[0151] Figure 7 The purification of BDG17.023 IgG is shown. The antibody was run in PBS buffer at 0.5 ml / min on a GE superdex 200 10 / 300 increase (CV=25 ml). The leading peak (0.38 CV) corresponds to a typical aggregate, and the second peak (0.51 CV) with a retention of approximately 12.9 ml is typical of normal human IgG.

[0152] Figure 8A-8B BDG17.023 IgG and hIL-2 ( Figure 8A ) and mIL-2( Figure 8B ) binding kinetics.

[0153] Figure 9 Receptor identification by the BDG17.023-IL-2 complex is presented as an SPR response trace. BDG17.023 was immobilized to a CM5 chip, and hIL-2 (60RU), CD122 (20RU), and CD25 (0RU) flowed as indicated by the arrows.

[0154] Figures 10A-10D Splenic immune cell populations of mice treated with JES6.1-mIL-2 complex and BDG17.023-hIL-2 complex are presented. Figure 10AShown are the percentages of immune cell populations from mice treated with JES6.1-mIL-2 complex. Figure 10B Shown are the memory phenotype effector T cell (MP) CD8+ / Treg ratios of mice treated with JES6.1-mIL-2 complex. Figure 10C Shown are the percentages of immune cell populations from mice treated with BDG17.023-hIL-2 complex. Figure 10D Shown are the MP CD8+ / Treg ratios of mice treated with BDG17.023-hIL-2 complex.

[0155] Figure 11 An alignment of the amino acid sequences of the heavy chain variable regions of JES6.1, clone 1 (17.021), clone 2 (17.022), clone 4 (17.023), clone 5 (17.030), and clone 6 (17.035) is presented.

[0156] Figure 12 An alignment of the amino acid sequences of the light chain variable regions of JES6.1, clone 1 (17.021), clone 2 (17.022), clone 4 (17.023), clone 5 (17.030), and clone 6 (17.035) is presented.

[0157] Figure 13A and Figure 13B The heavy chain variable regions of humanized clones 17.014, 17.038, 17.043, 17.053, and 17.054 are presented ( Figure 13A ) and light chain variable region ( Figure 13B ). Black triangles indicate IMGT CDR positions. Bold / italic fonts indicate ABR / CDR positions, respectively.

[0158] Figures 14A-14G . Binding kinetics of the indicated antibodies to human IL-2. Anti-IL-2 antibody clone BDG17.038 ( Figure 14A )、BDG 17.043( Figure 14B )、BDG17.053( Figure 14C )、BDG17.054( Figure 14D )、BDG17.066( Figure 14E )、BDG17.067( Figure 14F ) and BDG17.069( Figure 14G) to human IL-2. The binding kinetics of BDG 17.038, BDG 17.043, BDG 17.066, BDG 17.067, and BDG 17.069 were determined using a multi-cycle approach. The binding kinetics of BDG 17.053 and BDG 17.054 were determined using a single-cycle approach.

[0159] Figures 15A-15B . Binding kinetics of the indicated antibodies to cynomolgus monkey IL-2. Anti-IL-2 antibody clone BDG17.067 ( Figure 15A ) and BDG17.069( Figure 15B ) and cynomolgus monkey IL-2 binding kinetics.

[0160] Figures 16A-16G Differential scanning fluorimetry (DSF) analysis of the melting points of the indicated IgGs. The light green dashed line indicates T onset (Tonset), and the bold green dashed line indicates Tm1 and, where applicable, Tm2. The anti-IL-2 clone analyzed was BDG17.038 ( Figure 16A )、BDG17.043( Figure 16B )、BDG 17.053( Figure 16C )、BDG 17.054( Figure 16D )、BDG17.066( Figure 16E )、BDG17.067( Figure 16F ) and BDG17.069( Figure 16G ).

[0161] Figures 17A-17G Receptor identification of the indicated antibody / IL-2 complexes is presented by tracking SPR responses. Antibodies were immobilized to a CM5 chip, and hIL-2, CD122, and CD25 flowed as indicated by arrows. Figure 17A A schematic sequence of compound injections into the SPR chip is presented, representing sequential anti-IL-2 antibodies complexed with human IL-2 (hIL2), bound to CD122 but not CD25. Figures 17B-17G SPR responses are presented: BDG17.038 ( Figure 17B )、BDG017.043( Figure 17C )、BDG17.054( Figure 17D )、BDG17.066( Figure 17E )、BDG17.067( Figure 17F ) and BDG17.069(AU-007; Figure 17G).

[0162] Figure 18A and Figure 18B Anti-human IL-2 antibodies (clones 17.043 and 17.054) were shown to exhibit potent immunostimulatory effects in vivo. Anti-IL-2 antibody / hIL-2 complexes increased the population of effector cells, while no effect on regulatory T cells was observed. Figure 18A Shown are C57BL / 6 mice administered anti-IL-2 antibody (10 μg) pre-complexed with 0.5 μg of hIL-2 daily for four days. Figure 18B Show that C57BL / 6 mice are administered the anti-IL-2 antibody (25 μ g) of the hIL-2 pre-compound with 1.25 μ g every day and continue four days.At the 5th day, splenocytes are separated, and immune cell colony is analyzed by flow cytometry.The mean value of each experimental group (n=6 in every group) is presented.Lymphocyte is gated according to side scatter parameter and forward scatter parameter, and immune cell subpopulation is gated as follows subsequently: Treg (CD45+, CD3+, CD4+, CD25+, FoxP3+), CD8T cell (CD45+, CD3+, CD8+, CD122+, CD25-), NKT-cell (CD45+, CD3+, CD49b+, NK1.1+), NK cell (CD45+, CD3-, CD49b+, NK1.1+).

[0163] Figure 19A and Figure 19B It was shown that anti-human IL-2 antibodies (clones 17.043 and 17.054) exhibited potent dose-dependent immunostimulatory effects in vivo. Figure 19A Shown are C57BL / 6 healthy mice administered anti-IL-2 antibody / hIL-2 complex daily for four days (25 μg / 1.25 μg, respectively).On day 5, splenocytes were isolated and immune cell populations were analyzed by flow cytometry. Figure 19BShow that anti-human IL-2 antibody demonstrates effective in vivo immunostimulatory effect in a dose-dependent manner. As indicated, C57BL / 6 healthy mice are administered increasing doses of anti-IL-2 antibody / hIL-2 complex every day. At the 5th day, splenocytes are separated, and flow cytometry analysis of immune cell colonies is used. Lymphocytes are gated according to side scatter parameters and forward scatter parameters, and subsequent immune cell subpopulations are gated as follows: Treg (CD45+, CD3+, CD4+, CD25+, FoxP3+), CD8 T cells (CD45+, CD3+, CD8+, CD122+, CD25-), NKT-cells (CD45+, CD3+, CD49b+, NK1.1+), NK cells (CD45+, CD3-, CD49b+, NK1.1+).

[0164] Figure 20A and Figure 20B Anti-human IL-2 antibodies (clones 17.043 and 17.054) were shown to exhibit a safe dosing regimen in vivo. Figure 20A Shown are C57BL / 6 healthy mice administered anti-IL-2 antibody / hIL-2 complex daily for four days (10 μg / 0.5 μg, respectively). Figure 20B Show that C57BL / 6 healthy mice were administered anti-IL-2 antibody / hIL-2 complex (respectively 25 μ g / 1.25 μ g) every day for four days. At the end of the experiment, mice were weighed, and the percentage of body weight change was calculated relative to the body weight of each mouse at the beginning of the study. The average percentage of body weight (BW) change for each experimental group (n=6 per group) is presented.

[0165] Figure 21A and Figure 21B Mean tumor volume results are shown. Anti-IL-2 antibodies (clones 17.038, 17.043, 17.053 and 17.054) inhibited tumor growth in the I / O tolerance tumor model and had a tolerable safety profile. C57BL / 6 healthy mice were inoculated with B16F10 melanoma tumor cells on day 0. On day 5, mice were randomly divided into experimental groups (10 per group) and anti-IL-2 antibody / hIL-2 complex (20 μg / 1 μg, respectively) or PBS was administered daily for 4 days. 17.054 is the parent antibody of 17.069 (AU-007), but lacks the LALA effector silent mutation in the Fc domain, which has been engineered into AU-007. Figure 21A The changes in tumor volume for each experimental group are shown. Figure 21B The changes in body weight of experimental groups 17.043 and 17.054 are shown. The percentage change in body weight was calculated relative to the body weight of each mouse at the beginning of the study.

[0166] Figures 22A-22G Shown are the results of analyzing different preparations of the anti-IL-2 antibody clone BDG 17.069. Figure 22A BDG 17.069 parameters at T=0 are presented. Figure 22B BDG 17.069 appearance, pH, protein concentration and sub-visual particle formation at T=0 and after incubation at 40°C for 1 and 2 weeks are presented. Figure 22C BDG 17.069 SEC, caliper-SDS and capillary isoelectric focusing analyses at T=0 and after incubation at 40°C for 1 and 2 weeks are presented. Figure 22D Presented are the appearance, pH, protein concentration and sub-visual particle formation of BDG 17.069 at T=0 and after stirring at 300 rpm for 3 days. Figure 22E BDG 17.069 SEC, caliper-SDS and capillary isoelectric focusing analyses at T=0 and after stirring at 300 rpm for 3 days are presented. Figure 22F Presented are the appearance, pH, protein concentration, and subparticle formation of BDG 17.069 at T=0 and after five freeze / thaw cycles. Figure 22G SEC, caliper-SDS, and capillary isoelectric focusing analyses of BDG17.069 at T=0 and after five freeze / thaw cycles are presented.

[0167] Figure 23 The administration and dosage schedules of BDG17.069 Q2w monotherapy (left-A), combination BDG17.069+IL-2 loading dose (center-B), and combination BDG17.069 Q2w+IL-2 Q2w (right-C) are presented graphically.

[0168] Figure 24 Duration of treatment and efficacy details are presented. Patients in cohort 1 received 0.5 mg / kg BDG 17.069, while patients in cohort 2 received 1.5 mg / kg BDG 17.069.

[0169] Figure 25 Pharmacokinetic data showing early pharmacokinetic profiles of the first 3 patients administered monotherapy BGD 17.096 are presented.

[0170] Figures 26A-26D Pharmacodynamic (PD) data illustrating the effects and mechanisms of action of BDG 17.069 are presented graphically. Figure 26A The % change in CD4+ Treg population is presented. Figure 26B CD8 / Treg ratios are presented and include Figures 26A-26B Legend (key). Figure 26C Total circulating IFN-γ is presented. Figure 26D Eosinophil count and Figure 26C-Figure 26D Legend for (circles - patient 1; triangles - patient 2; squares - patient 3).

[0171] Figures 27A-27H Technical characteristics of the BDG17.069 (AU-007) antibody are presented, including a schematic diagram of the injection schedule of the SPR sensorgram trace. Figure 27A and Figures 27B-27D : AU-007 binds human IL-2 with high affinity and inhibits binding to hCD25 while retaining binding to hCD122. Affinity and epitope binding sites were evaluated using surface plasmon resonance (SPR). Figure 27A Presented are the SPR sensorgram traces and calculated binding kinetics of CM5 chip-bound AU-007 with hIL-2 as the analyte. Figure 27B An illustrative model of IL-2 binding as part of the human IL-2 trimeric receptor complex and, in comparison, AU-007 (yellow) blocking IL-2 access to CD25 is presented. Figure 27C and Figure 27D AU-007 epitope binding analysis is presented: AU-007 was captured on a CM5 chip and injected with soluble hIL-2 to form a complex. Subsequently, soluble hCD25 was injected, followed by soluble hCD122. The figure shows the SPR sensorgram trace of Ab / IL-2 / IL-2R complex formation. Arrows indicate the injection locations of hIL-2, hCD25, and hCD122. Figures 27E-27H : naIL-2 (hIL-2 / hCD25 conjugate) inhibits binding to hCD25 while retaining binding to hCD122. Figure 27E-Figure 27F A schematic diagram showing the capture of biotinylated hCD25 on a CM5 chip and subsequent injection of soluble hIL-2, soluble hCD25, and naIL-1 is presented. Figure 27G-27H A schematic diagram showing capture of Fc-tagged hCD122 on a CM5 chip and subsequent injection of soluble hIL-2, soluble hCD25, and soluble naIL-2 is presented. The figure represents the SPR sensorgram trace of IL-2R / cytokine complex formation. Arrows indicate the locations where hIL-2, hCD25, and hCD122 were injected.

[0172] Figures 28A-28H We demonstrate that AU-007 can capture and redirect endogenous IL-2 to disrupt autoinhibitory circuits in hPBMCs, whereas HD IL-2 or naIL-2 cannot. AU-007 promotes the expansion of NK and CD8 T cells while completely inhibiting the expansion of regulatory T cells. Figures 28A-28E (The legend is in Figure 28A (center): Naive hPBMCs were treated once on day 0 with 1 nM of NaIL-2 (purple) or with HD IL-2 (1 nM) in combination with 1 uM of isotype control Ab (black) or with 1 uM AU-007 (red) or 10 uM AU-007 (turquoise). Cultures were monitored for 7 days and immune cell subsets were analyzed daily by flow cytometry. Daily values were normalized to untreated samples (UNT). Although naIL-2 expanded NK cells similarly to AU-007, it did not inhibit Treg expansion. AU-007 completely inhibited Treg expansion in culture ( Figure 28A ) and significantly increased the Teff:Treg ratio ( Figure 28B ), without hindering NK( Figure 28C ). AU-007 down-regulates from Figure 28A The suppressive markers of CD4+ Tregs were expressed as defined by a significant decrease in the MFI of CD25 and FoxP3 ( Figure 28D-Figure 28E ). Figure 28F-28H (The legend is in Figure 28H (Center): AU-007 rescues activated lymphocyte viability reduced by treatment with HD IL-2. hPBMC cultures were stimulated once with anti-CD3 / anti-CD28 antibodies with or without 10 uM AU-007. Three (3) days after stimulation, all samples were given HD IL-2 (1 nM), and cell viability was monitored daily using flow cytometry.

[0173] Figures 29A-29K AU-007 is shown to bind endogenous IL-2 and disrupt the negative feedback loop in human PBMCs. Figures 29A-29E (The legend is in Figure 29E Middle): Naive hPBMCs were treated once with 1 uM AU-007 (red) or with an isotype control antibody (blue) on day 0. No exogenous IL-2 was added. Cultures were monitored for 7 days, and immune cell subsets were analyzed daily by flow cytometry. Daily values were normalized to the untreated sample (UNT). AU-007 completely inhibited Treg expansion ( Figure 29A ) and significantly increased the Teff:Treg ratio ( Figure 29B ), without hindering NK( Figure 29C ). AU-007 down-regulates from Figure 29A The suppressive markers of CD4+ Tregs were expressed as defined by a significant decrease in the mean fluorescence intensity (MFI) of CD25 and FoxP3 ( Figure 29D-Figure 29E ). Figure 29F-29KTotal hPBMCs were stimulated for 24 hours with anti-CD3 / anti-CD28 (stimulation only, green) or with anti-CD3 / anti-CD28 (red) or 200 nM isotype control mAb (blue) in the presence of 200 nM AU-007 mAb. No exogenous IL-2 was added. Immune cell subsets were analyzed by flow cytometry. AU-007 inhibited Tregs without hindering effector cells and NK cells. Figure 29F-Figure 29I ). AU-007 down-regulated the suppressive markers of CD4+ Treg from Figure G, as defined by a significant decrease in the MFI of CD25 and FoxP3 ( Figure 29J-29K ).

[0174] Figures 30A-30C Shown, AU-007 and naIL-2 do not hinder CD122 / CD132-STAT5 signaling activity.Stably express human IL-2 dimer receptor (CD122 / CD132) but do not express CD25 and drive secretory embryonic alkaline phosphatase (SEAP) expression under the STAT5 promoter HEK239-dimer-STAT5-SEAP reporter cell line for detecting IL-2 / IL-2-dimer receptor signaling. Figure 30A Flow cytometry was used to detect CD25, CD122, and CD132 expression levels, confirming the exclusive expression of the dimeric receptors. Figure 30B Dose response curves are presented for IL-2 alone (red circles) or in the presence of 200 nM AU-007 (blue squares) or 200 nM of an anti-IL-2 antibody that inhibits interaction with the dimeric receptor (green triangles). Figure 30C Dose response curves of hIL-2 (black circles) are presented. HEK239-dimmer-STAT5-SEAP reporter cells were treated with increasing concentrations of hIL-2 alone or together with the indicated anti-hIL-2 antibodies ( Figure 30B ) or together with increasing concentrations of naIL-2 ( Figure 30C ) were treated with 5% paraformaldehyde, and the accumulation level of SEAP was measured from the cell culture medium 24 h after treatment, and the functional EC-50 ( Figure 30B-Figure 30C ).

[0175] Figures 31A-31GShown, AU-007 captures endogenous IL-2 and even after single treatment 8 days also shows effective immunostimulatory effect in vivo.Transplant NOG-EXL mice with hPBMC from 3 human donors to check the variability between donors.10 days after hPBMC implantation, mice are randomly assigned to research group (9 mice per cohort, each cohort is made up of 3 mice from each donor, and colored dots represent the mean value of each donor of each cohort).Research group mice are treated once with 20mg / Kg AU-007 or with 20mg / Kg of isotype control Ab (hIgG1-LALA), without adding exogenous IL-2. Figure 31A .Experimental outline. Figure 31B . AU-007 / hIL-2 complex was detected in mouse serum using ELISA, n=9±SD. Figure 31C-31G Immune cell analysis of splenocytes was performed using flow cytometry, n = 9 ± SE. Human immune cells were defined as hCD45+ and hCD3+ from total lymphocytes, and subpopulations were defined using the indicated anti-human antibodies. Statistical analysis was performed using a two-way ANOVA test, *p < 0.05, ****p < 0.0001. (Figure legend in Figure 31F-31G below)

[0176] Figures 32A-32C It is presented that an IL-2 negative feedback loop caused by endogenous IL-2 limits the activity of modified IL-2-based therapeutics. Figure 32A Schematic diagram of the role of IL-2 as an immunomodulator in homeostasis and inflammation. Figure 32A The title in also indicates Figure 32B and Figure 32C Status / function. Figure 32B Exogenous administration of modified IL-2 with biased selectivity for dimer-expressing cells promoted the expansion of CD25-negative (CD25-) effector cells, but this was abrogated by endogenous IL-2 that pushed the system back to homeostasis. Figure 32C AU-007 captures and redirects endogenous IL-2, allowing it to expand CD25-negative (CD25-) effector cells while disrupting autoinhibitory circuits and amplifying inflammatory and immunostimulatory phases.

[0177] Figure 33A-Figure 33BHEK-293 cells expressing IL-2 dimer receptors were incubated with IL-2 (red) or IL2+AU-007 (17.069) (blue) or a control antibody with known dimer inhibition properties (green). The readout was the production of secretory embryonic alkaline phosphatase (SEAP) after stimulation. SEAP was detected using a phosphatase substrate and absorbance was measured at 620 nm. AU-007 did not inhibit the ability of IL-2 to stimulate the dimer receptor of IL-2 on HEK293 cells (EC50 for IL-2 alone = 0.71 pM and 0.78 pM in the presence of AU-007).

[0178] Figures 34A-34D The results of pSTAT5 IC50 assays in human peripheral blood mononuclear cells (PBMCs) are presented. Human PBMCs were incubated with IL-2 in the presence (blue line) or absence of AU-007 (black line). The data show that AU-007 inhibits IL-2 expression in regulatory T cells (CD3+CD4+CD25+CD127-FoxP3+) expressing the IL-2 trimeric receptor ( Figure 34A induced STAT5 phosphorylation in CD3-CD56+ NK cells ( Figure 34B ), memory phenotype (MP) CD8+ T cells ( Figure 34C ) or CD3+CD8+CD56+NKT cells ( Figure 34D ) in the ability to induce STAT5 phosphorylation.

[0179] Figures 35A-35E The results of the AU-007 / hIL-2 administration and the results of the AU-007 / hIL-2 administration from CD8 MP Teff ( Figure 35A )、NK( Figure 35B )、NKT( Figure 35C )、CD4+Treg( Figure 35D ) and CD8+Treg( Figure 35E ) cells were expressed as parental cells (%). Briefly, C57B1 / 6 mice were injected daily with AU-007, IL-2 + AU007, or IL2 + control antibody for 4 days, and spleens were harvested on day 5. Splenocytes were phenotypicly analyzed for percentages of lymphocyte populations. AU-007 significantly expanded CD8+ memory cells, NK cells, and NKT cells, but not regulatory T cells.

[0180] Figures 36A-36C The results of the BDG17.054 / hIL-2 administration and the results of the BDG17.054 / hIL-2 administration from CD8 MP Teff ( Figure 35A )、NK( Figure 36B ) and NKT( Figure 36C ) The percentage (%) of cells that are parental to the cells.

[0181] Figure 37A and Figure 37B Presented data showing that administration of BDG17.069 + human IL-2 to mice bearing MC38 colon cancer induced regression, and in the presence of anti-PD-1 ( Figure 37A ) or anti-PD-L1( Figure 37B ) inducing tumor eradication in the absence of .

[0182] Figure 38A and Figure 38B Data showing that administration of BDG17.069 + human IL-2 inhibits tumor growth in the LL / 2 (lung cancer) cancer model is presented. BDG17.069 (AU-007) reduced LL / 2 tumor growth by 74% relative to saline controls. Note that human IL-2 was administered with AU-007 because AU-007 does not bind to mouse IL-2.

[0183] Figure 39 A schematic diagram of the updated Phase I dose-escalation plan is presented. Solid green borders indicate completed and / or ongoing work. Dashed green lines indicate next steps in the progression (as of November 2023).

[0184] Figure 40 AU-007 monotherapy: treatment duration and best response as of October 13, 2023 (Arm 1A) are presented.

[0185] Figure 41A and Figure 41B Presents the results of the AU-007+IL-2(1B) arm (AU-007+1 loading dose of aldesleukin- Figure 41A ); 1C arm (AU-007+aldesleukin (Q2W)- Figure 41B ))Duration of treatment and optimal response.

[0186] Figure 42A 、 Figure 42B and Figure 42C Safety data are presented. Figure 42A Safety data for AU-007 monotherapy in Arm 1A (first 4 cohorts) are presented: mild toxicity profile. Figure 42BPopulation statistics for all three arms (1A, 1B, and 1C) as of October 13, 2023 are presented. *All grade 3 / 4 drug-related AEs were transient (3-7 days) lymphopenia. **A patient with cutaneous squamous cell carcinoma who received AU-007 + Q2W 135K IU / kg aldesleukin experienced a drug-related SAE of transient (~12 hours) grade 2 CRS. The patient developed fever and mild hypotension symptoms starting 6 hours after receiving the initial aldesleukin dose. The patient had pneumonia prior to treatment and received oral antibiotics for RUL consolidation. The patient continued treatment after receiving the second dose of AU-007 + aldesleukin with mild symptoms. Figure 42C Detailed graphs of drug-related adverse events for all three arms (1A, 1B, and 1C) as of October 13, 2023 are presented.

[0187] Figure 43A and Figure 43B Arm 1B presented (Aldesleukin): Mild toxicity profile of drug-related adverse events (AEs). Figure 43B In this study, dMMR indicated mismatch repair deficiency.

[0188] Figure 44A 、 Figure 44B and Figure 44C AU-007 objective responses are presented: waterfall chart showing ongoing results for patients with different cancers in the study as of October 13, 2023. Tumor status and subject participation in the trial, AU-007 monotherapy (Arm 1A): Best % change from baseline in Figure 44A Presented in. Figure 44B Shown are AU-007 + Aldesleukin: Best % change from baseline for all evaluable responding patients who received AU-007 + Aldesleukin. **Patient had a new brain lesion that was stabilized with radiation therapy. Figure 44C Data are presented for AU-007 + aldesleukin: Best % change from baseline immune-sensitive tumors (excluding GI cancers). This includes all response-evaluable patients with non-GI cancers who received AU-007 + aldesleukin.

[0189] Figure 45A and Figure 45B Presents the AU-007 spider diagram ( Figure 45A ) and AU-007+aldesleukin (IL-2) ( Figure 45B ):Percentage of tumor changes over time (%). Figure 45B All patients evaluable for response who received AU-007 + aldesleukin as of October 13, 2023, were included.

[0190] Figures 46A-46C Presented are tumor evaluations by computed tomography (baseline and 8-week scans) from melanoma patients whose cancer did not respond to anti-PD-1 and / or CTLA4 checkpoint inhibitors.

[0191] Figure 47 A plot of AU-007 concentration over time is presented. The left plot shows the first 60 hours of expansion, demonstrating Tmax and C-max. The right plot represents the dataset as of June 2023. Please note that not all cohorts have complete data, as this data is currently being acquired. Overall, the data demonstrate that AU007 exhibits a typical IgG1 therapeutic profile.

[0192] Figure 48A and Figure 48B AU-007 pharmacodynamic data are shown, showing that AU-007 continues to reduce peripheral blood circulating Tregs (as measured by flow cytometry) over time (days) after administration of AU-007. The percentage change in the absolute number of circulating regulatory T cells. Regulatory T cells are defined as CD3+CD4+CD25+CD127lo in CD45+ cells. Consistent with the mechanism of action of inhibiting the interaction of IL-2 with the trimeric receptor, regulatory T cells in the peripheral circulation are reduced. This was observed in both the monotherapy arm and the arm that also included Proleukin, and was consistent across patients. Figure 48A Data are presented for each individual patient (received AU-007 + / - at least one dose of Proleukin - see Figure 49 ), and Figure 48B Data are presented for each dose group (AU-007 only). These values represent the change in absolute (abs) cell counts from baseline.

[0193] Figure 49 Present with at least one dose The mean percentage change in peripheral blood Tregs per day for the cohorts receiving 4.5 mg / kg of AU-007 and 4.5 mg / kg of AU-007 was quantified. These values represent the change in absolute (abs) cell counts relative to baseline. Data points continue to be collected.

[0194] Figure 50A and Figure 50B Shown are changes in absolute peripheral blood CD8 cells over time (days) after administration of AU-007. Figure 50A Data for each individual patient are presented, and Figure 50B Data for each dose group are presented. These values represent the change in absolute (abs) cell counts relative to baseline. Data collection is ongoing.

[0195] Figure 51Shows acceptance (aldesleukin) and AU-007 administered cohorts. These values represent the change in absolute (abs) cell counts from baseline. Data collection is ongoing.

[0196] Figure 52A and Figure 52B Shown are changes in absolute peripheral blood NK cells over time (days) after administration of AU-007. Figure 52A Data for each individual patient are presented, and Figure 52B Data for each dose group are presented. These values represent the change in absolute (abs) cell counts relative to baseline. Data collection is ongoing.

[0197] Figure 53 Shows that after administration of AU-007 Changes in peripheral blood NK cells over time (days) for the cohort receiving (aldesleukin) . These values represent the change in absolute (abs) cell counts from baseline. Data as of September 2023

[0198] Figure 54A and Figure 54B Shown are the absolute numbers of eosinophils over time (days) over extended periods of time following administration of AU-007. Figure 54A Individual peripheral blood eosinophil counts in the AU-007 cohort only. Figure 54B Peripheral blood eosinophil counts of individuals in the AU-007+interleukin cohort. Figure 54A and Figure 54B Shown is the change in the number of circulating eosinophils over time. Figure 54A is the group that received AU-007 monotherapy only, and Figure 54B This is the group that received AU-007 with at least one dose of Proleukin. All but one patient in the AU-007 monotherapy arm and the AU-007 plus Proleukin arm had a decrease or no change in circulating eosinophil levels. One patient in the 9 mg / kg cohort developed severe seasonal allergies requiring treatment during AU-007 treatment and was consistent with a history of seasonal allergy treatment. The increase in eosinophils was attributed to an allergic reaction. All patients given AU-007 and Proleukin showed stabilization or decrease in circulating eosinophils. This is consistent with AU-007's mechanism of action of preventing IL-2 from interacting with the IL-2 trimeric receptor on eosinophils. These values represent the change from baseline in absolute (abs) cell counts. Data collection is ongoing.

[0199] Figures 55A-55CShown are the CD8:Treg ratios over time (days) in the periphery over extended periods of time following administration of AU-007. Figure 55A All available data for each individual patient are shown (see Figure legend for Figure 55C ). Figure 55B Data for each dose group are shown. Figure 55C Shows also accepted (aldesleukin) for each individual patient. Consistent with the results observed in the changes in Treg and CD8+ T cells, a trend towards an increase in the CD8+ / Treg ratio was observed with monotherapy. In the presence of Proleukin, an increase in the CD8+ / Treg ratio was observed, particularly in the presence of higher doses of Proleukin. Consistent with the mechanism of action, higher doses (compared to low-dose IL-2) and longer exposure tended to higher CD8+ / Treg ratios, and no drug-related toxicity was observed. It is expected that increasing the dose of Proleukin will further enhance peripheral responses. These values represent the change in absolute (abs) cell counts relative to baseline. Data collection is ongoing.

[0200] Figure 56A and Figure 56B Fold changes in IFN-γ expression in patients treated with AU-007 + / - Proleukin are presented. Heat map of changes in circulating interferon gamma (IFN-γ) levels relative to baseline. Light green represents 20%-2-fold changes, medium green represents 2-5-fold changes, and dark green represents >5-fold changes. These preliminary results suggest that the longer patients receive monotherapy ( Figure 56A ), the more likely the patient is to have increased circulating IFN-γ. This is consistent with observations in circulating cell populations, particularly Treg and NK cells. Figure 56B ) in the absence of IL-2, the addition of low-dose IL-2 sustained an increase in IFN-γ in the peripheral circulation.

[0201] Figure 57 A protocol for hPBMC expansion assay is presented.

[0202] Figure 58A and Figure 58B Presented are the fold changes in CD4+ Tregs from CD4+ cells in vitro over time in the presence of control antibody (black), AU-007 (red), or CD25-IL-2 conjugate (green). Figure 58A Results from low-dose administration (10 pM) are presented, and Figure 58BThe results of high-dose administration (1 nM) are presented. The data show that although both IL-2 alone or non-αCD25-conjugated IL2 expand the Treg population, there is no Treg expansion in the presence of AU-007. Since non-α IL-2 does not bind to the trimeric receptors on Treg, Treg population expansion is a direct result of IL-2 (endogenous IL-2) produced by T cells. AU-007 not only inhibits exogenous IL-2, but also inhibits endogenous IL-2.

[0203] Figure 59A and Figure 59B Fold changes in CD4+ Tregs from CD4+ cells in vitro over time in the presence of control antibody (blue) or AU-007 (green) and the absence of IL-2 are presented. Figure 59A ) and high doses ( Figure 59B ) Control wells for the IL-2 experiment, in which no IL-2 was added. The images were normalized to untreated wells (i.e., medium only). The results demonstrate that AU-007 inhibits the ability of Tregs to utilize low levels of endogenous IL-2.

[0204] Figure 60A and Figure 60B Figure 2 shows the expression of CD4+ regulatory T cells (CD3+CD4+CD127-CD25+FoxP3+) in vitro in the presence of control antibody (black), AU-007 (red) or CD25-IL-2 conjugate (green). Figure 60A ) and NK( Figure 60B ) Fold changes of cells over time. (IL-2 and CD25-IL-2 conjugates were 1 nM, isotype control antibody and AU-007 were 1 uM; legend in Figure 60B middle)

[0205] Figures 61A-61C The fold change of mean fluorescence intensity (MFI) over time is presented to show the suppressive markers of Treg (CD25 and FoxP3; respectively Figure 61A and Figure 61B ) and NK cell activation markers (CD56; Figure 61C ) changes. Observe 3-6 days after treatment.

[0206] Figure 62A and Figure 62B The results of the present study were in the presence of low doses of IL-2 (10 pM; Figure 62A ) and high-dose IL-2 (1 nM; Figure 62B ) in the case of ΔCD8+Teff:ΔTreg ratio.

[0207] Figure 63The experimental design for measuring restimulation-induced cell death (RICD) is presented.

[0208] Figure 64 Graphs are presented showing the percentage (%) of live lymphocytes over time (days) after administration of 1 nM IL-2 (black) or 1 nM IL-2 and 10 μM AU-007 (green).

[0209] Figure 65 The experimental design for the determination of tetanus toxoid (TT) is presented.

[0210] Figures 66A-66C The results of the TT assay at low doses (0.9 μg / ml TT; Figure 66A ) and high-dose TT (9.0 μg / ml TT; Figure 66B ) of the TT assay results. Figure 66C Shown are the fold changes in Tregs (CD25+) over time.

[0211] Figure 67A and Figure 67B The results of the experiments with the protein antigen tetanus toxoid (high dose - 9.0 μg / ml; Figure 67A and low dose -0.9 μg / ml; Figure 67B )-stimulated CD8+ cells in culture.

[0212] Figure 68 Toxicokinetics (TK) from acute toxicity studies in cynomolgus monkeys (cyno) are presented.

[0213] Figures 69A-69C AU-007 was shown to reduce CD4+CD25+ cells relative to placebo in a cynomolgus monkey study. Figure 69A -PBS (n=12) versus 5 mg / kg (n=8); Figure 69B - PBS (n=12) versus 25 mg / kg (n=8); and Figure 69C -PBS (n=12) versus 100 mg / kg (n=12). PBS values are plotted in black in each figure. Each point represents the pre-dose value for each dosing day. Mean values are + / - SEM. Each graph represents the percentage of CD25+ cells in the CD4+ population for each cohort.

[0214] Figures 70A-70C AU-007 was shown to increase peripheral NK cells relative to baseline in cynomolgus monkey studies. Figure 70A -PBS (n=12) versus 5 mg / kg (n=8); Figure 70B - PBS (n=12) versus 25 mg / kg (n=8); and Figure 70C -PBS (n=12) versus 100 mg / kg (n=12). PBS values are plotted in black in each graph. Each point represents the pre-dose value for each dosing day. Mean values are + / - SEM.

[0215] Figure 71 It was shown that IL-2 was increased in a dose-dependent manner in cynomolgus monkeys administered AU-007 compared to saline (PBS) controls.

[0216] Figure 72 Simulated human pharmacokinetic (PK) profiles following a Q2W dosing regimen are presented.

[0217] Figure 73 Simulated steady-state human PK profiles based on a Q2W dosing regimen are presented.

[0218] Figure 74 Table presenting human PK modeling details for Q2W AU-007 dosing and IL-2 coverage is shown. Reference superscripts - a: amount of IL-2 that can bind to AU-007 on a direct one-to-one molecular stoichiometry basis (IU / cc); b: amount of HD IL-2 that can bind to AU-007 on a direct one-to-one molecular stoichiometry basis (600,000 IU / kg) based on an 80 kg patient (5 L blood volume); c: between the 6th and 7th doses: 12-14 weeks; and d: amount of HD IL-2 that can bind to AU-007 based on an 80 kg patient (5 L blood volume) considering 2 IL-2 molecules to 1 AU-007 molecule (600,000 IU / kg).

[0219] Figure 75 The administration of AU-007 and subcutaneous administration of IL-2 ( The combination of IL-2 inhibitors (aldesleukin) and IL-3b (aldesleukin) is expected to deliver significantly more IL-2 per day to dimeric receptors on Teff and NK cells than competing products.

[0220] Details

[0221] The present disclosure provides engineered anti-human IL-2 antibodies that bind to human IL-2 with high affinity (e.g., 12.7 pM to 48 pM) to a predetermined binding epitope. Antibodies are combined with IL-2 in a manner that completely prevents CD25 from binding but retains the binding of IL-2 to CD122, thereby regulating the immune response to immunostimulation by directly activating and amplifying effector cells without interacting with CD25-expressing cells (e.g., regulatory T cells, short-lived cytotoxic T cells, lung endothelial cells, and vascular endothelial cells). Therefore, the antibody / IL-2 complex will, by amplification and activation of effector cells such as NK cells, central memory T cells, and virus or tumor-specific T cells, simultaneously suppress the cell death of short-lived CD25+ cytotoxic T cells that IL-2 activation induces for virus / tumor clearance and is important, to drive a robust immune response to clear viral load or tumor. The antibody / IL-2 complex will also reduce the immunosuppression caused by the regulatory arm of the immune system. In addition, the antibody / IL-2 complex will prevent the undesirable interaction of IL-2 with blood vessels and lung CD25-expressing cells, thereby preventing the severe syndrome of IL-2-induced vascular leakage and IL-2-induced pulmonary edema commonly seen in viral lung infection models. In some embodiments, the activity of the engineered anti-IL-2 antibodies described herein depends on the predetermined epitope to which they are designed to bind.

[0222] In some embodiments, the IL-2 antibodies disclosed herein block the binding of IL-2 to CD25. In some embodiments, the IL-2 antibodies disclosed herein bind to IL-2 and prevent newly secreted endogenous IL-2 from binding to Tregs, thereby effectively blocking the negative feedback loop of IL-2 and Tregs. In some embodiments, the IL-2 antibodies disclosed herein prevent Treg expansion. In some embodiments, the IL-2 antibodies disclosed herein block the binding of IL-2 to vascular endothelium. In some embodiments, the IL-2 antibodies disclosed herein block the binding of IL-2 to pulmonary endothelium. In some embodiments, the IL-2 antibodies disclosed herein block the binding of IL-2 to vascular endothelium and pulmonary endothelium.

[0223] The skilled artisan will understand that, in certain embodiments, the term "anti-IL-2 antibody," as used herein, is interchangeable with the term "anti-human IL-2 antibody," which has all the same properties and meanings. Similarly, as used throughout, in certain embodiments, the term "IL-2" is interchangeable with the term "human IL-2," which has all the same properties and meanings.

[0224] In some embodiments, the anti-human IL-2 antibodies described herein inhibit the binding of IL-2 to the IL-2 receptor α (IL-2Rα, i.e., CD25) subunit and, therefore, inhibit binding to the trimeric IL-2Rαβγ receptor. In certain embodiments, the anti-IL-2 antibodies that inhibit the binding of IL-2 to the trimeric IL-2 receptor (IL-2Rαβγ) do not inhibit the binding of IL-2 to the dimeric IL-2 receptor (IL-2Rβγ).

[0225] Figure 2 A schematic diagram of an anti-IL-2 antibody-directed immunotherapy is presented. Targeting IL-2 to different cell populations can be used to modulate immune responses to immunosuppression or immune activation. The anti-human IL-2 antibodies disclosed herein are designed to bind with high affinity to an IL-2 epitope that blocks IL-2 binding to CD25. Thus, IL-2 is prevented from binding to short-lived CD8+ cytotoxic T cells or regulatory T cells that express high levels of CD25, but is redirected to preferentially bind to effector T cells to stimulate an enhanced immune response to improve viral or bacterial clearance. In addition, since the binding of IL-2 to CD25-expressing endothelial cells is also blocked, IL-2-induced pulmonary edema and vascular leakage will also be prevented.

[0226] In one embodiment, the present disclosure provides a method of treating a disease (e.g., a viral infection, a bacterial infection, or a cancer) or condition (e.g., an undesirable condition caused by IL-2, such as, but not limited to, pulmonary edema) with an anti-IL-2 antibody designed to enhance T cell immune responses and prevent the severe edema symptoms of acute pneumonia caused by IL-2. The anti-IL-2 antibody will specifically bind to human IL-2 with high affinity at a predetermined epitope, blocking IL-2 binding to the α chain (CD25) of the IL-2 receptor while retaining binding to the receptor's primary signaling β chain and γ chain complex (CD122 / CD132). Thus, in the presence of such an antibody, IL-2 will be directed to immune cells responsible for viral / tumor clearance and away from cells that slow the immune response or cause edema. The formation of this IL-2 / antibody immune complex will guide IL-2 to exclusively bind to and activate naive T lymphocytes and memory T lymphocytes, NK cells and natural killer T lymphocytes, while preventing the activation of regulatory T cells and the apoptosis of short-lived CD25+ cytotoxic T effector cells. In short, the end result is an effective immune response, such as viral or tumor clearance. In addition, this treatment will prevent the toxicity caused by IL-2 binding to endothelial CD25-expressing cells. Therefore, in one embodiment, targeting IL-2 with the anti-IL-2 antibodies disclosed herein will be an effective treatment for respiratory diseases caused by viral or bacterial infections. In another embodiment, treatment with the anti-IL-2 antibodies disclosed herein will effectively prevent the toxicity caused by IL-2 binding to endothelial CD25-expressing cells, such as pulmonary edema or IL-2-induced vascular leakage. More importantly, enhancing IL-2 immune stimulation for general immune activation and amplification of immune effector cells independent of specific pathogens (e.g., viral antigens) will be an effective strategy to combat future viral or bacterial pandemics caused by unknown pathogens ( Figure 3A and Figure 3B ).

[0227] In one embodiment, the method disclosed herein will be useful for combating infections caused by SARS Co-V2. SARS Co-V2 binds to angiotensin-converting enzyme 2, a protein in lung cells that allows viral entry and replication. The immune response to viral infection of the lungs consists of both the innate and acquired arms of the immune system. As with many respiratory viruses, it is expected that the clearance of SARS-CoV2 from the lungs will depend on the T cell immune response. The cytokine IL-2 is crucial for the expansion of T cells and plays an important role in the immune response to the virus. However, in addition to the pro-stimulatory effect of IL-2, IL-2 also induces some adverse side effects such as pulmonary edema and vascular leakage syndrome by binding to the endothelium expressing the CD25 receptor.

[0228] Figure 3Aand Figure 3B A schematic diagram of the progression of COVID-19 infection and potential anti-IL-2 therapy as an adjunctive intervention is presented. Figure 3A It has been shown that the invading SARS Co-V2 causes mild symptoms and triggers a protective immune response after the incubation period. The successful elimination of the infection depends on the health status of the infected individual. Individuals with poor immune responses to the virus will find it difficult to clear the virus, while individuals with overly robust immune responses may lead to pulmonary edema and other cytokine-mediated adverse effects. Therefore, strategies to enhance the immune response and prevent pulmonary edema are desired. Although high concentrations of IL-2 will be beneficial for viral clearance, especially in the early stages, high levels of IL-2 can lead to IL-2-induced pulmonary edema and vascular leakage through the interaction of IL-2 and CD25-expressing endothelial cells. Figure 3B It is shown that anti-human IL-2 antibodies designed to bind and block the CD25 / IL-2 interaction are expected to enhance the expansion of immune effector cells to improve viral clearance and reduce the negative effects of IL-2 binding to CD25 expressed on endothelial cells, thereby preventing IL-2-induced pulmonary edema and vascular leakage.

[0229] Figure 1 A schematic diagram of the mechanism of action of IL-2 and its dual role in controlling the immune response is presented. The left figure shows that IL-2 is composed of three binding epitope sites (α, β, γ) that interact with different forms of IL2-R (CD25, CD122, and CD132) with different affinities. The right figure shows that different IL-2R complexes are expressed on different T cell populations, and their different affinities for IL2 allow for immunosuppression under conditions of low local concentrations of IL-2 and immune stimulation when local concentrations of IL-2 are elevated.

[0230] In the following detailed description, numerous specific details are set forth in order to provide a thorough understanding of the antibodies disclosed herein. However, those skilled in the art will appreciate that, in some cases, the preparation and use of the antibodies disclosed herein can be practiced without these specific details. In other cases, well-known methods, procedures, and components are not described in detail in order not to obscure the disclosure presented herein.

[0231] Throughout this application, various references or publications are cited, the disclosures of which are hereby incorporated by reference in their entireties into this application in order to more fully describe the state of the art to which this invention pertains.

[0232] As used herein, the term "antibody" can be used interchangeably with the term "immunoglobulin", which have all the same properties and meanings. An antibody binding domain or antigen binding site can be a fragment of an antibody or a genetically engineered product of one or more fragments of an antibody that participates in specific binding to a target antigen. "Specific binding" means that the binding is selective for the antigen of interest and can be distinguished from unwanted or nonspecific interactions. For example, when the equilibrium dissociation constant is ≤10 -5 M, 10 -6 M or 10 -7 M, the antibody is said to specifically bind to the IL-2 epitope. In some embodiments, the equilibrium dissociation constant may be ≤10 -8 M or 10 -9 M. In some further embodiments, the equilibrium dissociation constant may be ≤10 -10 M, 10 -11 M or 10 -12 M. In some embodiments, the equilibrium dissociation constant can be ≤10 -5 M to 10 -12 Within the range of M.

[0233] As used herein, the term "antibody" encompasses one or more antibody fragments that retain binding specificity, including but not limited to IgG, heavy chain variable region (VH), light chain variable region (VL), Fab fragment, F(ab')2 fragment, scFv fragment, Fv fragment, nanobody, minibody, diabody, triabody, tetrabody and single domain antibody (see, e.g., Hudson and Souriau, Nature Med. 9: 129-134 (2003)). Also encompassed are humanized, primatized and chimeric antibodies, as these terms are commonly understood in the art.

[0234] As used herein, the term "heavy chain variable region" can be used interchangeably with the term "VH domain" or the term "VH", which have all the same meanings and properties. As used herein, the term "light chain variable region" can be used interchangeably with the term "VL domain" or the term "VL", which have all the same meanings and properties. The skilled person will recognize that a "heavy chain variable region" or "VH" of an antibody encompasses a fragment of the heavy chain comprising three complementary determining regions (CDRs) between the flanking extensions known as framework regions. The framework regions are more highly conserved than CDRs and form a scaffold that supports the CDRs. Similarly, the skilled person will also recognize that a "light chain variable region" or "VL" of an antibody encompasses a fragment of the light chain comprising three CDRs between the framework regions.

[0235] As used herein, the term "complementarity determining region" or "CDR" refers to the hypervariable region of the heavy chain variable region or the light chain variable region. Starting from the N-terminus, each of the heavy chain polypeptide or the light chain polypeptide has three CDRs denoted as "CDR1", "CDR2" and "CDR3". Crystallographic analysis of many antigen-antibody complexes has demonstrated that the amino acid residues of the CDRs form extensive contacts with the bound antigen, with the most extensive antigen contact being with the heavy chain CDR3. Therefore, the CDR region is primarily responsible for the specificity of the antigen binding site. In one embodiment, the antigen binding site includes six CDRs, including CDRs from each of the heavy chain variable region and the light chain variable region.

[0236] As used herein, the term "framework region" or "FR" refers to the four flanking amino acid sequences that frame the CDRs of the heavy or light chain variable region. Some FR residues may contact the bound antigen; however, the FR residues are primarily responsible for folding the variable region into an antigen binding site. In some embodiments, the FR residues responsible for folding the variable region include residues directly adjacent to the CDRs. Within the FRs, certain amino acid residues and certain structural features are very highly conserved. In this regard, all variable region sequences contain an internal disulfide bond loop of approximately 90 amino acid residues. When the variable region folds into the antigen binding site, the CDRs are displayed as protruding loop motifs that form the antigen binding surface. It is generally recognized that there are conserved structural regions of FRs that influence the folded shape of the CDR loops into certain "standard" structures, regardless of the precise CDR amino acid sequence. In addition, certain FR residues are known to participate in non-covalent interdomain contacts that stabilize the interactions between the antibody heavy and light chains.

[0237] Wu and Kabat (Tai Te Wu, Elvin A. Kabat. An analysis of the sequences of the variable regions of bence jones proteins and myeloma light chains and their implications for antibody complementarity. Journal of Experimental Medicine, 132, 2, 8 (1970); Kabat EA, Wu TT, Bilofsky H, Reid-Miller M, Perry H. Sequence of proteins of immunological interest. Bethesda: National Institute of Health; 1983. 323 (1983)) pioneered the alignment of antibody peptide sequences, and their contributions in this regard are multifaceted: First, by studying the sequence similarity between variable domains, they identified corresponding residues that are more or less homologous in all antibodies across all vertebrate species because they adopt similar three-dimensional structures, play similar functional roles, interact similarly with neighboring residues, and exist in similar chemical environments. Second, they designed a peptide sequence numbering system in which homologous immunoglobulin residues are assigned the same position number. Those skilled in the art can clearly use the content that is now commonly referred to as Kabat numbering for any variable domain sequence, without relying on any experimental data except the sequence itself.Third, Kabat and Wu calculated the variability of the sequence position of each Kabat numbering, which means that when the variable domain sequence is compared, it is found that there are few or many possible amino acids.They identified three continuous high variability zones that are embedded in four less variable continuous regions.Kabat and Wu formally delineated the residues that constitute these variable segments (variabletract), and these are expressed as "complementarity determining regions" (CDRs), which refer to the chemical complementation between antibody and antigen.In the three-dimensional folding of variable domains rather than the effect in antigen recognition is attributed to the remaining less variable regions, these regions are now referred to as "framework regions".Fourth, Kabat and Wu have established a public database of antibody peptides and nucleotide sequences, which continues to be maintained and is well known to those skilled in the art.

[0238] Chothia and colleagues (Cyrus Chothia, Arthur M. Lesk. Canonical structures for the hypervariable regions of immunoglobulins. Journal of Molecular Biology, 196, 4, 8 (1987)) found that certain sub-segments within the Kabat CDRs adopt nearly identical peptide backbone conformations despite having significant diversity at the amino acid sequence level. These sub-segments are designated L1, L2, and L3 or H1, H2, and H3, where "L" and "H" represent the light chain and heavy chain regions, respectively. These regions are referred to as Chothia CDRs, which have overlapping boundaries with the Kabat CDRs.

[0239] More recent studies have shown that almost all antibody binding residues fall within regions of structural consistency. (Kunik, V. et al., PloS Computational Biology 8(2):e1002388 (February 2012)). In some embodiments, these regions are referred to as antibody binding regions. It has been shown that these regions can also be identified from antibody sequences. "Paratome," an embodiment of a structural method for identifying structurally consistent residues in antibodies, is used for this purpose. (Ofran, Y. et al., J. Immunol. 757:6230-6235 (2008)). Although the residues identified by Paratome cover almost all antibody binding sites, CDRs (as identified by commonly used CDR identification tools) miss a significant portion of them. Antibody binding residues identified by Paratome but not by any common CDR identification methods are referred to as Paratome-unique residues. Similarly, antibody binding residues identified by any common CDR identification methods but not by Paratome are referred to as CDR-unique residues. Paratome-unique residues make a crucial energetic contribution to the antibody-antigen interaction, whereas CDR-unique residues have a considerably smaller contribution. These results allow for better identification of antigen-binding sites.

[0240] yes (international ImMunoGeneTicsinformation (See, Nucleic Acids Res. 2015 Jan;43(Database Issue):D413-22. doi:10.1093 / nar / gku1056. Epub 2014 Nov 5 Free Article. PMID:25378316 LIGM:441 and Dev Comp Immunol. 2003 Jan;27(1):55-77). IMGT is a unique numbering system for immunoglobulin and T cell receptor variable domains and Ig superfamily V-like domains (Lefranc et al., Dev Comp Immunol. 27:55-77 (2003)). Based on alignment of 5 or more IG and TcR variable region sequences, taking into account and combining Kabat's definitions of FRs and CDRs, structural data, and Chothia's characterization of hypervariable loops, A unified numbering system for these IG and TcR variable domain sequences is provided. IMGT is considered well known in the art as a universal numbering scheme for antibodies.

[0241] In some embodiments, the identification of potential variant amino acid positions in the VH and VL domains uses the IMGT analysis system. In some embodiments, the identification of potential variant amino acid positions in the VH and VL domains uses the Paratome analysis system. In some embodiments, the identification of potential variant amino acid positions in the VH and VL domains uses the Kabat analysis system. In some embodiments, the identification of potential variant amino acid positions in the VH and VL domains uses the Clothia analysis system.

[0242] When describing variant amino acid positions present in the VH and VL domains, in some embodiments, IMGT numbering is used. When describing variant amino acid positions present in the VH and VL domains, in some embodiments, Paratome numbering is used. When describing variant amino acid positions present in the VH and VL domains, in some embodiments, Kabat numbering is used. When describing variant amino acid positions present in the VH and VL domains, in some embodiments, Clothia numbering is used.

[0243] Antigen binding sequences are typically located within the heavy and light chain variable regions of an antibody. In some cases, these heavy and light chain variable regions can be manipulated to create new binding sites, for example, to generate antibodies or fragments thereof that bind to different antigens or to different epitopes of the same antigen. In some embodiments, as described herein, manipulating the sequence of the heavy chain variable region or the sequence of the light chain variable region, or both, will generate new binding sites for a second antigen.

[0244] Antibodies can exist in various forms or have various domains, including but not limited to complementarity determining regions (CDRs), variable regions (Fv), VH domains, VL domains, single chain variable regions (scFv), and Fab fragments.

[0245] Those of ordinary skill in the art will appreciate that scFv is a fusion polypeptide comprising the variable heavy (VH) and variable light (VL) regions of an immunoglobulin, connected by a short linker peptide, which can have, for example, 10 to about 25 amino acids.

[0246] The skilled artisan will also understand that the term "Fab" with respect to an antibody generally encompasses that portion of an antibody consisting of a single light chain bound by disulfide bonds to the variable region and first constant region of a single heavy chain (both the variable and constant regions), whereas F(ab')2 includes fragments of a heavy chain comprising the VH domain and a light chain comprising the VL domain.

[0247] In some embodiments, antibodies encompass complete antibody molecules, including monoclonal antibodies and polyclonal antibodies. In some embodiments, antibodies encompass one or more antibody fragments that retain binding specificity, including but not limited to variable heavy chain (VH) fragments, variable light chain (VL) fragments, Fab fragments, F(ab')2 fragments, scFv fragments, Fv fragments, miniantibodies, diabodies, triabodies, and tetrabodies.

[0248] Engineered anti-IL-2 antibodies

[0249] In one embodiment, the present disclosure provides engineered anti-IL-2 antibodies produced by introducing amino acid variations into parent anti-IL-2 antibodies. In one embodiment, one or more of the amino acid variations are introduced into the CDR region. In another embodiment, one or more amino acid variations are introduced into the framework (FR) region. In yet another embodiment, amino acid variations are introduced into both the CDR region and the framework (FR) region. One of ordinary skill in the art will readily employ a variety of standard techniques known in the art to introduce amino acid variations into anti-IL-2 antibodies, and then test any changes in the resulting modified antibodies in combination with IL-2. Although standard techniques can be used, the binding pattern of the resulting newly generated antibodies is unpredictable and must be analyzed to determine functionality.

[0250] In certain embodiments, the present disclosure provides polypeptides comprising a VH domain and a VL domain that can dimerize under suitable conditions. For example, the VH domain and the VL domain can be combined in a suitable buffer and dimerize through appropriate interactions such as hydrophobic interactions. In another embodiment, the VH domain and the VL domain can be combined in a suitable buffer comprising an enzyme and / or a cofactor that can promote the dimerization of the VH domain and the VL domain. In another embodiment, the VH domain and the VL domain can be combined in a suitable vehicle that allows them to react with each other in the presence of a suitable reagent and / or catalyst.

[0251] In certain embodiments, the VH domain and the VL domain may be included in a longer polypeptide sequence, which may include, for example, but not limited to, a constant region, a hinge region, a linker region, an Fc region, or a disulfide bond binding region or any combination thereof. A constant domain is an immunoglobulin folding unit of the constant portion of an immunoglobulin molecule, also known as a domain of a constant region (e.g., CH1, CH2, CH3, CH4, Ck, Cl). In some embodiments, a longer polypeptide may include multiple copies of one or both of a VH domain and a VL domain produced according to the methods disclosed herein; for example, when the polypeptides produced herein are used to form a diabody or a triabody.

[0252] In some embodiments, the Fc region includes at least one mutation that reduces Fc-γ binding, i.e., binding to Fcγ receptors (FcγRs). In some embodiments, the reduced binding is a combination that is eliminated, and its binding to Fcγ receptors is undetectable. In some embodiments, the reduced binding reduces the binding affinity to Fcγ receptors. In some embodiments, the reduced binding reduces the binding rate (on rate) bound to Fcγ receptors. In some embodiments, the reduced binding reduces the dissociation rate (off rate) bound to Fcγ receptors. In some embodiments, the mutation that reduces Fc-γ binding includes L234A mutation, L235A mutation, also known as LALA mutation. In some embodiments, the mutation that reduces Fc-γ binding includes P329G mutation in addition to L234A mutation, L235A mutation. In some embodiments, the antibodies described herein include heavy chains containing mutations that reduce binding to Fcγ receptors.

[0253] In one embodiment, the present disclosure provides an engineered (or modified) anti-IL-2 antibody, wherein the antibody comprises a heavy chain variable region having a sequence of one of SEQ ID NO: 10, SEQ ID NO: 12, SEQ ID NO: 14, SEQ ID NO: 16, SEQ ID NO: 18, SEQ ID NO: 20, SEQ ID NO: 22, SEQ ID NO: 24, SEQ ID NO: 26, or SEQ ID NO: 36. In one embodiment, the engineered antibody can be IgG, IgA, IgM, IgE, IgD, Fv, scFv, Fab, or F(ab')2. IgG can be of the subclass IgG1, IgG2, IgG3, or IgG4. In another embodiment, the engineered antibody can be part of a minibody, diabody, or triabody.

[0254] In one embodiment, the present disclosure provides an engineered (or modified) anti-IL-2 antibody, wherein the antibody comprises a light chain variable region having a sequence of one of SEQ ID NO: 11, SEQ ID NO: 13, SEQ ID NO: 15, SEQ ID NO: 17, SEQ ID NO: 19, SEQ ID NO: 21, SEQ ID NO: 23, SEQ ID NO: 25, SEQ ID NO: 27, or SEQ ID NO: 37. In one embodiment, the engineered antibody can be IgG, IgA, IgM, IgE, IgD, Fv, scFv, Fab, or F(ab')2. IgG can be of the subclass IgG1, IgG2, IgG3, or IgG4. In another embodiment, the engineered antibody can be part of a minibody, diabody, or triabody.

[0255] In one embodiment, the disclosure provides an engineered (or modified) anti-IL-2 antibody, wherein the antibody comprises a heavy chain variable region and a light chain variable region having a sequence of one of: SEQ ID NO: 10 and SEQ ID NO: 11; SEQ ID NO: 12 and SEQ ID NO: 13; SEQ ID NO: 14 and SEQ ID NO: 15; SEQ ID NO: 16 and SEQ ID NO: 17; SEQ ID NO: 18 and SEQ ID NO: 19; SEQ ID NO: 20 and SEQ ID NO: 21; SEQ ID NO: 22 and SEQ ID NO: 23; SEQ ID NO: 24 and SEQ ID NO: 25; SEQ ID NO: 26 and SEQ ID NO: 27; or SEQ ID NO: 36 and SEQ ID NO: 37. In one embodiment, the engineered anti-IL-2 antibody comprises the sequence of SEQ ID NO: 10 and SEQ ID NO: 11. In one embodiment, the engineered anti-IL-2 antibody comprises the sequence of SEQ ID NO: 12 and SEQ ID NO: 13. In one embodiment, the engineered anti-IL-2 antibody comprises the sequence of SEQ ID NO: 14 and SEQ ID NO: 15. In one embodiment, the engineered anti-IL-2 antibody comprises the sequence of SEQ ID NO: 16 and SEQ ID NO: 17. In one embodiment, the engineered anti-IL-2 antibody comprises the sequence of SEQ ID NO: 18 and SEQ ID NO: 19. In one embodiment, the engineered anti-IL-2 antibody comprises the sequence of SEQ ID NO: 20 and SEQ ID NO: 21. In one embodiment, the engineered anti-IL-2 antibody comprises the sequence of SEQ ID NO: 22 and SEQ ID NO: 23. In one embodiment, the engineered anti-IL-2 antibody comprises the sequence of SEQ ID NO: 24 and SEQ ID NO: 25. In one embodiment, the engineered anti-IL-2 antibody comprises the sequence of SEQ ID NO: 26 and SEQ ID NO: 27. In one embodiment, the engineered anti-IL-2 antibody comprises the sequence of SEQ ID NO:36 and SEQ ID NO:37.

[0256] In some embodiments, the isolated anti-IL-2 antibody comprises a heavy chain variable region (VH) and a light chain variable region (VL), wherein the VH comprises heavy chain complementarity determining regions (HCDRs) HCDR1, HCDR2, and HCDR3, and the VL comprises light chain complementarity determining regions (LCDRs) LCDR1, LCDR2, and LCDR3, wherein the CDRs have the following amino acid sequences:

[0257] (a) HCDR1 comprises the amino acid sequence of SEQ ID NO: 38, HCDR2 comprises the amino acid sequence of SEQ ID NO: 39, HCDR3 comprises the amino acid sequence of SEQ ID NO: 40, LCDR1 comprises the amino acid sequence of SEQ ID NO: 41, LCDR2 comprises the amino acid sequence of YAS, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 43;

[0258] (b) HCDR1 comprises the amino acid sequence of SEQ ID NO: 44, HCDR2 comprises the amino acid sequence of SEQ ID NO: 45, HCDR3 comprises the amino acid sequence of SEQ ID NO: 46, LCDR1 comprises the amino acid sequence of SEQ ID NO: 47, LCDR2 comprises the amino acid sequence of YAS, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 49;

[0259] (c) HCDR1 comprises the amino acid sequence of SEQ ID NO: 50, HCDR2 comprises the amino acid sequence of SEQ ID NO: 51, HCDR3 comprises the amino acid sequence of SEQ ID NO: 52, LCDR1 comprises the amino acid sequence of SEQ ID NO: 53, LCDR2 comprises the amino acid sequence of YAS, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 55;

[0260] (d) HCDR1 comprises the amino acid sequence of SEQ ID NO: 56, HCDR2 comprises the amino acid sequence of SEQ ID NO: 57, HCDR3 comprises the amino acid sequence of SEQ ID NO: 58, LCDR1 comprises the amino acid sequence of SEQ ID NO: 59, LCDR2 comprises the amino acid sequence of YAS, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 61; or

[0261] (e) HCDR1 comprises the amino acid sequence of SEQ ID NO: 62, HCDR2 comprises the amino acid sequence of SEQ ID NO: 63, HCDR3 comprises the amino acid sequence of SEQ ID NO: 64, LCDR1 comprises the amino acid sequence of SEQ ID NO: 65, LCDR2 comprises the amino acid sequence of DAS, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 67.

[0262] In some embodiments, the isolated anti-IL-2 antibody comprises a heavy chain variable region (VH) and a light chain variable region (VL), wherein the VH comprises heavy chain complementarity determining regions (HCDRs) HCDR1, HCDR2, and HCDR3, and the VL comprises light chain complementarity determining regions (LCDRs) LCDR1, LCDR2, and LCDR3, wherein the CDRs have the following amino acid sequences:

[0263] (a) HCDR1 comprises the amino acid sequence of SEQ ID NO: 38, HCDR2 comprises the amino acid sequence of SEQ ID NO: 39, HCDR3 comprises the amino acid sequence of SEQ ID NO: 40, LCDR1 comprises the amino acid sequence of SEQ ID NO: 41, LCDR2 comprises the amino acid sequence of YAS, and LCDR3 comprises the amino acid sequence of SEQ ID NO: 43.

[0264] In some embodiments, the VH and VL have amino acid sequences, wherein the VH comprises the amino acid sequence of SEQ ID NO: 10 and the VL comprises the amino acid sequence of SEQ ID NO: 11; the VH comprises the amino acid sequence of SEQ ID NO: 12 and the VL comprises the amino acid sequence of SEQ ID NO: 13; the VH comprises the amino acid sequence of SEQ ID NO: 14 and the VL comprises the amino acid sequence of SEQ ID NO: 15; the VH comprises the amino acid sequence of SEQ ID NO: 16 and the VL comprises the amino acid sequence of SEQ ID NO: 17; the VH comprises the amino acid sequence of SEQ ID NO: 18 and the VL comprises the amino acid sequence of SEQ ID NO: 19; the VH comprises the amino acid sequence of SEQ ID NO: 20 and the VL comprises the amino acid sequence of SEQ ID NO: 21; the VH comprises the amino acid sequence of SEQ ID NO: 22 and the VL comprises the amino acid sequence of SEQ ID NO: 23; the VH comprises the amino acid sequence of SEQ ID NO: 24 and the VL comprises the amino acid sequence of SEQ ID NO: 25; the VH comprises the amino acid sequence of SEQ ID NO: 26 and the VL comprises the amino acid sequence of SEQ ID NO: or VH comprises the amino acid sequence of SEQ ID NO: 36 and VL comprises the amino acid sequence of SEQ ID NO: 37. In some embodiments, VH and VL have amino acid sequences, wherein VH comprises the amino acid sequence of SEQ ID NO: 26 and VL comprises the amino acid sequence of SEQ ID NO: 27.

[0265] In some embodiments, the antibody comprises a heavy chain sequence set forth in SEQ ID NO: 68 and a light chain sequence set forth in SEQ ID NO: 69; the heavy chain sequence set forth in SEQ ID NO: 70 and the light chain sequence set forth in SEQ ID NO: 71; or the heavy chain sequence set forth in SEQ ID NO: 72 and the light chain sequence set forth in SEQ ID NO: 73. In some embodiments, the antibody comprises a heavy chain sequence set forth in SEQ ID NO: 68 and a light chain sequence set forth in SEQ ID NO: 69. In some embodiments, the antibody comprises a heavy chain sequence set forth in SEQ ID NO: 70 and a light chain sequence set forth in SEQ ID NO: 71. In some embodiments, the antibody comprises a heavy chain sequence set forth in SEQ ID NO: 72 and a light chain sequence set forth in SEQ ID NO: 73.

[0266] In one embodiment, the engineered antibody can be IgG, IgA, IgM, IgE, IgD, Fv, scFv, Fab, or F(ab')2. IgG can be a subclass of IgG1, IgG2, IgG3, or IgG4. In another embodiment, the engineered antibody can be part of a minibody, diabody, or triabody.

[0267] In one embodiment, the present disclosure also provides an isolated polynucleotide sequence encoding the heavy chain variable region of an anti-IL-2 antibody, wherein the heavy chain variable region comprises an amino acid sequence of one of SEQ ID NO: 10, SEQ ID NO: 12, SEQ ID NO: 14, SEQ ID NO: 16, SEQ ID NO: 18, SEQ ID NO: 20, SEQ ID NO: 22, SEQ ID NO: 24, SEQ ID NO: 26, or SEQ ID NO: 36. In another embodiment, the present disclosure also provides a vector comprising the polynucleotide sequence mentioned above. Given the amino acid sequence disclosed herein, one of ordinary skill in the art will readily construct a vector or plasmid encoding the amino acid sequence. In another embodiment, the present disclosure also provides a host cell comprising the vector provided herein. Depending on the application and experimental conditions, one of ordinary skill in the art will readily employ a suitable host cell to carry and / or express the polynucleotide sequence mentioned above.

[0268] In one embodiment, the present disclosure also provides an isolated polynucleotide sequence encoding the light chain variable region of an anti-IL-2 antibody, wherein the light chain variable region comprises an amino acid sequence of one of SEQ ID NO: 11, SEQ ID NO: 13, SEQ ID NO: 15, SEQ ID NO: 17, SEQ ID NO: 19, SEQ ID NO: 21, SEQ ID NO: 23, SEQ ID NO: 25, SEQ ID NO: 27, or SEQ ID NO: 37. In another embodiment, the present disclosure also provides a vector comprising the polynucleotide sequence mentioned above. Given the amino acid sequence disclosed herein, one of ordinary skill in the art will readily construct a vector or plasmid encoding the amino acid sequence. In another embodiment, the present disclosure also provides a host cell comprising the vector provided herein. Depending on the application and experimental conditions, one of ordinary skill in the art will readily employ a suitable host cell to carry and / or express the polynucleotide sequence mentioned above.

[0269] Given the sequences of the heavy chain variable region and the light chain variable region disclosed herein, one of ordinary skill in the art will readily employ standard techniques known in the art to construct anti-IL-2 scFv. In one embodiment, the polynucleotide sequence encoding such an anti-IL-2 scFv may have a sequence of one of SEQ ID NOs: 1-5 or one of SEQ ID NOs: 31-35.

[0270] In certain embodiments, the isolated polynucleotide sequence disclosed herein encodes a heavy chain variable region of an anti-IL-2 antibody comprising the VH amino acid sequence set forth in any one of the amino acid sequences of SEQ ID NO: 10, SEQ ID NO: 12, SEQ ID NO: 14, SEQ ID NO: 16, SEQ ID NO: 18, SEQ ID NO: 20, SEQ ID NO: 22, SEQ ID NO: 24, SEQ ID NO: 26, or SEQ ID NO: 36. In some embodiments, the vector comprises the polynucleotide sequence of any one of SEQ ID NO: 10, SEQ ID NO: 12, SEQ ID NO: 14, SEQ ID NO: 16, SEQ ID NO: 18, SEQ ID NO: 20, SEQ ID NO: 22, SEQ ID NO: 24, SEQ ID NO: 26, or SEQ ID NO: 36. In some embodiments, the host cell contains a vector comprising a polynucleotide sequence of any one of SEQ ID NO: 10, SEQ ID NO: 12, SEQ ID NO: 14, SEQ ID NO: 16, SEQ ID NO: 18, SEQ ID NO: 20, SEQ ID NO: 22, SEQ ID NO: 24, SEQ ID NO: 26, or SEQ ID NO: 36.

[0271] In certain embodiments, the isolated polynucleotide sequence disclosed herein encodes a light chain variable region of an anti-IL-2 antibody comprising a VL amino acid sequence as set forth in the amino acid sequence of any one of SEQ ID NO: 11, SEQ ID NO: 13, SEQ ID NO: 15, SEQ ID NO: 17, SEQ ID NO: 19, SEQ ID NO: 21, SEQ ID NO: 23, SEQ ID NO: 25, SEQ ID NO: 27, or SEQ ID NO: 37. In some embodiments, the vector contains a polynucleotide sequence comprising the amino acid sequence of any one of SEQ ID NO: 11, SEQ ID NO: 13, SEQ ID NO: 15, SEQ ID NO: 17, SEQ ID NO: 19, SEQ ID NO: 21, SEQ ID NO: 23, SEQ ID NO: 25, SEQ ID NO: 27, or SEQ ID NO: 37. In some embodiments, the host cell contains a vector comprising a polynucleotide sequence encoding the amino acid sequence of any one of SEQ ID NO: 11, SEQ ID NO: 13, SEQ ID NO: 15, SEQ ID NO: 17, SEQ ID NO: 19, SEQ ID NO: 21, SEQ ID NO: 23, SEQ ID NO: 25, SEQ ID NO: 27, or SEQ ID NO: 37. In some embodiments, the isolated polynucleotide sequence encodes an anti-IL-2 scFv, wherein the polynucleotide sequence is set forth in SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 31, SEQ ID NO: 32, SEQ ID NO: 33, SEQ ID NO: 34, or SEQ ID NO: 35. In some embodiments, the vector comprises an isolated polynucleotide sequence encoding an anti-IL-2 scFv, wherein the polynucleotide sequence is set forth in SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 31, SEQ ID NO: 32, SEQ ID NO: 33, SEQ ID NO: 34, or SEQ ID NO: 35.In some embodiments, the host cell contains a vector comprising an isolated polynucleotide sequence encoding an anti-IL-2 scFv, wherein the polynucleotide sequence is set forth in SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 31, SEQ ID NO: 32, SEQ ID NO: 33, SEQ ID NO: 34, or SEQ ID NO: 35.

[0272] In another embodiment, the present disclosure also provides an isolated anti-IL-2 antibody, wherein the antibody comprises a heavy chain variable region comprising a complementarity determining region (CDR) 1, CDR2, and CDR3. In one embodiment, CDR1, CDR2, and CDR3 comprise the amino acid sequences of SEQ ID NOs: 38-40, 44-46, 50-52, 56-58, or 62-64, respectively. In one embodiment, the antibody can be an IgG, IgA, IgM, IgE, IgD, Fv, scFv, Fab, F(ab')2, minibody, diabody, or triabody. The IgG can be IgG1, IgG2, IgG3, or IgG4. In one embodiment, the present disclosure also encompasses a composition comprising the above-mentioned antibody and a pharmaceutically acceptable carrier.

[0273] In another embodiment, the present disclosure also provides an isolated anti-IL-2 antibody, wherein the antibody comprises a light chain variable region comprising a complementarity determining region (CDR) 1, CDR2, and CDR3. In one embodiment, CDR1, CDR2, and CDR3 comprise the amino acid sequences of SEQ ID NOs: 41-43, 47-49, 53-55, 59-61, or 65-67, respectively. In one embodiment, the antibody can be an IgG, IgA, IgM, IgE, IgD, Fv, scFv, Fab, F(ab')2, minibody, diabody, or triabody. The IgG can be IgG1, IgG2, IgG3, or IgG4. In one embodiment, the present disclosure also encompasses a composition comprising the above-mentioned antibody and a pharmaceutically acceptable carrier.

[0274] In another embodiment, the present disclosure also provides an isolated anti-IL-2 antibody, wherein the antibody comprises a heavy chain variable region comprising a complementarity determining region (CDR) 1, CDR2, and CDR3, and a light chain variable region comprising CDR1, CDR2, and CDR3. In one embodiment, the heavy chain CDR1, CDR2, and CDR3 comprise the amino acid sequences of SEQ ID NOs: 38-40, 44-46, 50-52, 56-58, or 62-64, respectively. In one embodiment, the light chain CDR1, CDR2, and CDR3 comprise the amino acid sequences of SEQ ID NOs: 41-43, 47-49, 53-55, 59-61, or 65-67, respectively. In one embodiment, the antibody can be an IgG, IgA, IgM, IgE, IgD, Fv, scFv, Fab, F(ab')2, miniantibody, diabody, or triabody. IgG can be IgG1, IgG2, IgG3, or IgG4. In one embodiment, the present disclosure also encompasses a composition comprising the above-mentioned antibodies and a pharmaceutically acceptable carrier.

[0275] Pharmaceutical composition

[0276] In some embodiments, disclosed herein are compositions for therapeutic use. In some embodiments, the compositions described herein comprise an anti-IL-2 antibody as disclosed herein and a pharmaceutically acceptable carrier.

[0277] As used herein, the terms "composition" and "pharmaceutical composition" may, in some embodiments, be used interchangeably and have all the same properties and meanings. Disclosed herein, in some embodiments, are pharmaceutical compositions for treating a condition or disease as described herein.

[0278] Disclosed herein, in some embodiments, are pharmaceutical compositions for use in combination therapy.

[0279] In another embodiment, disclosed herein are compositions for treating a disease or condition in a subject. In some embodiments, the disease comprises a viral infection, a bacterial infection, or cancer. In some embodiments, the condition comprises an IL-2-induced condition. In some embodiments, the IL-2-induced condition comprises pulmonary edema or vascular leakage.

[0280] Application

[0281] The VH polypeptides and / or VL polypeptides disclosed herein can be administered to a subject (e.g., a human or animal) alone or in combination with a carrier, i.e., a pharmaceutically acceptable carrier. Pharmaceutically acceptable means that it is not a biologically or otherwise undesirable material, i.e., the material can be administered to a subject without causing any undesirable biological effects or interacting in a harmful manner with any other component of a pharmaceutical composition containing it. As will be known to those of ordinary skill in the art, the carrier is selected to minimize any degradation of the polypeptides disclosed herein and to minimize any adverse side effects of the subject. The pharmaceutical composition can be prepared by methods well known in the pharmaceutical art.

[0282] The pharmaceutical compositions described above comprising polypeptides disclosed herein can be administered in any suitable manner (e.g., to a mammal, cell, or tissue) depending on whether local or systemic treatment is desired. For example, the composition can be administered topically (e.g., ocularly, vaginally, rectally, intranasally, transdermally, etc.), orally, by inhalation, or parenterally (including by intravenous instillation or subcutaneous, intracavitary, intraperitoneal, intradermal, or intramuscular injection). Topical intranasal administration refers to delivery of the composition to the nose and nasal passages through one or both nostrils. The composition can be delivered by a spray mechanism or a droplet mechanism or by atomization. Delivery can also be directed to any area of the respiratory system (e.g., lungs) via an intubation tube. Alternatively, administration can be intratumoral, such as by topical or intravenous injection.

[0283] If the composition is to be administered parenterally, administration is typically by injection. Injectables can be prepared in conventional forms as liquid solutions or suspensions, solid forms suitable for suspension in liquid prior to injection, or as emulsions. Additionally, parenteral administration can involve the preparation of slow-release or sustained-release systems to maintain a constant dose.

[0284] In some embodiments, the composition comprises an anti-IL-2 antibody comprising a heavy chain variable region having a sequence of one of SEQ ID NO: 10, SEQ ID NO: 12, SEQ ID NO: 14, SEQ ID NO: 16, SEQ ID NO: 18, SEQ ID NO: 20, SEQ ID NO: 22, SEQ ID NO: 24, SEQ ID NO: 26, or SEQ ID NO: 36. In some embodiments, the composition comprises an anti-IL-2 antibody comprising a light chain variable region having a sequence of one of SEQ ID NO: 11, SEQ ID NO: 13, SEQ ID NO: 15, SEQ ID NO: 17, SEQ ID NO: 19, SEQ ID NO: 21, SEQ ID NO: 23, SEQ ID NO: 25, SEQ ID NO: 27, or SEQ ID NO: 37. In some embodiments, the composition comprises an anti-IL-2 antibody comprising a heavy chain variable region and a light chain variable region having a sequence of one of the following: SEQ ID NO: 10 and SEQ ID NO: 11; SEQ ID NO: 12 and SEQ ID NO: 13; SEQ ID NO: 14 and SEQ ID NO: 15; SEQ ID NO: 16 and SEQ ID NO: 17; SEQ ID NO: 18 and SEQ ID NO: 19; SEQ ID NO: 20 and SEQ ID NO: 21; SEQ ID NO: 22 and SEQ ID NO: 23; SEQ ID NO: 24 and SEQ ID NO: 25; SEQ ID NO: 26 and SEQ ID NO: 27; or SEQ ID NO: 36 and SEQ ID NO: 37. In some embodiments, the composition comprises an anti-IL-2 antibody comprising a heavy chain variable domain comprising CDR1, CDR2, and CDR3 regions comprising the amino acid sequences of SEQ ID NOs: 38-40, respectively; SEQ ID NOs: 44-46, respectively; SEQ ID NOs: 50-52, respectively; SEQ ID NOs: 56-58, respectively; or SEQ ID NOs: 62-64, respectively.In some embodiments, the composition comprises an anti-IL-2 antibody comprising a light chain variable domain comprising a CDR1 region, a CDR2 region, and a CDR3 region comprising the amino acid sequence of SEQ ID NOs: 41-43, respectively; SEQ ID NOs: 47-49, respectively; SEQ ID NOs: 53-55, respectively; SEQ ID NOs: 59-61, respectively; or SEQ ID NOs: 65-67, respectively. In some embodiments, the composition comprises an anti-IL-2 antibody comprising a heavy chain variable domain comprising CDR1, CDR2, and CDR3 regions comprising the amino acid sequences of SEQ ID NOs: 38-40, 44-46, 50-52, 56-58, or 62-64, respectively, and a light chain variable domain comprising CDR1, CDR2, and CDR3 regions comprising the amino acid sequences of SEQ ID NOs: 41-43, 47-49, 53-55, 59-61, or 65-67, respectively.

[0285] In some embodiments, the composition comprises an anti-IL-2 antibody comprising any of clones BDG 17.014, BDG 17.023, BDG 17.038, BDG 17.043, BDG 17.053, BDG 17.054, BDG 17.066, BDG 17.067, and BDG 17.069.

[0286] In one embodiment, the methods disclosed herein administer a composition comprising an anti-IL-2 antibody disclosed herein as a monotherapy. In another embodiment, the methods disclosed herein administer a composition comprising an anti-IL-2 antibody disclosed herein and a single dose of IL-2. In another embodiment, the methods disclosed herein administer a combination therapy comprising a composition comprising an anti-IL-2 antibody disclosed herein and IL-2.

[0287] In some embodiments, when anti-IL2 antibodies and IL-2 are administered, they can be administered in the same composition. In some embodiments, when anti-IL2 antibodies and IL-2 are administered, they can be administered in separate compositions.

[0288] In some embodiments, IL-2 can be administered before, simultaneously with, or after the step of administering the anti-IL2 antibody. In some embodiments, IL-2 is administered before the anti-IL2 antibody is administered. In some embodiments, IL-2 is administered simultaneously with the administration of the anti-IL2 antibody. In some embodiments, IL-2 is administered after the step of administering the anti-IL2 antibody.

[0289] In some embodiments, the route of IL-2 administration is subcutaneous. In some embodiments, the dose of IL-2 administered subcutaneously is much lower and less frequent than the approved regimen for aldesleukin administered intravenously. In some embodiments, the route of anti-IL-2 administration is by intravenous injection. In some embodiments, where the subject receives both an anti-IL-2 antibody and IL-2, the route of administration of the anti-IL2 antibody is by intravenous injection, and the route of administration of the IL-2 is by subcutaneous injection.

[0290] In some embodiments, the administration of the anti-IL-2 antibody comprises monotherapy. In some embodiments, the administration of the anti-IL-2 antibody comprises combining a loading dose of IL-2 with the anti-IL-2 antibody monotherapy. In some embodiments, the administration of the anti-IL-2 antibody comprises combination therapy, wherein the anti-IL-2 antibody and IL-2 are administered to the subject at regular intervals.

[0291] In some embodiments, multiple doses of anti-IL2 antibodies are administered over a given time period. In some embodiments, a dose of anti-IL2 antibodies is administered weekly. In some embodiments, a dose of anti-IL2 antibodies is administered bi-weekly, once every two weeks. In some embodiments, a dose of anti-IL2 antibodies is administered once every three weeks. In some embodiments, a one-time dose of IL-2 may be administered before, simultaneously with, or after the first dose of an anti-IL2 antibody as described herein. In some embodiments, multiple doses of anti-IL2 antibodies and IL-2 are administered over a given time period. In some embodiments, a dose of anti-IL2 antibodies and IL-2 is administered once weekly. In some embodiments, a dose of anti-IL2 antibodies and IL-2 is administered bi-weekly, once every two weeks. In some embodiments, a dose of anti-IL2 antibodies and IL-2 is administered once every three weeks.

[0292] In some embodiments, the anti-IL2 antibody is administered once a week, once every two weeks, or once every three weeks, wherein IL-2 is administered as a one-time dose, once a week, once every two weeks, or once every three weeks, wherein the administration of the anti-IL-2 antibody and IL-2 is administered simultaneously. In some embodiments, the anti-IL2 antibody is administered once a week, once every two weeks, or once every three weeks, wherein IL-2 is administered as a one-time dose, once a week, once every two weeks, or once every three weeks, wherein the administration of the anti-IL-2 antibody and IL-2 is administered independently of each other, for example, but not limited to, the anti-IL2 antibody is administered once a week, once every two weeks, or once every three weeks, wherein IL-2 is administered as a one-time dose, or the anti-IL2 antibody is administered once a week, once every two weeks, or once every three weeks, wherein IL-2 is administered once a week, or the anti-IL2 antibody is administered once a week, once every two weeks, or once every three weeks, wherein IL-2 is administered once every two weeks, or the anti-IL2 antibody is administered once a week, once every two weeks, or once every three weeks, wherein IL-2 is administered once every three weeks.

[0293] In some embodiments, the therapeutic dose of anti-IL-2 is administered over a period of several months. In some embodiments, the therapeutic dose of anti-IL-2 is administered for up to 3 months. In some embodiments, the therapeutic dose of anti-IL-2 is administered for at least 3 months. In some embodiments, the therapeutic dose of anti-IL-2 is administered for up to 6 months. In some embodiments, the therapeutic dose of anti-IL-2 is administered for at least 6 months. In some embodiments, the therapeutic dose of anti-IL-2 is administered for up to 9 months. In some embodiments, the therapeutic dose of anti-IL-2 is administered for at least 9 months.

[0294] In some embodiments, the therapeutic dose of anti-IL-2 is administered over a period of up to one year. In some embodiments, the therapeutic dose of anti-IL-2 is administered over a period of at least one year.

[0295] In some embodiments, an anti-IL-2 antibody disclosed herein is administered in combination with IL-2, wherein the dose of IL-2 is considered a low dose of IL-2. In some embodiments, IL-2 is administered in a single dose (loading dose). In some embodiments, IL-2 is administered in the same time period as the anti-IL-2 antibody. In some embodiments, IL-2 is administered in multiple doses before, simultaneously with, or after the administration of the anti-IL-2 antibody.

[0296] In some embodiments, the dosage of an anti-IL-2 antibody disclosed herein is between about 0.5 mg / kg and 12 mg / kg. In some embodiments, the dosage of an anti-IL-2 antibody disclosed herein is about 0.5 mg / kg, 1.0 mg / kg, 1.5 mg / kg, 2.0 mg / kg, 2.5 mg / kg, 3.0 mg / kg, 3.5 mg / kg, 4.0 mg / kg, 4.5 mg / kg, 5.0 mg / kg, 5.5 mg / kg, 6.0 mg / kg, 6.5 mg / kg, 7.0 mg / kg, 7.5 mg / kg, 8.0 mg / kg, 8.5 mg / kg, 9.0 mg / kg, 9.5 mg / kg, 10.0 mg / kg, 10.5 mg / kg, 11.0 mg / kg, 11.5 mg / kg, and 12 mg / kg.

[0297] In some embodiments, the dose of an anti-IL-2 antibody disclosed herein is 0.5 mg / kg. In some embodiments, the dose of an anti-IL-2 antibody disclosed herein is 1.0 mg / kg. In some embodiments, the dose of an anti-IL-2 antibody disclosed herein is 1.5 mg / kg. In some embodiments, the dose of an anti-IL-2 antibody disclosed herein is 2.0 mg / kg. In some embodiments, the dose of an anti-IL-2 antibody disclosed herein is 2.5 mg / kg. In some embodiments, the dose of an anti-IL-2 antibody disclosed herein is 3.0 mg / kg. In some embodiments, the dose of an anti-IL-2 antibody disclosed herein is 3.5 mg / kg. In some embodiments, the dose of an anti-IL-2 antibody disclosed herein is 4.0 mg / kg. In some embodiments, the dose of an anti-IL-2 antibody disclosed herein is 4.5 mg / kg. In some embodiments, the dose of an anti-IL-2 antibody disclosed herein is 5.0 mg / kg. In some embodiments, the dose of an anti-IL-2 antibody disclosed herein is 5.5 mg / kg. In some embodiments, the dose of an anti-IL-2 antibody disclosed herein is 6.0 mg / kg. In some embodiments, the dose of an anti-IL-2 antibody disclosed herein is 6.5 mg / kg. In some embodiments, the dose of an anti-IL-2 antibody disclosed herein is 7.0 mg / kg. In some embodiments, the dose of an anti-IL-2 antibody disclosed herein is 7.5 mg / kg. In some embodiments, the dose of an anti-IL-2 antibody disclosed herein is 8.0 mg / kg. In some embodiments, the dose of an anti-IL-2 antibody disclosed herein is 8.5 mg / kg. In some embodiments, the dose of an anti-IL-2 antibody disclosed herein is 9.0 mg / kg. In some embodiments, the dose of an anti-IL-2 antibody disclosed herein is 9.5 mg / kg. In some embodiments, the dose of an anti-IL-2 antibody disclosed herein is 10.0 mg / kg. In some embodiments, the dose of an anti-IL-2 antibody disclosed herein is 10.5 mg / kg. In some embodiments, the dose of an anti-IL-2 antibody disclosed herein is 11.0 mg / kg. In some embodiments, the dose of an anti-IL-2 antibody disclosed herein is 11.5 mg / kg. In some embodiments, the dose of an anti-IL-2 antibody disclosed herein is 12.0 mg / kg. In some embodiments, the dose of an anti-IL-2 antibody disclosed herein is 12.5 mg / kg.

[0298] In some embodiments, the dose of IL-2 includes a low dose. It will be understood by those skilled in the art that a low dose of IL-2 can encompass dose levels lower than those provided in studies currently known in the art. In certain embodiments, at about 10×10 3 IU / Kg-300×10 3IU / Kg of IL-2 dosage encompasses low doses of IL-2. In certain embodiments, between about 10×10 3 IU / Kg-500×10 3 IL-2 doses between IU / Kg encompass low doses of IL-2.

[0299] In some embodiments, the dose of IL-2 is about 10×10 3 IU / Kg to 500×10 3 IU / Kg. In some embodiments, the dose of IL-2 is between about 10×10 3 IU / Kg to 300×10 3 IU / Kg. In some embodiments, the dose of IL-2 is about 15×10 3 IU / Kg to 270×10 3 IU / Kg. In some embodiments, the dose of IL-2 is about 10×10 3 , 15×10 3 , 20×10 3 , 25×10 3 , 30×10 3 , 35×10 3 , 40×10 3 , 45×10 3 , 50×10 3 55×10 3 , 60×10 3 , 65×10 3 , 70×10 3 , 75×10 3 ,80×10 3 ,85×10 3 ,90×10 3 95×10 3 , 100×10 3 , 125×10 3 , 150×10 3 , 175×10 3 , 200×10 3 , 225×10 3 , 250×10 3 , 275×10 3 , 300×10 3 , 325×10 3 , 350×10 3 ,375×10 3 , 400×10 3 , 425×10 3 , 450×10 3, 475×10 3 or 500×10 3 IU / Kg. In some embodiments, the dose of IL-2 is about 15×10 3 IU / Kg. In some embodiments, the dose of IL-2 is about 45×10 3 IU / Kg. In some embodiments, the dose of IL-2 is about 135×10 3 IU / kg. In some embodiments, the dose of IL-2 is about 270×10 3 IU / Kg. In some embodiments, the dose of IL-2 is about 300×10 3 IU / Kg. In some embodiments, the dose of IL-2 is about 400×10 3 IU / Kg. In some embodiments, the dose of IL-2 is about 500×10 3 IU / Kg. In some embodiments, the dose of IL-2 is 15×10 3 IU / Kg. In some embodiments, the dose of IL-2 is 45×10 3 IU / Kg. In some embodiments, the dose of IL-2 is 135×10 3 IU / kg. In some embodiments, the dose of IL-2 is 270×10 3 IU / Kg. In some embodiments, the dose of IL-2 is 300×10 3 IU / Kg. In some embodiments, the dose of IL-2 is 400×10 3 IU / Kg. In some embodiments, the dose of IL-2 is 500×10 3 IU / Kg.

[0300] In some embodiments, when an anti-IL-2 antibody disclosed herein is administered with a single loading dose of IL-2, the dose of the anti-IL-2 antibody is between about 0.1 mg / kg and 12 mg / kg of the anti-IL-2 antibody, and the dose of IL-2 is between about 10×10 3 IU / Kg-300×10 3 In some embodiments, when an anti-IL-2 antibody disclosed herein is administered with a single loading dose of IL-2, the dose of the anti-IL-2 antibody is between about 0.1 mg / kg and 12 mg / kg of the anti-IL-2 antibody, and the dose of IL-2 is between about 10×10 3 IU / Kg-500×10 3In some embodiments, when an anti-IL-2 antibody disclosed herein is administered with a single loading dose of IL-2, the dose of the anti-IL-2 antibody is 0.5 mg / kg and the dose of IL-2 is 15×10 3 In some embodiments, when an anti-IL-2 antibody disclosed herein is administered with a single loading dose of IL-2, the dose of the anti-IL-2 antibody is 0.5 mg / kg and the dose of IL-2 is 45×10 3 In some embodiments, when an anti-IL-2 antibody disclosed herein is administered with a single loading dose of IL-2, the dose of the anti-IL-2 antibody is 0.5 mg / kg and the dose of IL-2 is 135×10 3 In some embodiments, when an anti-IL-2 antibody disclosed herein is administered with a single loading dose of IL-2, the dose of the anti-IL-2 antibody is 0.5 mg / kg and the dose of IL-2 is 270×10 3 In some embodiments, when an anti-IL-2 antibody disclosed herein is administered with a single loading dose of IL-2, the dose of the anti-IL-2 antibody is 0.5 mg / kg and the dose of IL-2 is 300×10 3 In some embodiments, when an anti-IL-2 antibody disclosed herein is administered with a single loading dose of IL-2, the dose of the anti-IL-2 antibody is 0.5 mg / kg and the dose of IL-2 is 400×10 3 In some embodiments, when an anti-IL-2 antibody disclosed herein is administered with a single loading dose of IL-2, the dose of the anti-IL-2 antibody is 0.5 mg / kg and the dose of IL-2 is 500×10 3 IU / Kg.

[0301] In some embodiments, when an anti-IL-2 antibody disclosed herein is administered with a single loading dose of IL-2, the dose of the anti-IL-2 antibody is 1.5 mg / kg and the dose of IL-2 is 15×10 3 In some embodiments, when an anti-IL-2 antibody disclosed herein is administered with a single loading dose of IL-2, the dose of the anti-IL-2 antibody is 1.5 mg / kg and the dose of IL-2 is 45×10 3 In some embodiments, when an anti-IL-2 antibody disclosed herein is administered with a single loading dose of IL-2, the dose of the anti-IL-2 antibody is 1.5 mg / kg and the dose of IL-2 is 135×10 3 In some embodiments, when an anti-IL-2 antibody disclosed herein is administered with a single loading dose of IL-2, the dose of the anti-IL-2 antibody is 1.5 mg / kg and the dose of IL-2 is 270×103 In some embodiments, when an anti-IL-2 antibody disclosed herein is administered with a single loading dose of IL-2, the dose of the anti-IL-2 antibody is 1.5 mg / kg and the dose of IL-2 is 300×10 3 In some embodiments, when an anti-IL-2 antibody disclosed herein is administered with a single loading dose of IL-2, the dose of the anti-IL-2 antibody is 1.5 mg / kg and the dose of IL-2 is 400×10 3 In some embodiments, when an anti-IL-2 antibody disclosed herein is administered with a single loading dose of IL-2, the dose of the anti-IL-2 antibody is 1.5 mg / kg and the dose of IL-2 is 500×10 3 IU / Kg.

[0302] In some embodiments, when an anti-IL-2 antibody disclosed herein is administered with a single loading dose of IL-2, the dose of the anti-IL-2 antibody is 4.5 mg / kg and the dose of IL-2 is 15×10 3 In some embodiments, when an anti-IL-2 antibody disclosed herein is administered with a single loading dose of IL-2, the dose of the anti-IL-2 antibody is 4.5 mg / kg and the dose of IL-2 is 45×10 3 In some embodiments, when an anti-IL-2 antibody disclosed herein is administered with a single loading dose of IL-2, the dose of the anti-IL-2 antibody is 4.5 mg / kg and the dose of IL-2 is 135×10 3 In some embodiments, when an anti-IL-2 antibody disclosed herein is administered with a single loading dose of IL-2, the dose of the anti-IL-2 antibody is 4.5 mg / kg and the dose of IL-2 is 270×10 3 In some embodiments, when an anti-IL-2 antibody disclosed herein is administered with a single loading dose of IL-2, the dose of the anti-IL-2 antibody is 4.5 mg / kg and the dose of IL-2 is 300×10 3 In some embodiments, when an anti-IL-2 antibody disclosed herein is administered with a single loading dose of IL-2, the dose of the anti-IL-2 antibody is 4.5 mg / kg and the dose of IL-2 is 400×10 3 In some embodiments, when an anti-IL-2 antibody disclosed herein is administered with a single loading dose of IL-2, the dose of the anti-IL-2 antibody is 4.5 mg / kg and the dose of IL-2 is 500×10 3 IU / Kg.

[0303] In some embodiments, when an anti-IL-2 antibody disclosed herein is administered with a single loading dose of IL-2, the dose of the anti-IL-2 antibody is 9.0 mg / kg and the dose of IL-2 is 15×10 3 In some embodiments, when an anti-IL-2 antibody disclosed herein is administered with a single loading dose of IL-2, the dose of the anti-IL-2 antibody is 9.0 mg / kg and the dose of IL-2 is 45×10 3 In some embodiments, when an anti-IL-2 antibody disclosed herein is administered with a single loading dose of IL-2, the dose of the anti-IL-2 antibody is 9.0 mg / kg and the dose of IL-2 is 135×10 3 In some embodiments, when an anti-IL-2 antibody disclosed herein is administered with a single loading dose of IL-2, the dose of the anti-IL-2 antibody is 9.0 mg / kg and the dose of IL-2 is 270×10 3 In some embodiments, when an anti-IL-2 antibody disclosed herein is administered with a single loading dose of IL-2, the dose of the anti-IL-2 antibody is 9.0 mg / kg and the dose of IL-2 is 300×10 3 In some embodiments, when an anti-IL-2 antibody disclosed herein is administered with a single loading dose of IL-2, the dose of the anti-IL-2 antibody is 9.0 mg / kg and the dose of IL-2 is 400×10 3 In some embodiments, when an anti-IL-2 antibody disclosed herein is administered with a single loading dose of IL-2, the dose of the anti-IL-2 antibody is 9.0 mg / kg and the dose of IL-2 is 500×10 3 IU / Kg.

[0304] In some embodiments, when an anti-IL-2 antibody disclosed herein is administered with a single loading dose of IL-2, the dose of the anti-IL-2 antibody is 12 mg / kg and the dose of IL-2 is 15×10 3 In some embodiments, when an anti-IL-2 antibody disclosed herein is administered with a single loading dose of IL-2, the dose of the anti-IL-2 antibody is 12 mg / kg and the dose of IL-2 is 45×10 3 In some embodiments, when an anti-IL-2 antibody disclosed herein is administered with a single loading dose of IL-2, the dose of the anti-IL-2 antibody is 12 mg / kg and the dose of IL-2 is 135×10 3 In some embodiments, when an anti-IL-2 antibody disclosed herein is administered with a single loading dose of IL-2, the dose of the anti-IL-2 antibody is 12 mg / kg and the dose of IL-2 is 270×10 3In some embodiments, when an anti-IL-2 antibody disclosed herein is administered with a single loading dose of IL-2, the dose of the anti-IL-2 antibody is 12 mg / kg and the dose of IL-2 is 300×10 3 In some embodiments, when an anti-IL-2 antibody disclosed herein is administered with a single loading dose of IL-2, the dose of the anti-IL-2 antibody is 12 mg / kg and the dose of IL-2 is 400×10 3 In some embodiments, when an anti-IL-2 antibody disclosed herein is administered with a single loading dose of IL-2, the dose of the anti-IL-2 antibody is 12 mg / kg and the dose of IL-2 is 250×10 3 IU / Kg.

[0305] In some embodiments, when a combination therapy comprising an anti-IL-2 antibody disclosed herein and IL-2 is administered, wherein the dose of the anti-IL-2 antibody is between about 0.1 mg / kg and 12 mg / kg of the anti-IL-2 antibody, and the dose of IL-2 is between about 10×10 3 IU / Kg-300×10 3 In some embodiments, when a combination therapy comprising an anti-IL-2 antibody disclosed herein and IL-2 is administered, the dose of the anti-IL-2 antibody is between about 0.1 mg / kg and 12 mg / kg of the anti-IL-2 antibody, and the dose of IL-2 is between about 10×10 3 IU / Kg-500×10 3 In some embodiments, when a combination therapy comprising an anti-IL-2 antibody disclosed herein and IL-2 is administered, the dose of the anti-IL-2 antibody is between about 0.5 mg / kg and 12 mg / kg of the anti-IL-2 antibody, and the dose of IL-2 is between about 10×10 3 IU / Kg-300×10 3 In some embodiments, when a combination therapy comprising an anti-IL-2 antibody disclosed herein and IL-2 is administered, the dose of the anti-IL-2 antibody is between about 0.5 mg / kg and 12 mg / kg of the anti-IL-2 antibody, and the dose of IL-2 is between about 10×10 3 IU / Kg-500×10 3 Between IU / Kg.

[0306] In some embodiments, when a combination therapy comprising an anti-IL-2 antibody disclosed herein and IL-2 is administered, the dose of the anti-IL-2 antibody is 0.5 mg / kg and the dose of IL-2 is 15×10 3In some embodiments, when a combination therapy comprising an anti-IL-2 antibody disclosed herein and IL-2 is administered, the dose of the anti-IL-2 antibody is 0.5 mg / kg and the dose of IL-2 is 45×10 3 In some embodiments, when a combination therapy comprising an anti-IL-2 antibody disclosed herein and IL-2 is administered, the dose of the anti-IL2 antibody is 0.5 mg / kg and the dose of IL-2 is 135×10 3 In some embodiments, when a combination therapy comprising an anti-IL-2 antibody disclosed herein and IL-2 is administered, the dose of the anti-IL-2 antibody is 0.5 mg / kg and the dose of IL-2 is 270×10 3 In some embodiments, when a combination therapy comprising an anti-IL-2 antibody disclosed herein and IL-2 is administered, the dose of the anti-IL-2 antibody is 0.5 mg / kg and the dose of IL-2 is 300×10 3 In some embodiments, when a combination therapy comprising an anti-IL-2 antibody disclosed herein and IL-2 is administered, the dose of the anti-IL-2 antibody is 0.5 mg / kg and the dose of IL-2 is 400×10 3 In some embodiments, when a combination therapy comprising an anti-IL-2 antibody disclosed herein and IL-2 is administered, the dose of the anti-IL-2 antibody is 0.5 mg / kg and the dose of IL-2 is 500×10 3 IU / Kg.

[0307] In some embodiments, when a combination therapy comprising an anti-IL-2 antibody disclosed herein and IL-2 is administered, the dose of the anti-IL-2 antibody is 1.5 mg / kg and the dose of IL-2 is 15×10 3 In some embodiments, when a combination therapy comprising an anti-IL-2 antibody disclosed herein and IL-2 is administered, the dose of the anti-IL-2 antibody is 1.5 mg / kg and the dose of IL-2 is 45×10 3 In some embodiments, when a combination therapy comprising an anti-IL-2 antibody disclosed herein and IL-2 is administered, the dose of the anti-IL-2 antibody is 1.5 mg / kg and the dose of IL-2 is 135×10 3 In some embodiments, when a combination therapy comprising an anti-IL-2 antibody disclosed herein and IL-2 is administered, the dose of the anti-IL-2 antibody is 1.5 mg / kg and the dose of IL-2 is 270×10 3In some embodiments, when a combination therapy comprising an anti-IL-2 antibody disclosed herein and IL-2 is administered, the dose of the anti-IL-2 antibody is 1.5 mg / kg and the dose of IL-2 is 300×10 3 In some embodiments, when a combination therapy comprising an anti-IL-2 antibody disclosed herein and IL-2 is administered, the dose of the anti-IL-2 antibody is 1.5 mg / kg and the dose of IL-2 is 400×10 3 In some embodiments, when a combination therapy comprising an anti-IL-2 antibody disclosed herein and IL-2 is administered, the dose of the anti-IL-2 antibody is 1.5 mg / kg and the dose of IL-2 is 500×10 3 IU / Kg.

[0308] In some embodiments, when a combination therapy comprising an anti-IL-2 antibody disclosed herein and IL-2 is administered, the dose of the anti-IL-2 antibody is 4.5 mg / kg and the dose of IL-2 is about 10×10 3 In some embodiments, when a combination therapy comprising an anti-IL-2 antibody disclosed herein and IL-2 is administered, the dose of the anti-IL-2 antibody is 4.5 mg / kg and the dose of IL-2 is about 10×10 3 IU / Kg-500 IU / kg.

[0309] In some embodiments, when a combination therapy comprising an anti-IL-2 antibody disclosed herein and IL-2 is administered, the dose of the anti-IL-2 antibody is 4.5 mg / kg and the dose of IL-2 is 15×10 3 In some embodiments, when a combination therapy comprising an anti-IL-2 antibody disclosed herein and IL-2 is administered, the dose of the anti-IL-2 antibody is 4.5 mg / kg and the dose of IL-2 is 45×10 3 In some embodiments, when a combination therapy comprising an anti-IL-2 antibody disclosed herein and IL-2 is administered, the dose of the anti-IL2 antibody is 4.5 mg / kg and the dose of IL-2 is 135×10 3 In some embodiments, when a combination therapy comprising an anti-IL-2 antibody disclosed herein and IL-2 is administered, the dose of the anti-IL2 antibody is 4.5 mg / kg and the dose of IL-2 is 270×10 3 In some embodiments, when a combination therapy comprising an anti-IL-2 antibody disclosed herein and IL-2 is administered, the dose of the anti-IL-2 antibody is 4.5 mg / kg and the dose of IL-2 is 300×10 3In some embodiments, when a combination therapy comprising an anti-IL-2 antibody disclosed herein and IL-2 is administered, the dose of the anti-IL-2 antibody is 4.5 mg / kg and the dose of IL-2 is 400×10 3 In some embodiments, when a combination therapy comprising an anti-IL-2 antibody disclosed herein and IL-2 is administered, the dose of the anti-IL-2 antibody is 4.5 mg / kg and the dose of IL-2 is 500×10 3 In some embodiments, when a combination therapy comprising an anti-IL-2 antibody disclosed herein and IL-2 is administered, the dose of the anti-IL-2 antibody is 9.0 mg / kg and the dose of IL-2 is about 10×10 3 In some embodiments, when a combination therapy comprising an anti-IL-2 antibody disclosed herein and IL-2 is administered, the dose of the anti-IL-2 antibody is 9.0 mg / kg and the dose of IL-2 is about 10×10 3 IU / Kg-500 IU / kg.

[0310] In some embodiments, when a combination therapy comprising an anti-IL-2 antibody disclosed herein and IL-2 is administered, the dose of the anti-IL-2 antibody is 9.0 mg / kg and the dose of IL-2 is 15×10 3 In some embodiments, when a combination therapy comprising an anti-IL-2 antibody disclosed herein and IL-2 is administered, the dose of the anti-IL-2 antibody is 9.0 mg / kg and the dose of IL-2 is 45×10 3 In some embodiments, when a combination therapy comprising an anti-IL-2 antibody disclosed herein and IL-2 is administered, the dose of the anti-IL2 antibody is 9.0 mg / kg and the dose of IL-2 is 135×10 3 In some embodiments, when a combination therapy comprising an anti-IL-2 antibody disclosed herein and IL-2 is administered, the dose of the anti-IL-2 antibody is 9.0 mg / kg and the dose of IL-2 is 270×10 3 In some embodiments, when a combination therapy comprising an anti-IL-2 antibody disclosed herein and IL-2 is administered, the dose of the anti-IL-2 antibody is 9.0 mg / kg and the dose of IL-2 is 300×10 3 In some embodiments, when a combination therapy comprising an anti-IL-2 antibody disclosed herein and IL-2 is administered, the dose of the anti-IL-2 antibody is 9.0 mg / kg and the dose of IL-2 is 400×10 3In some embodiments, when a combination therapy comprising an anti-IL-2 antibody disclosed herein and IL-2 is administered, the dose of the anti-IL-2 antibody is 9.0 mg / kg and the dose of IL-2 is 500×10 3 IU / Kg.

[0311] In some embodiments, when a combination therapy comprising an anti-IL-2 antibody disclosed herein and IL-2 is administered, the dose of the anti-IL-2 antibody is 12 mg / kg and the dose of IL-2 is 15×10 3 In some embodiments, when a combination therapy comprising an anti-IL-2 antibody disclosed herein and IL-2 is administered, the dose of the anti-IL-2 antibody is 12 mg / kg and the dose of IL-2 is 45×10 3 In some embodiments, when a combination therapy comprising an anti-IL-2 antibody disclosed herein and IL-2 is administered, the dose of the anti-IL-2 antibody is 12 mg / kg and the dose of IL-2 is 135×10 3 In some embodiments, when a combination therapy comprising an anti-IL-2 antibody disclosed herein and IL-2 is administered, the dose of the anti-IL-2 antibody is 12 mg / kg and the dose of IL-2 is 270×10 3 In some embodiments, when a combination therapy comprising an anti-IL-2 antibody disclosed herein and IL-2 is administered, the dose of the anti-IL-2 antibody is 12 mg / kg and the dose of IL-2 is 300×10 3 In some embodiments, when a combination therapy comprising an anti-IL-2 antibody disclosed herein and IL-2 is administered, the dose of the anti-IL-2 antibody is 12 mg / kg and the dose of IL-2 is 400×10 3 In some embodiments, when a combination therapy comprising an anti-IL-2 antibody disclosed herein and IL-2 is administered, the dose of the anti-IL-2 antibody is 12 mg / kg and the dose of IL-2 is 500×10 3 IU / Kg.

[0312] In some embodiments, the dose of IL-2 is considered low when compared to other therapies. In some embodiments, the low dose of IL-2 is about 10×10 3 IU / Kg-500×10 3 IU / Kg. In some embodiments, the low dose of IL-2 is about 10×10 3 IU / Kg-500×10 3 In some embodiments, the low dose of IL-2 is between about 15×10 3 IU / Kg-500×103 In some embodiments, the low dose of IL-2 is between about 45×10 3 IU / Kg-500×10 3 In some embodiments, the low dose of IL-2 is equal to or less than about 500×10 3 IU / Kg. In some embodiments, the composition comprises an anti-IL-2 antibody comprising anti-IL-2 clone BDG 17.014. In some embodiments, the composition comprises an anti-IL-2 antibody comprising anti-IL-2 clone BDG 17.023. In some embodiments, the composition comprises an anti-IL-2 antibody comprising anti-IL-2 clone BDG 17.038. In some embodiments, the composition comprises an anti-IL-2 antibody comprising anti-IL-2 clone BDG 17.043. In some embodiments, the composition comprises an anti-IL-2 antibody comprising anti-IL-2 clone BDG 17.053. In some embodiments, the composition comprises an anti-IL-2 antibody comprising anti-IL-2 clone BDG 17.054. In some embodiments, the composition comprises an anti-IL-2 antibody comprising anti-IL-2 clone BDG 17.066. In some embodiments, the composition comprises an anti-IL-2 antibody comprising anti-IL-2 clone BDG 17.067. In some embodiments, the composition comprises an anti-IL-2 antibody comprising anti-IL-2 clone BDG 17.069.

[0313] In some embodiments, the composition comprises an anti-IL-2 antibody and a pharmaceutically acceptable carrier. In some embodiments, the composition comprises an anti-IL-2 antibody and IL-2 and a pharmaceutically acceptable carrier. In some embodiments, the composition comprises an anti-IL-2 antibody complexed with IL-2 and a pharmaceutically acceptable carrier.

[0314] In some embodiments, the anti-IL-2 antibody and IL-2 are contained in the same composition. In some embodiments, the anti-IL-2 antibody and IL-2 are contained in different compositions. In some embodiments, administration of the anti-IL-2 antibody and IL-2, or a combination thereof, is simultaneous. In some embodiments, administration of the anti-IL-2 antibody and IL-2, or a combination thereof, comprises administering the anti-IL-2 antibody or a combination thereof before IL-2, or a combination thereof. In some embodiments, administration of the anti-IL-2 antibody and IL-2, or a combination thereof, comprises administering the anti-IL-2 antibody or a combination thereof after administering IL-2, or a combination thereof.

[0315] The skilled artisan will understand that a "pharmaceutical composition" may encompass a preparation of one or more active ingredients described herein with other chemical components such as physiologically suitable carriers and excipients. The purpose of a pharmaceutical composition is to facilitate administration of an active agent, such as, but not limited to, an antibody or compound, to an organism.

[0316] In some embodiments, disclosed herein are pharmaceutical compositions for use in the treatment of a subject with a weakened immune system. In some embodiments, disclosed herein are pharmaceutical compositions for use in the treatment of a subject with a viral infection, a bacterial infection, or a cancer. In some embodiments, disclosed herein are pharmaceutical compositions for use as part of a combination therapy for treating a subject with a weakened immune system. In some embodiments, disclosed herein are pharmaceutical compositions for use as part of a combination therapy for treating a subject with a viral infection, a bacterial infection, or a cancer.

[0317] A skilled artisan will understand that the phrases "physiologically acceptable carrier," "pharmaceutically acceptable carrier," "physiologically acceptable excipient," and "pharmaceutically acceptable excipient" may be used interchangeably to encompass a carrier, excipient, or diluent that does not cause significant irritation to an organism and does not abrogate the biological activity and properties of the administered active ingredient.

[0318] The skilled artisan will understand that "excipients" can encompass inert substances added to pharmaceutical compositions to further facilitate administration of the active ingredient. In some embodiments, excipients include calcium carbonate, calcium phosphate, various sugars and types of starch, cellulose derivatives, gelatin, vegetable oils, and polyethylene glycols.

[0319] Techniques for the formulation and administration of drugs are found in "Remington's Pharmaceutical Sciences," Mack Publishing Co., Easton, PA, most recent edition, which is incorporated herein by reference.

[0320] In some embodiments, the compositions as disclosed herein comprise therapeutic compositions. In some embodiments, the compositions as disclosed herein comprise therapeutic efficacy.

[0321] Combination therapy

[0322] In some embodiments, the anti-IL-2 antibodies or compositions thereof disclosed herein are used as part of a combination therapy. In some embodiments, the anti-IL-2 antibodies or compositions thereof disclosed herein are used in combination with IL-2. In some embodiments, the anti-IL-2 antibodies or compositions thereof disclosed herein are used in combination with IL-2 and an immune checkpoint inhibitor. In some embodiments, the anti-IL-2 antibodies or compositions thereof disclosed herein are used in combination with an immune checkpoint inhibitor.

[0323] In some embodiments, anti-IL-2 antibodies or compositions thereof are used in combination with immune checkpoint inhibitors. In some embodiments, the term "immune checkpoint inhibitor" can encompass any compound or molecule that can inhibit the function of a checkpoint protein. In some embodiments, the term "immune checkpoint inhibitor" can encompass any compound or molecule that targets an immune checkpoint protein. Technicians will understand that "immune checkpoints" are key regulators of the immune system that, when stimulated, can suppress the immune response to immune stimulation. Checkpoint inhibitors can block inhibitory checkpoints and thereby restore immune system function. In some embodiments, one or more checkpoint inhibitors include immune checkpoint inhibitors.

[0324] It will be understood by those skilled in the art that the terms "immune checkpoint inhibitor" (ICI), "checkpoint inhibitors" and the like can be used interchangeably herein and have all the same properties and meanings, wherein immune checkpoint inhibitors encompass compounds that inhibit the activity of the immune system or control the mechanisms of the immune system. Immune system checkpoints or immune checkpoints are inhibitory pathways in the immune system that typically act to maintain self-tolerance or regulate the duration and amplitude of physiological immune responses to minimize incidental tissue damage. Checkpoint inhibitors can inhibit immune system checkpoints by inhibiting the activity of proteins in the pathway.

[0325] Immune checkpoint inhibitor targets include, but are not limited to, PD-1, PD-L1, CTLA-4, TIGIT, TIM-3, B7-H3, CD73, LAG3, CD27, CD70, 4-1BB, GITR, OX40, SIRP-α (CD47), CD39, ILDR2, VISTA, BTLA, and VTCN-1. In some embodiments, anti-IL-2 antibody therapy is used in combination with an immune checkpoint inhibitor, wherein the target of the immune checkpoint inhibitor includes PD-1, PD-L1, CTLA-4, TIGIT, TIM-3, B7-H3, CD73, LAG3, CD27, CD70, 4-1BB, GITR, OX40, SIRP-α (CD47), CD39, ILDR2, VISTA, BTLA, or VTCN-1, or any combination thereof.

[0326] Check point inhibitors can include antibodies or their antigen binding fragments, other binding proteins, biotherapeutics or small molecules that bind to and block or inhibit the activity of one or more of PD-1, PD-L1, CTLA-4, TIGIT, TIM-3, B7-H3, CD73, LAG3, CD27, CD70, 4-1BB, GITR, OX40, SIRP-α (CD47), CD39, ILDR2, VISTA, BTLA or VTCN-1. Illustrative check point inhibitors include but are not limited to those listed in Table 1 below.

[0327] Table 1: Non-limiting examples of checkpoint inhibitors and immune checkpoint inhibitor targets.

[0328]

[0329]

[0330] In some embodiments, checkpoint inhibitors include PD-1 inhibitors. In some embodiments, checkpoint inhibitors include PD-L1 inhibitors. In some embodiments, checkpoint inhibitors include CTLA-4 inhibitors. In some embodiments, checkpoint inhibitors include TIGIT inhibitors. In some embodiments, checkpoint inhibitors include TIM-3 inhibitors. In some embodiments, checkpoint inhibitors include B7-H3 inhibitors. In some embodiments, checkpoint inhibitors include CD73 inhibitors. In some embodiments, checkpoint inhibitors include LAG3 inhibitors. In some embodiments, checkpoint inhibitors include CD27 inhibitors. In some embodiments, checkpoint inhibitors include CD70 inhibitors. In some embodiments, checkpoint inhibitors include 4-1BB agonist conjugates. In some embodiments, checkpoint inhibitors include GITR agonist conjugates. In some embodiments, checkpoint inhibitors include OX40 agonist conjugates. In some embodiments, checkpoint inhibitors include SIRP-α (CD47) inhibitors. In some embodiments, checkpoint inhibitors include CD39 inhibitors. In some embodiments, checkpoint inhibitors include ILDR2 inhibitors. In some embodiments, checkpoint inhibitors include VISTA inhibitors. In some embodiments, the checkpoint inhibitor comprises a BTLA inhibitor. In some embodiments, the checkpoint inhibitor comprises a VTCN-1 inhibitor.

[0331] In some embodiments, the check point inhibitors include PD-1 inhibitors, PD-L1 inhibitors, CTLA-4 inhibitors, TIGIT inhibitors, TIM-3 inhibitors, B7-H3 inhibitors, CD73 inhibitors, LAG3 inhibitors, CD27 inhibitors, CD70 inhibitors, 4-1BB inhibitors, GITR inhibitors, OX40 inhibitors, SIRP-α (CD47) inhibitors, CD39 inhibitors, ILDR2 inhibitors, VISTA inhibitors, BTLA inhibitors, VTCN-1 inhibitors. In some embodiments, the check point inhibitors include at least two check point inhibitors, and the at least two check point inhibitors are selected from PD-1 inhibitors, PD-L1 inhibitors, CTLA-4 inhibitors, TIGIT inhibitors, TIM-3 inhibitors, B7-H3 inhibitors, CD73 inhibitors, LAG3 inhibitors, CD27 inhibitors, CD70 inhibitors, 4-1BB inhibitors, GITR inhibitors, OX40 inhibitors, SIRP-α (CD47) inhibitors, CD39 inhibitors, ILDR2 inhibitors, VISTA inhibitors, BTLA inhibitors and VTCN-1 inhibitors.

[0332] In some embodiments, a pharmaceutical composition as described herein for use in combination therapy comprises an effective amount of a checkpoint inhibitor as described herein and a pharmaceutically acceptable carrier.

[0333] In some embodiments, the compositions disclosed herein include a checkpoint inhibitor and a pharmaceutically acceptable carrier. In some embodiments, the compositions disclosed herein include a combination of a checkpoint inhibitor and a pharmaceutically acceptable carrier. In some embodiments, the compositions include a checkpoint inhibitor, including a PD-1 inhibitor, a PD-L1 inhibitor, a CTLA-4 inhibitor, a TIGIT inhibitor, a TIM-3 inhibitor, a B7-H3 inhibitor, a CD73 inhibitor, a LAG3 inhibitor, a CD27 inhibitor, a CD70 inhibitor, a 4-1BB inhibitor, a GITR inhibitor, an OX40 inhibitor, a SIRP-α (CD47) inhibitor, a CD39 inhibitor, an ILDR2 inhibitor, a VISTA inhibitor, a BTLA inhibitor, and a VTCN-1 inhibitor. In some embodiments, the check point inhibitor comprises at least two check point inhibitors and a pharmaceutically acceptable carrier, wherein the at least two check point inhibitors are selected from a PD-1 inhibitor, a PD-L1 inhibitor, a CTLA-4 inhibitor, a TIGIT inhibitor, a TIM-3 inhibitor, a B7-H3 inhibitor, a CD73 inhibitor, a LAG3 inhibitor, a CD27 inhibitor, a CD70 inhibitor, a 4-1BB inhibitor, a GITR inhibitor, an OX40 inhibitor, a SIRP-α (CD47) inhibitor, a CD39 inhibitor, an ILDR2 inhibitor, a VISTA inhibitor, a BTLA inhibitor, or a VTCN-1 inhibitor. In some embodiments, the composition comprises at least two check point inhibitors, wherein the at least two check point inhibitors are selected from PD-1 inhibitors, PD-L1 inhibitors, CTLA-4 inhibitors, TIGIT inhibitors, TIM-3 inhibitors, B7-H3 inhibitors, CD73 inhibitors, LAG3 inhibitors, CD27 inhibitors, CD70 inhibitors, 4-1BB inhibitors, GITR inhibitors, OX40 inhibitors, SIRP-α (CD47) inhibitors, CD39 inhibitors, ILDR2 inhibitors, VISTA inhibitors, BTLA inhibitors, and VTCN-1 inhibitors. In some embodiments, the check point inhibitor comprises at least two check point inhibitors and a pharmaceutically acceptable carrier, wherein the at least two check point inhibitors are selected from PD-1 inhibitors, PD-L1 inhibitors, CTLA-4 inhibitors, TIGIT inhibitors, TIM-3 inhibitors, B7-H3 inhibitors, CD73 inhibitors, LAG3 inhibitors, CD27 inhibitors, CD70 inhibitors, 4-1BB inhibitors, GITR inhibitors, OX40 inhibitors, SIRP-α (CD47) inhibitors, CD39 inhibitors, ILDR2 inhibitors, VISTA inhibitors, BTLA inhibitors and VTCN-1 inhibitors.

[0334] In certain embodiments, when more than one checkpoint inhibitor is used in the treatment methods described herein, each checkpoint inhibitor is contained in a separate composition. In certain embodiments, when more than one checkpoint inhibitor is used in the treatment methods described herein, the checkpoint inhibitors can be contained in the same composition.

[0335] In some embodiments, the combination therapy comprises the use of an anti-IL-2 antibody or composition thereof as described herein, and a checkpoint inhibitor or composition thereof. In some embodiments, the combination therapy comprises the use of an anti-IL-2 antibody or composition thereof as described herein and IL-2, and a checkpoint inhibitor or composition thereof. In some embodiments, the combination therapy comprises the use of an anti-IL-2 antibody or composition thereof complexed with IL-2 as described herein, and a checkpoint inhibitor or composition thereof.

[0336] In some embodiments, the combination therapy comprises the use of an anti-IL-2 antibody or composition thereof as described herein, and at least two checkpoint inhibitors or compositions thereof. In some embodiments, the combination therapy comprises the use of an anti-IL-2 antibody or composition thereof as described herein and IL-2, and at least two checkpoint inhibitors or compositions thereof. In some embodiments, the combination therapy comprises the use of an anti-IL-2 antibody or composition thereof complexed with IL-2 as described herein, and at least two checkpoint inhibitors or compositions thereof.

[0337] In some embodiments, the combination therapy includes a second composition comprising one or more checkpoint inhibitors as described herein.

[0338] In some embodiments, the combination therapy includes the use of anti-IL-2 antibody BDG17.069 or a composition thereof; and IL-2 as described herein; and a check point inhibitor or a composition thereof as described herein. In some embodiments, the combination therapy includes the use of BDG17.069 or a composition thereof; and a low dose of IL-2 as described herein; and a check point inhibitor or a composition thereof as described herein. In some embodiments, the combination therapy includes the use of BDG17.069 or a composition thereof; and a low dose of IL-2 as described herein; and a check point inhibitor or a composition thereof, wherein the check point inhibitor is selected from PD-1 inhibitors, PD-L1 inhibitors, CTLA-4 inhibitors, TIGIT inhibitors, TIM-3 inhibitors, B7-H3 inhibitors, CD73 inhibitors, LAG3 inhibitors, CD27 inhibitors, CD70 inhibitors, 4-1BB inhibitors, GITR inhibitors, OX40 inhibitors, SIRP-α (CD47) inhibitors, CD39 inhibitors, ILDR2 inhibitors, VISTA inhibitors, BTLA inhibitors and VTCN-1 inhibitors.

[0339] In some embodiments, the combination therapy includes the use of BDG17.069 or a composition thereof; and a low dose of IL-2 as described herein; and a checkpoint inhibitor or a composition thereof, wherein the checkpoint inhibitor includes PD-L1. In some embodiments, the combination therapy includes the use of BDG17.069 or a composition thereof; and a low dose of IL-2 (aldesleukin); and avelumab or a composition thereof. In certain embodiments of the combination therapy comprising BDG17.069 or a composition thereof; and a low dose of IL-2 (aldesleukin); and avelumab or a composition thereof, IL-2 is administered by subcutaneous injection.

[0340] In some embodiments of the combination therapy, the IL-2 administered comprises a low dose of IL-2. In some embodiments of the combination therapy, the IL-2 administered is administered subcutaneously. In some embodiments of the combination therapy, the IL-2 administered comprises a low dose of IL-2 administered by subcutaneous administration.

[0341] In some embodiments of the combination therapy, the anti-IL-2 antibody and IL-2 are contained in the same composition as the checkpoint inhibitor. In some embodiments, the anti-IL-2 antibody and IL-2 are contained in compositions that are different from each other and from the checkpoint inhibitor. In some embodiments, the anti-IL-2 antibody, IL-2, and the checkpoint inhibitor are contained in the same composition. In some embodiments, the anti-IL-2 antibody and IL-2 are contained in a composition, and the checkpoint inhibitor is contained in a different composition. In some embodiments, the anti-IL-2 antibody and the checkpoint inhibitor are contained in a composition, and the IL-2 is contained in a different composition.

[0342] In some embodiments of the combination therapy, BDG17.069 and aldesleukin are included in the same composition as the PD-L1 checkpoint inhibitor. In some embodiments of the combination therapy, BDG17.069 and aldesleukin are included in the same composition as avelumab. In some embodiments, BDG17.069 and aldesleukin are included in compositions that are different from each other and different from avelumab. In some embodiments, BDG17.069, aldesleukin, and avelumab are included in the same composition. In some embodiments, BDG17.069 and aldesleukin are included in a composition, and avelumab is included in a different composition. In some embodiments, BDG17.069 and avelumab are included in a composition, and aldesleukin is included in a different composition. In some embodiments of the combination therapy, the order of administration of the anti-IL-2 antibody or its composition and the checkpoint inhibitor or its composition can be any order. In some embodiments of the combination therapy, the order of administration of BDG17.069 or its composition and avelumab or its composition can be any order. In some embodiments of the combination therapy, the order of administration of anti-IL-2 antibodies or their compositions, IL-2 or their compositions, and checkpoint inhibitors or their compositions can be any order. In some embodiments of the combination therapy, the order of administration of BDG17.069 or its composition, aldesleukin or its composition, and avelumab or its composition can be any order. For example, but not limited to, the anti-IL-2 antibody can be administered before, simultaneously, or after the administration of the checkpoint inhibitor. Similarly, the combination of anti-IL-2 antibodies and IL-2 can be administered before, simultaneously, or after the administration of the checkpoint inhibitor. For example, but not limited to, BDG17.069 can be administered before, simultaneously, or after the administration of avelumab. Similarly, the combination of BDG17.069 and aldesleukin can be administered before, simultaneously, or after the administration of avelumab. In some embodiments, the anti-IL-2 antibody can be administered before, simultaneously, or after the administration of at least two checkpoint inhibitors. Similarly, the combination of an anti-IL-2 antibody and IL-2 can be administered before, simultaneously with, or after administration of at least two checkpoint inhibitors.

[0343] In some embodiments, administering the combination therapy with a checkpoint inhibitor comprises administering an anti-IL-2 antibody or a composition thereof and a checkpoint inhibitor simultaneously. In some embodiments, administering the combination therapy with a checkpoint inhibitor comprises administering BDG17.069 or a composition thereof and avelumab simultaneously. In some embodiments, administering the combination therapy with a checkpoint inhibitor comprises administering an anti-IL-2 antibody and IL-2 or a composition thereof and a checkpoint inhibitor simultaneously. In some embodiments, administering the combination therapy with a checkpoint inhibitor comprises administering BDG17.069 and aldesleukin or a composition thereof and avelumab simultaneously. In some embodiments, administering the combination therapy with a checkpoint inhibitor comprises administering an anti-IL-2 antibody or a composition thereof in advance before the checkpoint inhibitor. In some embodiments, administering the combination therapy with a checkpoint inhibitor comprises administering BDG17.069 or a composition thereof in advance before avelumab. In some embodiments, administering the combination therapy with a checkpoint inhibitor comprises administering an anti-IL-2 antibody and IL-2 or a composition thereof in advance before the checkpoint inhibitor. In some embodiments, the administration of the combination therapy with the checkpoint inhibitor comprises pre-administration of BDG17.069 and aldesleukin, or a composition thereof, before avelumab. In some embodiments, the administration of the combination therapy with the checkpoint inhibitor comprises post-administration of an anti-IL-2 antibody, or a composition thereof, after the administration of the checkpoint inhibitor. In some embodiments, the administration of the combination therapy with the checkpoint inhibitor comprises post-administration of BDG17.069, or a composition thereof, after the administration of avelumab. In some embodiments, the administration of the combination therapy with the checkpoint inhibitor comprises post-administration of an anti-IL-2 antibody and IL-2, or a composition thereof, after the administration of the checkpoint inhibitor. In some embodiments, the administration of the combination therapy with the checkpoint inhibitor comprises post-administration of BDG17.069 and aldesleukin, or a composition thereof, after the administration of avelumab.

[0344] In some embodiments, the combination therapy comprises the use of a checkpoint inhibitor and an anti-IL-2 antibody comprising a heavy chain variable region having a sequence of one of SEQ ID NO: 10, SEQ ID NO: 12, SEQ ID NO: 14, SEQ ID NO: 16, SEQ ID NO: 18, SEQ ID NO: 20, SEQ ID NO: 22, SEQ ID NO: 24, SEQ ID NO: 26, or SEQ ID NO: 36. In some embodiments, the combination therapy comprises the use of a checkpoint inhibitor and an anti-IL-2 antibody comprising a light chain variable region having a sequence of one of SEQ ID NO: 11, SEQ ID NO: 13, SEQ ID NO: 15, SEQ ID NO: 17, SEQ ID NO: 19, SEQ ID NO: 21, SEQ ID NO: 23, SEQ ID NO: 25, SEQ ID NO: 27, or SEQ ID NO: 37. In some embodiments, the combination therapy comprises the use of a checkpoint inhibitor and an anti-IL-2 antibody comprising a heavy chain variable region and a light chain variable region having a sequence of one of: SEQ ID NO: 10 and SEQ ID NO: 11; SEQ ID NO: 12 and SEQ ID NO: 13; SEQ ID NO: 14 and SEQ ID NO: 15; SEQ ID NO: 16 and SEQ ID NO: 17; SEQ ID NO: 18 and SEQ ID NO: 19; SEQ ID NO: 20 and SEQ ID NO: 21; SEQ ID NO: 22 and SEQ ID NO: 23; SEQ ID NO: 24 and SEQ ID NO: 25; SEQ ID NO: 26 and SEQ ID NO: 27; or SEQ ID NO: 36 and SEQ ID NO: 37. In some embodiments, the combination therapy comprises the use of a checkpoint inhibitor and an anti-IL-2 antibody comprising a heavy chain variable domain comprising CDR1, CDR2, and CDR3 regions comprising the amino acid sequence of SEQ ID NOs: 38-40, respectively; SEQ ID NOs: 44-46, respectively; SEQ ID NOs: 50-52, respectively; SEQ ID NOs: 56-58, respectively; or SEQ ID NOs: 62-64, respectively.In some embodiments, the combination therapy comprises the use of a checkpoint inhibitor and an anti-IL-2 antibody comprising a light chain variable domain comprising CDR1, CDR2, and CDR3 regions comprising the amino acid sequence of SEQ ID NOs: 41-43, respectively; SEQ ID NOs: 47-49, respectively; SEQ ID NOs: 53-55, respectively; SEQ ID NOs: 59-61, respectively; or SEQ ID NOs: 65-67, respectively. In some embodiments, the combination therapy comprises the use of a checkpoint inhibitor and an anti-IL-2 antibody comprising a heavy chain variable domain comprising CDR1, CDR2, and CDR3 regions comprising the amino acid sequences of SEQ ID NOs: 38-40, 44-46, 50-52, 56-58, or 62-64, respectively, and a light chain variable domain comprising CDR1, CDR2, and CDR3 regions comprising the amino acid sequences of SEQ ID NOs: 41-43, 47-49, 53-55, 59-61, or 65-67, respectively.

[0345] In some embodiments, the combination therapy comprises the use of a checkpoint inhibitor and an anti-IL-2 antibody comprising any one of clones BDG 17.014, BDG 17.023, BDG 17.038, BDG 17.043, BDG 17.053, BDG 17.054, BDG 17.066, BDG 17.067, and BDG 17.069. In some embodiments, the combination therapy comprises the use of a checkpoint inhibitor and an anti-IL-2 antibody comprising anti-IL-2 clone BDG 17.014. In some embodiments, the combination therapy comprises the use of a checkpoint inhibitor and an anti-IL-2 antibody comprising anti-IL-2 clone BDG 17.023. In some embodiments, the combination therapy comprises the use of a checkpoint inhibitor and an anti-IL-2 antibody comprising anti-IL-2 clone BDG 17.038. In some embodiments, the combination therapy comprises the use of a checkpoint inhibitor and an anti-IL-2 antibody comprising the anti-IL-2 clone BDG 17.043. In some embodiments, the combination therapy comprises the use of a checkpoint inhibitor and an anti-IL-2 antibody comprising the anti-IL-2 clone BDG 17.053. In some embodiments, the combination therapy comprises the use of a checkpoint inhibitor and an anti-IL-2 antibody comprising the anti-IL-2 clone BDG17.054. In some embodiments, the combination therapy comprises the use of a checkpoint inhibitor and an anti-IL-2 antibody comprising the anti-IL-2 clone BDG 17.066. In some embodiments, the combination therapy comprises the use of a checkpoint inhibitor and an anti-IL-2 antibody comprising the anti-IL-2 clone BDG 17.067. In some embodiments, the combination therapy comprises the use of a checkpoint inhibitor and an anti-IL-2 antibody comprising the anti-IL-2 clone BDG 17.069.

[0346] In some embodiments, the combination therapy comprises the use of a checkpoint inhibitor; and an anti-IL-2 antibody comprising any one of clones BDG 17.014, BDG 17.023, BDG 17.038, BDG 17.043, BDG 17.053, BDG 17.054, BDG 17.066, BDG 17.067, and BDG 17.069; and IL-2. In some embodiments, the combination therapy comprises the use of a checkpoint inhibitor and an anti-IL-2 antibody comprising anti-IL-2 clone BDG 17.014, and IL-2. In some embodiments, the combination therapy comprises the use of a checkpoint inhibitor and an anti-IL-2 antibody comprising anti-IL-2 clone BDG 17.023, and IL-2. In some embodiments, the combination therapy comprises the use of a checkpoint inhibitor and an anti-IL-2 antibody comprising anti-IL-2 clone BDG 17.038, and IL-2. In some embodiments, the combination therapy comprises the use of a checkpoint inhibitor and an anti-IL-2 antibody comprising the anti-IL-2 clone BDG17.043, and IL-2. In some embodiments, the combination therapy comprises the use of a checkpoint inhibitor and an anti-IL-2 antibody comprising the anti-IL-2 clone BDG 17.053, and IL-2. In some embodiments, the combination therapy comprises the use of a checkpoint inhibitor and an anti-IL-2 antibody comprising the anti-IL-2 clone BDG 17.054, and IL-2. In some embodiments, the combination therapy comprises the use of a checkpoint inhibitor and an anti-IL-2 antibody comprising the anti-IL-2 clone BDG 17.066, and IL-2. In some embodiments, the combination therapy comprises the use of a checkpoint inhibitor and an anti-IL-2 antibody comprising the anti-IL-2 clone BDG 17.067, and IL-2. In some embodiments, the combination therapy comprises the use of a checkpoint inhibitor and an anti-IL-2 antibody comprising the anti-IL-2 clone BDG17.069, and IL-2.

[0347] In certain embodiments, the use of the combination therapy is for the treatment of cancer or tumors. In some embodiments, the use of the combination therapy is for the treatment of solid cancers or solid tumors as described herein.

[0348] In some embodiments of the combination therapies disclosed herein, treating solid tumors includes treating primary tumors and secondary metastases of the tumor. In some embodiments of the combination therapies disclosed herein, treating solid tumors includes treating secondary metastases of the tumor. In some embodiments of the combination therapies disclosed herein, treating solid tumors includes second-line treatment of the tumor. In some embodiments of the combination therapies disclosed herein, treating solid tumors includes third-line treatment of the tumor. In some embodiments of the combination therapies disclosed herein, treating solid tumors includes second-line and third-line treatment of the tumor.

[0349] In some embodiments of the combination therapies disclosed herein, the solid tumors treated include metastatic cancers. In some embodiments of the combination therapies disclosed herein, the solid tumors treated include unresectable locally advanced cancers or tumors. In some embodiments of the combination therapies disclosed herein, the solid tumors treated include metastases. In some embodiments of the combination therapies disclosed herein, the solid tumors treated include unresectable locally advanced cancers or tumors.

[0350] In some embodiments of the combination therapies disclosed herein, the solid tumor treated comprises head and neck cancer, head and neck squamous cell carcinoma (HNSCC), pancreatic cancer, lung cancer, thyroid cancer, non-small cell lung cancer (NSCLC), nasopharyngeal carcinoma, melanoma, acral melanoma, uveal melanoma, colorectal cancer (CRC), bladder cancer, cholangiocarcinoma (bile duct cancer), uterine cancer, cervical cancer, gallbladder cancer, or renal cell carcinoma (RCC). In some embodiments of the combination therapies disclosed herein, the solid tumors treated include urothelial carcinoma, adrenocortical carcinoma, clear cell renal cell carcinoma (ccRCC), melanoma, triple-negative breast cancer, head and neck squamous cell carcinoma (HNSCC), gastric cancer or gastroesophageal cancer, esophageal squamous cell carcinoma, cutaneous squamous cell carcinoma (cSCC), pancreatic adenocarcinoma, bile duct carcinoma, hepatocellular carcinoma (HCC), colorectal cancer (CRC), epithelial ovarian cancer, cervical cancer, endometrial cancer, thyroid cancer (follicular or papillary histology), non-small cell lung cancer (NSCLC) or Merkel cell carcinoma. In some embodiments, urothelial carcinoma occurs in the bladder, renal pelvis, ureter or urethra, or any combination thereof. In certain embodiments, the cancer has progressed during or after anti-PDx therapy and, if appropriate, a platinum-containing regimen. In some embodiments, urothelial carcinoma occurs in the bladder, renal pelvis, ureter, or urethra, or any combination thereof, wherein the cancer has progressed during or after anti-PDx therapy and, if appropriate, a platinum-containing regimen. In some embodiments, adrenocortical carcinoma includes unresectable, locally advanced, or metastatic cancer. In some embodiments, clear cell renal cell carcinoma (ccRCC) includes cancer that has progressed during or after at least 2 approved treatment regimens (e.g., small molecule inhibitors, anti-PDx therapy). In some embodiments, melanoma includes locally unresectable or metastatic cancer, wherein the locally unresectable or metastatic cancer may encompass (a) BRAF wild type: wherein the cancer progresses after receiving anti-PD-1 therapy with or without anti-CTLA-4; or (b) BRAF mutant: wherein the cancer progresses after BRAF+MEK inhibitor. In some embodiments, triple-negative breast cancer includes cancer that is unresectable, locally advanced, or metastatic and refractory to standard first-line therapy, which may include, for example, but not limited to, cytotoxic chemotherapy alone and / or poly ADP ribose polymerase (PARP) inhibitors for breast cancer genes (BRCA), 1 or 2 mutations, and / or anti-PDx therapy in MSI-H / dMMR positive tumors. In some embodiments, head and neck squamous cell carcinoma (HNSCC) includes cancer that has progressed during or after treatment of metastatic or recurrent disease with, for example, but not limited to, anti-PDx (unless unsuitable, such as chemotherapy failure and PD-L1 combined positive score (CPS) <1) and platinum-based chemotherapy (unless unsuitable for platinum chemotherapy).In some embodiments, gastric cancer or gastroesophageal cancer includes cancer that progressed during or after cytotoxic chemotherapy (such as but not limited to paclitaxel, fluoropyrimidine, platinum) with or without trastuzumab (for HER2 overexpressing adenocarcinoma) and with or without anti-PD-1 inhibitor therapy. Patients with CPS ≥ 1 may have received an anti-PD-1-containing regimen (unless intolerant or unavailable for treatment). In some embodiments, esophageal squamous cell carcinoma includes cancer that progressed during or after cytotoxic chemotherapy (such as but not limited to paclitaxel, fluoropyrimidine, platinum) using anti-PD-1 therapy. Patients with CPS ≥ 10 may have received an anti-PD-1-containing regimen (unless intolerant or unavailable for treatment). In some embodiments, cutaneous squamous cell carcinoma (cSCC) includes recurrent or metastatic cSCC that cannot be cured by surgery or radiation. In some embodiments, pancreatic adenocarcinoma includes cancer that is unresectable, locally advanced, or metastatic and has received at least one chemotherapy (such as but not limited to FOLFIRINOX; unless inappropriate or infeasible). In some embodiments, cholangiocarcinoma includes unresectable, locally advanced, or metastatic cancer in patients who may have received ≥1 line of systemic chemotherapy, unless the patient is not suitable for chemotherapy. In some embodiments, hepatocellular carcinoma (HCC) includes cancer that progressed during or after an approved treatment regimen (unless not suitable). In some embodiments, colorectal cancer (CRC) comprises (a) K-Ras wild type: patients who have progressed during or after both irinotecan-based and oxaliplatin-based chemotherapy or are not suitable for both irinotecan-based and oxaliplatin-based chemotherapy and relapsed or refractory to at least 1 previous systemic therapy including anti-epidermal growth factor receptor (EGFR) antibodies (such as cetuximab or panitumumab); or (b) K-Ras mutant: patients who have progressed during or after both irinotecan-based and oxaliplatin-based chemotherapy (± bevacizumab) or are not suitable for both. In some embodiments, epithelial ovarian cancer includes cancer that progressed during or after at least one prior cytotoxic chemotherapy regimen (unless unsuitable) and subsequent poly ADP ribose polymerase (PARP) inhibitor therapy (unless unsuitable) in BRCA mutation-positive patients. In some embodiments, cervical cancer includes cancer that progressed during or after first-line cytotoxic chemotherapy and second-line cytotoxic chemotherapy or anti-PDx therapy in PD-L1 positive (CPS ≥ 1) or MSI-H / dMMR positive tumors (unless unsuitable). In some embodiments, endometrial cancer in patients includes cancer that progressed during or after cytotoxic chemotherapy (such as, but not limited to, ± trastuzumab) or hormone therapy and anti-PDx therapy in MSI-H / dMMR positive tumors.In some embodiments, thyroid cancer (follicular or papillary histology) includes iodine-refractory cancer. In some embodiments, non-small cell lung cancer (NSCLC) includes cancer that has progressed during or after treatment with platinum-based chemotherapy and anti-PDx therapy for unresectable, locally advanced or metastatic disease. In some embodiments, NSCLC carrying an activating EGFR mutation (excluding exon 20 insertion mutations) or anaplastic lymphoma kinase (ALK) rearrangement must have progressed after treatment with available EGFR or ALK targeted therapy and platinum-based chemotherapy (unless not suitable for platinum therapy). In some embodiments, Merkel cell carcinoma includes metastatic Merkel cell carcinoma that cannot be cured by surgery or radiation.

[0351] In some embodiments of the combination therapies disclosed herein, the solid tumors treated include, including solid tumors of head and neck cancer, pancreatic cancer or non-small cell lung cancer. In some embodiments of the combination therapies disclosed herein, the solid tumors treated include non-small cell lung cancer (NSCLC), melanoma, metastatic melanoma, primary melanoma and metastatic melanoma, cutaneous squamous cell carcinoma or renal cell carcinoma (RCC). In some embodiments of the combination therapies disclosed herein, melanoma includes acral melanoma or uveal melanoma. In some embodiments of the combination therapies disclosed herein, the solid tumors treated include head and neck cancer. In some embodiments, solid tumors include pancreatic cancer. In some embodiments of the combination therapies disclosed herein, the solid tumors treated include lung cancer. In some embodiments of the combination therapies disclosed herein, the solid tumors treated include thyroid cancer. In some embodiments of the combination therapies disclosed herein, the solid tumors treated include non-small cell lung cancer (NSCLC). In some embodiments of the combination therapies disclosed herein, the solid tumors treated include nasopharyngeal carcinoma. In some embodiments of the combination therapies disclosed herein, the solid tumors treated include melanoma. In some embodiments of the combination therapies disclosed herein, the solid tumors treated include acral melanoma. In some embodiments of the combination therapies disclosed herein, the solid tumors treated include uveal melanoma. In some embodiments of the combination therapies disclosed herein, the solid tumors treated include colorectal cancer (CRC). In some embodiments of the combination therapies disclosed herein, the solid tumors treated include bladder cancer. In some embodiments of the combination therapies disclosed herein, the solid tumors treated include cholangiocarcinoma (bile duct cancer). In some embodiments of the combination therapies disclosed herein, the solid tumors treated include uterine cancer. In some embodiments of the combination therapies disclosed herein, the solid tumors treated include cervical cancer. In some embodiments of the combination therapies disclosed herein, the solid tumors treated include gallbladder cancer. In some embodiments of the combination therapies disclosed herein, the solid tumors treated include renal cell carcinoma (RCC). In some embodiments of the combination therapies disclosed herein, the solid tumors treated include head and neck cancer, wherein the head and neck cancer is head and neck squamous cell carcinoma (HNSCC). In some embodiments of the combination therapies disclosed herein, the solid tumors treated include CRC, wherein the CRC has high MSI. In some embodiments of the combination therapies disclosed herein, the solid tumors treated include melanoma, wherein the melanoma has a wild-type BRAF gene or a mutant BRAF gene. In some embodiments of the combination therapies disclosed herein, the solid tumors treated include pancreatic cancer, wherein the pancreatic cancer is an adenocarcinoma. In some embodiments of the combination therapies disclosed herein, the solid tumors treated include non-small cell lung cancer (NSCLC), wherein the NSCLC is a squamous cell carcinoma.In some embodiments of the combination therapies disclosed herein, the solid tumor treated comprises NSCLC, wherein the NSCLC has a mutated epidermal growth factor receptor (EGFRm). In some embodiments of the combination therapies disclosed herein, the solid tumor treated comprises skin squamous cell carcinoma.

[0352] preparation

[0353] The pharmaceutical compositions disclosed herein comprising anti-IL-2 antibodies, or a combination of anti-IL-2 antibodies and IL-2, or checkpoint inhibitors can be conveniently provided as sterile liquid products, such as isotonic aqueous solutions, suspensions, emulsions, dispersions or viscous compositions, which can be buffered to a selected pH. Liquid products are generally easier to prepare than gels, other viscous compositions and solid compositions. In addition, liquid compositions are slightly easier to administer, especially by injection. On the other hand, viscous compositions can be formulated within an appropriate viscosity range to provide a longer contact period with a specific tissue. The liquid composition or viscous composition may include a vehicle, which may be a solvent or a dispersion medium, which includes, for example, water, saline, phosphate-buffered saline, a polyol (e.g., glycerol, propylene glycol, liquid polyethylene glycol, etc.) and a suitable mixture thereof.

[0354] Sterile injectable solutions can be prepared as needed by incorporating the anti-IL-2 antibodies described herein and used to practice the methods disclosed herein, or a combination of anti-IL-2 antibodies and IL-2, or checkpoint inhibitors into the desired amount of appropriate solvents and various amounts of other ingredients. Such preparations can be mixed with suitable carriers, diluents or excipients such as sterile water, normal saline, glucose, dextrose, etc. The preparation can also be lyophilized. The preparation may include auxiliary substances such as wetting agents, dispersants or emulsifiers (e.g., methylcellulose), pH buffers, gelling additives or viscosity enhancing additives, preservatives, flavorings, colorants, etc., depending on the route of administration and the desired product. Standard textbooks, such as "REMINGTON'S PHARMACEUTICAL SCIENCE," 17th edition, 1985, incorporated herein by reference, can be used to prepare suitable products without excessive experimentation.

[0355] Various additives that enhance the stability and sterility of the formulation may be added, including antimicrobial preservatives, antioxidants, chelating agents, and buffers. Protection against the action of microorganisms may be ensured by various antibacterial and antifungal agents, such as parabens, chlorobutanol, phenol, sorbic acid, and the like. Prolonged absorption of injectable pharmaceutical forms may be produced by the use of agents that delay absorption, such as aluminum monostearate and gelatin.

[0356] In certain embodiments, the terms "pharmaceutical composition," "composition," and "formulation" may be used interchangeably and have the same meaning and properties.

[0357] The compositions or formulations described herein can be isotonic, i.e., they can have the same osmotic pressure as blood and tears. The desired isotonicity of the compositions disclosed herein can be achieved using sodium chloride or other pharmaceutically acceptable agents such as dextrose, boric acid, sodium tartrate, propylene glycol, or other inorganic or organic solutes. For buffers containing sodium ions, sodium chloride may be preferred.

[0358] If necessary, a pharmaceutically acceptable thickening agent may be used to maintain the viscosity of the composition at a selected level.Methylcellulose may be preferred because it is readily and economically available and easy to use.

[0359] In one embodiment, the present invention provides the thickening agent of the present invention.Other suitable thickening agents comprise for example xanthan gum, carboxymethyl cellulose, hydroxypropyl cellulose, carbomer etc.The preferred concentration of thickening agent will depend on selected agent.Emphasis is to use the amount that will reach selected viscosity.Obviously, the selection of suitable carrier and other additives will depend on the character of definite route of administration and specific dosage form, for example liquid dosage form (for example, compositions is to be mixed with solution, suspension, gel or another kind of liquid form, such as timed release form or liquid-filled form).

[0360] In some embodiments, the composition is formulated to have a pH between about pH 5.0-6.0. In some embodiments, the composition is formulated to have a pH between about pH 5.0-7.0. In some embodiments, the composition is formulated to have a pH between about pH 5.0-6.5. In some embodiments, the composition is formulated to have a pH between about pH 5.0-5.5. In some embodiments, the composition is formulated to have a pH between about pH 5.5-6.0. In some embodiments, the composition is formulated to have a pH between about pH 5.5-6.5. In some embodiments, the composition is formulated to have a pH at about pH 5.0. In some embodiments, the composition is formulated to have a pH at about pH 5.5. In some embodiments, the composition is formulated to have a pH at about pH 6.0. In some embodiments, the composition is formulated to have a pH at about pH 6.5.

[0361] In some embodiments, the composition is formulated to have a pH between about pH 5.0 and 6.0 and comprises a buffer. In some embodiments, the buffer comprises a pharmaceutically acceptable buffer. In some embodiments, the buffer comprises a histidine buffer or a citrate buffer. In some embodiments, the buffer comprises a histidine buffer. In some embodiments, the buffer comprises a citrate buffer.

[0362] In some embodiments, the composition is formulated to have a pH between about pH 5.0-6.0 and comprises a buffer selected from a histidine buffer and a citrate buffer. In some embodiments, the composition is formulated to have a pH between about pH 5.0-6.0 and comprises a histidine buffer. In some embodiments, the composition is formulated to have a pH between about pH 5.0-6.0 and comprises a citrate buffer.

[0363] In some embodiments, the composition further comprises at least one of sucrose, methionine, or PS80, or any combination thereof. In some embodiments, the composition further comprises sucrose. In some embodiments, the composition further comprises methionine. In some embodiments, the composition further comprises PS80.

[0364] In some embodiments, the composition comprises an anti-IL-2 antibody as disclosed herein and is formulated to have a pH between about pH 5.0-6.0 and comprises a buffer selected from a histidine buffer and a citrate buffer. In some embodiments, the composition further comprises IL-2.

[0365] Those skilled in the art will recognize that the components of the composition or formulation should be selected to be chemically inert and will not affect the viability or efficacy of the early apoptotic cell population as described herein for use in the methods disclosed herein. From the present disclosure and the documents cited herein, this will not be a problem for those skilled in the art of chemical and pharmaceutical principles, or the problem can be easily avoided by reference to standard textbooks or by simple experiments that do not involve undue experimentation.

[0366] How to use

[0367] In one embodiment, the present disclosure provides a method for producing a heavy chain variable region of an anti-IL-2 antibody, comprising culturing a host cell under conditions conducive to the expression of a vector encoding the heavy chain variable region, thereby producing the heavy chain variable region of the anti-IL-2 antibody.

[0368] In one embodiment, the present disclosure provides a method for producing a light chain variable region of an anti-IL-2 antibody, comprising culturing a host cell under conditions conducive to the expression of a vector encoding the light chain variable region, thereby producing the light chain variable region of the anti-IL-2 antibody.

[0369] The VH polypeptides and / or VL polypeptides disclosed herein can be used in therapeutic methods. In one embodiment, the polypeptides of the present disclosure can be used as immunotherapeutic agents, for example, for differential activation of immune cells as described herein. The polypeptides of the present invention can be administered to a subject directly or by administering to the subject a nucleic acid sequence encoding the polypeptide, which can be carried by a vector.

[0370] The exact amount of the polypeptide of the present invention or its composition required for the desired effect will vary with the subject, depending on the species, age, sex, body weight and general condition of the subject, specific polypeptide, route of administration and whether other drugs are included in the scheme. Therefore, it is impossible to specify an exact amount for each composition. However, the appropriate amount can be determined by those of ordinary skill in the art using routine experiments. Dosage can vary, and the polypeptide can be used with one or more doses (e.g., two or more doses, three or more doses, four or more doses or five or more doses) per day for one or more days. Guidance on selecting appropriate dosage for the antibody can be easily found in the literature.

[0371] In some embodiments of the methods of use of the anti-IL-2 antibodies described herein, the subject comprises a mammalian subject. In some embodiments, the subject comprises a human subject. In some embodiments, the subject suffers from an immunodeficiency problem. In some embodiments, treatment of an immunodeficient subject will include prophylactic treatment.

[0372] In one embodiment, the present disclosure provides a method for promoting differential growth of immune cells in a subject, the method comprising the steps of preparing a composition comprising an anti-IL-2 antibody disclosed herein, and administering the composition to a subject, thereby promoting differential growth of immune cells in the subject. In one embodiment, the present disclosure provides a method for promoting differential growth of immune cells in a subject, the method comprising preparing a composition comprising IL-2 and an anti-IL-2 antibody disclosed herein, and administering the composition to a subject, thereby promoting differential growth of immune cells in the subject. In one embodiment, the subject can be an animal or a human. In one embodiment, the immune cells can be CD8 + cells or NK cells.

[0373] In some embodiments, disclosed herein is a method of treating a disease or condition in a subject, comprising administering to the subject a composition comprising an anti-IL-2 antibody as disclosed herein, wherein the antibody promotes differential growth of immune cell subsets and reduces undesirable effects caused by IL-2, thereby treating the disease or condition in the subject. In some embodiments, the methods of treating a disease disclosed herein comprise the use of a composition comprising an anti-IL-2 antibody and IL-2, or an anti-IL-2 antibody complexed with IL-2. In some embodiments, the methods of treating a disease include treating a viral infection, a bacterial infection, or cancer. In some embodiments, the methods of treating a condition include treating a weakened immune system and treating prophylactically to enhance the immune system.

[0374] In some embodiments of the method of treating a disease or condition, the condition comprises a genetic predisposition that increases the likelihood of cancer in the subject. In some embodiments, the genetic predisposition comprises an alteration in the expression or activity of a gene product. In some embodiments, the genetic predisposition that increases the likelihood of cancer comprises a mutation in a tumor suppressor gene or a mismatch repair (MMR) gene, or a combination thereof.

[0375] Many hereditary cancers are known in the art, non-limiting examples include but are not limited to hereditary breast and ovarian cancer (HBOC) syndrome, Lynch syndrome (hereditary nonpolyposis colorectal cancer), and Li-Fraumeni syndrome.

[0376] In some embodiments, a genetic predisposition increases the likelihood of HBOC. HBOC is associated with mutations in the BRAC1 and BRAC2 genes. HBOC is associated with many different cancers, not just breast cancer, including but not limited to fallopian tube cancer, primary peritoneal cancer, male breast cancer, pancreatic cancer, and prostate cancer. In some embodiments, a genetic predisposition increases the likelihood of developing any one of breast cancer, ovarian cancer, fallopian tube cancer, primary peritoneal cancer, male breast cancer, pancreatic cancer, or prostate cancer, or a combination thereof.

[0377] In some embodiments, a genetic predisposition increases the likelihood of developing hereditary non-polyposis colorectal cancer (HNPCC). HNPCC is associated with mutations in genes including, but not limited to, MLH1, MSH2, MSH6, PMS1, and PMS2. HNPCC is associated with a higher risk of developing endometrial cancer, as well as ovarian cancer, stomach cancer, small intestine cancer, pancreatic cancer, kidney cancer, brain cancer, ureteral cancer, and bile duct cancer. In some embodiments, a genetic predisposition increases the likelihood of developing any of hereditary non-polyposis colorectal cancer, ovarian cancer, stomach cancer, small intestine cancer, pancreatic cancer, kidney cancer, brain cancer, ureteral cancer, and bile duct cancer.

[0378] In some embodiments, genetic susceptibility increases the likelihood of Ley-Fraumeni syndrome. Ley-Fraumeni syndrome is associated with mutations in genes including, but not limited to, TP53 and CHEK2, or a combination thereof. Ley-Fraumeni syndrome is associated with cancers including sarcomas, osteosarcomas, soft tissue sarcomas, leukemias, brain (central nervous system) cancer, adrenocortical carcinoma, and breast cancer, or a combination thereof. In some embodiments, genetic susceptibility increases the likelihood of any one of sarcomas, osteosarcomas, soft tissue sarcomas, leukemias, brain (central nervous system) cancer, adrenocortical carcinoma, and breast cancer, or a combination thereof.

[0379] The anti-IL2 antibodies described and exemplified herein bind to the portion of IL-2 that interacts with the α (CD25) receptor subunit, which is a component of the IL-2 trimeric receptor (CD25 / CD132 / CD122, sometimes expressed as α / β / γ) found on Treg cells, eosinophils, and lung and vascular endothelial cells. In some embodiments, the anti-IL2 antibodies disclosed herein prevent the activation of the trimeric IL-2 receptor found on Treg, eosinophils, and lung and vascular endothelial cells. In some embodiments, the anti-IL2 antibodies disclosed herein that bind to IL-2 activate signaling by IL-2 dimer receptors (CD132 / CD122, sometimes expressed as β / γ) found on naive Teff cells, NK cells, and natural killer T (NKT) cells.

[0380] In some embodiments, the condition being treated in the subject comprises treating a subject with a genetic predisposition comprising altered expression or activity of a gene product comprising BRCA1, BRAC2, MLH1, MSH2, MSH6, PMS1, PMS2, TP53, or CHEK2, or a combination thereof.

[0381] As described herein, the complex of IL-2 and the disclosed anti-IL-2 antibodies is effective in inducing memory phenotype effector T cells (MP) CD8 + cells and NK cells showed significant effects on the proliferation of CD4 + Tregs have a much smaller effect. Therefore, the engineered anti-IL-2 antibodies disclosed herein will be useful for modulating immune cell populations and inducing differential expansion of certain immune effector cells. In one embodiment, such differential expansion of immune effector cells will lead to robust activation of the immune system and can be used for the treatment of tumors.

[0382] In some embodiments, treatment includes treating solid tumors. In some embodiments, treatment includes treating non-solid tumors. In some embodiments, treatment includes treating solid tumors and / or non-solid tumors, such as, but not limited to, melanoma, renal cell carcinoma, small cell lung cancer, or other cancer conditions. In another embodiment, the methods disclosed herein can be used to treat viral infections or bacterial infections. In another embodiment, the methods disclosed herein can be used to treat or prevent conditions caused by IL-2 binding to endothelial CD25 expressing cells, such as pulmonary edema or IL-2 induced vascular leakage.

[0383] In some embodiments, solid tumors are treated using the methods described for treating a disease or condition. In some embodiments, solid tumors include head and neck cancer, head and neck squamous cell carcinoma (HNSCC), pancreatic cancer, lung cancer, thyroid cancer, non-small cell lung cancer (NSCLC), nasopharyngeal carcinoma, melanoma, acral melanoma, uveal melanoma, colorectal cancer (CRC), bladder cancer, bile duct cancer (bile duct cancer), uterine cancer, cervical cancer, gallbladder cancer, skin squamous cell carcinoma, or renal cell carcinoma (RCC). In some embodiments, solid tumors include head and neck cancer, pancreatic cancer, or non-small cell lung cancer. In some embodiments, solid tumors include non-small cell lung cancer (NSCLC), melanoma, metastatic melanoma, primary melanoma, and metastatic melanoma or renal cell carcinoma (RCC). In some embodiments, solid tumors include head and neck cancer. In some embodiments, solid tumors include pancreatic cancer. In some embodiments, solid tumors include lung cancer. In some embodiments, solid tumors include thyroid cancer. In some embodiments, solid tumors include non-small cell lung cancer (NSCLC). In some embodiments, solid tumors include nasopharyngeal carcinoma. In some embodiments, solid tumors include melanomas. In some embodiments, melanomas include acral melanomas or uveal melanomas. In some embodiments, solid tumors include colorectal cancer (CRC). In some embodiments, solid tumors include bladder cancer. In some embodiments, solid tumors include bile duct cancer (bile duct cancer). In some embodiments, solid tumors include uterine cancer. In some embodiments, solid tumors include cervical cancer. In some embodiments, solid tumors include gallbladder cancer. In some embodiments, solid tumors include renal cell carcinoma (RCC). In some embodiments, head and neck cancer is head and neck squamous cell carcinoma (HNSCC). In some embodiments, CRC has high MSI. In some embodiments, melanomas have wild-type BRAF genes. In some embodiments, melanomas have mutant BRAF genes. In some embodiments, pancreatic cancer is adenocarcinoma. In some embodiments, non-small cell lung cancer (NSCLC) is squamous carcinoma. In some embodiments, NSCLC has mutated epidermal growth factor receptor (EGFRm). In some embodiments, solid tumors include cutaneous squamous cell carcinoma.

[0384] In some embodiments, the method for treating cancer includes treating solid tumors. In certain embodiments, the solid tumors treated include urothelial carcinoma, adrenocortical carcinoma, clear cell renal cell carcinoma (ccRCC), melanoma, triple negative breast cancer, head and neck squamous cell carcinoma (HNSCC), gastric cancer or gastroesophageal cancer, esophageal squamous cell carcinoma, skin squamous cell carcinoma (cSCC), pancreatic adenocarcinoma, bile duct cancer, hepatocellular carcinoma (HCC), colorectal cancer (CRC), epithelial ovarian cancer, cervical cancer, endometrial cancer, thyroid cancer (follicular or papillary histology), non-small cell lung cancer (NSCLC) or Merkel cell carcinoma. In some embodiments, urothelial carcinoma occurs in bladder, renal pelvis, ureter or urethra or any combination thereof. In certain embodiments, cancer is in anti-PDx therapy and, if applicable, during or after platinum-containing regimens have progressed. In some embodiments, urothelial carcinoma occurs in the bladder, renal pelvis, ureter, or urethra, or any combination thereof, wherein the cancer has progressed during or after anti-PDx therapy and, if appropriate, a platinum-containing regimen. In some embodiments, adrenocortical carcinoma includes unresectable, locally advanced, or metastatic cancer. In some embodiments, clear cell renal cell carcinoma (ccRCC) includes cancer that has progressed during or after at least 2 approved treatment regimens (e.g., small molecule inhibitors, anti-PDx therapy). In some embodiments, melanoma includes locally unresectable or metastatic cancer, wherein the locally unresectable or metastatic cancer may encompass (a) BRAF wild type: wherein the cancer progresses after receiving anti-PD-1 therapy with or without anti-CTLA-4 therapy; or (b) BRAF mutant: wherein the cancer progresses after BRAF+MEK inhibitor. In some embodiments, triple-negative breast cancer includes cancer that is unresectable, locally advanced, or metastatic and refractory to standard first-line therapy, which may include, for example, but not limited to, cytotoxic chemotherapy alone and / or poly ADP ribose polymerase (PARP) inhibitors for breast cancer genes (BRCA), 1 or 2 mutations, and / or anti-PDx therapy in MSI-H / dMMR positive tumors. In some embodiments, head and neck squamous cell carcinoma (HNSCC) includes cancer that has progressed during or after treatment of metastatic or recurrent disease with, for example, but not limited to, anti-PDx (unless unsuitable, such as chemotherapy failure and PD-L1 combined positive score (CPS) <1) and platinum-based chemotherapy (unless unsuitable for platinum chemotherapy). In some embodiments, gastric cancer or gastroesophageal cancer includes cancer that has progressed during or after treatment with cytotoxic chemotherapy (such as, but not limited to, paclitaxel, fluoropyrimidine, platinum) with or without trastuzumab (for HER2 overexpressing adenocarcinoma) and with or without anti-PD-1 inhibitor therapy. Patients with a CPS ≥1 may have received an anti-PD-1 containing regimen (unless intolerant or unavailable).In some embodiments, esophageal squamous cell carcinoma includes cancer that progressed during or after cytotoxic chemotherapy (such as, but not limited to, paclitaxel, fluoropyrimidine, platinum) using anti-PD-1 therapy. Patients with CPS ≥ 10 may have received a regimen containing anti-PD-1 (unless intolerant or unavailable for treatment). In some embodiments, cutaneous squamous cell carcinoma (cSCC) includes recurrent or metastatic cSCC that cannot be cured by surgery or radiation. In some embodiments, pancreatic adenocarcinoma includes cancer that is unresectable, locally advanced, or metastatic and has received at least one line of chemotherapy (such as, but not limited to, FOLFIRINOX; unless unsuitable or infeasible). In some embodiments, cholangiocarcinoma includes cancer that is unresectable, locally advanced, or metastatic in patients who may have received ≥ 1 line of systemic chemotherapy, unless the patient is not suitable for chemotherapy. In some embodiments, hepatocellular carcinoma (HCC) includes cancer that progressed during or after an approved treatment regimen (unless unsuitable). In some embodiments, colorectal cancer (CRC) comprises (a) K-Ras wild type: patients who have progressed during or after both irinotecan-based and oxaliplatin-based chemotherapy or are not suitable for both irinotecan-based and oxaliplatin-based chemotherapy and have relapsed or are refractory to at least 1 prior systemic therapy including an anti-epidermal growth factor receptor (EGFR) antibody (such as cetuximab or panitumumab); or (b) K-Ras mutant: patients who have progressed during or after both irinotecan-based and oxaliplatin-based chemotherapy (± bevacizumab) or are not suitable for both. In some embodiments, epithelial ovarian cancer comprises cancer that progressed during or after at least one prior cytotoxic chemotherapy regimen (unless not suitable) and subsequent poly ADP ribose polymerase (PARP) inhibitor therapy (unless not suitable) in BRCA mutation-positive patients. In some embodiments, cervical cancer includes cancer that has progressed during or after first-line cytotoxic chemotherapy and second-line cytotoxic chemotherapy or anti-PDx therapy in PD-L1 positive (CPS ≥ 1) or MSI-H / dMMR positive tumors (unless unsuitable). In some embodiments, endometrial cancer in patients includes cancer that has progressed during or after cytotoxic chemotherapy (such as, but not limited to, ± trastuzumab) or hormone therapy and anti-PDx therapy in MSI-H / dMMR positive tumors. In some embodiments, thyroid cancer (follicular or papillary histology) includes iodine-refractory cancer. In some embodiments, non-small cell lung cancer (NSCLC) includes cancer that has progressed during or after treatment with platinum-based chemotherapy and anti-PDx therapy for unresectable, locally advanced, or metastatic disease.In some embodiments, NSCLC harboring an activating EGFR mutation (excluding exon 20 insertion mutations) or an anaplastic lymphoma kinase (ALK) rearrangement must have progressed after treatment with available EGFR or ALK targeted therapy and platinum-based chemotherapy (unless not suitable for platinum therapy). In some embodiments, Merkel cell carcinoma includes metastatic Merkel cell carcinoma that is not curable by surgery or radiation.

[0385] In some embodiments, treating a solid tumor comprises treating a primary tumor and a secondary metastasis of the tumor. In some embodiments, treating a solid tumor comprises treating a secondary metastasis of the tumor. In some embodiments, treating a solid tumor comprises treating a second-line treatment of the tumor. In some embodiments, treating a solid tumor comprises treating a third-line treatment of the tumor. In some embodiments, treating a solid tumor comprises treating a second-line and third-line treatment of the tumor.

[0386] As used throughout, the terms "cancer" and "tumor" are used interchangeably in some embodiments and have the same meaning and properties.

[0387] In some embodiments, the solid tumor being treated comprises a metastatic cancer. In some embodiments, the solid tumor being treated comprises an unresectable locally advanced cancer or tumor. In some embodiments, the solid tumor being treated comprises a metastasis. In some embodiments, the solid tumor being treated comprises an unresectable locally advanced cancer or tumor.

[0388] In some embodiments, the subject treated by the methods disclosed herein has any of 19 solid tumors. In some embodiments, the solid tumor comprises an unresectable locally advanced or metastatic cancer. In some embodiments, the solid tumor comprises an unresectable locally advanced cancer. In some embodiments, the solid tumor comprises a metastatic cancer. In some embodiments, the subject treated by the methods disclosed herein is not suitable for treatment with standard and / or approved therapies. In some embodiments, the subject being treated is human.

[0389] In some embodiments, the subject treated by the methods disclosed herein has or suffers from urothelial carcinoma, adrenocortical carcinoma, clear cell renal cell carcinoma (ccRCC), melanoma, triple-negative breast cancer, head and neck squamous cell carcinoma (HNSCC), gastric cancer or gastroesophageal cancer, esophageal squamous cell carcinoma, cutaneous squamous cell carcinoma (cSCC), pancreatic adenocarcinoma, bile duct carcinoma, hepatocellular carcinoma (HCC), colorectal cancer (CRC), epithelial ovarian cancer, cervical cancer, endometrial cancer, thyroid cancer (follicular or papillary histology), non-small cell lung cancer (NSCLC) or Merkel cell carcinoma. In some embodiments, urothelial carcinoma occurs in the bladder, renal pelvis, ureter or urethra, or any combination thereof. In certain embodiments, the cancer has progressed during or after anti-PDx therapy and, if applicable, a platinum-containing regimen. In some embodiments, urothelial carcinoma occurs in the bladder, renal pelvis, ureter or urethra, or any combination thereof, wherein the cancer has progressed during or after anti-PDx therapy and, if applicable, a platinum-containing regimen. In some embodiments, adrenocortical carcinoma includes unresectable, locally advanced, or metastatic cancer. In some embodiments, clear cell renal cell carcinoma (ccRCC) includes cancer that has progressed during or after at least 2 approved treatment regimens (e.g., small molecule inhibitors, anti-PDx therapy). In some embodiments, melanoma includes locally unresectable or metastatic cancer, wherein the locally unresectable or metastatic cancer may encompass (a) BRAF wild type: wherein the cancer progresses after receiving anti-PD-1 therapy with or without anti-CTLA-4; or (b) BRAF mutant: wherein the cancer progresses after BRAF+MEK inhibitor. In some embodiments, triple-negative breast cancer includes cancer that is unresectable, locally advanced, or metastatic and is refractory to standard first-line therapy, which may include, for example, but not limited to, cytotoxic chemotherapy alone and / or poly ADP ribose polymerase (PARP) inhibitors for breast cancer genes (BRCA), 1 or 2 mutations, and / or anti-PDx therapy in MSI-H / dMMR positive tumors. In some embodiments, head and neck squamous cell carcinoma (HNSCC) includes cancers that have progressed during or after treatment with, for example, but not limited to, anti-PDx (unless not suitable, e.g., patients who have failed chemotherapy and have a PD-L1 combined positive score (CPS) <1) and platinum-based chemotherapy (unless not suitable for platinum chemotherapy) for metastatic or recurrent disease. In some embodiments, gastric or gastroesophageal cancer includes cancers that have progressed during or after treatment with cytotoxic chemotherapy (e.g., but not limited to, paclitaxel, fluoropyrimidine, platinum) with or without trastuzumab (for HER2-overexpressing adenocarcinoma) and with or without anti-PD-1 inhibitor therapy. Patients with a CPS ≥ 1 may have received an anti-PD-1 containing regimen (unless intolerant or unavailable).In some embodiments, esophageal squamous cell carcinoma includes cancer that progressed during or after cytotoxic chemotherapy (such as, but not limited to, paclitaxel, fluoropyrimidine, platinum) using anti-PD-1 therapy. Patients with CPS ≥ 10 may have received a regimen containing anti-PD-1 (unless intolerant or unavailable for treatment). In some embodiments, cutaneous squamous cell carcinoma (cSCC) includes recurrent or metastatic cSCC that cannot be cured by surgery or radiation. In some embodiments, pancreatic adenocarcinoma includes cancer that is unresectable, locally advanced, or metastatic and has received at least one line of chemotherapy (such as, but not limited to, FOLFIRINOX; unless unsuitable or infeasible). In some embodiments, cholangiocarcinoma includes cancer that is unresectable, locally advanced, or metastatic in patients who may have received ≥ 1 line of systemic chemotherapy, unless the patient is not suitable for chemotherapy. In some embodiments, hepatocellular carcinoma (HCC) includes cancer that progressed during or after an approved treatment regimen (unless unsuitable). In some embodiments, colorectal cancer (CRC) comprises (a) K-Ras wild type: patients who have progressed during or after both irinotecan-based and oxaliplatin-based chemotherapy or are not suitable for both irinotecan-based and oxaliplatin-based chemotherapy and have relapsed or are refractory to at least 1 prior systemic therapy including an anti-epidermal growth factor receptor (EGFR) antibody (such as cetuximab or panitumumab); or (b) K-Ras mutant: patients who have progressed during or after both irinotecan-based and oxaliplatin-based chemotherapy (± bevacizumab) or are not suitable for both. In some embodiments, epithelial ovarian cancer comprises cancer that progressed during or after at least one prior cytotoxic chemotherapy regimen (unless not suitable) and subsequent poly ADP ribose polymerase (PARP) inhibitor therapy (unless not suitable) in BRCA mutation-positive patients. In some embodiments, cervical cancer includes cancer that has progressed during or after first-line cytotoxic chemotherapy and second-line cytotoxic chemotherapy or anti-PDx therapy in PD-L1 positive (CPS ≥ 1) or MSI-H / dMMR positive tumors (unless unsuitable). In some embodiments, endometrial cancer in patients includes cancer that has progressed during or after cytotoxic chemotherapy (such as, but not limited to, ± trastuzumab) or hormone therapy and anti-PDx therapy in MSI-H / dMMR positive tumors. In some embodiments, thyroid cancer (follicular or papillary histology) includes iodine-refractory cancer. In some embodiments, non-small cell lung cancer (NSCLC) includes cancer that has progressed during or after treatment with platinum-based chemotherapy and anti-PDx therapy for unresectable, locally advanced, or metastatic disease.In some embodiments, NSCLC harboring an activating EGFR mutation (excluding exon 20 insertion mutations) or an anaplastic lymphoma kinase (ALK) rearrangement must have progressed after treatment with available EGFR or ALK targeted therapy and platinum-based chemotherapy (unless not suitable for platinum therapy). In some embodiments, Merkel cell carcinoma includes metastatic Merkel cell carcinoma that is not curable by surgery or radiation.

[0390] In some embodiments of the method of treating a disease or condition, the immune cells exhibiting differential growth include one or more of naive T cells, memory T cells, CD8+ T cells, NK cells, or natural killer T cells. In some embodiments of the method of treating a disease or condition, the undesirable effects caused by IL-2 include one or more of activation of regulatory T cells, apoptosis of CD25+ T effector cells, IL-2-induced pulmonary edema, IL-2-induced pneumonitis, or IL-2-induced vascular leakage. In some embodiments of the method of treating a disease or condition, an anti-IL-2 antibody disclosed herein inhibits the binding of IL-2 to CD25.

[0391] In some embodiments, treatment of cancer includes maintenance therapy. In some embodiments, maintenance therapy is administered to maintain the absence of cancer or tumor. In some embodiments, maintenance therapy is administered to maintain the absence of metastasis of cancer or tumor. In some embodiments, maintenance therapy is administered to inhibit metastasis of cancer or tumor. In some embodiments, maintenance therapy is administered to maintain the absence of growth of cancer or tumor. In some embodiments, maintenance therapy is administered to inhibit the growth of cancer or tumor.

[0392] In some embodiments, treating a solid cancer in a subject reduces the size of the tumor, inhibits or reduces the growth of the tumor, or inhibits or reduces metastasis of the tumor, or any combination thereof.

[0393] In some embodiments, treatment of cancer includes prophylactic treatment, such as, but not limited to, subjects carrying one or more genetic markers that are at high risk for developing cancer. In some embodiments, the genetic markers include mutations in the BRCA1 gene.

[0394] In some embodiments of the method of promoting differential growth of immune cells in a subject, the method comprises the steps of preparing and administering a composition comprising an anti-IL-2 antibody disclosed herein.

[0395] In some embodiments of the method for promoting differential growth of immune cells in a subject, the method comprises the steps of preparing and administering a composition comprising IL-2 and an anti-IL-2 antibody disclosed herein, and administering the combination of the anti-IL-2 antibody and IL-2, or a composition thereof, is simultaneous. In some embodiments of the method for promoting differential growth of immune cells in a subject, the method comprises the steps of preparing and administering a composition comprising IL-2 and an anti-IL-2 antibody, and administering the anti-IL-2 antibody and IL-2, or a composition thereof, comprises administering the anti-IL-2 antibody or a composition thereof before IL-2, or a composition thereof. In some embodiments of the method for promoting differential growth of immune cells in a subject, the method comprises the steps of preparing and administering a composition comprising IL-2 and an anti-IL-2 antibody, and administering the anti-IL-2 antibody and IL-2, or a composition thereof, comprises administering the anti-IL-2 antibody or a composition thereof after administering IL-2, or a composition thereof.

[0396] In some embodiments, the present disclosure provides a method for treating a subject having a disease or condition by inducing differential growth of immune cells. In one embodiment, the disease can be a viral infection, a bacterial infection, or cancer. In one embodiment, the condition can be IL-2-induced pulmonary edema or IL-2-induced vascular leakage. The method comprises the following steps:

[0397] (a) preparing a composition comprising an anti-IL-2 antibody as disclosed herein; and

[0398] (b) administering the composition from (a) to the subject, thereby treating the subject by differential growth of immune cells in the subject.In certain embodiments, any of the engineered anti-IL-2 antibodies disclosed herein can be used in the methods of treatment as described.

[0399] In some embodiments, the present disclosure provides a method for treating a subject having a disease or condition by inducing differential growth of immune cells. In one embodiment, the disease can be a viral infection, a bacterial infection, or cancer. In one embodiment, the condition can be IL-2-induced pulmonary edema or IL-2-induced vascular leakage. The method comprises the following steps:

[0400] (a) preparing a composition comprising IL-2 and an anti-IL-2 antibody as disclosed herein; and

[0401] (b) administering the composition from (a) to the subject, thereby treating the subject by the differential growth of immune cells in the subject. In addition to promoting the expansion of immune effector cell subsets, the antibody / IL-2 complex will also reduce the undesirable effects caused by IL-2 (e.g., IL-2-induced pulmonary edema or IL-2-induced vascular leakage). In one embodiment, the subject can be an animal or a human. In certain embodiments, any engineered anti-IL-2 antibody disclosed herein can be used in a method of treatment as described.

[0402] In one embodiment, the present disclosure provides a method of treating a disease or condition in a subject (e.g., an animal or human), the method comprising administering to the subject a composition comprising an anti-IL-2 antibody, wherein the antibody promotes the expansion of a subset of immune cells and reduces undesirable effects caused by IL-2, thereby treating the disease or condition in the subject. In some embodiments of the method of treating a disease or condition in a subject, the method comprises the steps of preparing and administering a composition comprising an anti-IL-2 antibody disclosed herein, and administering a composition comprising an anti-IL-2 antibody. In one embodiment, the composition comprises IL-2 and an anti-IL-2 antibody as disclosed herein, or the composition comprises an anti-IL-2 antibody complexed with IL-2.

[0403] In some embodiments of the method of treating a disease or condition in a subject, the method comprises the steps of preparing and administering a composition comprising IL-2 and an anti-IL-2 antibody disclosed herein, wherein administering the combination of the anti-IL-2 antibody and IL-2, or a composition thereof, is simultaneous. In some embodiments of the method of treating a disease or condition in a subject, the method comprises the steps of preparing and administering a composition comprising IL-2 and an anti-IL-2 antibody, wherein administering the anti-IL-2 antibody and IL-2, or a composition thereof, comprises administering the anti-IL-2 antibody or a composition thereof before administering the IL-2 or a composition thereof. In some embodiments of the method of treating a disease or condition in a subject, the method comprises the steps of preparing and administering a composition comprising IL-2 and an anti-IL-2 antibody, wherein administering the anti-IL-2 antibody and IL-2, or a composition thereof, comprises administering the anti-IL-2 antibody or a composition thereof after administering the IL-2 or a composition thereof.

[0404] In one embodiment, the method of treatment will be effective for treating conditions such as IL-2 induced pulmonary edema or IL-2 induced vascular leakage. In another embodiment, the method of treatment will be effective for treating pulmonary edema (mild or chronic) caused by viral infection or bacterial infection.

[0405] In one embodiment, the disease can be a viral infection, a bacterial infection, a cancer, an autoimmune disease, or an immune disorder. In one embodiment, the disease can be an upper respiratory tract viral infection, an early lung infection, or a late lung infection. Many diseases and cancers are known to be caused by viruses. Examples of pathogenic viruses include, but are not limited to, norovirus, rotavirus, hepatitis A virus, hepatitis B virus, hepatitis C virus, hepatitis D virus, or hepatitis E virus, rabies virus, West Nile virus, enterovirus, echovirus, coxsackievirus, herpes simplex virus (HSV), HSV-2, varicella-zoster virus, mosquito-borne virus, arbovirus, St. Louis encephalitis virus, California encephalitis virus, lymphocytic choriomeningitis virus, human immunodeficiency virus (HIV), poliovirus, Zika virus, rubella virus, cytomegalovirus, human papillomavirus (HPV), enterovirus D68, severe acute respiratory syndrome (SARS) coronavirus, Middle East respiratory syndrome coronavirus, SARS coronavirus 2, Epstein-Barr virus, influenza virus, respiratory syncytial virus, polyomavirus (such as JC virus, BK virus), Ebola virus, dengue virus, or any combination thereof. In one embodiment, the viral infection is caused by SARS CoV-2. In another embodiment, the cancer can be, but is not limited to, melanoma or renal cell carcinoma.

[0406] In one embodiment, immune cells expanded by treatment with anti-IL-2 antibodies include naive T cells, memory T cells, CD8 + In one embodiment, treatment with an anti-IL-2 antibody will reduce one or more undesirable effects caused by IL-2, such as activation of regulatory T cells, CD25 + Apoptosis of T effector cells, pulmonary edema, pneumonia, and IL-2-induced vascular leakage.

[0407] In one embodiment, the anti-IL-2 antibody administered in the above method is an engineered or modified anti-IL-2 antibody that can inhibit the binding of IL-2 to CD25. In some embodiments, the engineered or modified anti-IL-2 antibody comprises a heavy chain variable region having a sequence of one of SEQ ID NO: 10, SEQ ID NO: 12, SEQ ID NO: 14, SEQ ID NO: 16, SEQ ID NO: 18, SEQ ID NO: 20, SEQ ID NO: 22, SEQ ID NO: 24, SEQ ID NO: 26, or SEQ ID NO: 36. In some embodiments, the engineered or modified anti-IL-2 antibody comprises a light chain variable region having a sequence of one of SEQ ID NO: 11, SEQ ID NO: 13, SEQ ID NO: 15, SEQ ID NO: 17, SEQ ID NO: 19, SEQ ID NO: 21, SEQ ID NO: 23, SEQ ID NO: 25, SEQ ID NO: 27, or SEQ ID NO: 37. In some embodiments, the engineered or modified anti-IL-2 antibody comprises a heavy chain variable region and a light chain variable region having a sequence of one of the following: SEQ ID NO: 10 and SEQ ID NO: 11; SEQ ID NO: 12 and SEQ ID NO: 13; SEQ ID NO: 14 and SEQ ID NO: 15; SEQ ID NO: 16 and SEQ ID NO: 17; SEQ ID NO: 18 and SEQ ID NO: 19; SEQ ID NO: 20 and SEQ ID NO: 21; SEQ ID NO: 22 and SEQ ID NO: 23; SEQ ID NO: 24 and SEQ ID NO: 25; SEQ ID NO: 26 and SEQ ID NO: 27; or SEQ ID NO: 36 and SEQ ID NO: 37.

[0408] In another embodiment, the engineered or modified anti-IL-2 antibody comprises a heavy chain variable region comprising a complementarity determining region (CDR) 1, CDR2, and CDR3. In one embodiment, the heavy chain CDR1, CDR2, and CDR3 comprise the amino acid sequences of SEQ ID NOs: 38-40, 44-46, 50-52, 56-58, or 62-64, respectively.

[0409] In another embodiment, the engineered or modified anti-IL-2 antibody comprises a light chain variable region comprising a complementarity determining region (CDR) 1, CDR2, and CDR3. In one embodiment, the light chain CDR1, CDR2, and CDR3 comprise the amino acid sequences of SEQ ID NOs: 41-43, 47-49, 53-55, 59-61, or 65-67, respectively.

[0410] In some embodiments, the engineered anti-IL-2 antibody can be IgG, IgA, IgM, IgE, IgD, Fv, scFv, Fab, or F(ab')2. IgG can be of the subclass IgG1, IgG2, IgG3, or IgG4. In some embodiments, the engineered antibody can be part of a minibody, diabody, or triabody.

[0411] In some embodiments, a polynucleotide sequence encoding an engineered anti-IL-2 antibody is used in a method of treating a subject having a disease or condition as described herein, wherein the polynucleotide encodes an antibody comprising a heavy chain variable region having an amino acid sequence of one of SEQ ID NO: 10, SEQ ID NO: 12, SEQ ID NO: 14, SEQ ID NO: 16, SEQ ID NO: 18, SEQ ID NO: 20, SEQ ID NO: 22, SEQ ID NO: 24, SEQ ID NO: 26, or SEQ ID NO: 36. In some embodiments, a polynucleotide sequence encoding an engineered anti-IL-2 antibody is used in a method of treating a subject having a disease or condition as described herein, wherein the polynucleotide encodes an antibody comprising a light chain variable region having an amino acid sequence of one of SEQ ID NO: 11, SEQ ID NO: 13, SEQ ID NO: 15, SEQ ID NO: 17, SEQ ID NO: 19, SEQ ID NO: 21, SEQ ID NO: 23, SEQ ID NO: 25, SEQ ID NO: 27, or SEQ ID NO: 37. In some embodiments, a polynucleotide sequence encoding an engineered anti-IL-2 antibody is used in a method of treating a subject having a disease or condition as described herein, wherein the polynucleotide encodes an antibody comprising a heavy chain variable region and a light chain variable region having an amino acid sequence of one of the following: SEQ ID NO: 10 and SEQ ID NO: 11; SEQ ID NO: 12 and SEQ ID NO: 13; SEQ ID NO: 14 and SEQ ID NO: 15; SEQ ID NO: 16 and SEQ ID NO: 17; SEQ ID NO: 18 and SEQ ID NO: 19; SEQ ID NO: 20 and SEQ ID NO: 21; SEQ ID NO: 22 and SEQ ID NO: 23; SEQ ID NO: 24 and SEQ ID NO: 25; SEQ ID NO: 26 and SEQ ID NO: 27; or SEQ ID NO: 36 and SEQ ID NO: 37.

[0412] In some embodiments of the methods of using polynucleotides to treat a disease or condition as described above, the polynucleotide encodes an engineered anti-IL-2 antibody that can be IgG, IgA, IgM, IgE, IgD, Fv, scFv, Fab, or F(ab')2. IgG can be of the subclass IgG1, IgG2, IgG3, or IgG4. In some embodiments, the polynucleotide encodes an engineered antibody that is part of a minibody, diabody, or triabody.

[0413] In some embodiments, a polynucleotide sequence encoding an engineered anti-IL-2 antibody is used in a method of treating a subject having a disease or condition as described herein, wherein the polynucleotide sequence comprises the sequence of one of SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 31, SEQ ID NO: 32, SEQ ID NO: 33, SEQ ID NO: 34, or SEQ ID NO: 35.

[0414] In some embodiments of the methods of treating a disease or condition as described herein, the immune effector cells activated by the treatment are CD8+ cells or NK cells. In one embodiment, an anti-IL-2 antibody disclosed herein, or a complex of IL-2 and an anti-IL-2 antibody disclosed herein, induces MP CD8+ cells. + cells and NK cells showed significant effects on the proliferation of CD4 + Tregs have a much smaller effect. In certain embodiments, CD4 + Tregs had no effect.

[0415] In certain embodiments, the methods of use of the anti-IL-2 antibodies disclosed herein provide a pro-stimulatory effect. A skilled artisan will appreciate that the use of the anti-IL-2 antibodies described and exemplified herein, such as in Example 1, clearly demonstrates a pro-stimulatory effect, rather than an anti-stimulatory effect or a pro-regulatory effect.

[0416] In some embodiments, a pro-stimulatory immune effect is provided in a subject in need thereof using an engineered or modified anti-IL-2 antibody comprising a heavy chain variable region having a sequence of one of SEQ ID NO: 10, SEQ ID NO: 12, SEQ ID NO: 14, SEQ ID NO: 16, SEQ ID NO: 18, SEQ ID NO: 20, SEQ ID NO: 22, SEQ ID NO: 24, SEQ ID NO: 26, or SEQ ID NO: 36. In some embodiments, a pro-stimulatory immune effect is provided in a subject in need thereof using an engineered or modified anti-IL-2 antibody comprising a light chain variable region having a sequence of one of SEQ ID NO: 11, SEQ ID NO: 13, SEQ ID NO: 15, SEQ ID NO: 17, SEQ ID NO: 19, SEQ ID NO: 21, SEQ ID NO: 23, SEQ ID NO: 25, SEQ ID NO: 27, or SEQ ID NO: 37. In some embodiments, an engineered or modified anti-IL-2 antibody comprising a heavy chain variable region and a light chain variable region having a sequence of one of SEQ ID NO: 10 and SEQ ID NO: 11; SEQ ID NO: 12 and SEQ ID NO: 13; SEQ ID NO: 14 and SEQ ID NO: 15; SEQ ID NO: 16 and SEQ ID NO: 17; SEQ ID NO: 18 and SEQ ID NO: 19; SEQ ID NO: 20 and SEQ ID NO: 21; SEQ ID NO: 22 and SEQ ID NO: 23; SEQ ID NO: 24 and SEQ ID NO: 25; SEQ ID NO: 26 and SEQ ID NO: 27; or SEQ ID NO: 36 and SEQ ID NO: 37 is used to provide a pro-stimulatory immune effect in a subject in need thereof. In some embodiments, the use comprises an anti-IL-2 antibody. In some embodiments, the use comprises an anti-IL-2 antibody and IL-2. In some embodiments, the use comprises a complex of an anti-IL-2 antibody and IL-2.

[0417] In some embodiments, an engineered or modified anti-IL-2 antibody comprising a heavy chain variable region having a sequence of one of SEQ ID NO: 10, SEQ ID NO: 12, SEQ ID NO: 14, SEQ ID NO: 16, SEQ ID NO: 18, SEQ ID NO: 20, SEQ ID NO: 22, SEQ ID NO: 24, SEQ ID NO: 26, or SEQ ID NO: 36 provides a pro-stimulatory immune effect, rather than an anti-stimulatory or pro-regulatory effect, in a subject in need thereof. In some embodiments, an engineered or modified anti-IL-2 antibody comprising a light chain variable region having a sequence of one of SEQ ID NO: 11, SEQ ID NO: 13, SEQ ID NO: 15, SEQ ID NO: 17, SEQ ID NO: 19, SEQ ID NO: 21, SEQ ID NO: 23, SEQ ID NO: 25, SEQ ID NO: 27, or SEQ ID NO: 37 provides a pro-stimulatory immune effect, rather than an anti-stimulatory or pro-regulatory effect, in a subject in need thereof. In some embodiments, the use of an engineered or modified anti-IL-2 antibody comprising a heavy chain variable region and a light chain variable region having a sequence of one of SEQ ID NO: 10 and SEQ ID NO: 11; SEQ ID NO: 12 and SEQ ID NO: 13; SEQ ID NO: 14 and SEQ ID NO: 15; SEQ ID NO: 16 and SEQ ID NO: 17; SEQ ID NO: 18 and SEQ ID NO: 19; SEQ ID NO: 20 and SEQ ID NO: 21; SEQ ID NO: 22 and SEQ ID NO: 23; SEQ ID NO: 24 and SEQ ID NO: 25; SEQ ID NO: 26 and SEQ ID NO: 27; or SEQ ID NO: 36 and SEQ ID NO: 37 provides a pro-stimulatory immune effect, rather than an anti-stimulatory or pro-regulatory effect, in a subject in need thereof. In some embodiments, the use comprises an anti-IL-2 antibody. In some embodiments, the use comprises an anti-IL-2 antibody and IL-2. In some embodiments, the use comprises a complex of an anti-IL-2 antibody and IL-2.

[0418] In some embodiments, an anti-IL-2 antibody comprising a heavy chain variable region comprising a heavy chain CDR1, CDR2, and CDR3 as set forth in the amino acid sequences of SEQ ID NOs: 38-40, 44-46, 50-52, 56-58, or 62-64, is used to provide a pro-stimulatory immune effect in a subject in need thereof. In some embodiments, an anti-IL-2 antibody comprising a light chain variable region comprising a light chain CDR1, CDR2, and CDR3 as set forth in the amino acid sequences of SEQ ID NOs: 41-43, 47-49, 53-55, 59-61, or 65-67, is used to provide a pro-stimulatory immune effect in a subject in need thereof. In some embodiments, an anti-IL-2 antibody comprising a heavy chain variable region comprising a heavy chain CDR1, CDR2, and CDR3 as set forth in the amino acid sequences of SEQ ID NOs: 38-40, 44-46, 50-52, 56-58, or 62-64, and a light chain variable region comprising a light chain CDR1, CDR2, and CDR3 as set forth in the amino acid sequences of SEQ ID NOs: 41-43, 47-49, 53-55, 59-61, or 65-67, provides a pro-stimulatory immune effect in a subject in need thereof. In some embodiments, the use comprises an anti-IL-2 antibody. In some embodiments, the use comprises an anti-IL-2 antibody and IL-2. In some embodiments, the use comprises a complex of an anti-IL-2 antibody and IL-2.

[0419] In some embodiments, an anti-IL-2 antibody comprising a heavy chain variable region comprising a heavy chain CDR1, CDR2, and CDR3 as set forth in the amino acid sequences of SEQ ID NOs: 38-40, 44-46, 50-52, 56-58, or 62-64, provides a pro-stimulatory immune effect, rather than an anti-stimulatory or pro-regulatory effect, in a subject in need thereof. In some embodiments, an anti-IL-2 antibody comprising a light chain variable region comprising a light chain CDR1, CDR2, and CDR3 as set forth in the amino acid sequences of SEQ ID NOs: 41-43, 47-49, 53-55, 59-61, or 65-67, provides a pro-stimulatory immune effect, rather than an anti-stimulatory or pro-regulatory effect, in a subject in need thereof. In some embodiments, the use of an anti-IL-2 antibody comprising a heavy chain variable region comprising a heavy chain CDR1, CDR2, and CDR3 as set forth in the amino acid sequences of SEQ ID NOs: 38-40, 44-46, 50-52, 56-58, or 62-64, and a light chain variable region comprising a light chain CDR1, CDR2, and CDR3 as set forth in the amino acid sequences of SEQ ID NOs: 41-43, 47-49, 53-55, 59-61, or 65-67, provides a pro-stimulatory immune effect, rather than an anti-stimulatory or pro-regulatory effect, in a subject in need thereof. In some embodiments, the use comprises an anti-IL-2 antibody. In some embodiments, the use comprises an anti-IL-2 antibody and IL-2. In some embodiments, the use comprises a complex of an anti-IL-2 antibody and IL-2.

[0420] Therefore, the engineered anti-IL-2 antibodies disclosed herein will be useful in methods of modulating immune cell populations and inducing differential expansion of certain immune effector cells in treating diseases (such as viral infections, bacterial infections, or cancer) or treating conditions (such as IL-2-induced pulmonary edema or IL-2-induced vascular leakage).

[0421] In some embodiments, disclosed herein is a method of immunizing a subject, wherein the immunization comprises administering a vaccine comprising an adjuvant, the adjuvant comprising an IL-2 antibody adjuvant. In some embodiments, the IL-2 antibody adjuvant comprises an anti-IL-2 antibody and IL-2, or comprises an anti-IL-2 antibody complexed with IL-2. In some embodiments, the IL-2 antibody adjuvant comprises an anti-IL-2 antibody and IL-2. In some embodiments, the IL-2 antibody adjuvant comprises an anti-IL-2 antibody complexed with IL-2. In some embodiments, the IL-2 antibody adjuvant comprises an anti-IL-2 antibody.

[0422] In some embodiments, the subject being immunized is a mammalian subject. In some embodiments, the subject being immunized is a human. In some embodiments, the subject being immunized has a weakened immune system.

[0423] In some embodiments of the immunization method, the anti-IL-2 antibody comprises a heavy chain variable region having a sequence of one of SEQ ID NO: 10, SEQ ID NO: 12, SEQ ID NO: 14, SEQ ID NO: 16, SEQ ID NO: 18, SEQ ID NO: 20, SEQ ID NO: 22, SEQ ID NO: 24, SEQ ID NO: 26, or SEQ ID NO: 36. In some embodiments of the immunization method, the anti-IL-2 antibody comprises a light chain variable region having a sequence of one of SEQ ID NO: 11, SEQ ID NO: 13, SEQ ID NO: 15, SEQ ID NO: 17, SEQ ID NO: 19, SEQ ID NO: 21, SEQ ID NO: 23, SEQ ID NO: 25, SEQ ID NO: 27, or SEQ ID NO: 37. In some embodiments of the immunization method, the anti-IL-2 antibody includes an anti-IL-2 antibody comprising a heavy chain variable region and a light chain variable region, wherein the heavy chain variable region and the light chain variable region have a sequence of one of the following: SEQ ID NO: 10 and SEQ ID NO: 11; SEQ ID NO: 12 and SEQ ID NO: 13; SEQ ID NO: 14 and SEQ ID NO: 15; SEQ ID NO: 16 and SEQ ID NO: 17; SEQ ID NO: 18 and SEQ ID NO: 19; SEQ ID NO: 20 and SEQ ID NO: 21; SEQ ID NO: 22 and SEQ ID NO: 23; SEQ ID NO: 24 and SEQ ID NO: 25; SEQ ID NO: 26 and SEQ ID NO: 27; or SEQ ID NO: 36 and SEQ ID NO: 37.

[0424] In some embodiments of the immunization method, the anti-IL-2 antibody comprises an anti-IL-2 antibody comprising a heavy chain variable region comprising a complementarity determining region (CDR) 1, a CDR2, and a CDR3, wherein the CDR1, CDR2, and CDR3 comprise the amino acid sequences of SEQ ID NOs: 38-40, 44-46, 50-52, 56-58, or 62-64, respectively. In some embodiments of the immunization method, the anti-IL-2 antibody comprises an anti-IL-2 antibody comprising a light chain variable region comprising a complementarity determining region (CDR) 1, a CDR2, and a CDR3, wherein the CDR1, CDR2, and CDR3 comprise the amino acid sequences of SEQ ID NOs: 41-43, 47-49, 53-55, 59-61, or 65-67, respectively. In some embodiments of the immunization method, the anti-IL-2 antibody includes an anti-IL-2 antibody comprising a heavy chain variable region and a light chain variable region, each of which comprises a complementarity determining region (CDR) 1, CDR2, and CDR3, wherein the heavy chain CDR1, CDR2, and CDR3 comprise the amino acid sequence of SEQ ID NOs: 38-40, respectively; SEQ ID NOs: 44-46, respectively; SEQ ID NOs: 50-52, respectively; SEQ ID NOs: 56-58, respectively; or SEQ ID NOs: 62-64, respectively, wherein the light chain CDR1, CDR2, and CDR3 comprise the amino acid sequence of SEQ ID NOs: 41-43, respectively; SEQ ID NOs: 47-49, respectively; SEQ ID NOs: 53-55, respectively; SEQ ID NOs: 59-61, respectively; or SEQ ID NOs: 65-67, respectively.

[0425] In some embodiments of the method of immunizing a subject, the immunization comprises administering a vaccine comprising an adjuvant, the adjuvant comprising an IL-2 antibody adjuvant, the anti-IL-2 antibody comprising an anti-IL-2 antibody as disclosed herein. In certain embodiments, the IL-2 antibody adjuvant comprises an anti-IL-2 antibody and IL-2, or comprises an anti-IL-2 antibody complexed with IL-2. In some embodiments of the method of immunizing a subject, the subject has a weakened immune system.

[0426] In some embodiments, the subject immunized with a vaccine comprising an IL-2 antibody adjuvant includes a subject having a condition that includes a genetic predisposition that increases the likelihood of cancer in the subject. In some embodiments, the genetic predisposition includes an alteration in the expression or activity of a gene product. In some embodiments, the genetic predisposition that increases the likelihood of cancer includes a mutation in a tumor suppressor gene or a mismatch repair (MMR) gene, or a combination thereof. Many hereditary cancers are known in the art, non-limiting examples of which include, but are not limited to, hereditary breast and ovarian cancer (HBOC) syndrome, Lynch syndrome (hereditary non-polyposis colorectal cancer), and Leigh-Fraumeni syndrome.

[0427] In some embodiments, the subject treated by the method for treating a disease or condition disclosed herein is also treated with one or more immune checkpoint inhibitors targeting one or more immune checkpoints. In some embodiments, the subject is treated with the immune checkpoint inhibitor while being treated with the anti-IL-2 antibody, before being treated with the anti-IL-2 antibody, or after being treated with the anti-IL-2 antibody. In some embodiments of the methods of treatment disclosed herein, immune checkpoint inhibitors include PD-1, PD-L1, CTLA-4, TIGIT, TIM-3, B7-H3, CD73, LAG3, CD27, CD70, 4-1BB, GITR, OX40, SIRP-α (CD47), CD39, ILDR2, VISTA, BTLA or VTCN-1 or a combination thereof.

[0428] As discussed above, in some embodiments, the therapeutic treatment methods as disclosed herein further include an additional active agent comprising a checkpoint inhibitor. It will be understood by those skilled in the art that a combination therapy comprising an anti-IL-2 antibody therapy in the presence or absence of IL-2 and additionally comprising a checkpoint inhibitor can utilize any of the therapeutic compositions or formulations comprising an anti-IL-2 antibody + / - IL-2 and a checkpoint inhibitor as provided herein. In some embodiments, at least two checkpoint inhibitors are used in the combination therapy.

[0429] The embodiments of the present application include:

[0430] An isolated anti-IL-2 antibody, wherein the antibody comprises a heavy chain variable region comprising the sequence of one of SEQ ID NO: 10, SEQ ID NO: 12, SEQ ID NO: 14, SEQ ID NO: 16, SEQ ID NO: 18, SEQ ID NO: 20, SEQ ID NO: 22, SEQ ID NO: 24, SEQ ID NO: 26, or SEQ ID NO: 36.

[0431] Antibodies include those comprising IgG, IgA, IgM, IgE, IgD, Fv, scFv, Fab, F(ab')2, minibodies, diabodies, or triabodies.

[0432] An IgG comprising

[0433] (a) IgG1, IgG2, IgG3 or IgG4;

[0434] (b) a heavy chain comprising a mutation that reduces binding to an Fcγ receptor (FcγR); or

[0435] (c) a lambda or kappa light chain; or

[0436] (d) Any combination of (a)-(c).

[0437] A composition comprising an isolated anti-IL-2 antibody and a pharmaceutically acceptable carrier.

[0438] An isolated anti-IL-2 antibody, wherein the antibody comprises a light chain variable region comprising the sequence of one of SEQ ID NO: 11, SEQ ID NO: 13, SEQ ID NO: 15, SEQ ID NO: 17, SEQ ID NO: 19, SEQ ID NO: 21, SEQ ID NO: 23, SEQ ID NO: 25, SEQ ID NO: 27, or SEQ ID NO: 37.

[0439] SEQ ID NO: 19; SEQ ID NO: 20 and SEQ ID NO: 21; SEQ ID NO: 22 and SEQ ID NO: 23; SEQ ID NO: 24 and SEQ ID NO: 25; SEQ ID NO: 26 and SEQ ID NO: 27; or SEQ ID NO: 36 and SEQ ID NO: 37.

[0440] An isolated anti-IL-2 antibody, wherein the antibody comprises a heavy chain variable region comprising a complementarity determining region 1 (CDR1), a CDR2, and a CDR3, wherein the CDR1, CDR2, and CDR3 comprise the amino acid sequences of SEQ ID NOs: 38-40, respectively; SEQ ID NOs: 44-46, respectively; SEQ ID NOs: 50-52, respectively; SEQ ID NOs: 56-58, respectively; or SEQ ID NOs: 62-64, respectively.

[0441] An isolated anti-IL-2 antibody, wherein the antibody comprises a light chain variable region comprising a complementarity determining region 1 (CDR1), a CDR2, and a CDR3, wherein the CDR1, CDR2, and CDR3 comprise the amino acid sequences of SEQ ID NOs: 41-43, respectively; SEQ ID NOs: 47-49, respectively; SEQ ID NOs: 53-55, respectively; SEQ ID NOs: 59-61, respectively; or SEQ ID NOs: 65-67, respectively.

[0442] An isolated anti-IL-2 antibody, wherein the antibody comprises: a heavy chain variable region comprising a complementarity determining region 1 (CDR1), a CDR2, and a CDR3, wherein the CDR1, CDR2, and CDR3 comprise the amino acid sequence of SEQ ID NOs: 38-40, respectively; SEQ ID NOs: 44-46, respectively; SEQ ID NOs: 50-52, respectively; SEQ ID NOs: 56-58, respectively; or SEQ ID NOs: 62-64, respectively; and a light chain variable region having a complementarity determining region 1 (CDR1), a CDR2, and a CDR3, wherein the CDR1, CDR2, and CDR3 comprise the amino acid sequence of SEQ ID NOs: 41-43, respectively; SEQ ID NOs: 47-49, respectively; SEQ ID NOs: 53-55, respectively; SEQ ID NOs: 59-61, respectively; or SEQ ID NOs: 65-67, respectively.

[0443] An isolated polynucleotide sequence encoding a heavy chain variable region of an anti-IL-2 antibody, wherein the heavy chain variable region comprises the amino acid sequence of one of SEQ ID NO: 10, SEQ ID NO: 12, SEQ ID NO: 14, SEQ ID NO: 16, SEQ ID NO: 18, SEQ ID NO: 20, SEQ ID NO: 22, SEQ ID NO: 24, SEQ ID NO: 26, or SEQ ID NO: 36.

[0444] A vector comprising a polynucleotide sequence described herein. A host cell comprising a vector described herein.

[0445] An isolated polynucleotide sequence encoding a light chain variable region of an anti-IL-2 antibody, wherein the light chain variable region comprises the amino acid sequence of one of SEQ ID NO: 11, SEQ ID NO: 13, SEQ ID NO: 15, SEQ ID NO: 17, SEQ ID NO: 19, SEQ ID NO: 21, SEQ ID NO: 23, SEQ ID NO: 25, SEQ ID NO: 27, or SEQ ID NO: 37.

[0446] An isolated polynucleotide sequence encoding a heavy chain variable region of an anti-IL-2 antibody and encoding a light chain variable region of an anti-IL-2 antibody, wherein the heavy chain variable region comprises the amino acid sequence of one of SEQ ID NO: 10, SEQ ID NO: 12, SEQ ID NO: 14, SEQ ID NO: 16, SEQ ID NO: 18, SEQ ID NO: 20, SEQ ID NO: 22, SEQ ID NO: 24, SEQ ID NO: 26, or SEQ ID NO: 36, and wherein the light chain variable region comprises the amino acid sequence of one of SEQ ID NO: 11, SEQ ID NO: 13, SEQ ID NO: 15, SEQ ID NO: 17, SEQ ID NO: 19, SEQ ID NO: 21, SEQ ID NO: 23, SEQ ID NO: 25, SEQ ID NO: 27, or SEQ ID NO: 37.

[0447] An isolated polynucleotide sequence encoding an scFv, the polynucleotide sequence comprising a sequence of one of SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 31, SEQ ID NO: 32, SEQ ID NO: 33, SEQ ID NO: 34 or SEQ ID NO: 35.

[0448] A method for producing a heavy chain variable region of an anti-IL-2 antibody, comprising culturing a host cell comprising a vector disclosed herein under conditions conducive to expression of the vector in the host cell, thereby producing the heavy chain variable region of the anti-IL-2 antibody.

[0449] A method for producing a light chain variable region of an anti-IL-2 antibody, comprising culturing a host cell under conditions conducive to the expression of the vector in the host cell, thereby producing the light chain variable region of the anti-IL-2 antibody.

[0450] A method for producing an anti-IL-2 antibody comprising a heavy chain variable region and a light chain variable region of the anti-IL-2 antibody, the method comprising culturing a host cell under conditions conducive to expression of the vector in the host cell, thereby producing the heavy chain variable region and the light chain variable region of the anti-IL-2 antibody.

[0451] A method for promoting differential growth of immune cells in a subject, the method comprising administering a composition comprising an anti-IL-2 antibody, thereby promoting differential growth of immune cells in the subject. In some embodiments, the composition comprises the anti-IL-2 antibody and IL-2, or the anti-IL-2 antibody complexed with IL-2.

[0452] A method of treating a subject having cancer by inducing differential growth of immune cells, the method comprising the step of administering a composition comprising an anti-IL-2 antibody, thereby treating the subject having cancer.

[0453] A method of treating a disease or condition in a subject, comprising the step of administering to the subject a composition comprising an anti-IL-2 antibody, wherein the antibody promotes expansion of immune cell subsets and reduces undesirable effects caused by IL-2, thereby treating the disease or condition in the subject.

[0454] In some embodiments, the disease comprises a viral infection, a bacterial infection, or a cancer. In some embodiments, the viral infection is caused by SARS CoV-2; norovirus; rotavirus; hepatitis A virus, hepatitis B virus, hepatitis C virus, hepatitis D virus, or hepatitis E virus; rabies virus; West Nile virus; enterovirus; echovirus; coxsackievirus; herpes simplex virus (HSV); HSV-2; varicella-zoster virus; mosquito-borne virus; arbovirus; St. Louis encephalitis virus; California encephalitis virus; lymphocytic choriomeningitis virus; human immunodeficiency virus (HIV); poliovirus; Zika virus; rubella virus; cytomegalovirus; human papillomavirus (HPV); enterovirus D68; severe acute respiratory syndrome (SARS) coronavirus; Middle East respiratory syndrome coronavirus; Epstein-Barr virus; influenza virus; respiratory syncytial virus; polyomavirus (including JC virus; BK virus); Ebola virus; dengue virus; or any combination thereof. In some embodiments, the condition comprises a weakened immune system, and the treatment prophylactically boosts the immune system.

[0455] In some embodiments, the condition comprises IL-2 induced pulmonary edema.

[0456] In some embodiments of the methods disclosed herein, the immune cells include one or more of naive T cells, memory T cells, CD8+ T cells, NK cells, or natural killer T cells.

[0457] In some embodiments, the undesirable effects caused by IL-2 include one or more of activation of regulatory T cells, apoptosis of CD25+ T effector cells, IL-2-induced pulmonary edema, pneumonitis, or IL-2-induced vascular leakage.

[0458] In some embodiments, the anti-IL-2 antibodies disclosed herein inhibit the binding of IL-2 to CD25.

[0459] A method of immunizing a subject, wherein the immunization comprises administering a vaccine comprising an adjuvant, the adjuvant comprising an IL-2 antibody adjuvant.

[0460] In some embodiments, the IL-2 antibody adjuvant comprises an anti-IL-2 antibody and IL-2, or comprises an anti-IL-2 antibody complexed with IL-2.

[0461] In some embodiments, the subject is an animal or a human.In some embodiments, the subject has a weakened immune system.

[0462] In some embodiments of the methods disclosed herein, the immune cell is a CD8+ cell or a NK cell.

[0463] In some embodiments of the methods disclosed herein, the anti-IL-2 antibody comprises a heavy chain variable region having a sequence of one of SEQ ID NO: 10, SEQ ID NO: 12, SEQ ID NO: 14, SEQ ID NO: 16, SEQ ID NO: 18, SEQ ID NO: 20, SEQ ID NO: 22, SEQ ID NO: 24, SEQ ID NO: 26, or SEQ ID NO: 36.

[0464] In some embodiments of the methods disclosed herein, the anti-IL-2 antibody comprises a light chain variable region having a sequence of one of SEQ ID NO: 11, SEQ ID NO: 13, SEQ ID NO: 15, SEQ ID NO: 17, SEQ ID NO: 19, SEQ ID NO: 21, SEQ ID NO: 23, SEQ ID NO: 25, SEQ ID NO: 27, or SEQ ID NO: 37.

[0465] In some embodiments of the methods disclosed herein, the anti-IL-2 antibody comprises a heavy chain variable region and a light chain variable region having a sequence of one of: SEQ ID NO: 10 and SEQ ID NO: 11; SEQ ID NO: 12 and SEQ ID NO: 13; SEQ ID NO: 14 and SEQ ID NO: 15; SEQ ID NO: 16 and SEQ ID NO: 17; SEQ ID NO: 18 and SEQ ID NO: 19; SEQ ID NO: 20 and SEQ ID NO: 21; SEQ ID NO: 22 and SEQ ID NO: 23; SEQ ID NO: 24 and SEQ ID NO: 25; SEQ ID NO: 26 and SEQ ID NO: 27; or SEQ ID NO: 36 and SEQ ID NO: 37.

[0466] In some embodiments of the methods disclosed herein, the anti-IL-2 antibody comprises a heavy chain variable region comprising a complementarity determining region (CDR) 1, CDR2, and CDR3 comprising the amino acid sequence of SEQ ID NOs: 38-40, respectively; SEQ ID NOs: 44-46, respectively; SEQ ID NOs: 50-52, respectively; SEQ ID NOs: 56-58, respectively; or SEQ ID NOs: 62-64, respectively.

[0467] In some embodiments of the methods disclosed herein, the anti-IL-2 antibody comprises a light chain variable region comprising a complementarity determining region (CDR) 1, CDR2, and CDR3 comprising the amino acid sequence of SEQ ID NOs: 41-43, respectively; SEQ ID NOs: 47-49, respectively; SEQ ID NOs: 53-55, respectively; SEQ ID NOs: 59-61, respectively; or SEQ ID NOs: 65-67, respectively.

[0468] In some embodiments of the methods disclosed herein, the anti-IL-2 antibody comprises a heavy chain variable region and a light chain variable region, each of which comprises a complementarity determining region (CDR) 1, CDR2, and CDR3,

[0469] wherein the heavy chain CDR1, CDR2, and CDR3 comprise the amino acid sequences of SEQ ID NOs: 38-40, 44-46, 50-52, 56-58, or 62-64, respectively;

[0470] wherein the light chain CDR1, CDR2, and CDR3 comprise the amino acid sequences of SEQ ID NOs: 41-43, 47-49, 53-55, 59-61, or 65-67, respectively.

[0471] As used herein, the singular forms "a," "an," and "the" include plural referents unless the context clearly dictates otherwise. For example, the term "a compound" or "at least one compound" may include more than one compound, including mixtures thereof.

[0472] Throughout this application, various embodiments of the present invention can be presented in range format. It should be understood that the description in range format is only for convenience and brevity and should not be interpreted as an unchangeable limitation to the scope of the present invention. Therefore, the description of the scope should be considered to have specifically disclosed all possible sub-ranges and the independent numerical values within the scope. For example, the description of the range, such as from 1 to 6, should be considered to have specifically disclosed sub-ranges such as from 1 to 3, from 1 to 4, from 1 to 5, from 2 to 4, from 2 to 6, from 3 to 6, etc., and the independent numerals such as 1, 2, 3, 4, 5 and 6 within the scope. Regardless of the width of the scope, this applies.

[0473] Whenever a numerical range is indicated herein, it is meant to include any stated numeral (fractional or integer) within the indicated range. The phrases "ranging / ranges between" a first indicated numeral and a second indicated numeral and "ranging / ranges from" a first indicated numeral "to" a second indicated numeral are used interchangeably herein and are meant to include the first indicated numeral and the second indicated numeral and all fractional and integer values therebetween.

[0474] A skilled artisan will understand that the term "about" can encompass deviations of between 0.0001% and 5% from the indicated number or numerical range. In some cases, the term "about" can encompass deviations of between 1% and 10% from the indicated number or numerical range. In some cases, the term "about" can encompass deviations of up to 25% from the indicated number or numerical range. Example

[0475] Example 1

[0476] This example describes the generation of modified anti-IL-2 antibodies based on embodiments of the generated antibodies. The exemplary generation of modified anti-IL-2 antibodies is based on the antibody subpopulations disclosed herein. The description and results presented in Example 1 are exemplary and do not limit the generation of modified anti-IL-2 antibodies disclosed throughout this application.

[0477] Library design

[0478] The library is designed to introduce mutations into the sequence of JES6.1. The amino acid sequences of the heavy chain variable region and light chain variable region of JES6.1 are shown in SEQ ID NO:6 and SEQ ID NO:7, respectively. In short, three positions are changed so that codons encode all amino acids (codons NNS). The design of this library allows one mutation in both CDR L3 and CDR H3, as well as one mutation in one of the following CDRs: H1, H2, or L2. CDRs are defined by meeting IMGT or ABR (Kunik et al., 2012) definitions. CDR residues that are conserved (based on Blast searches for the PDB database) or do not form specific interactions with mouse IL-2 (mIL-2) in the crystal structure of the mIL2-JES6.1 complex (PDB 4YQX) are excluded from the mutation. The theoretical size of this library is 1.38E+7 variants.

[0479] Library selection

[0480] Screening and selection using yeast surface display

[0481] The yeast-displayed scFv library was grown in SDCAA selective medium and induced to express overnight with 2% w / v galactose at 30°C according to a pre-defined protocol. The library was incubated with 100 nM of recombinant human IL-2 (hIL-2-His) (Reprokine, Israel) with a 6xhis tag in PBS 0.1% BSA for 1 hour, then washed three times with PBS 0.1% BSA, and labeled with fluorescently labeled antibodies mouse anti-Myc-FITC (Santa Cruze, USA) and mouse monoclonal anti-His APC (Miltenyi Biotec, Germany. Catalog number 0020130-119-782). After labeling, the library was sorted for high-affinity binding to recombinant human IL-2 on a BioRad S3e fluorescence-activated cell sorter. Isolated clones from the final sorting were sequenced by extracting plasmid DNA from yeast clones using the Zymoprep kit (Zymo Research, USA), and the DNA was sequenced.

[0482] Koff selection

[0483] To select for binders with improved off-rates, clones selected in round 2 were incubated with 10 nM 6xhis tag (hIL-2) for 15 min, then the yeast were washed 3 times with 1 ml PBS 0.1% BSA and incubated with 100 nM unlabeled IL-2 for 5 minutes, 4 hours, 6 hours, and 24 hours. At the indicated time points, yeast were washed and labeled with Myc-FITC (Santa Cruze, USA) and monoclonal anti-His APC (Miltenyi Biotec, Germany. Cat. No. 0020130-119-782) and sorted on Se3 as described above.

[0484] IgG production

[0485] JES6.1 w.t. was purchased from Thermo Fisher (Cat. No. 16-7022-81). JES6.1.RMC was cloned as a rat Fv with a mouse IgG2a constant region and produced by GeneScript Antibody Production Services (Genscript NJ, USA). BDG17.014 was cloned into a human IgG1 constant region and produced by GeneScript Antibody Production Services. The amino acid sequences of the heavy and light chain variable regions of JES6.1.RMC are shown in SEQ ID NO: 8 and SEQ ID NO: 9, respectively. All other antibodies were produced as described below.

[0486] Reformatting

[0487] The scFv clone selected is reformatted into human IgG1 form. The sequence of light chain (LC) variable region and heavy chain (HC) variable region is optimized for mammalian codon use and is ordered from IDT (Integrated DNA Technologies.Coralville, Iowa USA) as genblocks (GB). GB is cloned into pSF-CMV-HuIgG1_HC (HC plasmid) and pSF-CMV-HuLambda_LC (LC plasmid) (Oxford genetics, Oxford UK) using standard cloning techniques. When indicated, variable heavy chain is cloned into pSF-CMV-HuIgG1_HC_LALA (HC plasmid), wherein the DNA of L234 and L235 of encoding heavy chain is mutated into alanine codon (L234A, L235A).

[0488] IgG expression

[0489] Expi-CHO cells (Thermo Fisher Scientific, USA) were transfected with LC plasmid and HC plasmid at a ratio of 2:1 and expressed according to the manufacturer's instructions. Briefly, 50 ml of Expi-CHO cells were cultured at 37°C, 120 rpm, 8% CO2 to a concentration of 6 × 10 6 The cells were purified by ELISA.Then, 50 μg of heavy chain expression plasmid and light chain expression plasmid at a ratio of 1: 2 were transfected into CHO cells.After transfection, booster enhancer and feed were added to the culture, and growth conditions were changed to 32 ° C, 120 rpm, 5% CO . Cells were harvested 10 days after transfection. IgG was purified from the supernatant using protein A beads (Tosoh Bioscience GmbH, Germany), and subsequently size exclusion chromatography (SEC) purification was performed on a superdex 200 10 / 300 increase column using PBS as a mobile phase (GE healthcare, USA).

[0490] sequence

[0491] The DNA sequence encoding the scFv of clone 1 (17.021) is shown in SEQ ID NO: 1. The DNA sequence encoding the scFv of clone 2 (17.022) is shown in SEQ ID NO: 2. The DNA sequence encoding the scFv of clone 4 (17.023) is shown in SEQ ID NO: 3. The DNA sequence encoding the scFv of clone 5 (17.030) is shown in SEQ ID NO: 4. The DNA sequence encoding the scFv of clone 6 (17.035) is shown in SEQ ID NO: 5.

[0492] The amino acid sequences of the original JES6_1 starting sequence and the heavy and light chain variable regions of various anti-IL-2 clones are given in the table below and in Figure 11 and Figure 12 Shown in.

[0493] Table 2: VH and VL amino acid sequences of anti-IL-2 clone subsets.

[0494] BDG clone Heavy chain variable region Light chain variable region 17.021 SEQ ID NO: 10 SEQ ID NO:11 17.022 SEQ ID NO:12 SEQ ID NO:13 17.023 SEQ ID NO:14 SEQ ID NO:15 17.030 SEQ ID NO:16 SEQ ID NO: 17 17.035 SEQ ID NO: 18 SEQ ID NO: 19

[0495] Measurement of IgG binding to human IL-2

[0496] SPR analysis was performed on a Biacore 200 (GE healthcare, USA) on a CM5 chip (Cat. No. br10005-30, GE healthcare, USA). The chip was cross-linked to 8000 RU of target with a primary capture Ab for human IgG (Cat. No. br-1008-39, GE healthcare, USA) or a primary capture antibody for mouse IgG (Cat. No. BR-1008-38, GE healthcare, USA). After cross-linking of the primary Ab, mouse and human test antibodies were immobilized on the primary Ab to approximately another 500 RU of target. JES6.1 was directly cross-linked to the CM5 chip. Human IL-2 (Cat. No. 60568, Reprokine, Israel) analyte was flowed in a series of two-fold or three-fold dilutions in HEB-EP or PBS 0.05% tween-20 (PBS-T) buffer at concentrations ranging from 128 nM to 0.03 nM, with one concentration per cycle. Mouse IL-2 (Cat. No.: RKP04351, Reprokine, Israel) was flowed in HEB-EP or PBS-T buffer at concentrations ranging from 0.5 nM to 40 nM. At the end of each cycle, 3 M MgCl was used to strip the analyte and test antibody from the chip, and a new test Ab was loaded onto the chip as described above. When indicated, instead of stripping the antibody, kinetics was determined by a single-cycle kinetic method by injecting a range of analyte concentrations in one cycle. Binding kinetics was determined using Biacore T200 evaluation software using a 1:1 binding model.

[0497] Binding of IgG to cynomolgus monkey IL-2

[0498] SPR analysis was performed on a Biacore 200 (GE healthcare, USA) on a CM5 chip (Cat. No. br10005-30, GE healthcare, USA). The chip was cross-linked to 5000 RU of target with a primary capture Ab against human IgG (Cat. No. br-1008-39, GE healthcare, USA), and cynomolgus monkey IL-2 (cIL-2) was tested by a multi-cycle method under the same conditions as described above.

[0499] SEC analysis

[0500] For IgG analysis, 100 μg of sample was loaded onto a Superdex 200 10 / 300 increase column (GE healthcare, USA) on a GE AKTA Explorer chromatography system (GE healthcare, USA) at a flow rate of 0.8 ml / min. Antibody retention time was monitored at 280 nm.

[0501] Testing for specific binding to CD25 and CD122

[0502] To test specific binding to CD25, BDG17.023 was immobilized on a CM5 chip to a target RU of approximately 300 RU as described above. Subsequently, 50 nM IL-2 was injected until BDG17.023 or control antibody was saturated. The Ab-IL-2 complex was then washed with PBS-T buffer for 10 seconds, and 1000 nM CD25 was injected and the response was monitored.

[0503] To test specific binding to CD122, as described above, BDG17.023 was fixed to a CM5 chip to a target RU of approximately 300-500 RU. Subsequently, 50 nM hIL-2 was injected until the BDG17.023 antibody was saturated with hIL-2. Next, the Ab-IL-2 complex was washed with PBS-T buffer for 10 seconds, and 1000 nM CD122 was injected and the response was monitored.

[0504] In order to test the specific binding of humanized antibody IL-2 compound and CD122 and CD25, as described above, antibody BDG17.038, BDG17.043, BDG17.053, BDG17.054, BDG17.067, BDG17.069 (sequence information of these clones is referring to table 6 and table 7 of embodiment 2) is fixed to the capture antibody that is attached to CM5 chip channel to target RU is about 300RU. Subsequently, 50nM IL-2 is injected, until corresponding antibody is saturated by hIL-2.Then Ab-IL-2 compound is washed with PBS-T buffer solution for 60 seconds, and the CD25 of 1000nM is injected and monitors response. Subsequently, running buffer is injected for 60 seconds to reach stable baseline, and then the CD122 of 1000nM is injected for 30 seconds with the flow rate of 30 μ l / min. To test CD122 binding, the same experiment was repeated in the reverse order, where CD122 was injected first followed by CD25.

[0505] DSF analysis of IgG Tm

[0506] To determine the T-start and Tm of humanized anti-hIL-2 antibodies, antibodies were diluted to 0.5 mg / ml in PBS and analyzed using a NanoDSF Prometheus NT.48 (Nanotemper, Germany) at a heating rate of 1°C / min.

[0507] In vivo experiments

[0508] Treatment of mice with IL-2 / Ab complex

[0509] Groups of six 7-8 week old male C57BL / 6 mice were injected intraperitoneally (ip) with BDG17.023 / hIL-2 or JES6.1 / mIL-2 immune complexes daily for four consecutive days. PBS and free hIL-2 or mIL-2 were used as controls. At the end of the fourth day, mice were sacrificed, spleens harvested, and homogenized to a single cell suspension. Cells were filtered, centrifuged (400 g for 5 minutes), and resuspended in 5 ml of PBS to a final concentration of 5 × 10 6 The experiment was performed in accordance with the guidelines of the Institutional Animal Care and Use Committee (IACUC) of Israel.

[0510] Groups of six 7-8 week old male C57BL / 6 mice were injected intraperitoneally (IP) daily for four consecutive days with BDG17.038 / hIL-2, BDG17.043 / hIL-2, BDG17.054 / hIL-2, BDG17.038 / hIL-2 or isotype control / hIL-2 immune complexes. To form the complex, 10 μg of antibody was pre-incubated with 0.5 μg of hIL-2 at 37°C for 30 minutes prior to injection. At the end of the fourth day, mice were sacrificed, spleens were harvested and homogenized to a single cell suspension. Cells were filtered, centrifuged (400 g for 5 minutes), and resuspended in 5 ml of PBS to a final concentration of 5×10 6 The experiment was performed in accordance with the guidelines of the Institutional Animal Care and Use Committee (IACUC) of Israel.

[0511] B16F10 murine melanoma tumor xenograft model

[0512] Female C57BL / 6 mice were inoculated subcutaneously in the right hind flank region with (2×10 5 Five days after inoculation, when the tumor volume reached 30 mm 3 -50 mm 3At 4 hr, mice were randomly divided into experimental groups (n=10 per group) and injected intraperitoneally with a single dose of 10 μg anti-IL-2 antibody / 1 μg hIL-2 complex of the indicated antibodies or PBS control daily for four consecutive days. The mice were monitored for tumor volume growth, weight loss, and nonspecific clinical signs throughout the experiment.

[0513] Identification of immune cell populations by FACS

[0514] To identify immune cell populations, splenic lymphocytes were labeled with the antibodies described below according to the manufacturer's instructions. Regulatory T cells (Tregs) were designated as those labeled with CD45. + / CD3 + / CD4 + / CD25 + / FoxP3 + Memory phenotype effector T cells (MP CD8 + ):CD45 + / CD3 + / CD8 + / CD44 + / IL-2RB (CD122) + Natural Killer (NK) cells: CD45 + / CD3 - / CD49b + / NK1.1 (CD161). Natural killer T cells (NKT): CD45 + / CD3 + / CD49b + / NK1.1(CD161). The frequency and number of positive cells were calculated from the raw data obtained by flow cytometry.

[0515] Table 3: Markers / labeled antibodies

[0516]

[0517]

[0518] result

[0519] JES6.1 strongly binds mouse IL-2 but not human IL-2

[0520] JES6.1 has been reported to have a K of 5.6 nM. DBinding to mouse IL-2 (mIL-2). To test whether JES6.1 can bind to human IL-2 (hIL-2), the JES6.1 antibody was tested by SPR on a Biacore T200. JES6.1 was directly cross-linked to a CM5 chip, and human IL-2 analyte or mouse IL-2 analyte was flowed at concentrations ranging from 0.5 nM to 128 nM or 0.5 nM to 16 nM, respectively. Figure 4A As can be seen in the Figure 3, the response units (RU) of JES6.1 did not show significant changes when tested with human IL-2. On the other hand, a robust response was evident when mouse IL-2 was used as the analyte ( Figure 4B ), indicating that JES6.1 strongly binds to mouse IL-2 but not to human IL-2. The experiment was repeated with the JES6.1RMC antibody chimera, which was expressed as a JES6.1 rat FV with mouse constant regions as described herein. JES6.1RMC was immobilized on a CM5 chip using the GE antibody capture kit. Flowing hIL-2 at concentrations up to 100 nM did not result in a change in RU, indicating no binding to human IL-2 ( Figure 4C To test whether the JES6.1RMC chimera retained its mIL-2 binding properties as JES6.1 above, its binding to mouse IL-2 was tested. Flowing mIL-2 at concentrations ranging from 0.5 nM to 320 nM resulted in large changes in RU, indicating robust binding ( Figure 4D These results indicate that JES6.1 and JES6.1RMC strongly bind to mo...

Claims

1. A method for treating solid cancer in a subject, comprising administering to the subject a composition comprising an anti-IL-2 antibody, wherein the IL-2 antibody comprises a heavy chain variable region (VH) and a light chain variable region (VL), wherein the VH comprises heavy chain complementarity determining regions (HCDRs) HCDR1, HCDR2, and HCDR3, and the VL comprises light chain complementarity determining regions (LCDRs) LCDR1, LCDR2, and LCDR3, wherein the CDRs have the following amino acid sequences: (a) the HCDR1 comprises the amino acid sequence of SEQ ID NO: 62, the HCDR2 comprises the amino acid sequence of SEQ ID NO: 63, the HCDR3 comprises the amino acid sequence of SEQ ID NO: 64, the LCDR1 comprises the amino acid sequence of SEQ ID NO: 65, the LCDR2 comprises the amino acid sequence of DAS, and the LCDR3 comprises the amino acid sequence of SEQ ID NO: 67; (b) the HCDR1 comprises the amino acid sequence of SEQ ID NO: 38, the HCDR2 comprises the amino acid sequence of SEQ ID NO: 39, the HCDR3 comprises the amino acid sequence of SEQ ID NO: 40, the LCDR1 comprises the amino acid sequence of SEQ ID NO: 41, the LCDR2 comprises the amino acid sequence of YAS, and the LCDR3 comprises the amino acid sequence of SEQ ID NO: 43; (c) the HCDR1 comprises the amino acid sequence of SEQ ID NO: 44, the HCDR2 comprises the amino acid sequence of SEQ ID NO: 45, the HCDR3 comprises the amino acid sequence of SEQ ID NO: 46, the LCDR1 comprises the amino acid sequence of SEQ ID NO: 47, the LCDR2 comprises the amino acid sequence of YAS, and the LCDR3 comprises the amino acid sequence of SEQ ID NO: 49; (d) the HCDR1 comprises the amino acid sequence of SEQ ID NO: 50, the HCDR2 comprises the amino acid sequence of SEQ ID NO: 51, the HCDR3 comprises the amino acid sequence of SEQ ID NO: 52, the LCDR1 comprises the amino acid sequence of SEQ ID NO: 53, the LCDR2 comprises the amino acid sequence of YAS, and the LCDR3 comprises the amino acid sequence of SEQ ID NO: 55; or (e) the HCDR1 comprises the amino acid sequence of SEQ ID NO: 56, the HCDR2 comprises the amino acid sequence of SEQ ID NO: 57, the HCDR3 comprises the amino acid sequence of SEQ ID NO: 58, the LCDR1 comprises the amino acid sequence of SEQ ID NO: 59, the LCDR2 comprises the amino acid sequence of YAS, and the LCDR3 comprises the amino acid sequence of SEQ ID NO: 61, wherein the dosage of the anti-IL-2 antibody is between about 0.5 mg / kg and 12 mg / kg of the subject, and wherein the antibody promotes the differential growth of immune cell subsets and reduces undesirable effects caused by IL-2, thereby treating the cancer in the subject.

2. The method of claim 1, wherein multiple doses of the composition comprising the anti-IL2 antibody are administered.

3. The method of claim 1 or claim 2, wherein the method further comprises administering at least a single low dose of IL-2, wherein the low dose of IL-2 comprises about 15×10 3 IU / Kg-500×10 3 IU / Kg between the subjects. The method of claim 3 , wherein the administration of IL-2 comprises subcutaneous administration.

5. The method of claim 3 or claim 4, wherein the IL-2 is administered before, simultaneously with, or after administration of the anti-IL-2 antibody.

6. The method according to any one of claims 3 to 5, wherein the IL-2 is administered in multiple doses.

7. The method of claim 6, wherein the multiple doses of IL-2 are administered before, simultaneously with, or after administration of the anti-IL-2 antibody, or any combination thereof.

8. The method of any one of claims 3-7, wherein the method further comprises administering a checkpoint inhibitor.

9. The method of claim 8, wherein the checkpoint inhibitor comprises PD-L1, PD-1, CTLA-4, TIGIT, TIM-3, B7-H3, CD73, LAG3, CD27, CD70, 4-1BB, GITR, OX40, SIRP-α (CD47), CD39, ILDR2, VISTA, BTLA, or VTCN-1.

10. The method of any one of claims 1 to 9, wherein the solid cancer comprises melanoma, metastatic melanoma, primary melanoma and metastatic melanoma, renal cell carcinoma (RCC), non-small cell lung cancer (NSCLC), urothelial carcinoma, adrenocortical carcinoma, clear cell renal cell carcinoma (ccRCC), triple-negative breast cancer, head and neck squamous cell carcinoma (HNSCC), gastric or gastroesophageal cancer, esophageal squamous cell carcinoma, cutaneous squamous cell carcinoma (cSCC), pancreatic adenocarcinoma, bile duct cancer, hepatocellular carcinoma (HCC), colorectal cancer (CRC), epithelial ovarian cancer, cervical cancer, endometrial cancer, thyroid cancer (follicular or papillary histology), or Merkel cell carcinoma.

11. The method of any one of claims 1-10, wherein the method comprises second-line therapy or third-line therapy or a combination thereof.

12. The method of any one of claims 1-11, wherein treating the subject reduces the size of the tumor, inhibits or reduces the growth of the tumor, or inhibits or reduces metastasis of the tumor, or any combination thereof.

13. The method according to any one of claims 1 to 12, wherein the VH and VL have the following amino acid sequences (a) the VH comprises the amino acid sequence of SEQ ID NO: 26, and the VL comprises the amino acid sequence of SEQ ID NO: 27; (b) the VH comprises the amino acid sequence of SEQ ID NO: 20, and the VL comprises the amino acid sequence of SEQ ID NO: 21; (c) the VH comprises the amino acid sequence of SEQ ID NO: 22, and the VL comprises the amino acid sequence of SEQ ID NO: 23; (d) the VH comprises the amino acid sequence of SEQ ID NO: 24, and the VL comprises the amino acid sequence of SEQ ID NO: 25; or (e) the VH comprises the amino acid sequence of SEQ ID NO: 36, and the VL comprises the amino acid sequence of SEQ ID NO:

37.

14. The method of any one of claims 1-13, wherein the antibody comprises an IgG, IgA, IgM, IgE, IgD, Fv, scFv, Fab, F(ab')2, a minibody, a diabody, or a triabody.

15. The method of any one of claims 1-14, wherein the antibody comprises a heavy chain comprising a mutation that reduces binding to an Fcγ receptor. The method according to claim 15 , wherein the mutations include L234A and L235A mutations.

17. The method according to any one of claims 1 to 16, wherein When the HCDR1 comprises the amino acid sequence of SEQ ID NO: 62, the HCDR2 comprises the amino acid sequence of SEQ ID NO: 63, the HCDR3 comprises the amino acid sequence of SEQ ID NO: 64, the LCDR1 comprises the amino acid sequence of SEQ ID NO: 65, the LCDR2 comprises the amino acid sequence of DAS, and the LCDR3 comprises the amino acid sequence of SEQ ID NO: 67, the amino acid sequence of the full-length heavy chain is shown in SEQ ID NO: 72, and the amino acid sequence of the full-length light chain is shown in SEQ ID NO: 73; - when the HCDR1 comprises the amino acid sequence of SEQ ID NO: 44, the HCDR2 comprises the amino acid sequence of SEQ ID NO: 45, the HCDR3 comprises the amino acid sequence of SEQ ID NO: 46, the LCDR1 comprises the amino acid sequence of SEQ ID NO: 47, the LCDR2 comprises the amino acid sequence of YAS, and the LCDR3 comprises the amino acid sequence of SEQ ID NO: 49, the amino acid sequence of the full-length heavy chain is shown in SEQ ID NO: 68, and the amino acid sequence of the full-length light chain is shown in SEQ ID NO: 69; and When the HCDR1 comprises the amino acid sequence of SEQ ID NO: 50, the HCDR2 comprises the amino acid sequence of SEQ ID NO: 51, the HCDR3 comprises the amino acid sequence of SEQ ID NO: 52, the LCDR1 comprises the amino acid sequence of SEQ ID NO: 53, the LCDR2 comprises the amino acid sequence of YAS, and the LCDR3 comprises the amino acid sequence of SEQ ID NO: 55, the amino acid sequence of the full-length heavy chain is shown in SEQ ID NO: 70, and the amino acid sequence of the full-length light chain is shown in SEQ ID NO:

71.

18. A method of treating cancer in a subject, comprising administering to the subject an anti-IL-2 antibody and IL-2, wherein the IL-2 antibody comprises a heavy chain variable region (VH) and a light chain variable region (VL), wherein the VH comprises heavy chain complementarity determining regions (HCDRs) HCDR1, HCDR2, and HCDR3, and the VL comprises light chain complementarity determining regions (LCDRs) LCDR1, LCDR2, and LCDR3, wherein the CDRs have the following amino acid sequences: (a) the HCDR1 comprises the amino acid sequence of SEQ ID NO: 62, the HCDR2 comprises the amino acid sequence of SEQ ID NO: 63, the HCDR3 comprises the amino acid sequence of SEQ ID NO: 64, the LCDR1 comprises the amino acid sequence of SEQ ID NO: 65, the LCDR2 comprises the amino acid sequence of DAS, and the LCDR3 comprises the amino acid sequence of SEQ ID NO: 67; (b) the HCDR1 comprises the amino acid sequence of SEQ ID NO: 38, the HCDR2 comprises the amino acid sequence of SEQ ID NO: 39, the HCDR3 comprises the amino acid sequence of SEQ ID NO: 40, the LCDR1 comprises the amino acid sequence of SEQ ID NO: 41, the LCDR2 comprises the amino acid sequence of YAS, and the LCDR3 comprises the amino acid sequence of SEQ ID NO: 43; (c) the HCDR1 comprises the amino acid sequence of SEQ ID NO: 44, the HCDR2 comprises the amino acid sequence of SEQ ID NO: 45, the HCDR3 comprises the amino acid sequence of SEQ ID NO: 46, the LCDR1 comprises the amino acid sequence of SEQ ID NO: 47, the LCDR2 comprises the amino acid sequence of YAS, and the LCDR3 comprises the amino acid sequence of SEQ ID NO: 49; (d) the HCDR1 comprises the amino acid sequence of SEQ ID NO: 50, the HCDR2 comprises the amino acid sequence of SEQ ID NO: 51, the HCDR3 comprises the amino acid sequence of SEQ ID NO: 52, the LCDR1 comprises the amino acid sequence of SEQ ID NO: 53, the LCDR2 comprises the amino acid sequence of YAS, and the LCDR3 comprises the amino acid sequence of SEQ ID NO: 55; or (e) the HCDR1 comprises the amino acid sequence of SEQ ID NO: 56, the HCDR2 comprises the amino acid sequence of SEQ ID NO: 57, the HCDR3 comprises the amino acid sequence of SEQ ID NO: 58, the LCDR1 comprises the amino acid sequence of SEQ ID NO: 59, the LCDR2 comprises the amino acid sequence of YAS, and the LCDR3 comprises the amino acid sequence of SEQ ID NO: 61, wherein the IL-2 is administered by subcutaneous injection, wherein the dose of the anti-IL-2 antibody is between about 0.5 mg / kg-12 mg / kg of the subject, and wherein the antibody promotes differential growth of immune cell subsets and reduces undesirable effects caused by IL-2, thereby treating the cancer in the subject.

19. A method for treating solid cancer in a subject, comprising administering to the subject an anti-IL-2 antibody, IL-2, and a checkpoint inhibitor, wherein the IL-2 antibody comprises a heavy chain variable region (VH) and a light chain variable region (VL), wherein the VH comprises heavy chain complementarity determining regions (HCDRs) HCDR1, HCDR2, and HCDR3, and the VL comprises light chain complementarity determining regions (LCDRs) LCDR1, LCDR2, and LCDR3, wherein the CDRs have the following amino acid sequences: (a) the HCDR1 comprises the amino acid sequence of SEQ ID NO: 62, the HCDR2 comprises the amino acid sequence of SEQ ID NO: 63, the HCDR3 comprises the amino acid sequence of SEQ ID NO: 64, the LCDR1 comprises the amino acid sequence of SEQ ID NO: 65, the LCDR2 comprises the amino acid sequence of DAS, and the LCDR3 comprises the amino acid sequence of SEQ ID NO: 67; (b) the HCDR1 comprises the amino acid sequence of SEQ ID NO: 38, the HCDR2 comprises the amino acid sequence of SEQ ID NO: 39, the HCDR3 comprises the amino acid sequence of SEQ ID NO: 40, the LCDR1 comprises the amino acid sequence of SEQ ID NO: 41, the LCDR2 comprises the amino acid sequence of YAS, and the LCDR3 comprises the amino acid sequence of SEQ ID NO: 43; (c) the HCDR1 comprises the amino acid sequence of SEQ ID NO: 44, the HCDR2 comprises the amino acid sequence of SEQ ID NO: 45, the HCDR3 comprises the amino acid sequence of SEQ ID NO: 46, the LCDR1 comprises the amino acid sequence of SEQ ID NO: 47, the LCDR2 comprises the amino acid sequence of YAS, and the LCDR3 comprises the amino acid sequence of SEQ ID NO: 49; (d) the HCDR1 comprises the amino acid sequence of SEQ ID NO: 50, the HCDR2 comprises the amino acid sequence of SEQ ID NO: 51, the HCDR3 comprises the amino acid sequence of SEQ ID NO: 52, the LCDR1 comprises the amino acid sequence of SEQ ID NO: 53, the LCDR2 comprises the amino acid sequence of YAS, and the LCDR3 comprises the amino acid sequence of SEQ ID NO: 55; or (e) the HCDR1 comprises the amino acid sequence of SEQ ID NO: 56, the HCDR2 comprises the amino acid sequence of SEQ ID NO: 57, the HCDR3 comprises the amino acid sequence of SEQ ID NO: 58, the LCDR1 comprises the amino acid sequence of SEQ ID NO: 59, the LCDR2 comprises the amino acid sequence of YAS, and the LCDR3 comprises the amino acid sequence of SEQ ID NO: 61, wherein the IL-2 is administered by subcutaneous injection, wherein the dose of the anti-IL-2 antibody is between about 0.5 mg / kg-12 mg / kg of the subject, wherein the antibody promotes differential growth of immune cell subsets and reduces undesirable effects caused by IL-2, and wherein the checkpoint inhibitor comprises PD-L1, PD-1, CTLA-4, TIGIT, TIM-3, B7-H3, CD73, LAG3, CD27, CD70, 4-1BB, GITR, OX40, SIRP-α (CD47), CD39, ILDR2, VISTA, BTLA or VTCN-1, thereby treating the cancer in the subject.

20. The method of claim 19, wherein the checkpoint inhibitor comprises a PD-L1 checkpoint inhibitor.

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