Agonist antibodies that bind human CD137 and uses thereof

Abstract

To provide compounds (e.g., antibodies, or antigen-binding fragments thereof) that bind to an epitope of CD137 and agonize CD137, and use of the compounds in methods for treating or ameliorating one or more symptoms of cancer.SOLUTION: In some aspects, the disclosure provides an isolated monoclonal antibody or an antigen binding portion thereof that specifically binds to human CD137, wherein the antibody or antigen binding portion binds to human CD137 with an affinity (KD) of about 40 nM to about 100 nM.SELECTED DRAWING: None

Description

[Technical Field]

[0001] Related Applications This application is a continuation of U.S. Provisional Patent Application No. 62 / 531,259, filed July 11, 2017; U.S. Provisional Patent Application No. 62 / 531,190, filed July 11, 2017; U.S. Provisional Patent Application No. 62 / 531,190, filed October 4, 2017. This application claims the benefit of U.S. Provisional Patent Application No. 62 / 568,231, filed October 26, 2017; U.S. Provisional Patent Application No. 62 / 577,257, filed October 26, 2017; and U.S. Provisional Patent Application No. 62 / 577,259, filed October 26, 2017, the entire contents of which are incorporated herein by reference. [Background technology]

[0002] In recent years, increasing evidence has suggested that the immune system acts as an important barrier to tumor formation and progression. The idea that naturally occurring T cells with antitumor potential or activity exist in cancer patients has led to the development of immunotherapeutic approaches in the oncology field. This provides a rationale for the development of immune cells, such as T cells, macrophages, and natural killers. Immune cells such as killer cells can exhibit antitumor activity and effectively control the development and growth of malignant tumors. Tumor-specific or tumor-associated antigens can induce immune cells to recognize and eliminate malignant tumors (Chen & Mellman, (2013) Immunity 39(1):1-10). Despite the existence of tumor-specific immune responses, malignant tumors are also affected by various immune responses. Tumors often escape or evade immune attack through these mechanisms, resulting in failure to control tumor initiation and progression (Motz & Coukos, (2013) Immunity 39(1):61-730). Indeed, a hallmark of cancer emergence is the exploitation of these immune regulatory mechanisms and the subversion of antitumor immune responses, allowing tumors to evade and escape immune killing (Hanahan and Weinberg (2011) Cell 144(5):646-674).

[0003] New approaches to cancer immunotherapy involve counteracting these immune evasion and escape mechanisms and inducing the endogenous immune system to reject tumors. Tumor necrosis factor receptor superfamily member 9 (TNFRSF9), also known as 4-1BB and "induced by lymphocyte activation" (ILA), is a transmembrane costimulatory receptor protein belonging to the tumor necrosis factor superfamily. CD137 is a T cell costimulatory receptor induced by TCR activation (Nam et al., (2005) Curr Cancer Drug Targets 5:357-363; Watts et al., (2005) Annu Rev Immunol 23:23-68). In addition to its expression on activated CD4+ and CD8+ T cells, CD137 is also expressed on CD4+CD25+ regulatory T cells, activated It is also expressed on natural killer (NK) T cells and NK-T cells, monocytes, neutrophils, and dendritic cells. It will be revealed.

[0004] Under physiological conditions, CD137 ligates to CD137 ligand (CD137L), an agonistic membrane molecule present on antigen-presenting cells, including B cells, monocytes, macrophages, and dendritic cells. Upon interaction with its ligand, CD137 mediates TCR-induced T cell proliferation, cytokine production, and functional success (Watts et al., (2005) Annu Rev Immunol 23:23-68). This results in increased maturation and prolonged survival of CD8+ T cells, which may contribute to the immune system's ability to attack tumors. Various agonists (e.g., agonistic antibodies, recombinant CD137L protein, and CD137 specific The potential of CD137 costimulation using specific aptamers has been reported in a number of models. (Dharmadhikari et al., (2016) Oncoimmunology 5(4):e1113367 and (References therein). A recent report on the clinical evaluation of an agonistic CD137 antibody (Urelumab, BMS-663513; Bristol-Myers Squibb) demonstrated anti-CD137 activity in human subjects. Treatment-related adverse events, including evidence of severe hepatotoxicity (hypertransaminasemia) correlated with the total body dose, have been reported (Segal et al., (2016) Clin Cancer Res 23(8):1929-1936). In contrast, another agonistic CD137 antibody (utomilumab, PF-05082566; Pfizer) was tested in combination with an anti-PD-1 antibody (pembrolizumab) without any dose-limiting toxicities, but showed results comparable to anti-PD-1 antibody monotherapy (Tolcher, A. et al., (2017) Clin Cancer Res 23(18): 5349-5357). These results suggest that CD137 agonists can bind to human CD137 and provide safe and effective treatments for patients with a variety of diseases and conditions, including cancer, that are suitable for treatment with CD137 agonists. This highlights the continuing unmet need for novel agonistic antibodies that are well characterized for development into therapeutic agents. [Prior art documents] [Non-patent literature]

[0005] [Non-Patent Document 1] Chen & Mellman, (2013) Immunity 39(1):1-10 [Non-patent document 2] Motz & Coukos, (2013) Immunity 39(1):61-730 [Non-patent document 3] Hanahan and Weinberg (2011) Cell 144(5):646-674 [Non-patent document 4] Nam et al., (2005) Curr Cancer Drug Targets 5:357-363 [Non-patent document 5] Watts et al., (2005) Annu Rev Immunol 23:23-68 [Non-patent document 6] Dharmadhikari et al., (2016) Oncoimmunology 5(4):e1113367 [Non-Patent Document 7] Segal et al., (2016) Clin Cancer Res 23(8):1929-1936 [Non-patent document 8] Tolcher, A. et al., (2017) Clin Cancer Res 23(18): 5349-5357 Summary of the Invention [Means for solving the problem]

[0006] The present disclosure is based, at least in part, on the development of novel agonistic anti-CD137 antibodies that confer protective anti-tumor immunity in animals. In particular, the antibodies described herein are effective against a variety of tumors. The antibodies described herein are effective against a wide range of tumor types and across a broad dose range. As further illustrated in the Examples, the antibodies described herein are therapeutically effective against very large tumors. For example, treating tumor-bearing mice with the agonistic anti-CD137 antibodies described herein resulted in This makes it 1,800mm 3Treatment of these mice resulted in complete regression of tumors of up to 100% tumor size. As shown in Figure 15, treatment of these mice also resulted in protective immunity. Consistent with the observed efficacy, there were positive immunophenotypic changes in the tumor microenvironment, such as increased immune cell infiltration accompanied by a decrease in regulatory T cell and exhausted T cell populations (see, e.g., Figures 22A-22D).

[0007] As mentioned above, agonism of CD137 has been shown to induce certain adverse effects in humans, including hepatotoxicity-related death. associated with events (e.g., Segal et al. (2017) Clin Cancer Res 23(8) : 1929-1935). Similar toxicities resulting from treatment with agonistic anti-CD137 antibodies (e.g., 3H3 antibody) have also been observed in animal models (see, e.g., Bartkowiak et al. (2018) Clin Cancer Res 24(5) :1138-1151). The agonistic anti-CD137 antibodies described herein may inhibit, for example, liver enzymes (e.g., alanine aminotransferase). liver, as determined by plasma levels of ALT and immune cell infiltration. For example, there is no evidence of increased immune cell infiltration in the liver or spleen in mice treated with the antibodies. Thus, the antibodies described herein are not only highly effective, but also reduce certain toxicities associated with CD137 agonism.

[0008] Although the present disclosure is not limited to any particular theory or mechanism of action, it is believed that the superior therapeutic and toxicity-reducing properties of the antibodies described herein are derived in part from one or both of the antibody's affinity and the novel epitope to which it binds. That is, the antibodies described herein share a common novel epitope that is distinct from other agonistic anti-CD137 antibodies. And, as illustrated in the Examples, the association of the antibodies described herein with the epitope allows for the binding of different CD137 epitopes. Compared to agonistic antibodies that bind to the target peptide, + Different in vitro activities occur, such as effects on cytokine production by T cells and macrophages and intracellular signaling. Furthermore, the affinity range (sweet spot) of antibodies has been shown to be particularly optimal for antitumor activity. For example, antibodies with intermediate affinity have been shown to be more effective against large tumors than antibodies with higher or lower affinity.

[0009] In view of the foregoing, in some embodiments, the present disclosure provides a method for the production of antibodies that specifically bind to human CD137. The present invention provides an isolated monoclonal antibody or antigen-binding portion thereof, wherein the antibody or antigen-binding portion has an affinity (K D In some embodiments, the present disclosure provides an isolated monoclonal antibody or antibody that specifically binds to human CD137. or an antigen-binding portion thereof, wherein the antibody or antigen-binding portion has an affinity (K D ) binds to human CD137. In embodiments, the affinity of the anti-CD137 antibody for human CD137 is at least 2-fold (e.g., at least 3, 4, 5, 6, 7, 8, 9, or 10-fold) higher than the affinity of mAb10 for mouse CD137. In some embodiments, the affinity of the anti-CD137 antibody is 500, 450, 400, 350, 300, 250, 200, 250, 200, 175, 150, 125, 110, or 100 nM or less. In some embodiments, the affinity of the anti-CD137 antibody for human CD137 is at least 2-fold (e.g., at least 3, 4, 5, 6, 7, 8, 9, or 10-fold) higher than the affinity of mAb10 for mouse CD137. at least 2-fold (e.g., at least 3, 4, 5, 6, 7, 8, 9, or 10-fold) higher than 500 , 450, 400, 350, 300, 250, 200, 250, 200, 175, 150, 125, 110 or 100 nM or less do.

[0010] In some aspects, the present disclosure provides an isolated monoclonal antibody or antibody fragment that specifically binds to human CD137. In some embodiments, the present disclosure provides an isolated monoclonal antibody or antigen-binding portion thereof that specifically binds to human CD137, wherein the antibody or antigen-binding portion thereof binds to an epitope on human CD137 that includes one or more (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, or all 25) amino acids 111-132 of SEQ ID NO:3. In some embodiments, the present disclosure provides an isolated monoclonal antibody or antigen-binding portion thereof that specifically binds to human CD137, wherein the antibody or antigen-binding portion thereof binds to an epitope within amino acids 111-132 of SEQ ID NO:3. In an embodiment, the present disclosure provides an isolated monoclonal antibody that specifically binds to human CD137 or and an antigen-binding portion thereof, wherein the antibody or antigen-binding portion is of SEQ ID NO: 3. In some embodiments, the epitope binds to all or part of amino acids 111-132 of SEQ ID NO: 3. In some embodiments, the epitope binds to K114 of SEQ ID NO: 3. In some embodiments, the epitope includes residues E111, T113, and K114 of SEQ ID NO: 3. In some embodiments, the epitope binds to all or part of amino acids 111-132 ... , K114, N126, and I132 residues. In some embodiments, the epitope comprises residues E111, T113, K114, and P135 of SEQ ID NO: 3. In some embodiments, the epitope comprises residues E111, T113, K114, N126, I132, and P135 of SEQ ID NO: 3. In some embodiments, the antibody or antigen-binding portion thereof binds to human CD137 with an affinity of about 30 nM to about 100 nM (e.g., about 30 nM to about 110 nM).

[0011] In some embodiments, the present disclosure provides an isolated monoclonal antibody that specifically binds to human CD137. The antibody or antigen-binding portion thereof has an affinity (K D ) binds to human CD137 and and binds to an epitope on human CD137 comprising K114 of SEQ ID NO: 3. In some embodiments, the present disclosure provides an isolated monoclonal antibody or antigen-binding portion thereof that specifically binds to human CD137. The antibody or antigen-binding portion has a concentration of about 30 nM to 100 nM (e.g., about 30 nM ~100 nM) affinity (K D ) binds to human CD137 and contains K114 of SEQ ID NO:3 In some embodiments, the epitope comprises residues E111, T113, and K114 of SEQ ID NO: 3. In some embodiments, the epitope comprises residues E111, T113, K114, N126, and I132 of SEQ ID NO: 3. In some embodiments, the epitope comprises residues E111, T113, K114, N126, and I132 of SEQ ID NO: 3. In some embodiments, the epitope comprises residues E111, T113, K114 and P135 of number 3. It includes residues E111, T113, K114, N126, I132 and P135 of SEQ ID NO:3.

[0012] In some embodiments, the present disclosure provides an isolated monoclonal antibody that specifically binds to human CD137. The antibody or antigen-binding portion thereof has an affinity (K D ) binds to human CD137 and a sequence of one or more amino acid residues corresponding to amino acids 111 to 135 of SEQ ID NO: 3; In some embodiments, the epitope binds to an epitope on human CD137 comprising the sequence It contains 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, or 25 amino acid residues corresponding to amino acids 111 to 135 of No. 3.

[0013] In some embodiments, the present disclosure provides an isolated monoclonal antibody that specifically binds to human CD137. The antibody or antigen-binding portion thereof has an affinity (K D ) binds to human CD137 and and binds to an epitope on human CD137 located within amino acid residues 111 to 135 of SEQ ID NO: 3. In some embodiments, the epitope is at least 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, or 15 amino acids. In some embodiments, the epitope is fewer than 25, 24, 23, 22, 21, 20, 19, 18, 17, 16, 15, 14, 13, 12, 11, 10, 9, 8, 7, 6, or 5 amino acids.

[0014] In some embodiments, the present disclosure provides an isolated monoclonal antibody that specifically binds to human CD137. The antibody or antigen-binding portion thereof has an affinity (K D ) binds to human CD137 and and binds to an epitope on human CD137 that includes ELTK (corresponding to amino acid residues 111-114 of SEQ ID NO: 3). In some embodiments, the epitope further includes one or more of residues N126, I132, and P135 of SEQ ID NO: 3.

[0015] In any of the foregoing embodiments, the epitope is a non-linear epitope. In any of the foregoing embodiments, mutation of residue K114 of SEQ ID NO: 3 abolishes binding of the antibody, or antigen-binding portion thereof, to human CD137.

[0016] In any of the foregoing embodiments, the antibodies or antigen-binding portions thereof described herein have an affinity (K D In some embodiments, the antibody or antigen-binding portion thereof binds to human CD137 at a non- In some embodiments, the antibody or antigen-binding portion thereof does not inhibit the interaction of CD137 with CD137L. In some embodiments, the non-ligand binding region binds to the ligand binding region. In any of the foregoing embodiments, the protein comprises a CRD III and a CRD IV. and the antibody or antigen-binding portion thereof does not inhibit trimer formation of CD137:CD137L monomers. stomach.

[0017] In some embodiments, the present disclosure provides an isolated monoclonal antibody that specifically binds to human CD137. providing an antibody or antigen-binding portion thereof, wherein the antibody or antigen-binding portion: (i) an affinity (K) of about 30 to 100 nM (e.g., about 30 nM to about 100 nM); D ) binds to human CD137; (ii) binds to a non-ligand binding region of the extracellular domain of human CD137; and (iii) binds to an epitope on human CD137 that includes K114 of SEQ ID NO:3. In some aspects, the present disclosure provides an isolated monoclonal antibody or antigen-binding portion thereof that specifically binds to human CD137, wherein the antibody or antigen-binding portion is: (i) an affinity (K) of about 30 to 100 nM (e.g., about 30 nM to about 100 nM); D ) binds to human CD137; (ii) does not inhibit the interaction between human CD137 and human CD137 ligand; and (iii) binds to an epitope on human CD137 that includes K114 of SEQ ID NO:3. In some embodiments, the present disclosure provides an isolated monoclonal antibody that specifically binds to human CD137. or an antigen-binding portion thereof, wherein the antibody or antigen-binding portion is ) affinity (K D ) that binds to human CD137 and (ii) does not inhibit the formation of CD137:CD137L monomer trimers (i.e., CD137:CD137L trimer:trimer complexes). In some embodiments, the present disclosure provides antibodies specific for human CD137. and (i) an affinity of about 30 nM to 100 nM (e.g., about 30 nM to about 100 nM) that binds to a target molecule. Nity (K D (ii) binds to human CD137 at a non-ligand binding region of the extracellular domain of human CD137. In some embodiments, the present disclosure provides antibodies that specifically bind to human CD137. The present invention features an isolated monoclonal antibody or antigen-binding portion thereof, wherein the antibody or antigen-binding portion has (i) an affinity (K D ) binds to human CD137, and (ii) does not inhibit the interaction between human CD137 and CD137 ligand.

[0018] In any of the foregoing embodiments, the antibody or antigen-binding portion thereof comprises a heavy chain CDR3 comprising the amino acid sequence DXXXXLXXXXYXYYX (SEQ ID NO: 126), where X is any amino acid. In some embodiments, the antibody or antigen-binding portion thereof has the amino acid sequence DXPFXLDXXYYYYYX (sequence wherein X is any amino acid. In any of the foregoing embodiments, mutation of residues D95, L100, Y100E, Y100G, Y100H, or combinations thereof, in the heavy chain CDR3 to alanine results in loss of binding to human CD137. In any of the above, the P97, F98, D100A, Y100D, Y100F residues or combinations thereof are Mutation of the nucleotides to the nucleotides results in reduced binding to human CD137. wherein the antibody or antigen-binding portion thereof comprises a heavy chain CDR and a light chain CDR, and the heavy chain CDR3 comprises the amino acid sequence set forth in SEQ ID NO:68.

[0019] In another aspect, the present disclosure provides an isolated monoclonal antibody that specifically binds to human CD137. or an antigen-binding portion thereof, wherein (i) The antibody or antigen-binding portion thereof has an affinity of about 30 to 100 nM (e.g., about 30 nM to about 100 nM). Nity (K D and (ii) the antibody or antigen-binding portion thereof binds to human CD137; and a heavy chain CDR3 comprising the amino acid sequence DXXXXLXXXXYXYYX (SEQ ID NO: 126), where X is any amino acid. In some embodiments, X is any amino acid except alanine.

[0020] In another aspect, the present disclosure provides an isolated monoclonal antibody that specifically binds to human CD137. or an antigen-binding portion thereof, wherein (i) The antibody or antigen-binding portion thereof has an affinity of about 30 to 100 nM (e.g., about 30 nM to about 100 nM). Nity (K D ) binds to human CD137; and (ii) an antibody, or antigen-binding portion thereof, having the amino acid sequence DX1X2X3X4LX5X6X7X8YX9YYX 10 (SEQ ID NO: 128), wherein X1 is any amino acid and wherein X2 is a non-polar X is an amino acid, wherein X is a non-polar amino acid, wherein X is any amino acid, wherein X is a polar amino acid, wherein X is any amino acid, wherein X is a polar amino acid, wherein X is a polar amino acid, and wherein X 10 In some embodiments, X2 is proline and X3 is phenyl. X5 is phenylalanine or tryptophan, X5 is aspartic acid or glutamic acid, X8 is tyrosine, and X9 is tyrosine.

[0021] In another aspect, the present disclosure provides an isolated monoclonal antibody that specifically binds to human CD137. or an antigen-binding portion thereof, wherein (i) The antibody or antigen-binding portion thereof has an affinity of about 30 to 100 nM (e.g., about 30 nM to about 100 nM). Nity (K D ) binds to human CD137; and (ii) an antibody or antigen-binding portion thereof, comprising E111, T113, K114, N126, I132 and SEQ ID NO: 3; It specifically binds to an epitope on human CD137 that includes one or more of the P135 and P136 residues.

[0022] In another aspect, the present disclosure provides an isolated monoclonal antibody that specifically binds to human CD137. or an antigen-binding portion thereof, wherein (i) The antibody or antigen-binding portion thereof has an affinity of about 30 to 100 nM (e.g., about 30 nM to about 100 nM). Nity (K D ) binds to human CD137; (ii) an antibody or antigen-binding portion thereof, comprising E111, T113, K114, N126, I132 and SEQ ID NO: 3; It specifically binds to an epitope on human CD137 that includes one or more of the P135 and P136 residues. (iii) the antibody, or antigen-binding portion thereof, has the amino acid sequence DXXXXLXXXXYXYYX (SEQ ID NO: 126). wherein X is any amino acid; or (iv) combinations thereof. In some embodiments, X is any amino acid except alanine. It is an amino acid.

[0023] In another aspect, the present disclosure provides an isolated monoclonal antibody that specifically binds to human CD137. or an antigen-binding portion thereof, wherein (i) The antibody or antigen-binding portion thereof has an affinity of about 30 to 100 nM (e.g., about 30 nM to about 100 nM). Nity (K D ) binds to human CD137; (ii) an antibody or antigen-binding portion thereof, comprising E111, T113, K114, N126, I132 and SEQ ID NO: 3; It specifically binds to an epitope on human CD137 that includes one or more of the P135 and P136 residues. (iii) an antibody, or antigen-binding portion thereof, having the amino acid sequence DX1X2X3X4LX5X6X7X8YX9YYX 10 (Distribution wherein X1 is any amino acid and X2 is a non-polar X is a polar amino acid, wherein X is a nonpolar amino acid, wherein X is any amino acid, wherein X is a polar amino acid, wherein X is any amino acid, wherein X is a polar amino acid, wherein X is a polar amino acid, and wherein X is a 10 is any amino acid; or (iv) Combinations thereof. In some embodiments, X2 is proline, X3 is phenylalanine or tryptophan, X5 is aspartic acid or glutamic acid, X8 is tyrosine, and X9 is tyrosine.

[0024] In another aspect, the present disclosure provides an isolated monoclonal antibody that specifically binds to human CD137. or an antigen-binding portion thereof, wherein (i) The antibody or antigen-binding portion thereof has an affinity of about 30 to 100 nM (e.g., about 30 nM to about 100 nM). Nity (K D ) binds to human CD137; (ii) an antibody or antigen-binding portion thereof, comprising E111, T113, K114, N126, I132 and SEQ ID NO: 3; specifically binds to an epitope on human CD137 that includes one or more P135 residues; and (iii) the antibody, or antigen-binding portion thereof, has the amino acid sequence DXXXXLXXXXYXYYX (SEQ ID NO: 126). wherein X is any amino acid. In some embodiments, X is any amino acid except alanine.

[0025] In another aspect, the present disclosure provides an isolated monoclonal antibody that specifically binds to human CD137. or an antigen-binding portion thereof, wherein (i) The antibody or antigen-binding portion thereof has an affinity of about 30 to 100 nM (e.g., about 30 nM to about 100 nM). Nity (K D ) binds to human CD137; (ii) an antibody or antigen-binding portion thereof, comprising E111, T113, K114, N126, I132 and SEQ ID NO: 3; specifically binds to an epitope on human CD137 that includes one or more P135 residues; and (iii) an antibody, or antigen-binding portion thereof, having the amino acid sequence DX1X2X3X4LX5X6X7X8YX9YYX 10 (Distribution wherein X1 is any amino acid and X2 is a non-polar X is a polar amino acid, wherein X is a nonpolar amino acid, wherein X is any amino acid, wherein X is a polar amino acid, wherein X is any amino acid, wherein X is a polar amino acid, wherein X is a polar amino acid, and wherein X is a 10 is any amino acid. In some embodiments, X2 is proline and X3 is X is phenylalanine or tryptophan, X5 is aspartic acid or glutamic acid, X8 is tyrosine, and X9 is tyrosine.

[0026] In any of the foregoing embodiments, the epitope comprises K114. In any of the foregoing embodiments, the epitope comprises E111, T113 and K114. In any of the foregoing embodiments, the epitope comprises E11, T113, K114, N126 and I132. In any of the foregoing embodiments, Thus, the epitope includes residues E111, T113, K114, N126, I132 and P135 of SEQ ID NO:3.

[0027] In another aspect, the present disclosure provides an isolated monoclonal antibody that specifically binds to human CD137. or an antigen-binding portion thereof, wherein (i) The antibody or antigen-binding portion thereof has an affinity of about 30 to 100 nM (e.g., about 30 nM to about 100 nM). Nity (K D ) binds to human CD137; and (ii) an antibody or antigen-binding portion thereof, which corresponds to amino acids 111 to 135 of SEQ ID NO: 3; It specifically binds to an epitope comprising a sequence of one or more amino acid residues.

[0028] In another aspect, the present disclosure provides an isolated monoclonal antibody that specifically binds to human CD137. or an antigen-binding portion thereof, wherein (i) The antibody or antigen-binding portion thereof has an affinity of about 30 to 100 nM (e.g., about 30 nM to about 100 nM). Nity (K D ) binds to human CD137; (ii) an antibody or antigen-binding portion thereof, which corresponds to amino acids 111 to 135 of SEQ ID NO: 3; It specifically binds to an epitope comprising a sequence of one or more amino acid residues. (iii) the antibody, or antigen-binding portion thereof, has the amino acid sequence DXXXXLXXXXYXYYX (SEQ ID NO: 126). wherein X is any amino acid; or (iv) combinations thereof. In some embodiments, X is any amino acid except alanine. It is an amino acid.

[0029] In another aspect, the present disclosure provides an isolated monoclonal antibody that specifically binds to human CD137. or an antigen-binding portion thereof, wherein (i) The antibody or antigen-binding portion thereof has an affinity of about 30 to 100 nM (e.g., about 30 nM to about 100 nM). Nity (K D ) binds to human CD137; (ii) an antibody or antigen-binding portion thereof, which corresponds to amino acids 111 to 135 of SEQ ID NO: 3; It specifically binds to an epitope comprising a sequence of one or more amino acid residues. (iii) an antibody, or antigen-binding portion thereof, having the amino acid sequence DX1X2X3X4LX5X6X7X8YX9YYX 10 (Distribution wherein X1 is any amino acid and X2 is a non-polar X is a polar amino acid, wherein X is a nonpolar amino acid, wherein X is any amino acid, wherein X is a polar amino acid, wherein X is any amino acid, wherein X is a polar amino acid, wherein X is a polar amino acid, and wherein X is a 10 is any amino acid; or (iv) Combinations thereof. In some embodiments, X2 is proline, X3 is phenylalanine or tryptophan, X5 is aspartic acid or glutamic acid, X8 is tyrosine, and X9 is tyrosine.

[0030] In another aspect, the present disclosure provides an isolated monoclonal antibody that specifically binds to human CD137. or an antigen-binding portion thereof, wherein (i) The antibody or antigen-binding portion thereof has an affinity of about 30 to 100 nM (e.g., about 30 nM to about 100 nM). Nity (K D ) binds to human CD137; (ii) an antibody or antigen-binding portion thereof, which corresponds to amino acids 111 to 135 of SEQ ID NO: 3; specifically binds to an epitope comprising a sequence of one or more amino acid residues; and (iii) the antibody, or antigen-binding portion thereof, has the amino acid sequence DXXXXLXXXXYXYYX (SEQ ID NO: 126). wherein X is any amino acid. In some embodiments, X is any amino acid except alanine.

[0031] In another aspect, the present disclosure provides an isolated monoclonal antibody that specifically binds to human CD137. or an antigen-binding portion thereof, wherein (i) The antibody or antigen-binding portion thereof has an affinity of about 30 to 100 nM (e.g., about 30 nM to about 100 nM). Nity (K D ) binds to human CD137; (ii) an antibody or antigen-binding portion thereof, which corresponds to amino acids 111 to 135 of SEQ ID NO: 3; specifically binds to an epitope comprising a sequence of one or more amino acid residues; and (iii) an antibody, or antigen-binding portion thereof, having the amino acid sequence DX1X2X3X4LX5X6X7X8YX9YYX 10 (Distribution wherein X1 is any amino acid and X2 is a non-polar X is a polar amino acid, wherein X is a nonpolar amino acid, wherein X is any amino acid, wherein X is a polar amino acid, wherein X is any amino acid, wherein X is a polar amino acid, wherein X is a polar amino acid, and wherein X is a 10 is any amino acid. In some embodiments, X2 is proline and X3 is X is phenylalanine or tryptophan, X5 is aspartic acid or glutamic acid, X8 is tyrosine, and X9 is tyrosine.

[0032] In any of the foregoing embodiments, the epitope is selected from amino acids 111 to 135 of SEQ ID NO: 3. and 25 amino acid residues corresponding to:

[0033] In some embodiments, the present disclosure provides an isolated monoclonal antibody that specifically binds to human CD137. providing a human body or an antigen-binding portion thereof, wherein: (i) The antibody or antigen-binding portion thereof has an affinity of about 30 to 100 nM (e.g., about 30 nM to about 100 nM). binds to human CD137 with high affinity; and (ii) an antibody or antigen-binding portion thereof, which is a nucleotide sequence encoding ELTK (amino acid residues 111 to 114 of SEQ ID NO: 3); It specifically binds to an epitope containing the

[0034] In some embodiments, the present disclosure provides an isolated monoclonal antibody that specifically binds to human CD137. providing a human body or an antigen-binding portion thereof, wherein: (i) The antibody or antigen-binding portion thereof has an affinity of about 30 to 100 nM (e.g., about 30 nM to about 100 nM). Nity (K D ) binds to human CD137; (ii) an antibody or antigen-binding portion thereof, which is a nucleotide sequence encoding ELTK (amino acid residues 111 to 114 of SEQ ID NO: 3); specifically binds to an epitope containing (iii) the antibody, or antigen-binding portion thereof, has the amino acid sequence DXXXXLXXXXYXYYX (SEQ ID NO: 126). wherein X is any amino acid; or (iv) combinations thereof. In some embodiments, X is any amino acid except alanine. It is an amino acid.

[0035] In some embodiments, the present disclosure provides an isolated monoclonal antibody that specifically binds to human CD137. providing a human body or an antigen-binding portion thereof, wherein: (i) The antibody or antigen-binding portion thereof has an affinity of about 30 to 100 nM (e.g., about 30 nM to about 100 nM). Nity (K D ) binds to human CD137; (ii) the antibody, or antigen-binding portion thereof, specifically binds to an epitope comprising ELTK (corresponding to amino acid residues 111 to 114 of SEQ ID NO: 3); (iii) an antibody, or antigen-binding portion thereof, having the amino acid sequence DX1X2X3X4LX5X6X7X8YX9YYX 10 (Distribution wherein X1 is any amino acid and X2 is a non-polar X is a polar amino acid, wherein X is a nonpolar amino acid, wherein X is any amino acid, wherein X is a polar amino acid, wherein X is any amino acid, wherein X is a polar amino acid, wherein X is a polar amino acid, and wherein X is a 10 is any amino acid; or (iv) Combinations thereof. In some embodiments, X2 is proline, X3 is phenylalanine or tryptophan, X5 is aspartic acid or glutamic acid, X8 is tyrosine, and X9 is tyrosine.

[0036] In some embodiments, the present disclosure provides an isolated monoclonal antibody that specifically binds to human CD137. providing a human body or an antigen-binding portion thereof, wherein: (i) The antibody or antigen-binding portion thereof has an affinity of about 30 to 100 nM (e.g., about 30 nM to about 100 nM). Nity (K D ) binds to human CD137; (ii) an antibody or antigen-binding portion thereof, which is a nucleotide sequence encoding ELTK (amino acid residues 111 to 114 of SEQ ID NO: 3); specifically binds to an epitope containing an epitope corresponding to (iii) the antibody, or antigen-binding portion thereof, has the amino acid sequence DXXXXLXXXXYXYYX (SEQ ID NO: 126). wherein X is any amino acid. In some embodiments, X is any amino acid except alanine.

[0037] In some embodiments, the present disclosure provides an isolated monoclonal antibody that specifically binds to human CD137. providing a human body or an antigen-binding portion thereof, wherein: (i) The antibody or antigen-binding portion thereof has an affinity of about 30 to 100 nM (e.g., about 30 nM to about 100 nM). Nity (K D ) binds to human CD137; (ii) an antibody or antigen-binding portion thereof, which is a nucleotide sequence encoding ELTK (amino acid residues 111 to 114 of SEQ ID NO: 3); specifically binds to an epitope containing an epitope corresponding to (iii) an antibody, or antigen-binding portion thereof, having the amino acid sequence DX1X2X3X4LX5X6X7X8YX9YYX 10 (Distribution wherein X1 is any amino acid and X2 is a non-polar X is a polar amino acid, wherein X is a nonpolar amino acid, wherein X is any amino acid, wherein X is a polar amino acid, wherein X is any amino acid, wherein X is a polar amino acid, wherein X is a polar amino acid, and wherein X is a 10 is any amino acid. In some embodiments, X2 is proline and X3 is X is phenylalanine or tryptophan, X5 is aspartic acid or glutamic acid, X8 is tyrosine, and X9 is tyrosine.

[0038] In any of the foregoing embodiments, the epitope includes ELTK residues of SEQ ID NO: 3 (corresponding to amino acid residues 111-114 of SEQ ID NO: 3). In some embodiments, the epitope includes ELTK of SEQ ID NO: 3 (corresponding to amino acid residues 111-114 of SEQ ID NO: 3) and residues N126, I132, and P135 of SEQ ID NO: 3.

[0039] In any of the foregoing embodiments, the epitope is a non-linear epitope. In some embodiments, a mutation at the K114 residue of human CD137 (SEQ ID NO: 3) is made to an antibody or abolishes binding of the antigen-binding moiety.

[0040] In any of the foregoing embodiments, the antibody, or antigen-binding portion thereof, comprises a heavy chain CDR3 comprising the amino acid sequence DXPFXLDXXYYYYYX (SEQ ID NO: 128), where X is any amino acid. In some embodiments, mutation of residues D95, L100, Y100E, Y100G, Y100H, or combinations thereof, in the heavy chain CDR3 of an antibody or antigen-binding portion thereof described herein enhances binding to human CD137. In some embodiments, the alteration of residues P97, F98, D100A, Y100D, Y100F, or a combination thereof, in the heavy chain CDR3 of an antibody or antigen-binding portion thereof described herein results in a loss of binding. Mutation of the amino acid sequence to the amino acid sequence results in reduced binding to human CD137. P97, F98, D100A, Y100D, Y100F of the heavy chain CDR3 of the antibody or antigen-binding portion thereof Mutation of these residues or their combinations to any residue other than alanine resulted in a significant increase in the activity of human CD137. This results in increased binding.

[0041] In any of the foregoing embodiments, the antibody or antigen-binding portion thereof has a (K D ) binds to human CD137. In any of the foregoing embodiments, the antibody or its antigen-binding The moiety is a (K D ) to human CD137 Combine.

[0042] In any of the foregoing embodiments, the antibody or antigen-binding portion thereof comprises a heavy chain CDR and a light chain CDR, wherein the heavy chain CDR3 comprises the amino acid sequence set forth in SEQ ID NO:68.

[0043] In any of the foregoing embodiments, the antibody or antigen-binding portion thereof comprises heavy chain and light chain CDRs selected from the group consisting of: (a) heavy chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 48, 56 and 68, respectively; and light chain CDR1, CDR2 and CDR3 set forth in SEQ ID NOs: 69, 78 and 89, respectively; and (b) Heavy chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 51, 108 and 68, respectively, and light chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 69, 78 and 89, respectively.

[0044] In any of the foregoing embodiments, the antibody or antigen-binding portion thereof comprises a heavy chain variable region and a light chain variable region, wherein the heavy chain variable region comprises an amino acid sequence selected from the group consisting of SEQ ID NOs: 4 and 101, and wherein the light chain variable region comprises the amino acid sequence of SEQ ID NO: 6.

[0045] In any of the foregoing embodiments, the antibody or antigen-binding portion thereof comprises a heavy chain variable region and a light chain variable region comprising an amino acid sequence selected from the group consisting of: (a) SEQ ID NOs: 4 and 6, respectively; and (b) SEQ ID NOs: 101 and 6, respectively.

[0046] In any of the foregoing embodiments, the antibody or antigen-binding portion thereof comprises a heavy chain variable region and a light chain variable region, wherein the heavy chain variable region comprises an amino acid sequence at least 90% identical to an amino acid sequence selected from the group consisting of SEQ ID NOs: 4 and 101. and in this case, the light chain variable region has at least one amino acid sequence different from that of SEQ ID NO:6. The amino acid sequences are at least 90% identical.

[0047] In any of the foregoing embodiments, the antibody or antigen-binding portion thereof may comprise a heavy chain containing an amino acid sequence that is at least 90% identical to an amino acid sequence selected from the group consisting of: The variable and light chain variable regions include: (a) SEQ ID NOs: 4 and 6, respectively; and (b) SEQ ID NOs: 101 and 6, respectively.

[0048] In any of the preceding embodiments, the antibody or antigen-binding portion of any one of claims 1 to 27, wherein the antibody or antigen-binding portion comprises a heavy chain and a light chain comprising an amino acid sequence selected from the group consisting of: (a) SEQ ID NOs: 129 and 133, respectively; and (b) SEQ ID NOs: 131 and 133, respectively.

[0049] In any of the foregoing embodiments, the isolated monoclonal antibody or antigen-binding portion thereof described herein is an agonist of human CD137 activity.

[0050] In any of the foregoing aspects, the isolated monoclonal antibody or antigen-binding portion thereof described herein is selected from the group consisting of mAb1 and an antigen-binding portion thereof, for binding to an epitope of human CD137. Competes with the original binding fragment.

[0051] In some embodiments, the present disclosure provides an isolated monoclonal antibody or antibody that specifically binds to CD137. or an antigen-binding portion thereof, wherein the antibody or antigen-binding portion comprises a heavy chain CDR and a light chain CDR selected from the group consisting of: (a) heavy chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 48, 56 and 68, respectively; and light chain CDR1, CDR2 and CDR3 set forth in SEQ ID NOs: 69, 78 and 89, respectively; (b) heavy chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 48, 56 and 68, respectively; and light chain CDR1, CDR2 and CDR3 set forth in SEQ ID NOs: 70, 79 and 90, respectively; (c) heavy chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 48, 56 and 68, respectively; and light chain CDR1, CDR2 and CDR3 set forth in SEQ ID NOs: 71, 80 and 91, respectively; (d) heavy chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 48, 56 and 68, respectively; and light chain CDR1, CDR2 and CDR3 set forth in SEQ ID NOs: 72, 81 and 92, respectively; (e) heavy chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 48, 56 and 68, respectively; and light chain CDR1, CDR2 and CDR3 set forth in SEQ ID NOs: 73, 82 and 91, respectively; (f) heavy chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 48, 56 and 68, respectively; and light chain CDR1, CDR2 and CDR3 set forth in SEQ ID NOs: 74, 83 and 93, respectively; (g) heavy chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 48, 56 and 68, respectively; and light chain CDR1, CDR2 and CDR3 set forth in SEQ ID NOs: 75, 84 and 91, respectively; (h) heavy chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 48, 56 and 68, respectively; and light chain CDR1, CDR2 and CDR3 set forth in SEQ ID NOs: 74, 85 and 94, respectively; (i) heavy chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 48, 56 and 68, respectively; and light chain CDR1, CDR2 and CDR3 set forth in SEQ ID NOs: 76, 86 and 95, respectively; (j) heavy chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 48, 56 and 68, respectively; and light chain CDR1, CDR2 and CDR3 set forth in SEQ ID NOs: 77, 87 and 93, respectively; (k) heavy chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 48, 56 and 68, respectively; and light chain CDR1, CDR2 and CDR3 set forth in SEQ ID NOs: 69, 88 and 90, respectively; (l) heavy chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 49, 57 and 68, respectively; and light chain CDR1, CDR2 and CDR3 set forth in SEQ ID NOs: 69, 78 and 89, respectively; (m) heavy chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 49, 58 and 68, respectively; and light chain CDR1, CDR2 and CDR3 set forth in SEQ ID NOs: 69, 78 and 89, respectively; (n) heavy chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 49, 59 and 68, respectively; and light chain CDR1, CDR2 and CDR3 set forth in SEQ ID NOs: 69, 78 and 89, respectively; (o) heavy chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 49, 60 and 68, respectively; and light chain CDR1, CDR2 and CDR3 set forth in SEQ ID NOs: 69, 78 and 89, respectively; (p) heavy chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 50, 61 and 68, respectively; and light chain CDR1, CDR2 and CDR3 set forth in SEQ ID NOs: 69, 78 and 89, respectively; (q) heavy chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 50, 58 and 68, respectively; and light chain CDR1, CDR2 and CDR3 set forth in SEQ ID NOs: 69, 78 and 89, respectively; (r) heavy chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 51, 62 and 68, respectively; and light chain CDR1, CDR2 and CDR3 set forth in SEQ ID NOs: 69, 78 and 89, respectively; (s) heavy chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 52, 63 and 68, respectively; and light chain CDR1, CDR2 and CDR3 set forth in SEQ ID NOs: 69, 78 and 89, respectively; (t) heavy chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 50, 64 and 68, respectively; and light chain CDR1, CDR2 and CDR3 set forth in SEQ ID NOs: 69, 78 and 89, respectively; (u) heavy chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 50, 65 and 68, respectively; and light chain CDR1, CDR2 and CDR3 set forth in SEQ ID NOs: 69, 78 and 89, respectively; (v) heavy chain CDR1, CDR2, and CDR3 sequences set forth in SEQ ID NOs: 51, 108, and 68, respectively, and light chain CDR1, CDR2, and CDR3 sequences set forth in SEQ ID NOs: 69, 78, and 89, respectively; (w) heavy chain CDR1, CDR2, and CDR3 sequences set forth in SEQ ID NOs: 107, 56, and 68, respectively, and light chain CDR1, CDR2, and CDR3 sequences set forth in SEQ ID NOs: 69, 78, and 89, respectively; and (x) heavy chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 48, 56 and 68, respectively; and light chain CDR1, CDR2 and CDR3 set forth in SEQ ID NOs: 109, 110 and 92, respectively.

[0052] In another aspect, the present disclosure provides an isolated monoclonal antibody that specifically binds to human CD137. or an antigen-binding portion thereof, wherein the antibody or antigen-binding portion thereof comprises a heavy chain variable region and a light chain variable region, wherein the heavy chain variable region comprises an amino acid sequence selected from the group consisting of SEQ ID NOs: 4, 8, 10, 12, 14, 16, 18, 20, 22, 24, 26, 101, and 103, and the light chain variable region comprises an amino acid sequence selected from the group consisting of SEQ ID NOs: 6, 28, 30, 32, 34, 36, 38, 40, 42, 44, 46, and 105.

[0053] In another aspect, the present disclosure provides an isolated monoclonal antibody that specifically binds to human CD137. or an antigen-binding portion thereof, wherein the antibody or antigen-binding portion thereof comprises a heavy chain variable region and a light chain variable region encoded by a nucleotide sequence selected from the group consisting of: (a) SEQ ID NOs: 5 and 7, respectively; and (b) SEQ ID NOs: 102 and 7, respectively.

[0054] In another aspect, the present disclosure provides an isolated monoclonal antibody that specifically binds to human CD137. or an antigen-binding portion thereof, wherein the antibody or antigen-binding portion thereof comprises a heavy chain variable region and a light chain variable region encoded by a nucleotide sequence selected from the group consisting of: (a) SEQ ID NOs: 5 and 7, respectively; (b) SEQ ID NOs: 5 and 29, respectively; (c) SEQ ID NOs: 5 and 31, respectively; (d) SEQ ID NOs: 5 and 33, respectively; (e) SEQ ID NOs: 5 and 35, respectively; (f) SEQ ID NOs: 5 and 37, respectively; (g) SEQ ID NOs: 5 and 39, respectively; (h) SEQ ID NOs: 5 and 41, respectively; (i) SEQ ID NOs: 5 and 43, respectively; (j) SEQ ID NOs: 5 and 45, respectively; (k) SEQ ID NOs: 5 and 47, respectively; (l) SEQ ID NOs: 9 and 7, respectively; (m) SEQ ID NOs: 11 and 7, respectively; (n) SEQ ID NOs: 13 and 7, respectively; (o) SEQ ID NOs: 15 and 7, respectively; (p) SEQ ID NOs: 17 and 7, respectively; (q) SEQ ID NOs: 19 and 7, respectively; (r) SEQ ID NOs: 21 and 7, respectively; (s) SEQ ID NOs: 23 and 7, respectively; (t) SEQ ID NOs: 25 and 7, respectively; (u) SEQ ID NOs: 27 and 7, respectively; (v) SEQ ID NOs: 102 and 7, respectively; (w) SEQ ID NOs: 104 and 7, respectively; and (x) SEQ ID NOs: 5 and 106, respectively.

[0055] In yet another aspect, the present disclosure provides an isolated monoclonal antibody that specifically binds to human CD137. The present invention provides a human antibody or antigen-binding portion thereof, wherein the antibody or antigen-binding portion thereof comprises a heavy chain CDR and a light chain CDR, and the heavy chain CDR3 comprises the amino acid sequence set forth in SEQ ID NO:68.

[0056] In another aspect, the present disclosure provides an isolated monoclonal antibody that specifically binds to human CD137. or an antigen-binding portion thereof, wherein the antibody or antigen-binding portion thereof comprises a heavy chain CDR and a light chain CDR, and the heavy chain CDR3 comprises the amino acid sequence DXXXXLXXXXYXYYX (SEQ ID NO: 126), where X is any amino acid. In some embodiments, X is any amino acid except alanine.

[0057] In some embodiments, the present disclosure provides an isolated monoclonal antibody that specifically binds to human CD137. In some embodiments, the antibody or antigen-binding portion thereof is provided, wherein the antibody or antigen-binding portion thereof comprises a heavy chain CDR and a light chain CDR, and the heavy chain CDR3 comprises the amino acid sequence DXPFXLDXXYYYYYX (SEQ ID NO: 127), where X is any amino acid. In some embodiments, X is any amino acid except alanine.

[0058] In yet another aspect, the present disclosure provides an isolated monoclonal antibody that specifically binds to human CD137. The present invention provides a human antibody or antigen-binding portion thereof, the antibody or antigen-binding portion thereof comprising a heavy chain CDR and a light chain CDR, wherein the heavy chain CDR3 is a CDR having the amino acid sequence DXXXXLXXXXYXYYX (SEQ ID NO: 1). No. 126), wherein X is any amino acid, and mutation of residues D95, L100, Y100E, Y100G, Y100H, or a combination thereof, results in loss of binding to human CD137. .

[0059] In another aspect, the present disclosure provides an isolated monoclonal antibody that specifically binds to human CD137. or an antigen-binding portion thereof, wherein the antibody or antigen-binding portion thereof comprises a heavy chain CDR and a light chain CDR, wherein the heavy chain CDR3 comprises the amino acid sequence DXPFXLDXXYYYYYX (SEQ ID NO: 127), where X is any amino acid, and mutation of residues P97, F98, D100A, Y100D, Y100F, or combinations thereof, to alanine results in reduced binding to human CD137. Can.

[0060] In some embodiments, the present disclosure provides an isolated monoclonal antibody that specifically binds to human CD137. The present invention provides an antibody or antigen-binding portion thereof, wherein the antibody or antigen-binding portion thereof comprises a heavy chain CDR and a light chain CDR, wherein the heavy chain CDR3 comprises the amino acid sequence DXPFXLDXXYYYYYX (SEQ ID NO: 127), where X is any amino acid, and mutation of residues P97, F98, D100A, Y100D, Y100F, or a combination thereof, to any residue other than alanine provides an antibody or antigen-binding portion thereof against human CD137. This results in increased binding.

[0061] In yet another aspect, the present disclosure provides an isolated monoclonal antibody that specifically binds to human CD137. The present invention provides a novel antibody or antigen-binding portion thereof, comprising a heavy chain CDR and a light chain CDR, wherein the heavy chain CDR3 comprises the amino acid sequence DX1X2X3X4LX5X6X7X8YX9YYX10 (SEQ ID NO: 128), where X1 is any amino acid and X2 is a non-polar amino acid. In some embodiments, X2 is a proline, X3 is phenylalanine or tryptophan, and X5 is aspartic acid or glutamic acid, wherein X3 is a nonpolar amino acid, X4 is any amino acid, X5 is a polar amino acid, X6 is any amino acid, X7 is any amino acid, X8 is a polar amino acid, X9 is a polar amino acid, and X10 is any amino acid. glutamic acid, wherein X8 is tyrosine, and wherein X9 is tyrosine.

[0062] In some embodiments, the present disclosure provides an isolated monoclonal antibody that specifically binds to human CD137. The present invention provides an antibody or antigen-binding portion thereof, wherein the antibody or antigen-binding portion thereof comprises a heavy chain variable region and a light chain variable region comprising an amino acid sequence selected from the group consisting of: (a) SEQ ID NOs: 4 and 6, respectively; (b) SEQ ID NOs: 4 and 28, respectively; (c) SEQ ID NOs: 4 and 30, respectively; (d) SEQ ID NOs: 4 and 32, respectively; (e) SEQ ID NOs: 4 and 34, respectively; (f) SEQ ID NOs: 4 and 36, respectively; (g) SEQ ID NOs: 4 and 38, respectively; (h) SEQ ID NOs: 4 and 40, respectively; (i) SEQ ID NOs: 4 and 42, respectively; (j) SEQ ID NOs: 4 and 44, respectively; (k) SEQ ID NOs: 4 and 46, respectively; (l) SEQ ID NOs: 8 and 6, respectively; (m) SEQ ID NOs: 10 and 6, respectively; (n) SEQ ID NOs: 12 and 6, respectively; (o) SEQ ID NOs: 14 and 6, respectively; (p) SEQ ID NOs: 16 and 6, respectively; (q) SEQ ID NOs: 18 and 6, respectively; (r) SEQ ID NOs: 20 and 6, respectively; (s) SEQ ID NOs: 22 and 6, respectively; (t) SEQ ID NOs: 24 and 6, respectively; (u) SEQ ID NOs: 26 and 6, respectively; (v) SEQ ID NOs: 101 and 6, respectively; (w) SEQ ID NOs: 103 and 6, respectively; and (x) SEQ ID NOs: 4 and 105, respectively.

[0063] In another aspect, the present disclosure provides an isolated monoclonal antibody that specifically binds to human CD137. or an antigen-binding portion thereof, wherein the antibody or antigen-binding portion thereof comprises a heavy chain variable region and a light chain variable region, wherein the heavy chain variable region comprises an amino acid sequence at least 90% identical to an amino acid sequence selected from the group consisting of SEQ ID NOs: 4, 8, 10, 12, 14, 16, 18, 20, 22, 24, 26, 101, and 103, and the light chain variable region comprises an amino acid sequence at least 90% identical to an amino acid sequence selected from the group consisting of SEQ ID NOs: 4, 8, 10, 12, 14, 16, 18, 20, 22, 24, 26, 101, and 103, The present invention also includes an amino acid sequence that is at least 90% identical to an amino acid sequence selected from the group consisting of amino acid sequences of Nos. 6, 28, 30, 32, 34, 36, 38, 40, 42, 44, 46 and 105.

[0064] In some embodiments, the present disclosure provides an isolated monoclonal antibody that specifically binds to human CD137. and (c) providing an antibody or antigen-binding portion thereof, wherein the antibody or antigen-binding portion thereof is an amino acid sequence that is at least 90% identical to an amino acid sequence selected from the group consisting of: The heavy chain variable region and light chain variable region comprise the sequences: (a) SEQ ID NOs: 4 and 6, respectively; (b) SEQ ID NOs: 4 and 28, respectively; (c) SEQ ID NOs: 4 and 30, respectively; (d) SEQ ID NOs: 4 and 32, respectively; (e) SEQ ID NOs: 4 and 34, respectively; (f) SEQ ID NOs: 4 and 36, respectively; (g) SEQ ID NOs: 4 and 38, respectively; (h) SEQ ID NOs: 4 and 40, respectively; (i) SEQ ID NOs: 4 and 42, respectively; (j) SEQ ID NOs: 4 and 44, respectively; (k) SEQ ID NOs: 4 and 46, respectively; (l) SEQ ID NOs: 8 and 6, respectively; (m) SEQ ID NOs: 10 and 6, respectively; (n) SEQ ID NOs: 12 and 6, respectively; (o) SEQ ID NOs: 14 and 6, respectively; (p) SEQ ID NOs: 16 and 6, respectively; (q) SEQ ID NOs: 18 and 6, respectively; (r) SEQ ID NOs: 20 and 6, respectively; (s) SEQ ID NOs: 22 and 6, respectively; (t) SEQ ID NOs: 24 and 6, respectively; (u) SEQ ID NOs: 26 and 6, respectively; (v) SEQ ID NOs: 101 and 6, respectively; (w) SEQ ID NOs: 103 and 6, respectively; and (x) SEQ ID NOs: 4 and 105, respectively.

[0065] In some embodiments, the present disclosure provides an isolated monoclonal antibody that specifically binds to human CD137. The present invention provides an antibody or antigen-binding portion thereof, wherein the antibody or antigen-binding portion thereof comprises a heavy chain sequence and a light chain sequence comprising an amino acid sequence selected from the group consisting of: (a) SEQ ID NOs: 129 and 133, respectively; and (b) SEQ ID NOs: 131 and 133, respectively.

[0066] In some embodiments, the present disclosure provides an isolated monoclonal antibody that specifically binds to human CD137. The present invention provides an antibody or antigen-binding portion thereof, wherein the antibody or antigen-binding portion thereof comprises heavy and light chain sequences having the amino acid sequences set forth in SEQ ID NOs: 129 and 133, respectively.

[0067] In some embodiments, the present disclosure provides an isolated monoclonal antibody that specifically binds to human CD137. The present invention provides an antibody or antigen-binding portion thereof, wherein the antibody or antigen-binding portion thereof comprises heavy and light chain sequences having the amino acid sequences set forth in SEQ ID NOs: 131 and 133, respectively.

[0068] In any of the preceding embodiments, the antibody or antigen-binding portion thereof is specific for human CD137. binds to and agonizes

[0069] In any of the foregoing aspects, the isolated monoclonal antibody or antigen-binding portion thereof exhibits at least one or more of the following properties selected from the group consisting of: (a) Induction or enhancement of CD137 trimer dimerization; (b) induction or enhancement of multimerization of CD137 trimers; (c) induction or enhancement of T cell activation; (d) induction or enhancement of cytotoxic T cell responses; (e) induction or enhancement of T-cell proliferation; (f) induction or enhancement of cytokine production; and (g) Any combination of characteristics (a) through (f).

[0070] In any of the foregoing embodiments, the isolated monoclonal antibody, or antigen-binding portion thereof, exhibits, relative to a reference antibody that binds to human CD137, one of the following characteristics selected from the group consisting of: exhibits at least one or more of the following: (a) does not induce or enhance intrahepatic T cell activation; (b) does not induce or enhance intrahepatic T-cell proliferation; (c) does not induce or enhance intrasplenic T cell activation; (d) does not induce or enhance intrasplenic T-cell proliferation; (e) does not induce or enhance macrophage activation; (f) does not induce or enhance macrophage differentiation; (g) does not induce or enhance alanine aminotransferase (ALT) activity; and (h) Any combination of properties (a)-(g). In some embodiments, the reference antibody is urelumab.

[0071] In any of the foregoing embodiments, the isolated monoclonal antibody, or antigen-binding portion thereof, induces or enhances human CD137-mediated T cell activation in the tumor microenvironment, but does not induce or enhance human CD137-mediated T cell activation to a significant extent in the spleen and / or liver.

[0072] In any of the foregoing embodiments, the isolated monoclonal antibody, or antigen-binding portion thereof, induces or enhances T cell activation in the tumor microenvironment, but not in the spleen and / or liver. does not significantly induce or enhance T cell activation.

[0073] In any of the foregoing embodiments, the isolated monoclonal antibody, or antigen-binding portion thereof, induces or enhances a human CD137-mediated cytotoxic T cell response in the tumor microenvironment, but does not significantly induce or enhance a human CD137-mediated cytotoxic T cell response in the spleen and / or liver.

[0074] In any of the foregoing embodiments, the isolated monoclonal antibody or antigen-binding portion thereof induces or enhances a cytotoxic T cell response in the tumor microenvironment, but not in the spleen and / or does not significantly induce or enhance cytotoxic T cell responses in the liver.

[0075] In any of the foregoing embodiments, the isolated monoclonal antibody, or antigen-binding portion thereof, induces human CD137-mediated T cell proliferation in the tumor microenvironment, but does not induce significant human CD137-mediated T cell proliferation in the spleen and / or liver.

[0076] In any of the foregoing embodiments, the isolated monoclonal antibody, or antigen-binding portion thereof, induces T cell proliferation in the tumor microenvironment, but to a lesser extent in the spleen and / or liver.

[0077] In any of the foregoing embodiments, the isolated monoclonal antibody, or antigen-binding portion thereof, induces human CD137-mediated T cell infiltration in the tumor microenvironment, but does not induce significant human CD137-mediated T cell infiltration in the spleen and / or liver.

[0078] In any of the foregoing embodiments, the isolated monoclonal antibody, or antigen-binding portion thereof, induces T cell infiltration in the tumor microenvironment, but not to a significant extent in the spleen and / or liver.

[0079] In any of the foregoing embodiments, the isolated monoclonal antibody or antigen-binding fragment thereof induces or enhances human CD137-mediated cytokine production in the tumor microenvironment but not in the spleen. Human CD137-mediated cytokine production was not significantly induced in the liver and / or kidney. Do not strengthen.

[0080] In any of the foregoing embodiments, the properties of the antibodies or antigen-binding portions thereof described herein are not dependent on Fc gamma receptor binding. In some embodiments, the properties of the antibodies or antigen-binding portions thereof described herein are enhanced by Fc gamma receptor binding.

[0081] In any of the foregoing embodiments, the isolated monoclonal antibody, or antigen-binding portion thereof, cross-competes with mAb1 (i.e., an antibody comprising the heavy chain variable and light chain variable sequences of SEQ ID NOs: 4 and 6, respectively). In some embodiments, the isolated monoclonal antibody, or antigen-binding portion thereof, cross-competes with mAb1 (i.e., an antibody comprising the heavy chain variable and light chain variable sequences of SEQ ID NOs: 4 and 6, respectively), mab8 (i.e., an antibody comprising the heavy chain variable and light chain variable sequences of SEQ ID NOs: 101 and 6, respectively), or mAb10 (i.e., an antibody comprising the heavy chain variable and light chain variable sequences of SEQ ID NOs: 26 and 6, respectively). In some embodiments, the isolated monoclonal antibody, or antigen-binding portion thereof, cross-competes with mAb8 (i.e., an antibody comprising the heavy chain variable and light chain variable sequences of SEQ ID NOs: 101 and 6, respectively). and 6) an antibody comprising the heavy chain variable sequence and the light chain variable sequence of In this regard, the isolated monoclonal antibody or antigen-binding portion thereof is designated mAb10 (i.e., each sequence It cross-competes with antibodies containing the heavy and light chain variable sequences of sequences 26 and 6.

[0082] In any of the foregoing embodiments, the isolated monoclonal antibody, or antigen-binding portion thereof, comprises at least a functional characteristic of mAb1 (i.e., an antibody comprising the heavy and light chain variable sequences of SEQ ID NOs: 4 and 6, respectively). In some embodiments, the isolated monoclonal antibody, or antigen-binding portion thereof, comprises at least a functional characteristic of mAb1 (i.e., an antibody comprising the heavy and light chain variable sequences of SEQ ID NOs: 4 and 6, respectively), mab8 (i.e., an antibody comprising the heavy and light chain variable sequences of SEQ ID NOs: 101 and 6, respectively), or mAb10 (i.e., an antibody comprising the heavy and light chain variable sequences of SEQ ID NOs: 26 and 27, respectively). and 6 heavy chain variable sequences and light chain variable sequences) In some embodiments, the isolated monoclonal antibody, or antigen-binding portion thereof, comprises at least a functional characteristic of mab8 (i.e., an antibody comprising the heavy and light chain variable sequences of SEQ ID NOs: 101 and 6, respectively). In some embodiments, the isolated monoclonal antibody, or antigen-binding portion thereof, comprises at least a functional characteristic of mAb10 (i.e., an antibody comprising the heavy and light chain variable sequences of SEQ ID NOs: 26 and 6, respectively).

[0083] In any of the foregoing embodiments, the isolated monoclonal antibody, or antigen-binding portion thereof, has a K at least equivalent to that of mAb1 (i.e., an antibody comprising the heavy chain variable sequences and light chain variable sequences of SEQ ID NOs: 4 and 6, respectively). D In some embodiments, the isolated monoclonal antibody or antigen-binding portion thereof has a K value at least equivalent to that of mAb1 (i.e., an antibody comprising the heavy and light chain variable sequences of SEQ ID NOs: 4 and 6, respectively), mab8 (i.e., an antibody comprising the heavy and light chain variable sequences of SEQ ID NOs: 101 and 6, respectively), or mAb10 (i.e., an antibody comprising the heavy and light chain variable sequences of SEQ ID NOs: 26 and 6, respectively). D In some embodiments, the isolated monoclonal antibody or antigen-binding portion thereof has a K value at least equivalent to that of mab8 (i.e., an antibody comprising the heavy chain variable sequences and light chain variable sequences of SEQ ID NOs: 101 and 6, respectively). D In some embodiments, the isolated monoclonal antibody or antigen-binding portion thereof has a K value at least equivalent to that of mAb10 (i.e., an antibody comprising the heavy chain variable sequence and light chain variable sequence of SEQ ID NOs: 26 and 6, respectively). D It has a value.

[0084] In any of the foregoing embodiments, the isolated monoclonal antibody, or antigen-binding portion thereof, cross-reacts with cynomolgus monkey CD137 and / or mouse CD137.

[0085] In any of the foregoing embodiments, the isolated monoclonal antibody or antigen-binding portion thereof is selected from the group consisting of IgG1, IgG2, IgG3, IgG4, IgM, IgA1, IgA2, IgD, and IgE antibodies. In some embodiments, the isolated monoclonal antibody or antigen-binding portion thereof is an IgG1 antibody or an IgG4 antibody.

[0086] In any of the foregoing embodiments, the isolated monoclonal antibody comprises a heavy chain constant region of a wild-type IgG1 or a wild-type IgG4. In some embodiments, the isolated monoclonal antibody comprises a heavy chain constant region of a mutant IgG1. In some embodiments, the isolated monoclonal antibody comprises a heavy chain constant region of a mutant IgG4. In some embodiments, the heavy chain constant region of the mutant IgG4 comprises a substitution at Ser228. In some embodiments, the heavy chain constant region of the mutant IgG4 comprises a S228P substitution.

[0087] In any of the foregoing embodiments, the isolated monoclonal antibody, or antigen-binding portion thereof, binds to an epitope of CD137, and the amino acid residues comprising the epitope bound by the antibody are The amino acid residues comprising the paratope of the mAb1 antibody described herein are within 4 angstroms of each other. Located inside.

[0088] In any of the foregoing embodiments, the isolated monoclonal antibody, or antigen-binding portion thereof, binds to an epitope of CD137, and a mutation in the epitope bound by the antibody is and mAb1 antibodies.

[0089] In any of the foregoing embodiments, the isolated antibody or antigen-binding portion thereof is fully human or humanized (i.e., a fully human antibody or fully humanized antibody or antigen-binding portion thereof).

[0090] In some aspects, the present disclosure provides a pharmaceutical composition comprising an isolated monoclonal antibody or antigen-binding portion thereof described herein and a pharmaceutically acceptable carrier.

[0091] In other aspects, the present disclosure provides a nucleic acid comprising a nucleotide sequence encoding the light chain, the heavy chain, or both the light and heavy chains of an isolated monoclonal antibody, or antigen-binding portion thereof, described herein. In some embodiments, the nucleic acid comprises SEQ ID NOs: 5 and 7. In some embodiments, the nucleic acid comprises SEQ ID NOs: 102 and 7. In some embodiments, the present disclosure provides an expression vector comprising the nucleic acid described herein. In other aspects, the present disclosure provides a cell transformed with the expression vector described herein.

[0092] In another aspect, the present disclosure provides an isolated monoclonal antibody that specifically binds to human CD137. In some embodiments, the present invention provides a method for producing a monoclonal antibody or an antigen-binding portion thereof, the method comprising maintaining a cell described herein under conditions that allow for expression of the monoclonal antibody or antigen-binding portion thereof. The method of producing a clonal antibody or antigen-binding portion thereof further comprises obtaining the monoclonal antibody or antigen-binding portion thereof.

[0093] In yet another aspect, the present disclosure provides a method for inducing dimerization of human CD137 trimers in a subject. The present invention provides a method for inducing or enhancing a tumor suppressor gene, the method comprising administering to a subject in need thereof an effective amount of an isolated monoclonal antibody or antigen-binding portion thereof described herein, or a pharmaceutical composition described herein.

[0094] In another aspect, the present disclosure provides a method for inducing or administering to a subject multimerization of human CD137 trimers. provides a method of enhancing a tumor suppressor comprising administering to a subject in need thereof an effective amount of an isolated monoclonal antibody or antigen-binding portion thereof described herein, or a pharmaceutical composition described herein.

[0095] In other aspects, the present disclosure provides methods of inducing or enhancing human CD137-mediated T cell activation in a subject, the method comprising administering to a subject in need thereof an effective amount of an isolated monoclonal antibody or antigen-binding portion thereof described herein, or a pharmaceutical composition described herein. Activation occurs in the tumor microenvironment. In other embodiments, T cell activation is measured in the spleen of a subject. and / or occurs less frequently in the liver.

[0096] In another aspect, the present disclosure provides a method of inducing or enhancing a cytotoxic T cell response mediated by human CD137 in a subject, the method comprising administering to a subject in need thereof an effective amount of an isolated monoclonal antibody or antigen-binding portion thereof described herein, or a pharmaceutical composition described herein. In some embodiments, the cytotoxic T cell response is generated in the tumor microenvironment. In other embodiments, the cytotoxic T cell response is not significantly generated in the spleen and / or liver of the subject.

[0097] In some embodiments, the present disclosure provides a method for the production of human CD137-mediated cytokines in a subject. In some embodiments, the cytokines produced are IL-2, TNFα, IL-13, IFNγ, or a combination thereof. In some embodiments, the cytokine produced is IL-2. In some embodiments, the cytokine produced is TNFα. In some embodiments, the cytokine produced is In some embodiments, the cytokine produced is IL-13. In some embodiments, the cytokine produced is IFNγ. In some embodiments, the cytokines produced are IL-2 and TNFα. In some embodiments, the cytokines produced are IL-2 and IL-13. In some embodiments, the cytokines produced are IL-2 and IFNγ. In some embodiments, the cytokines produced are TNFα and IL-13. In some embodiments, the cytokines produced are TNFα and IFNγ. In some embodiments, the cytokines produced are IL-13 and IFNγ. In some embodiments, the cytokines produced are IL-2, TNF In some embodiments, the cytokines produced are IL-2, TNFα, and IFNγ. In some embodiments, the cytokines produced are IFNγ, TNFα, and IL-13. and IL-13. In other embodiments, cytokine production occurs in the tumor microenvironment. In still other embodiments, cytokine production does not occur significantly in the spleen and / or liver of the subject.

[0098] In another aspect, the present disclosure provides a method of inducing or enhancing human CD137-mediated T cell proliferation in a subject, the method comprising administering to a subject in need thereof an effective amount of an isolated monoclonal antibody or antigen-binding portion thereof described herein, or a pharmaceutical composition described herein. In other embodiments, T cell proliferation is measured in the spleen and / or occurs less frequently in the liver.

[0099] In another aspect, the present disclosure provides a method of reducing or inhibiting tumor growth, the method comprising administering to a subject in need thereof an effective amount of an isolated monoclonal antibody or antigen-binding portion thereof described herein, or a pharmaceutical composition described herein.

[0100] In yet another aspect, the present disclosure provides a method for treating a disorder mediated by human CD137 in a subject. The present invention provides a method for treating rheumatoid arthritis, the method comprising administering to a subject in need thereof an effective amount of an isolated monoclonal antibody or antigen-binding portion thereof described herein, or a pharmaceutical composition described herein.

[0101] In some embodiments, the present disclosure provides a method of treating cancer in a subject, the method comprising administering to a subject in need thereof an effective amount of an isolated monoclonal antibody or antigen-binding portion thereof described herein, or a pharmaceutical composition described herein. In some embodiments, the cancer is selected from the group consisting of melanoma, glioma, renal cancer, breast cancer, hematological cancer, and head and neck cancer. In some embodiments, the hematological cancer is B-cell lymphoma. It is a pancreatic cancer.

[0102] In some aspects, the present disclosure provides a method of inducing an anti-tumor memory immune response, the method comprising administering to a subject in need thereof an effective amount of an isolated monoclonal antibody or antigen-binding portion thereof described herein, or a pharmaceutical composition described herein.

[0103] In any of the foregoing embodiments, after administration of the antibody or antigen-binding portion, infiltration of immune cells into the tumor microenvironment is increased. In some embodiments, the immune cells express CD45.

[0104] In any of the foregoing embodiments, the amount of T regulatory (Treg) cells is reduced in the tumor microenvironment following administration of the antibody or antigen-binding portion. In some embodiments, the Treg cells express CD4, FOXP-3, and CD24.

[0105] In any of the foregoing embodiments, the amount of macrophage cells is reduced in the tumor microenvironment after administration of the monoclonal antibody or antigen-binding portion. In some embodiments, the macrophages express CD45 and CD11b.

[0106] In any of the preceding embodiments, T cell depletion is low after administration of the antibody or antigen-binding portion. In some embodiments, the reduction in T cell exhaustion is due to the expression of TIGIT, PD-1, LAG-3, or their respective In some embodiments, the reduced T cell exhaustion comprises a reduction in the expression of a combination of TIGIT and PD-1.

[0107] In any of the foregoing embodiments, depletion of CD4+ T cells, CD8+ T cells, natural killer cells, or a combination thereof reduces the effectiveness of the antibody or antigen-binding portion thereof.

[0108] In another embodiment, the present disclosure provides a method for detecting the presence or absence of human CD137 in a biological sample. The present invention provides a method for producing a fluororesin comprising: (a) contacting a biological sample with an antibody or antigen-binding portion described herein; wherein the antibody or antigen-binding portion is labeled with a detectable substance; and (b) detecting the antibody or antigen-binding portion that binds to human CD137, thereby detecting the presence or absence of human CD137 in the biological sample.

[0109] In another aspect, the disclosure provides a kit comprising a container containing an antibody or antigen-binding portion described herein and optionally a pharmaceutically acceptable carrier, or a pharmaceutical composition described herein, and a package insert containing instructions for administering the antibody or pharmaceutical composition to treat or delay the progression of cancer, or to reduce or inhibit tumor growth in a subject in need thereof.

[0110] In another aspect, the disclosure provides a kit comprising a container containing an antibody or antigen-binding portion described herein and optionally a pharmaceutically acceptable carrier, or a pharmaceutical composition described herein, and a package insert containing instructions for administering the antibody or pharmaceutical composition, alone or in combination with another agent, to treat or delay the progression of cancer, or to reduce or inhibit tumor growth, in a subject in need thereof.

[0111] In another aspect, the present disclosure provides use of an isolated monoclonal antibody, or antigen-binding portion thereof, described herein for inducing or enhancing human CD137-mediated T cell activation in a subject. In another aspect, the present disclosure provides use of an isolated monoclonal antibody, or antigen-binding portion thereof, described herein for inducing or enhancing multimerization of human CD137 trimers in a subject. In another aspect, the present disclosure provides the use of an isolated monoclonal antibody or antigen-binding portion thereof described herein for inducing or enhancing a cytotoxic T cell response mediated by human CD137 in a subject. In another aspect, the present disclosure provides the use of an isolated monoclonal antibody or antigen-binding portion thereof described herein for inducing or enhancing cytokine production mediated by human CD137 in a subject. In another aspect, the present disclosure provides a use of the isolated monoclonal antibody, or antigen-binding portion thereof, described herein for inducing or enhancing human CD137-mediated T cell proliferation in a subject.

[0112] In another aspect, the present disclosure provides a use of an isolated monoclonal antibody or antigen-binding portion thereof described herein to reduce or inhibit tumor growth in a subject in need thereof. In another aspect, the present disclosure provides a use of an isolated monoclonal antibody or antigen-binding portion thereof described herein to treat a disorder mediated by human CD137 in a subject in need thereof. In another aspect, the present disclosure provides a use of an isolated monoclonal antibody, or antigen-binding portion thereof, described herein to treat cancer in a subject in need thereof.

[0113] In another aspect, the disclosure provides for the use of an isolated monoclonal antibody, or antigen-binding portion thereof, described herein for the manufacture of a medicament for treating or delaying the progression of cancer, or reducing or inhibiting tumor growth in a subject in need thereof. In another aspect, the disclosure provides for an isolated monoclonal antibody, or antigen-binding portion thereof, described herein in the manufacture of a medicament for treating or delaying the progression of cancer, or reducing or inhibiting tumor growth in a subject in need thereof. In another aspect, the disclosure provides for an isolated monoclonal antibody, or antigen-binding portion thereof, described herein for use as a medicament. The present invention provides, for example, the following items. (Item 1) An isolated monoclonal antibody or antigen-binding portion thereof that specifically binds to human CD137. The antibody or antigen-binding portion has an affinity (K D ) an isolated monoclonal antibody or antigen-binding portion thereof that binds to human CD137. (Item 2) 2. The isolated monoclonal antibody or antigen-binding portion of item 1, wherein the antibody or antigen-binding portion binds to an epitope on human CD137 comprising K114 of SEQ ID NO: 3. (Item 3) 3. The isolated monoclonal antibody or antigen-binding portion of item 2, wherein the epitope comprises residues E111, T113 and K114 of SEQ ID NO:3. (Item 4) the epitope comprises residues E111, T113, K114, N126 and I132 of SEQ ID NO: 3. Item 4. The isolated monoclonal antibody or antigen-binding portion thereof according to any one of Items 2 to 3. (Item 5) The epitope comprises residues E111, T113, K114, N126, I132 and P135 of SEQ ID NO:3. 5. The isolated monoclonal antibody or antigen-binding portion according to any one of items 2 to 4. (Item 6) The epitope comprises one of residues E111, T113, K114, N126, I132 and P135 of SEQ ID NO: 3. 2. The isolated monoclonal antibody or antigen-binding portion of item 1, comprising: (Item 7) 2. The isolated antibody of claim 1, wherein the antibody or antigen-binding portion binds to an epitope comprising a sequence of one or more amino acid residues corresponding to amino acids 111 to 135 of SEQ ID NO: 3. Monoclonal antibodies or antigen-binding portions. (Item 8) The epitope is selected from the group consisting of amino acids 2, 3, 4, 5, 6, and 7, which correspond to amino acids 111 to 135 of SEQ ID NO: 3. 8. The isolated monoclonal antibody or antigen-binding portion of item 7, comprising 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24 or 25 amino acid residues. (Item 9) The antibody or antigen-binding portion thereof is a polypeptide corresponding to amino acid residues 111 to 114 of ELTK (SEQ ID NO: 3). 2. The isolated monoclonal antibody according to item 1, which binds to an epitope comprising its antigen-binding portion. (Item 10) The epitope further comprises one or more of residues N126, I132 and P135 of SEQ ID NO:3. 10. The isolated monoclonal antibody or antigen-binding portion of item 9. (Item 11) 11. The isolated monoclonal antibody or antigen-binding portion of any one of items 2 to 10, wherein the epitope is a non-linear epitope. (Item 12) Mutation of residue K114 of SEQ ID NO: 3 abolishes binding of the antibody or antigen-binding portion thereof. 12. The isolated monoclonal antibody or antigen-binding portion according to any one of items 2 to 11. (Item 13) the antibody or antigen-binding portion has an affinity (K D Item 14. The isolated monoclonal antibody or antigen-binding portion according to any one of items 1 to 12, which binds to human CD137 at the nucleotide sequence (SEQ ID NO: 1). the antibody or antigen-binding portion thereof binds to a non-ligand binding region of the extracellular domain of human CD137. 14. The isolated monoclonal antibody or antigen-binding portion of any one of items 1 to 13, which binds to (Item 15) An isolated monoclonal antibody or antigen-binding portion thereof that specifically binds to human CD137. wherein the antibody or antigen-binding portion comprises: (i) Affinity (K) of approximately 30–100 nM D ) binds to human CD137; (ii) binds to a non-ligand binding region of the extracellular domain of human CD137; and (iii) an isolated monoclonal antibody that binds to an epitope on human CD137 that includes K114 of SEQ ID NO: 3; A cloned antibody or antigen-binding portion thereof. (Item 16) An isolated monoclonal antibody or antigen-binding portion thereof that specifically binds to human CD137. wherein the antibody or antigen-binding portion comprises: (i) Affinity (K) of approximately 30–100 nM D ) binds to human CD137; (ii) does not inhibit the interaction between human CD137 and human CD137 ligand; and (iii) an isolated monoclonal antibody that binds to an epitope on human CD137 that includes K114 of SEQ ID NO: 3; A cloned antibody or antigen-binding portion thereof. (Item 17) 16. The isolated monoclonal antibody or antigen-binding portion of any one of paragraphs 1 to 15, wherein the antibody or antigen-binding portion does not inhibit the interaction between CD137 and CD137L. (Item 18) The non-ligand binding region is located in cysteine-rich domain (CRD) III and CRD IV. 16. The isolated monoclonal antibody or its antigen-binding fragment thereof according to any one of items 14 to 15. portion. (Item 19) 19. The antibody or antigen-binding portion of any one of items 1 to 18, wherein the antibody or antigen-binding portion comprises a heavy chain CDR3 comprising the amino acid sequence DXXXXLXXXXYXYYX (SEQ ID NO: 126), wherein X is any amino acid. An isolated monoclonal antibody or antigen-binding portion thereof. (Item 20) 20. The isolated monoclonal antibody or antigen-binding portion of any one of items 1, 2, 3, and 11-19, wherein the epitope comprises E111, T113, K114, and P135 of SEQ ID NO:3. (Item 21) 20. The antibody or antigen-binding portion of any one of items 1 to 19, wherein the antibody or antigen-binding portion comprises a heavy chain CDR3 comprising the amino acid sequence DXPFXLDXXYYYYYX (SEQ ID NO: 127), wherein X is any amino acid. An isolated monoclonal antibody or antigen-binding portion. (Item 22) 22. The isolated monoclonal antibody or antigen-binding portion of any one of paragraphs 19 to 21, wherein mutation of residues D95, L100, Y100E, Y100G, Y100H, or combinations thereof, in the heavy chain CDR3 to alanine results in loss of binding to human CD137. (Item 23) Mutation of residues P97, F98, D100A, Y100D, Y100F, or their combinations to alanine 23. The isolated monoclonal antibody or antigen-binding portion of any one of items 19 to 22, which results in reduced binding to human CD137. (Item 24) the antibody or antigen-binding portion does not inhibit trimer formation of CD137:CD137L monomers; 24. The isolated monoclonal antibody or antigen-binding portion according to any one of items 1 to 23. (Item 25) The antibody or antigen-binding portion is located within amino acid residues 111 to 135 of SEQ ID NO:3. 25. The isolated monoclonal antibody or antigen-binding portion thereof according to any one of items 1, 13, 14, 17 to 19 and 21 to 24, which binds to an epitope (Item 26) 26. The isolated monoclonal antibody or antigen-binding portion of item 25, wherein the epitope is a non-linear epitope. (Item 27) 27. The isolated monoclonal antibody or antigen-binding portion of any one of items 1 to 26, wherein the antibody or antigen-binding portion comprises a heavy chain CDR and a light chain CDR, and the heavy chain CDR3 comprises the amino acid sequence set forth in SEQ ID NO: 68. (Item 28) the antibody or antigen-binding portion thereof, (a) heavy chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 48, 56 and 68, respectively; and the light chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 69, 78 and 89, respectively; and (b) The isolated monoclonal antibody or antigen-binding portion of any one of items 1 to 27, comprising heavy chain CDRs and light chain CDRs selected from the group consisting of heavy chain CDR1, CDR2, and CDR3 sequences set forth in SEQ ID NOs: 51, 108, and 68, respectively, and light chain CDR1, CDR2, and CDR3 sequences set forth in SEQ ID NOs: 69, 78, and 89, respectively. (Item 29) 28. The antibody or antigen-binding portion thereof according to any one of items 1 to 27, wherein the antibody or antigen-binding portion comprises a heavy chain variable region and a light chain variable region, the heavy chain variable region comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 4 and 101, and the light chain variable region comprising the amino acid sequence of SEQ ID NO: 6. An isolated monoclonal antibody or antigen-binding portion thereof. (Item 30) the antibody or antigen-binding portion thereof, (a) SEQ ID NOs: 4 and 6, respectively; and (b) a group comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 101 and 6, respectively; 28. The isolated monoclonal antibody or antigen-binding portion of any one of items 1 to 27, comprising a light chain variable region and a light chain variable region. (Item 31) 28. The antibody or antigen-binding portion thereof according to any one of items 1 to 27, wherein the antibody or antigen-binding portion comprises a heavy chain variable region and a light chain variable region, wherein the heavy chain variable region comprises an amino acid sequence that is at least 90% identical to an amino acid sequence selected from the group consisting of SEQ ID NOs: 4 and 101, and the light chain variable region comprises an amino acid sequence that is at least 90% identical to the amino acid sequence of SEQ ID NO: 6. 2. An isolated monoclonal antibody or antigen-binding portion thereof according to claim 1. (Item 32) the antibody or antigen-binding portion thereof, (a) SEQ ID NOs: 4 and 6, respectively; and (b) to an amino acid sequence selected from the group consisting of SEQ ID NOs: 101 and 6, respectively: comprising a heavy chain variable region and a light chain variable region comprising amino acid sequences that are at least 90% identical; 28. The isolated monoclonal antibody or antigen-binding portion according to any one of items 1 to 27. (Item 33) the antibody or antigen-binding portion thereof, (a) SEQ ID NOs: 129 and 133, respectively; and (b) The isolated monoclonal antibody or antigen-binding portion thereof according to any one of items 1 to 27, comprising a heavy chain and a light chain comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 131 and 133, respectively. (Item 34) An isolated monoclonal antibody or antigen-binding portion thereof that specifically binds to human CD137. wherein the antibody or antigen-binding portion thereof is (a) heavy chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 48, 56 and 68, respectively; and the light chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 69, 78 and 89, respectively; (b) heavy chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 48, 56 and 68, respectively; and the light chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 70, 79 and 90, respectively; (c) heavy chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 48, 56 and 68, respectively; and the light chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 71, 80 and 91, respectively; (d) heavy chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 48, 56 and 68, respectively; and the light chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 72, 81 and 92, respectively; (e) heavy chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 48, 56 and 68, respectively; and the light chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 73, 82 and 91, respectively; (f) heavy chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 48, 56 and 68, respectively; and the light chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 74, 83 and 93, respectively; (g) heavy chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 48, 56 and 68, respectively; and the light chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 75, 84 and 91, respectively; (h) heavy chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 48, 56 and 68, respectively; and the light chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 74, 85 and 94, respectively; (i) heavy chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 48, 56 and 68, respectively; and the light chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 76, 86 and 95, respectively; (j) heavy chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 48, 56 and 68, respectively; and the light chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 77, 87 and 93, respectively; (k) heavy chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 48, 56 and 68, respectively; and the light chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 69, 88 and 90, respectively; (l) heavy chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 49, 57 and 68, respectively; and the light chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 69, 78 and 89, respectively; (m) heavy chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 49, 58 and 68, respectively; and the light chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 69, 78 and 89, respectively; (n) heavy chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 49, 59 and 68, respectively; and the light chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 69, 78 and 89, respectively; (o) heavy chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 49, 60 and 68, respectively; and the light chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 69, 78 and 89, respectively; (p) heavy chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 50, 61 and 68, respectively; and the light chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 69, 78 and 89, respectively; (q) heavy chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 50, 58 and 68, respectively; and the light chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 69, 78 and 89, respectively; (r) heavy chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 51, 62 and 68, respectively; and the light chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 69, 78 and 89, respectively; (s) heavy chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 52, 63 and 68, respectively; and the light chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 69, 78 and 89, respectively; (t) heavy chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 50, 64 and 68, respectively; and the light chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 69, 78 and 89, respectively; (u) heavy chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 50, 65 and 68, respectively; and the light chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 69, 78 and 89, respectively; (v) heavy chain CDR1, CDR2, and CDR3 sequences set forth in SEQ ID NOs: 51, 108, and 68, respectively, and light chain CDR1, CDR2, and CDR3 sequences set forth in SEQ ID NOs: 69, 78, and 89, respectively; (w) heavy chain CDR1, CDR2, and CDR3 sequences set forth in SEQ ID NOs: 107, 56, and 68, respectively, and light chain CDR1, CDR2, and CDR3 sequences set forth in SEQ ID NOs: 69, 78, and 89, respectively; and (x) heavy chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 48, 56 and 68, respectively; and light chain CDR1, CDR2 and CDR3 set forth in SEQ ID NOs: 109, 110 and 92, respectively. (Item 35) An isolated monoclonal antibody or antigen-binding portion thereof that specifically binds to human CD137. wherein the antibody or antigen-binding portion thereof comprises a heavy chain variable region and a light chain variable region, wherein the heavy chain variable region comprises an amino acid sequence selected from the group consisting of SEQ ID NOs: 4, 8, 10, 12, 14, 16, 18, 20, 22, 24, 26, 101, and 103, and the light chain variable region comprises an amino acid sequence selected from the group consisting of SEQ ID NO: 6 , 28, 30, 32, 34, 36, 38, 40, 42, 44, 46 and 105 2. An isolated monoclonal antibody or antigen-binding portion thereof, comprising a nucleotide sequence. (Item 36) An isolated monoclonal antibody or antigen-binding portion thereof that specifically binds to human CD137. wherein the antibody or antigen-binding portion thereof comprises a heavy chain CDR and a light chain CDR, and the heavy chain CDR3 comprises the amino acid sequence set forth in SEQ ID NO: 68. (Item 37) An isolated monoclonal antibody or antigen-binding portion thereof that specifically binds to human CD137. wherein the antibody or antigen-binding portion thereof comprises a heavy chain CDR and a light chain CDR, wherein the heavy chain CDR3 comprises the amino acid sequence DXXXXLXXXXYXYYX (SEQ ID NO: 126), where X is any amino acid; An isolated monoclonal antibody or antigen-binding portion thereof. (Item 38) An isolated monoclonal antibody or antigen-binding portion thereof that specifically binds to human CD137. wherein the antibody or antigen-binding portion thereof comprises a heavy chain CDR and a light chain CDR, wherein the heavy chain CDR3 comprises the amino acid sequence DXPFXLDXXYYYYYX (SEQ ID NO: 128), where X is any amino acid; An isolated monoclonal antibody or antigen-binding portion thereof. (Item 39) the antibody or antigen-binding portion thereof, (a) SEQ ID NOs: 5 and 7, respectively; and (b) a nucleotide sequence selected from the group consisting of SEQ ID NOs: 102 and 7, respectively; 28. The isolated monoclonal antibody or antigen-binding portion of any one of items 1 to 27, comprising a heavy chain variable region and a light chain variable region encoded by the same. (Item 40) 40. The isolated monoclonal antibody or antigen-binding portion of any one of paragraphs 37 to 39, wherein mutation of residues D95, L100, Y100E, Y100G, Y100H, or combinations thereof, to alanine results in loss of binding to human CD137. (Item 41) Mutation of residues P97, F98, D100A, Y100D, Y100F, or their combinations to alanine 41. The isolated monoclonal antibody or antigen-binding portion of any one of paragraphs 37 to 40, which results in reduced binding to human CD137. (Item 42) the antibody or antigen-binding portion thereof, (a) SEQ ID NOs: 5 and 7, respectively; and (b) for a nucleotide sequence selected from the group consisting of SEQ ID NOs: 102 and 7, respectively and a heavy chain variable region encoded by a nucleotide sequence having at least 90% identity thereto. 28. The isolated monoclonal antibody or antigen-binding portion of any one of items 1 to 27, comprising a light chain variable region and a light chain variable region. (Item 43) 43. The isolated monoclonal antibody or antigen-binding portion of claim 42, wherein the antibody or antigen-binding portion comprises a heavy chain variable region and a light chain variable region encoded by a nucleotide sequence having at least 90% identity to SEQ ID NOs: 5 and 7, respectively. (Item 44) An isolated monoclonal antibody or antigen-binding portion thereof that specifically binds to human CD137. wherein the antibody or antigen-binding portion thereof is (a) SEQ ID NOs: 5 and 7, respectively; (b) SEQ ID NOs: 5 and 29, respectively; (c) SEQ ID NOs: 5 and 31, respectively; (d) SEQ ID NOs: 5 and 33, respectively; (e) SEQ ID NOs: 5 and 35, respectively; (f) SEQ ID NOs: 5 and 37, respectively; (g) SEQ ID NOs: 5 and 39, respectively; (h) SEQ ID NOs: 5 and 41, respectively; (i) SEQ ID NOs: 5 and 43, respectively; (j) SEQ ID NOs: 5 and 45, respectively; (k) SEQ ID NOs: 5 and 47, respectively; (l) SEQ ID NOs: 9 and 7, respectively; (m) SEQ ID NOs: 11 and 7, respectively; (n) SEQ ID NOs: 13 and 7, respectively; (o) SEQ ID NOs: 15 and 7, respectively; (p) SEQ ID NOs: 17 and 7, respectively; (q) SEQ ID NOs: 19 and 7, respectively; (r) SEQ ID NOs: 21 and 7, respectively; (s) SEQ ID NOs: 23 and 7, respectively; (t) SEQ ID NOs: 25 and 7, respectively; (u) SEQ ID NOs: 27 and 7, respectively; (v) SEQ ID NOs: 102 and 7, respectively; (w) SEQ ID NOs: 104 and 7, respectively; and (x) An isolated monoclonal antibody or antigen-binding portion thereof, comprising a heavy chain variable region and a light chain variable region encoded by a nucleotide sequence selected from the group consisting of SEQ ID NOs: 5 and 106, respectively. (Item 45) An isolated monoclonal antibody or antigen-binding portion thereof that specifically binds to human CD137. wherein the antibody or antigen-binding portion thereof is (a) SEQ ID NOs: 4 and 6, respectively; (b) SEQ ID NOs: 4 and 28, respectively; (c) SEQ ID NOs: 4 and 30, respectively; (d) SEQ ID NOs: 4 and 32, respectively; (e) SEQ ID NOs: 4 and 34, respectively; (f) SEQ ID NOs: 4 and 36, respectively; (g) SEQ ID NOs: 4 and 38, respectively; (h) SEQ ID NOs: 4 and 40, respectively; (i) SEQ ID NOs: 4 and 42, respectively; (j) SEQ ID NOs: 4 and 44, respectively; (k) SEQ ID NOs: 4 and 46, respectively; (l) SEQ ID NOs: 8 and 6, respectively; (m) SEQ ID NOs: 10 and 6, respectively; (n) SEQ ID NOs: 12 and 6, respectively; (o) SEQ ID NOs: 14 and 6, respectively; (p) SEQ ID NOs: 16 and 6, respectively; (q) SEQ ID NOs: 18 and 6, respectively; (r) SEQ ID NOs: 20 and 6, respectively; (s) SEQ ID NOs: 22 and 6, respectively; (t) SEQ ID NOs: 24 and 6, respectively; (u) SEQ ID NOs: 26 and 6, respectively; (v) SEQ ID NOs: 101 and 6, respectively; (w) SEQ ID NOs: 103 and 6, respectively; and (x) a group comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 4 and 105, An isolated monoclonal antibody or antigen-binding portion thereof, comprising a chain variable region and a light chain variable region. (Item 46) An isolated monoclonal antibody or antigen-binding portion thereof that specifically binds to human CD137. wherein the antibody or antigen-binding portion thereof comprises a heavy chain variable region and a light chain variable region, wherein the heavy chain variable region has an amino acid sequence that is at least 90% identical to an amino acid sequence selected from the group consisting of SEQ ID NOs: 4, 8, 10, 12, 14, 16, 18, 20, 22, 24, 26, 101, and 103. and the light chain variable region comprises SEQ ID NOs: 6, 28, 30, 32, 34, 36, 38, 40, 42, 44 1. An isolated monoclonal antibody or antigen-binding portion thereof, comprising an amino acid sequence that is at least 90% identical to an amino acid sequence selected from the group consisting of: 46 and 105. (Item 47) An isolated monoclonal antibody or antigen-binding portion thereof that specifically binds to human CD137. wherein the antibody or antigen-binding portion thereof is (a) SEQ ID NOs: 4 and 6, respectively; (b) SEQ ID NOs: 4 and 28, respectively; (c) SEQ ID NOs: 4 and 30, respectively; (d) SEQ ID NOs: 4 and 32, respectively; (e) SEQ ID NOs: 4 and 34, respectively; (f) SEQ ID NOs: 4 and 36, respectively; (g) SEQ ID NOs: 4 and 38, respectively; (h) SEQ ID NOs: 4 and 40, respectively; (i) SEQ ID NOs: 4 and 42, respectively; (j) SEQ ID NOs: 4 and 44, respectively; (k) SEQ ID NOs: 4 and 46, respectively; (l) SEQ ID NOs: 8 and 6, respectively; (m) SEQ ID NOs: 10 and 6, respectively; (n) SEQ ID NOs: 12 and 6, respectively; (o) SEQ ID NOs: 14 and 6, respectively; (p) SEQ ID NOs: 16 and 6, respectively; (q) SEQ ID NOs: 18 and 6, respectively; (r) SEQ ID NOs: 20 and 6, respectively; (s) SEQ ID NOs: 22 and 6, respectively; (t) SEQ ID NOs: 24 and 6, respectively; (u) SEQ ID NOs: 26 and 6, respectively; (v) SEQ ID NOs: 101 and 6, respectively; (w) SEQ ID NOs: 103 and 6, respectively; and (x) to an amino acid sequence selected from the group consisting of SEQ ID NOs: 4 and 105, respectively; comprising a heavy chain variable region and a light chain variable region comprising amino acid sequences that are at least 90% identical; An isolated monoclonal antibody or antigen-binding portion thereof. (Item 48) An isolated monoclonal antibody or antigen-binding portion thereof that specifically binds to human CD137. wherein the antibody or antigen-binding portion thereof is (a) SEQ ID NOs: 129 and 133, respectively; and (b) an isolated monoclonal antibody or antigen-binding portion thereof, comprising a heavy chain sequence and a light chain sequence comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 131 and 133, respectively. (Item 49) An isolated monoclonal antibody or antigen-binding portion thereof that specifically binds to human CD137. wherein the antibody or antigen-binding portion thereof comprises a heavy chain sequence and a light chain sequence having the amino acid sequences set forth in SEQ ID NOs: 129 and 133, respectively. (Item 50) An isolated monoclonal antibody or antigen-binding portion thereof that specifically binds to human CD137. wherein the antibody or antigen-binding portion thereof comprises a heavy chain sequence and a light chain sequence having the amino acid sequences set forth in SEQ ID NOs: 131 and 133, respectively. (Item 51) the antibody or antigen-binding portion specifically binds to and agonizes human CD137. 51. The isolated monoclonal antibody or antigen-binding portion according to any one of Items 1 to 50. (Item 52) the antibody or antigen-binding portion thereof, (a) Induction or enhancement of dimerization of CD137 trimers; (b) induction or enhancement of multimerization of CD137 trimers; (c) induction or enhancement of T cell activation; (d) induction or enhancement of cytotoxic T cell responses; (e) induction or enhancement of T cell proliferation; (f) induction or enhancement of immune cell cytokine production; and (g) any combination of characteristics (a) through (f). 52. The isolated monoclonal antibody or antigen-binding portion of any one of items 1 to 51, exhibiting at least one or more of: (Item 53) The antibody or antigen-binding portion exhibits, compared to a reference antibody that binds to human CD137: (a) does not induce or enhance intrahepatic T cell activation; (b) does not induce or enhance intrahepatic T-cell proliferation; (c) does not induce or enhance intrasplenic T cell activation; (d) does not induce or enhance intrasplenic T-cell proliferation; (e) does not induce or enhance macrophage activation; (f) does not induce or enhance macrophage differentiation; (g) does not induce or enhance alanine aminotransferase (ALT) activity; and Beauty (h) any combination of properties (a) to (g). Antibodies or antigen-binding portions. (Item 54) 54. The isolated monoclonal antibody or antigen-binding portion of claim 53, wherein the reference antibody is urelumab. (Item 55) 55. The isolated monoclonal antibody or antigen-binding portion of any one of paragraphs 52 to 54, wherein the properties of the antibody or antigen-binding portion are not dependent on Fc receptor binding. (Item 56) 55. The isolated monoclonal antibody or antigen-binding portion of any one of paragraphs 52 to 54, wherein the properties of the antibody or antigen-binding portion are enhanced by Fc receptor binding. (Item 57) 57. The isolated monoclonal antibody or antigen-binding portion of any one of paragraphs 1 to 56, wherein the antibody or antigen-binding portion cross-reacts with cynomolgus monkey CD137, mouse CD137, or both. (Item 58) The antibodies include IgG1, IgG2, IgG3, IgG4, IgM, IgA1, IgA2, IgD, and IgE antibodies. 58. The isolated monoclonal antibody or antigen-binding portion according to any one of items 1 to 47 and 51 to 57, selected from the group consisting of: (Item 59) 59. The isolated monoclonal antibody or antigen-binding portion of item 58, wherein the antibody is an IgG1 antibody or an IgG4 antibody. (Item 60) Items 1 to 47. The antibody comprises a heavy chain constant region of wild-type human IgG1 or wild-type human IgG4. 58. The isolated monoclonal antibody or antigen-binding portion of any one of items 51 to 57. (Item 61) 58. The isolated monoclonal antibody or antigen-binding portion according to any one of items 1 to 47 and 51 to 57, wherein the antibody comprises a mutated IgG1 heavy chain constant region. (Item 62) 58. The isolated monoclonal antibody or antigen-binding portion according to any one of items 1 to 47 and 51 to 57, wherein the antibody comprises a mutated IgG4 heavy chain constant region. (Item 63) 63. The isolated monoclonal antibody or antigen-binding portion thereof of item 62, wherein the mutated IgG4 heavy chain constant region comprises a substitution at Ser228. (Item 64) 64. The isolated monoclonal antibody of claim 63, wherein the mutated IgG4 heavy chain constant region comprises a S228P substitution. A cloned antibody or antigen-binding portion thereof. (Item 65) 65. A pharmaceutical composition comprising the isolated monoclonal antibody or antigen-binding portion thereof according to any one of items 1 to 64, and a pharmaceutically acceptable carrier. (Item 66) 65. A nucleic acid comprising a nucleotide sequence encoding the light chain, the heavy chain, or both the light and heavy chains of the isolated monoclonal antibody or antigen-binding portion of any one of items 1 to 64. (Item 67) 67. An expression vector comprising the nucleic acid of item 66. (Item 68) A cell transformed with the expression vector according to Item 67. (Item 69) Producing a monoclonal antibody or antigen-binding portion thereof that specifically binds to human CD137 70. A method comprising maintaining the cell of item 68 under conditions that allow expression of the monoclonal antibody or antigen-binding portion thereof. (Item 70) 70. The method of claim 69, further comprising obtaining the monoclonal antibody or antigen-binding portion thereof. (Item 71) A method for inducing or enhancing dimerization of human CD137 trimers in a subject, comprising: 66. A method comprising administering to a subject in need thereof an effective amount of the isolated monoclonal antibody or antigen-binding portion thereof according to any one of items 1 to 64, or the pharmaceutical composition according to item 65. (Item 72) A method for inducing or enhancing multimerization of human CD137 trimers in a subject, comprising: 66. A method comprising administering to a subject in need thereof an effective amount of the isolated monoclonal antibody or antigen-binding portion thereof according to any one of items 1 to 64, or the pharmaceutical composition according to item 65. (Item 73) A method of inducing or enhancing T cell activation in a subject in need thereof. 66. A method comprising administering to a subject a therapeutically effective amount of the isolated monoclonal antibody or antigen-binding portion thereof according to any one of items 1 to 64, or the pharmaceutical composition according to item 65, (Item 74) 74. The method of claim 73, wherein the T cell activation occurs in a tumor microenvironment. (Item 75) A method of inducing or enhancing a cytotoxic T cell response in a subject, comprising administering to said subject a therapeutically effective amount of a cytotoxic T cell response to said subject. 66. A method comprising administering to a subject an effective amount of the isolated monoclonal antibody or antigen-binding portion thereof according to any one of items 1 to 64, or the pharmaceutical composition according to item 65. (Item 76) 76. The method of claim 75, wherein the cytotoxic T cell response is generated in the tumor microenvironment. . (Item 77) 66. A method for inducing or enhancing immune cell cytokine production in a subject, comprising administering to a subject in need thereof an effective amount of the isolated monoclonal antibody or antigen-binding portion thereof according to any one of items 1 to 64, or the pharmaceutical composition according to item 65. (Item 78) The cytokines produced are IL-2, TNFα, IL-13, IFNγ, or a combination thereof. Item 78. The method according to Item 77. (Item 79) 79. The method of claim 77 or 78, wherein the cytokine production occurs in a tumor microenvironment. (Item 80) A method of inducing or enhancing T cell proliferation in a subject, comprising administering to a subject in need thereof 66. A method comprising administering an effective amount of the isolated monoclonal antibody or antigen-binding portion thereof according to any one of items 1 to 64, or the pharmaceutical composition according to item 65. (Item 81) 81. The method of claim 80, wherein the T cell proliferation occurs in a tumor microenvironment. (Item 82) 1. A method for reducing or inhibiting tumor growth, comprising administering to a subject in need thereof a compound according to any one of items 1 to 64. 66. A method comprising administering an effective amount of the isolated monoclonal antibody or antigen-binding portion of any one of claims 61 to 65, or the pharmaceutical composition of claim 65. (Item 83) A method of treating a disorder mediated by human CD137 in a subject, comprising administering to a subject in need thereof 66. A method comprising administering to a subject an effective amount of the isolated monoclonal antibody or antigen-binding portion thereof according to any one of items 1 to 64, or the pharmaceutical composition according to item 65. (Item 84) A method of treating cancer in a subject, comprising administering to a subject in need thereof any of the compounds of items 1 to 64. 65. The isolated monoclonal antibody or antigen-binding portion thereof according to any one of claims 1 to 4, or the isolated monoclonal antibody or antigen-binding portion thereof according to claim 4. The method comprises administering an effective amount of the pharmaceutical composition described above. (Item 85) 85. The method of any one of paragraphs 82-84, wherein administration of the isolated monoclonal antibody or antigen-binding portion increases infiltration of immune cells into the tumor microenvironment. (Item 86) 86. The method of claim 85, wherein the immune cells express CD45. (Item 87) and (b) administering the isolated monoclonal antibody or antigen-binding portion thereof to a subject in need thereof. 87. The method of any one of items 82 to 86, wherein the amount of thymic (Treg) cells is reduced. (Item 88) 88. The method of claim 87, wherein the Treg cells express CD4, FOXP-3, and CD25. (Item 89) 89. The method of any one of paragraphs 82-88, wherein the amount of macrophages in the tumor microenvironment is reduced after administration of the isolated monoclonal antibody or antigen-binding portion. (Item 90) 89. The method of claim 89, wherein the macrophages express CD45 and CD11b. (Item 91) T cells in the tumor microenvironment after administration of the isolated monoclonal antibody or antigen-binding portion. 91. The method according to any one of items 82 to 90, wherein wasting is reduced. (Item 92) Decreased T cell exhaustion includes decreased expression of TIGIT, PD-1, LAG-3, or a combination thereof. Item 91. (Item 93) 93. The method according to any one of items 84 to 92, wherein the cancer is selected from the group consisting of melanoma, glioma, renal cancer, breast cancer, blood cancer and head and neck cancer. (Item 94) 84. The method of claim 83, wherein the hematological cancer is B-cell lymphoma. (Item 95) A method for inducing an anti-tumor memory immune response, comprising administering to a subject in need thereof a compound according to any one of items 1 to 64. 66. The isolated monoclonal antibody or antigen-binding portion of any one of items 64 to 65. A method comprising administering an effective amount of the pharmaceutical composition described. (Item 96) 96. Any one of items 71 to 95, wherein the antibody or antigen-binding portion binds to an Fc gamma receptor. The method described in paragraph . (Item 97) 96. The antibody of any one of items 82 to 95, wherein depletion of CD4+ T cells, CD8+ T cells, natural killer cells, or a combination thereof reduces the efficacy of the antibody or antigen-binding portion thereof. How to post. (Item 98) 1. A method for detecting the presence or absence of human CD137 in a biological sample, comprising: (i) contacting a biological sample with the antibody or antigen-binding portion according to any one of items 1 to 64; contacting, wherein the antibody or antigen-binding portion is labeled with a detectable substance; and (ii) detecting said antibody or antigen-binding portion bound to human CD137, thereby detecting said antibody or antigen-binding portion bound to human CD137; Detecting the presence or absence of human CD137 in a biological sample. (Item 99) 66. A kit comprising a container containing the isolated monoclonal antibody or antigen-binding portion according to any one of items 1 to 64 and optionally a pharmaceutically acceptable carrier, or the pharmaceutical composition according to item 65, and a package insert containing instructions for administering the antibody or pharmaceutical composition for treating or delaying the progression of cancer, or for reducing or inhibiting tumor growth in a subject in need thereof. (Item 100) 66. A kit comprising a container containing the isolated monoclonal antibody or antigen-binding portion according to any one of items 1 to 64 and optionally a pharmaceutically acceptable carrier, or the pharmaceutical composition according to item 65, and a package insert containing instructions for administering the antibody or pharmaceutical composition alone, or in combination with another agent, for treating or delaying the progression of cancer, or for reducing or inhibiting tumor growth in a subject in need thereof. (Item 101) 65. Use of the isolated monoclonal antibody or antigen-binding portion of any one of paragraphs 1 to 64 for the manufacture of a medicament for treating or delaying the progression of cancer, or reducing or inhibiting tumor growth in a subject in need thereof. (Item 102) 65. The isolated monoclonal antibody or antigen-binding portion of any one of paragraphs 1 to 64 in the manufacture of a medicament for treating or delaying the progression of cancer, or reducing or inhibiting tumor growth in a subject in need thereof. (Item 103) 65. The isolated monoclonal antibody or antigen-binding portion of any one of items 1 to 64 for use as a medicament. [Brief explanation of the drawings]

[0114] [Figure 1] FIG. 1 presents a graph showing the binding affinity distribution for affinity matured clones of the parent anti-CD137 antibody mAb1. [Figure 2] Figure 2 presents a schematic showing the results of mAb1 CDRH3 alanine scanning as measured by binding affinity (KD) to human or mouse CD137. [Figure 3A] FIG. 3A shows the amino acid sequence of human CD137, with residues containing the epitopes bound by mAb1, mAb4, or mAb5 shown in bold. [Figure 3B] FIG. 3B is a graph showing kinetic binding data of mAb1 to the extracellular domain of mouse and rat CD137 as determined by surface plasmon resonance. [Figure 3C] Figure 3C: X-ray crystallography image of human CD137 bound to CD137L (shown in gray). Residues E111, T113, K114, and P135 are circled. [Figure 3D] Figure 3D. X-ray crystallography image of human CD137 bound to trimerized CD137L (shown in gray). Residues E111, T113, K114, and P135 are circled. [Figure 4-1] Figure 4A presents scatter plots of flow cytometry data showing an increase in the expression of TIGIT (top) or PD-1 (bottom) on CD44+ T cells in response to anti-CD137 antibody. [Figure 4-2]Figure 4B presents graphs showing quantification of CD8+CD44+ T cells expressing TIGIT (top) or PD-1 (bottom) in mouse spleens after treatment with anti-CD137 antibodies. Figure 4C presents graphs showing quantification of CD8+ T cells in mouse spleens after treatment with anti-CD137 antibodies, as a percentage of CD45+ cells (left) or as the number of cells per spleen (right). [Figure 5-1] FIG. 5A presents a graph showing individual CT26 tumor volumes in mice after treatment with the indicated doses of anti-CD137 antibody. [Figure 5-2] Figure 5B is a graph showing the mean tumor volume presented in Figure 5A. Figure 5C is a Kaplan-Meier graph showing overall survival of tumor-bearing mice after treatment with anti-CD137 antibody. Figure 5D is a graph showing tumor volume in mice re-challenged with tumorigenic CT26 cells. [Figure 6A] FIG. 6A presents a graph showing individual CT26 tumor volumes in mice after treatment with parental and affinity-matured anti-CD137 antibodies. [Figure 6B] FIG. 6B is a graph showing the mean tumor volumes presented in FIG. 6A. [Figure 7] FIG. 7 presents graphs showing the percentage of CD8+ or CD4+ T cells from splenic T cells (top) and tumor-infiltrating leukocytes (bottom) after treatment with anti-CD137 antibody at the indicated doses. [Figure 8] Figure 8 presents graphs showing individual tumor volumes when mice were treated with mAb1 with or without lymphodepleting antibodies. CD4+ T cells were depleted with GK1.5 (middle graph). CD8+ T cells were depleted with YTS169.4 (second graph from the right). NK cells were depleted with anti-asialo-GM1 antibody (far right graph). [Figure 9] Figure 9 presents a graph showing individual tumor volumes in mice bearing either CT26 tumors (colon cancer), EMT-6 tumors (breast cancer), A20 tumors (B-cell lymphoma), or MC38 tumors (colon cancer) and treated with mAb8 or an isotype control antibody. [Figure 10]10A-10C show the in vivo anti-tumor efficacy of anti-CD137 antibodies administered at 150 μg / mouse. Individual tumor volumes are shown in 10A. Mean tumor volumes are shown in 10B. Survival rates are shown in 10C. [Figure 11] 11A-11C show the in vivo anti-tumor efficacy of anti-CD137 antibodies administered at 20 μg / mouse. Individual tumor volumes are shown in 11A. Mean tumor volumes are shown in 11B. Survival rates are shown in 11C. [Figure 12] FIG. 12 presents a graph showing individual tumor volumes in mice bearing CT26 tumors and treated with various doses of mAb1 (i.e., 12.5, 25, 50, 100, or 200 μg) or isotype control. [Figure 13A] Figures 13A and 13B show the contribution of Fc binding to the anti-tumor efficacy of mAb1. Figure 13A shows mAb1 as an IgG4 isotype or an IgG4 aglycosylated isotype. The mean tumor volume is shown above and individual tumor volumes are shown below. Figure 13B shows mAb1 as an IgG4 isotype or an IgG1 aglycosylated isotype. The mean tumor volume is shown above and individual tumor volumes are shown below. [Figure 13B] Same as above. [Figure 14-1] 14A-14D show the in vivo anti-tumor efficacy of anti-CD137 antibodies in mice bearing large, established tumors (i.e., 500 mm3) before treatment. Individual tumor volumes are shown in 14A and 14D. Mean tumor volumes are shown in 14B. Survival fractions are shown in 14C. [Figure 14-2] Same as above. [Figure 15] Figure 15 presents Kaplan-Meier survival graphs showing protective anti-tumor immunity in mice previously treated and considered cured with mAb1, mAb8 or isotype control from Figures 14A-14C and re-challenged with CT26 cells in the opposite flank. [Figure 16A] FIG. 16A presents scatter plots of flow cytometry data showing CD45+ intrahepatic T cell expansion after treatment with anti-CD137 antibody at the indicated doses. [Figure 16B] FIG. 16B presents graphs showing quantification of intrahepatic CD8+ T cells (left) and CD4+ T cells (right) after treatment with anti-CD137 antibody at the indicated doses. [Figure 17A] FIG. 17A presents a graph showing the percentage of CD3+, CD4+, or CD8+ T cells from splenic T cells after treatment of mice with affinity-matured anti-CD137 antibodies. [Figure 17B] FIG. 17B presents a graph showing the percentage of CD3+, CD4+, or CD8+ T cells from liver T cells after treatment of mice with affinity-matured anti-CD137 antibodies. [Figure 18A] Figure 18A presents a graph showing the percentage of splenic CD8+CD44+ T cells expressing TIGIT, PD-1, or LAG3 after treatment of mice with affinity-matured anti-CD137 antibodies. [Figure 18B] Figure 18B presents a graph showing the percentage of liver CD8+CD44+ T cells expressing TIGIT, PD-1, or LAG3 after treatment of mice with affinity-matured anti-CD137 antibodies. [Figure 19A] Figure 19A presents a graph showing the percentage of splenic CD4+CD44+ T cells expressing TIGIT, PD-1, or LAG3 after treatment of mice with affinity-matured anti-CD137 antibodies. [Figure 19B] Figure 19B presents a graph showing the percentage of liver CD4+CD44+ T cells expressing TIGIT, PD-1, or LAG3 after treatment of mice with affinity-matured anti-CD137 antibodies. [Figure 20-1] Figures 20A-20C show graphs of in vivo toxicity indicators resulting from multiple administration of anti-CD137 antibodies mAb1, mAb8, or 3H3 at various doses. Figure 20A shows a graph showing the percentage of CD8+ T cells in the liver after administration of anti-CD137 antibodies. Figure 20B shows a graph showing alanine aminotransferase (ALT) activity in the plasma of mice administered anti-CD137 antibodies. Figure 20C shows a graph showing TNFα levels in the plasma of mice administered anti-CD137 antibodies. [Figure 20-2] Same as above. [Figure 21] Figure 21 presents representative images of hematoxylin and eosin (H&E) stained liver sections from mice treated with mAb1, mAb8, 3H3, or isotype control as described in Figures 20A-20C. Arrows indicate infiltrating immune cells. [Figure 22-1] Figures 22A-22D present representative FACS plots showing immune cell reprogramming in the tumor microenvironment. Mice bearing CT26 tumors were administered multiple doses of mAb8 or isotype control (days 0, 3, 6, and 9). Figure 22A shows overall immune cell infiltration based on CD45 expression. Figure 22B shows a reduction in Treg cells, as measured by FOXP-3 and CD25 expression. Figure 22C shows a reduction in T cell exhaustion, as measured by PD-1 and TIGIT expression. Figure 22D shows a reduction in tumor-associated macrophages, as measured by F4 / 80 and CD11b expression. [Figure 22-2] Same as above. [Figure 23] FIG. 23 shows immunophenotypic analysis of spleens from mice bearing CT26 tumors and treated with either the anti-CD137 antibodies mAb1 and 3H3 or an isotype control. [Figure 24] Figure 24 is a graph showing the concentration of IL-2 (pg / ml) produced by mouse T cells upon stimulation with the indicated anti-CD137 antibodies in an OVA stimulation assay. Atezolizumab (an anti-PD-L1 antibody) was used with mouse anti-PD-1 (RMP1-14) as a comparator. [Figure 25] Figures 25A and 25B are graphs showing the percentage of mouse CD8+ T cells expressing either CD25 (25A) or TIGIT (25B) upon stimulation with the indicated anti-CD137 antibodies in an OVA stimulation assay. Atezolizumab (an anti-PD-L1 antibody) was used along with mouse anti-PD-1 (RMP1-14) and mouse anti-CD137 (3H3) as comparators. [Figure 26]Figure 26 presents a bar graph showing quantification of cytokines (IL-2, TNFα, IL-13, and IFNγ) produced by CD3+ T cells after incubation with plate-bound anti-CD137 antibody. Cytokine levels are shown as fold increase over baseline activation with anti-CD3 antibody. [Figure 27-1] 27A-27C present graphs showing the dose response of IFNγ production in mixed lymphocyte responses after treatment with anti-CD137 antibodies. Anti-PD1 antibodies (Keytruda; Merck) were used as controls. [Figure 27-2] Same as above. [Figure 28] Figure 28 is a graph showing IFNγ production from human T cells co-cultured with CHO cells engineered to express CD32 (CHO-CD32 cells) in the presence of anti-CD137 antibodies mAb1, mAb8, mAb4, or mAb5, or an isotype control. [Figure 29] Figure 29 is a graph showing the proliferation of Treg cells co-cultured with CHO cells engineered to express CD32 (CHO-CD32 cells) in the presence or absence of anti-CD137 antibodies mAb1, mAb8, mAb4, or mAb5, or an isotype control. [Figure 30] FIG. 30 presents graphs showing NFκβ and SRF signaling in CCL-119 cells transduced with luciferase reporters for NFκβ or SRF in the presence of various concentrations of mAb1, mAB8, mAb4, or mAb5. [Figure 31] Figure 31 presents graphs showing the induction of IL-6, TNFα or IL-27 by bone marrow-derived murine macrophages stimulated with the TLR9 agonist CpG in the presence of anti-CD137 antibodies mAb1, 3H3 or LOB12.3 or an isotype control. [Figure 32]FIG. 32 presents a graph showing the induction of TNFα by human monocyte-derived murine macrophages stimulated with LPS in the presence of anti-CD137 antibodies mAb1, mAb4, or mAb5 or an isotype control. [Figure 33] Figure 33 presents a graph showing the effect of anti-CD137 antibodies on macrophage differentiation, as determined by CD64 expression, of THP1 monocytes cultured with PMA in the presence of anti-CD137 antibodies mAb1, mAb4, or mAb5 or an isotype control. [Figure 34] 34A-34C present graphs showing the percentage of hCD45+, hCD8+, or hCD4+ in human PBMCs and immunocompetent mice administered anti-CD137 antibodies mAb1, mAb4, or mAb5, or an isotype control. DETAILED DESCRIPTION OF THE INVENTION

[0115] Cancer therapy with an agonistic anti-CD137 antibody induces immune-mediated tumor rejection in mice This type of analogue has been shown to induce the formation of polyclonal antibodies in the liver, and this type of analogue is currently being tested in cancer patients. It has been shown that a significant accumulation of T lymphocyte infiltration can be induced (Dubrot et al., (2010) Cancer Immunology, Immunotherapy 59(8):1223-1233), which suggests that liver Possible causes of liver inflammation and drug-induced hepatotoxicity have been suggested. A recent report on clinical evaluation (Urelumab, BMS-663513; Bristol-Myers Squibb) showed severe hepatotoxicity (hypertransferase) in human subjects that correlated with antibody dose. Treatment-related adverse events, including signs of vasodilatory steroids (Segal et al., 2011), have been reported (Segal et al., 2011). et al., (2016) Clin Cancer Res 23(8):1929-1936).

[0116] The present disclosure provides an isolated monoclonal antibody, or antigen-binding portion thereof, that specifically binds to an epitope on human CD137 and agonizes human CD137. In some embodiments, the antibody or antigen-binding portion thereof competes with mAb1 for binding to the epitope on human CD137. In some embodiments, the anti-CD137 agonistic antibody of the present disclosure is an anti-mouse CD137 3H3 antibody. (Melero et al. (1997) Nature Medicine 3(6):682-685; Uno et al. (2006) Nature Medicine 12(6):693-696) and at least two anti-human CD137s in clinical development. Antibodies (BMS-663513 / Urelumab, Bristol-Meyers Squibb and PF-05082566 / Utomilumab) Compared with IFN-γ-α (Pfizer), cytokine production and CD8+ T cell expansion in the tumor microenvironment were significantly enhanced. and induce protective anti-tumor immunity in vivo while simultaneously reducing the likelihood of toxicity-related events.

[0117] definition Terms used in the claims and specification, unless otherwise specified, are defined as set forth below.

[0118] It should be noted that as used in the specification and the appended claims, the singular forms (a, an, and the) include plural referents unless the context clearly dictates otherwise. Further, unless the context requires otherwise, singular terms shall include plurals and plural terms shall include the singular.

[0119] As used herein, "about" will be understood by one of ordinary skill in the art and will vary to some extent depending on the context in which it is used. If there are uses of the term that are not clear to persons of ordinary skill in the art given the context in which it is used, "about" will mean up to plus or minus 10% of the particular value.

[0120] As used herein, the term "agonist" refers to an agent that partially or fully stimulates the biological activity of a native polypeptide (e.g., CD137) disclosed herein. Agonistic activity refers to any molecule that induces, elevates, and / or activates a target protein. Suitable agonistic molecules include, among others, agonistic antibodies or antibody fragments, fragments or amino acid sequence variants of natural polypeptides, peptides, antisense oligonucleotides, small molecules, etc. In some embodiments, activation in the presence of an agonist is observed in a dose-dependent manner. In some embodiments, the measured signal (e.g., biological activity) is at least about 5%, at least about 10%, at least about 15%, at least about 20%, at least about 25%, at least about 30%, or less than the signal measured using a negative control under comparable conditions. At least about 35%, at least about 40%, at least about 45%, at least about 50%, at least about 55%, at least about 60%, at least about 65%, at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, or at least about 100% higher. Also disclosed herein are methods for identifying agonists suitable for use in the disclosed methods. For example, these methods include enzyme-linked immunosorbent assay (ELISA), Forte Bio(c) systems and binding assays such as radioimmunoassays (RIAs) These assays include, but are not limited to, determining the ability of an agonist to bind to a polypeptide of interest (e.g., a receptor or ligand, e.g., CD137). A functional assay is an indication of the ability of an agonist to promote, increase, or activate the activity of a polypeptide. The efficacy of an agonist, such as the ability of an agonist to activate or promote the function of a polypeptide, can also be determined using a functional assay. For example, a functional assay may involve contacting a polypeptide with a candidate agonist molecule and measuring a detectable change in one or more biological activities normally associated with the polypeptide. The efficacy of an agonist is typically measured by its EC 50 value (the concentration required to activate 50% of the agonist response) It is defined by EC 50 The lower the value, the more potent the agonist and the lower the concentration required to activate the maximal biological response.

[0121] As used herein, the term "alanine scanning" refers to a technique used to determine the contribution of a particular wild-type residue to the stability or function (e.g., binding affinity, etc.) of a given protein or polypeptide. This technique involves substituting an alanine residue for the wild-type residue in a polypeptide, then evaluating the stability or function (e.g., binding affinity, etc.) of the alanine-substituted derivative or mutant polypeptide and comparing it with the wild-type polypeptide. Techniques for substituting alanine for wild-type residues in polypeptides are known in the art.

[0122] The term "ameliorate" refers to any therapeutically beneficial outcome, including prevention, attenuation of the severity or progression, remission, or cure, in the treatment of a disease state, such as cancer.

[0123] As used herein, the term "amino acid" refers to naturally occurring and synthetic amino acids, as well as amino acid analogs and amino acid mimetics that function in a manner similar to the naturally occurring amino acids. Naturally occurring amino acids are those encoded by the genetic code, as well as those amino acids that are later modified, such as hydroxyproline, γ-carboxyglutamate, and O-phosphoserine. Amino acid analogs have the same basic chemical structure as naturally occurring amino acids, i.e., carbons bonded to a hydrogen, a carboxyl group, an amino group, and an R group. Such analogs include compounds with modified R groups (e.g., homoserine, norleucine, methionine sulfoxide, and methionine methylsulfonium). norleucine), or a modified peptide backbone, but retain the same basic chemical structure as a naturally occurring amino acid. Amino acid mimetics refer to chemical compounds that have a structure that is different from the general chemical structure of an amino acid, but that functions in a manner similar to a naturally occurring amino acid.

[0124] Amino acids may be referred to herein by either their commonly known three letter symbols or by the one-letter symbols recommended by the IUPAC-IUB Biochemical Nomenclature Commission. Nucleotides may also be referred to by their commonly accepted one-letter codes. As used herein, "polar amino acids" refer to "A" refers to an amino acid containing a side chain that preferentially exists in an aqueous environment. In some embodiments, the polar amino acid is selected from the group consisting of arginine, asparagine, aspartic acid, glutamic acid, glutamine, histidine, lysine, serine, threonine, and tyrosine. The polar amino acid may be positively charged, negatively charged, or neutrally charged. As used herein, "non-polar amino acid" refers to an amino acid selected from the group consisting of alanine, cysteine, glycine, isoleucine, leucine, methionine, phenylalanine, proline, tryptophan, and valine.

[0125] As used herein, an "amino acid substitution" refers to the replacement of at least one existing amino acid residue in a defined amino acid sequence (the amino acid sequence of the starting polypeptide) with a second, different "replacement" amino acid residue. An "amino acid insertion" refers to the incorporation of at least one additional amino acid into a defined amino acid sequence. Insertions usually consist of the insertion of one or two amino acid residues, although larger "peptide insertions" can also be made. For example, about 3 Insertions of up to about 5, or even up to about 10, 15, or 20 amino acid residues. The inserted residues may be natural or non-natural residues as described above. "Amino acid deletion" refers to the removal of at least one amino acid residue from a defined amino acid sequence.

[0126] As used herein, the terms "amount" or "level" refer to a detectable quantity, level, or abundance of a substance. When referring to polypeptides such as those described herein, the terms "level of expression" or "expression level" are generally used interchangeably and generally refer to a detectable amount of the polypeptide in a biological sample (e.g., on a cell surface).

[0127] As used herein, the term "anti-CD137 agonistic antibody" (used interchangeably with the term "anti-CD137 antibody") refers to an antibody that specifically binds to CD137. and partially or completely promote, induce, enhance, or inhibit the biological activity, response, and / or downstream pathways mediated by CD137 signaling, or other CD137-mediated functions. In some embodiments, an anti-CD137 agonist In some embodiments, an anti-CD137 agonistic antibody binds to CD137 and induces CD137 multimerization. In some embodiments, an anti-CD137 agonistic antibody binds to CD137 and induces dimerization of CD137 trimers. In some embodiments, the anti-CD137 agonistic antibody binds to CD137 and inhibits CD137 trimerization. Examples of anti-CD137 agonistic antibodies are provided herein. Methods for detecting the formation of trimeric:trimeric complexes are known to those skilled in the art. For example, electron microscopy has been shown to detect such complexes. See, for example, Won, E. The Journal of Biological Chemistry, Vol. 285 (12): 9202-9210 (2010).

[0128] As used herein, the term "anti-CD137 mAb1" (interchangeable with "mAb1") The variable heavy chain (V H ) Amino acid sequence: EVQLLESGGGLVQPGGSLRLSCAASGFTFSSYAMSWVRQAPGKGLEWVSAISGSGGSTYYADSVKGRFTISRDNSKNTLYLQMNSLRAEDTAVYYCAKDSPFLLDDYYYYYYMDVWGKGTTVTVSS (SEQ ID NO: 4), and the following variable light chains (V L ) Amino acid sequence: Exemplary anti-CD137 agonistic antibodies include: DIQMTQSPSSVSASVGDRVTITCRASQGISSWLAWYQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQGHLFPITFGGGTKVEIK (SEQ ID NO: 6). vinegar.

[0129] As used herein, the term "anti-CD137 mAb8" (interchangeable with "mAb8") The variable heavy chain (V H ) Amino acid sequence: EVQLLESGGGLVQPGGSLRLSCAASGFTFRNYAMSWVRQAPGKGLEWVSAISGSGDTTYYADSVKGRFTISRDNSKNTLYLQMNSLRAEDTAVYYCAKDSPFLLDDYYYYYYMDVWGKGTTVTVSS (SEQ ID NO: 101), and the following variable light chains (V L ) Amino acid sequence: Exemplary anti-CD137 agonistic antibodies include: DIQMTQSPSSVSASVGDRVTITCRASQGISSWLAWYQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQGHLFPITFGGGTKVEIK (SEQ ID NO: 6). vinegar.

[0130] As used herein, the term "anti-CD137 mAb10" (interchangeable with "mAb10") The variable heavy chain (V H ) Amino acid sequence: EVQLLESGGGLVQPGGSLRLSCAASGFTFYGYAMSWVRQAPGKGLEWVAAISGSGDSTYYADSVKGRFTISRDNSKNTLYLQMNSLRAEDTAVYYCAKDSPFLLDDYYYYYYMDVWGKGTTVTVSS (SEQ ID NO: 26), and the following variable light chains (V L ) Amino acid sequence: Exemplary anti-CD137 agonistic antibodies include: DIQMTQSPSSVSASVGDRVTITCRASQGISSWLAWYQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQGHLFPITFGGGTKVEIK (SEQ ID NO: 6). vinegar.

[0131] As used herein, the term "antibody" refers to a whole antibody comprising two light polypeptide chains and two heavy polypeptide chains. Whole antibodies include various antibody isotypes, including IgM, IgG, IgA, IgD, and IgE antibodies. The term "antibody" refers to a polyclonal antibody. Antibodies include monoclonal antibodies, chimeric or chimeric antibodies, humanized antibodies, primatized antibodies, deimmunized antibodies, and fully human antibodies. Antibodies can be derived from any of a variety of species. The antibody may be produced in or derived from a mammal, such as a human, a non-human primate (e.g., an orangutan, baboon, or chimpanzee), a horse, a cat, a pig, a sheep, a goat, a dog, a cat, a rabbit, a guinea pig, a gerbil, a hamster, a rat, and a mouse. The antibody may be a purified antibody or a recombinant antibody.

[0132] As used herein, the terms "antibody fragment," "antigen-binding fragment," "antigen-binding portion," or similar terms refer to an antibody fragment that has the ability to bind to a target antigen (e.g., CD137). and inhibits the activity of the target antigen. Such fragments include, for example, single-chain antibodies, single-chain Fv fragments (scFv), Fd fragments, Fab fragments, Fab' fragments, or F(ab')2 fragments. and scFv fragments. An scFv fragment is a single polypeptide chain that contains both the heavy and light chain variable regions of the antibody from which the scFv is derived. Additionally, intrabodies, minibodies, triabodies, and diabodies are included within the definition of antibody and are suitable for use in the methods described herein. See, e.g., Todorovska et al., (2001) J. Immunol. Methods 248(1):47-66; Hudson and Kortt, (1999) J. Immunol. Methods 231(1):177-189; Poljak, (1994) Structure 2(12):1121-1123; Rondon and Marasco, (1997) Annu. Rev. Microbiol. 51:257-283, the disclosures of each of which are incorporated by reference in their entirety. and is incorporated herein by reference in its entirety.

[0133] As used herein, the term "antibody fragment" also includes single-domain antibodies, such as camelized single-domain antibodies. See, for example, Muyldermans et al., (2001) Trends Biochem. Sci. 26:230-235; Nuttall et al., (2000) Curr. Pharm. Biotech. 1:253-263; Reichmann et al., (1999) J. Immunol. Meth. 231:25-38; PCT International Patent Application Publications WO 94 / 04678 and WO 94 / 25591, and U.S. Patent No. 6,005,079, all of which are incorporated herein by reference in their entireties. In some embodiments, the present disclosure provides single-domain antibodies comprising two VH domains with modifications to form single-domain antibodies.

[0134] In some embodiments, the antigen-binding fragment comprises the variable region of a heavy chain polypeptide and the variable region of a light chain polypeptide. In some embodiments, the antigen-binding fragment described herein comprises the CDRs of the light and heavy chain polypeptides of the antibody.

[0135] The term "antigen-presenting cell" or "APC" is a cell that presents foreign antigen complexed with MHC on its surface. T cells recognize this complex using the T cell receptor (TCR). Examples of APCs include, but are not limited to, dendritic cells (DCs), peripheral blood mononuclear cells (PBMCs), monocytes (e.g., THP-1), B lymphoblastoid cells (e.g., C1R.A2, 1518 B-LCL), and monocyte-derived dendritic cells (DCs). Some APCs are capable of phagocytosis or receptor-mediated cell migration. The antigen is internalized by either localized endocytosis or by endogenous endocytosis.

[0136] The term "antigen presentation" refers to the process by which APCs capture antigens and make them recognized by T cells, for example as components of MHC-I and / or MHC-II complexes.

[0137] As used herein, the term "apoptosis" refers to the process of programmed cell death that occurs in multicellular organisms (e.g., humans). Highly regulated biochemical and molecular events result in apoptosis, resulting in characteristic observable morphological changes, including membrane blebbing, cell volume reduction, chromatin DNA compaction and fragmentation, and mRNA decay. It can cause changes in cells, including T cells, that are undergoing apoptosis. A common method for detecting apoptotic cells is to expose them to a fluorophore-binding protein (Annexin V), which is commonly used to detect apoptotic cells due to its ability to bind to phosphatidylserine on the outer leaflet of the cell membrane, providing an early indicator that cells are undergoing the apoptotic process.

[0138] As used herein, the term "binds to immobilized CD137" refers to The disclosed human antibodies can be used to bind to CD137, e.g., expressed on a cell surface, or attached to a solid support. This refers to the ability of the antibody to bind to CD137, which is known to be a target of immunoglobulin A.

[0139] As used herein, the term "bispecific" or "bifunctional antibody" refers to an artificial hybrid antibody having two different heavy / light chain pairs and having two different binding sites. Bispecific antibodies can be synthesized by hybridoma fusion or by linking Fab' fragments. These antibodies can be prepared by various methods, including fusion. See, for example, Songsivilai & Lachmann (1990) Clin. Exp. Immunol. 79:315-321; Kostelny et al. (1992) See J. Immunol. 148:1547-1553.

[0140] Traditionally, recombinant production of bispecific antibodies has been based on the co-expression of two immunoglobulin heavy / light chain pairs, each with different specificities (Milstein and Cuello, (1983) Nature 305:537-539). Antibody variable domains (containing antibody binding sites and antigen binding sites) may be fused to immunoglobulin constant domain sequences. Fusion of a heavy chain variable region with an immunoglobulin heavy chain constant domain containing at least part of the hinge, CH2, and CH3 regions is preferred. For further details regarding exemplary methods currently known for producing bispecific antibodies, see, e.g., Suresh et al., (1986) Methods Enzymol. 121:210; PCT International Patent Application Publication No. WO 96 / 27011; Brennan et al., (1985) Science 229:81; Shalaby et al., J. Exp. Med. (1992) 175:217-225; See Kostelny et al., (1992) J. Immunol. 148(5):1547-1553; Hollinger et al., (1993) Proc. Natl. Acad. Sci. USA 90:6444-6448; Gruber et al., (1994) J. Immunol. 152:5368; and Tutt et al., (1991) J. Immunol. 147:60. Bispecific antibodies also include cross-linked or heteroconjugate antibodies. Heteroconjugate antibodies can be prepared by any convenient method. Suitable cross-linking agents are known in the art and are disclosed in U.S. Patent No. 4,676,980, along with a number of cross-linking methods.

[0141] Various methods for producing and isolating bispecific antibody fragments directly from recombinant cell culture have also been reported. For example, bispecific antibodies have been produced using leucine zippers. See, e.g., Kostelny et al. (1992) J Immunol 148(5):1547-1553. Leucine zipper peptides derived from the Fos and Jun proteins have been used to synthesize two different antibodies by gene fusion. The Fab' portions of the antibodies can be linked together. The antibody homodimers can be reduced at the hinge region to form monomers. The antibody fragments can be formed and re-oxidized to form heterodimeric antibodies. This method can also be used to generate homodimeric antibodies. The "diabody" method described by Hollinger et al. (1993) Proc Natl Acad Sci USA 90:6444-6448 provides another mechanism for generating bispecific antibody fragments. The fragments comprise a heavy chain variable domain (VH) connected by a linker to a light chain variable domain (VL). The linker can be used to generate bispecific antibody fragments on the same chain. The fragments are too short to allow pairing between the two domains. Therefore, the VH and VL domains of one fragment are forced to pair with the complementary VL and VH domains of another fragment, thereby forming two antigen-binding sites. Another strategy for making bispecific antibodies using single-chain Fv (scFv) dimers has also been reported. See, e.g., Gruber et al. (1994) J Immunol 152:5368. Alternatively, the antibodies may be "linear antibodies," as described, for example, in Zapata et al. (1995) Protein Eng. 8(10):1057-1062. Briefly, these antibodies comprise a pair of tandem Fd segments (VH-CH1-VH-CH1), which These form a pair of antigen-binding regions. Linear antibodies can be bispecific or monospecific.

[0142] Antibodies with more than two valencies (e.g., trispecific antibodies) are also contemplated, see, e.g., Tutt et al. (1991) J Immunol 147:60.

[0143] The present disclosure also encompasses variant forms of multispecific antibodies, such as dual variable domain immunoglobulin (DVD-Ig) molecules, described, for example, in Wu et al. (2007) Nat Biotechnol 25(11): 1290-1297. DVD-Ig molecules are constructed by combining two different light chain variable domains (VL) from two different parent antibodies in tandem, either directly or via a short linker, by recombinant DNA techniques. Similarly, the heavy chain is designed to have two different heavy chain variable domains (VH) linked in tandem, followed by the constant domains CH1 and and Fc regions. Methods for generating DVD-Ig molecules from two parent antibodies are described, for example, in PCT International Publication No. PCT / US99 / 023094. In some embodiments, the bispecific antibody is a Fab-in-Tandem immunoglobulin, which is a second specific antibody. In one embodiment, the antibody is an immunoglobulin in which a light chain variable region having the same function as the antibody is fused to a heavy chain variable region of a whole antibody. Such antibodies are described, for example, in International Patent Application Publication WO2015 / 103072.

[0144] As used herein, a "cancer antigen" refers to (i) a tumor-specific antigen; (ii) a tumor-associated antigen; (iii) cells expressing tumor-specific antigens, (iv) cells expressing tumor-associated antigens, (v) embryonic antigens on tumors, (vi) autologous tumor cells, (vii) tumor-specific membrane antigens, (viii) tumor-associated membrane antigens, (ix) growth factor receptors, (x) growth factor ligands, and (xi) cancer-associated antigens. The term "antigen-presenting cell" refers to any other type of antigen or antigen-presenting cell or substance.

[0145] The term "cancer" is art-recognized and refers to malignant tumors of epithelial and endocrine tissues, including cancers of the respiratory system, gastrointestinal system, genitourinary system, testicular cancer, breast cancer, prostate cancer, endocrine system cancer, and melanoma.

[0003] These compounds can be used to treat patients who have, are suspected of having, or may be at high risk of developing any type of cancer, including renal cancer or melanoma. Exemplary cancers include those forming from tissue of the cervix, lung, prostate, breast, head and neck, colon, and ovary. The term also includes carcinosarcomas, which include malignant tumors composed of carcinomatous and sarcomatous tissue. "Adenocarcinoma" refers to a cancer that originates in glandular tissue or in which the tumor cells form recognizable glandular structures.

[0146] As used herein, the term "compete," when used in the context of antigen-binding proteins (e.g., immunoglobulins, antibodies, or antigen-binding fragments thereof) that compete for binding to the same epitope, refers to an interaction between antigen-binding proteins as determined by an assay (e.g., competitive binding assay, cross-inhibition assay, etc.), where a test antigen-binding protein (e.g., a test antibody) is more efficient than a reference antigen-binding protein (e.g., a reference antibody, e.g., mAb1) in specific binding to a common antigen (e.g., CD137 or a fragment thereof). inhibit (e.g., decrease or prevent)

[0147] In some embodiments, the antibodies described herein cross-compete with mAb1 (i.e., an antibody comprising the heavy and light chain variable sequences of SEQ ID NOs: 4 and 6, respectively), mab8 (i.e., an antibody comprising the heavy and light chain variable sequences of SEQ ID NOs: 101 and 6, respectively), or mAb10 (i.e., an antibody comprising the heavy and light chain variable sequences of SEQ ID NOs: 26 and 6, respectively).

[0148] A polypeptide or amino acid sequence "derived from" a specified polypeptide or protein refers to the origin of the polypeptide. Preferably, a polypeptide or amino acid sequence derived from a particular sequence has an amino acid sequence that is essentially identical to that sequence or a portion thereof, where the portion consists of at least 10-20 amino acids, preferably at least 20-30 amino acids, more preferably at least 30-50 amino acids, or has an amino acid sequence that is recognizable to one of skill in the art as having that origin in sequence. A polypeptide derived from another peptide may have one or more mutations compared to the starting polypeptide, for example, one or more amino acid residues substituted for other amino acid residues, or one or more amino acid residues inserted or deleted.

[0149] Polypeptides may contain amino acid sequences that are not naturally occurring. Such variants necessarily have less than 100% sequence identity or sequence similarity with the starting molecule. In certain embodiments, In this case, variants have about 75% to less than 100% identical amino acid sequence to the starting polypeptide, more preferably or about 80% to less than 100%, more preferably about 85% to less than 100%, more preferably about 90% to less than 100%, (e.g., 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%), and most preferably about 95% to less than 100% amino acid sequence identity or similarity across the full length of the variant molecule. has over the years.

[0150] In certain embodiments, there is one amino acid sequence between the starting polypeptide sequence and the sequence derived therefrom. There are amino acid differences. Identity or similarity with respect to sequences is defined herein as the percentage of amino acid residues in a candidate sequence that are identical (i.e., the same residues) to the starting amino acid residues after aligning the sequences and inserting gaps, if necessary, to achieve the maximum percentage of sequence identity. In certain embodiments, a polypeptide consists of, consists essentially of, or comprises an amino acid sequence selected from the sequences set forth in Table 3 or Table 4. In certain embodiments, a polypeptide comprises an amino acid sequence that is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% identical to an amino acid sequence selected from the sequences set forth in Table 3 or Table 4. In certain embodiments, the polypeptide has at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, or more amino acid residues to a contiguous amino acid sequence selected from the sequences set forth in Table 3 or Table 4. , 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% identical In certain embodiments, the polypeptide comprises an amino acid sequence having at least 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, 100, 200, 300, 400, or 500 (or any integer within these numbers) consecutive amino acids of an amino acid sequence selected from the sequences set forth in Table 3 or Table 4.

[0151] In certain embodiments, the antibodies of the present disclosure are encoded by a nucleotide sequence. The nucleotide sequences of the present invention may be useful for many applications, including cloning, gene therapy, protein expression and purification, mutagenesis, DNA vaccination of a host in need thereof, antibody generation for, e.g., passive immunization, PCR, primer and probe generation, and the like. In certain embodiments, the nucleotide sequences of the present invention consist of, consist essentially of, or comprise a nucleotide sequence selected from a sequence set forth in Table 3 or Table 4. In certain embodiments, the nucleotide sequence comprises a nucleotide sequence that is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to a nucleotide sequence selected from a sequence set forth in Table 3 or Table 4. In certain embodiments, the nucleotide sequence is selected from a sequence set forth in Table 3 or Table 4. or a contiguous nucleotide sequence that is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% identical to a contiguous nucleotide sequence selected from the sequences set forth in Table 4. The polypeptide may comprise at least 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, 100, 200, 300, 400, or 500 (or any integer within these numbers) consecutive nucleotides of a nucleotide sequence selected from the sequences set forth in Table 3 or Table 4. It includes a nucleotide sequence having the following structure:

[0152] Those skilled in the art will also understand that antibodies suitable for use in the methods disclosed herein may be modified to differ in sequence from the native or natural sequence from which they are derived while retaining the desired activity of the native sequence. For example, nucleotide or amino acid substitutions leading to conservative substitutions or changes of "non-essential" amino acid residues may be made. Mutations may be made using targeted techniques such as, for example, site-directed mutagenesis and PCR-mediated mutagenesis. It can be introduced by standard methods.

[0153] Antibodies suitable for use in the methods disclosed herein may contain conservative amino acid substitutions at one or more amino acid residues, e.g., essential or non-essential amino acid residues. A "conservative amino acid substitution" is one in which an amino acid residue is replaced with an amino acid residue having a similar side chain. Families of amino acid residues with similar side chains have been defined in the art, including basic side chains (e.g., lysine, arginine, histidine), acidic side chains (e.g., aspartic acid, glutamic acid), uncharged polar side chains (e.g., glycine, asparagine, glutamine, serine, threonine, tyrosine, cysteine), nonpolar side chains (e.g., alanine, valine, leucine, isoleucine, proline, phenylalanine, methionine, tryptophan), beta-branched side chains (e.g., threonine, valine, isoleucine), and aromatic side chains (e.g., tyrosine, phenylalanine, tryptophan, histidine). Thus, non-essential amino acid residues in a binding polypeptide are preferably replaced with another amino acid residue from the same side chain family. In certain embodiments, a consecutive sequence of amino acids may be replaced with a structurally similar consecutive sequence that differs in the order and / or composition of side chain family members. Alternatively, in certain embodiments, mutations may be randomly introduced along all or part of the coding sequence, e.g., by saturation mutagenesis, and the resulting mutations may be incorporated into a binding polypeptide of the invention and screened for the ability to bind to a desired target.

[0154] As used herein, the term antigen "cross-presentation" refers to the presentation of foreign protein antigens to T cells via MHC class I and class II molecules on APCs.

[0155] As used herein, the term "cross-reactivity" refers to the ability of an antibody of the present disclosure to bind to CD137 from a different species. For example, an antibody of the present disclosure that binds to human CD137 may also bind to CD137 from another species. As used herein, cross-reactivity refers to the ability of an antibody of the present disclosure to bind to CD137 from a different species. Measured by detecting specific reactivity with purified antigen in a ELISA (e.g., SPR, ELISA) or bind to or otherwise functionally bind to cells that physiologically express CD137. The cross-reactivity is measured by detecting the interaction between the two proteins. For example, a Biacore™ 2000 SPR instrument (Biacore AB, Uppsala, Sweden) is used to determine the cross-reactivity. Standard binding assays such as those described herein, such as Biacore™ surface plasmon resonance (SPR) analysis or flow cytometry, were used.

[0156] As used herein, the term "cytotoxic T lymphocyte (CTL) response" refers to an immune response induced by cytotoxic T cells. CTL responses are primarily CD8 + Mediated by T cells There is.

[0157] As used herein, the term "dimerization" refers to the formation of two, usually non-covalent, bonds. "Dimerization" refers to the formation of a macromolecular complex by polymers, such as proteins or protein multimers, that are covalently linked together. Homodimerization refers to the process of dimerization when the properties of the polymers (e.g., proteins) are identical. Heterodimerization refers to the process of dimerization when the properties of the polymers (e.g., proteins) are not identical. Methods for measuring dimerization are known to those skilled in the art. For example, but not limited to, yeast two-hybrid assays are known. Assay, Fluorescence Resonance Energy Transfer (FRET), Bioluminescence Resonance Energy Transfer (BRET), Tandem Protein mass spectrometry, evanescent wave spectroscopy, size exclusion chromatography, analytical ultracentrifugation, scattering, NMR spectroscopy, isothermal titration calorimetry, fluorescence anisotropy, fluorescence correlation spectroscopy (FCS), fluorescence recovery after photobleaching (FRAP), proximity imaging (PRIM), and fluorescent tandem imaging. Protein reconstitution (BiFC) (see e.g. Gell DA, Grant RP, Mackay JP (2012) The Detection and Quantitation of Protein Oligomerization. In: Matthews JM (eds) Protein Dimerization and Oligomerization in Biology. Advances in Experimental Medicine and Biology, vol. 747. Springer, New York, NY; and Xie, Q. et al. Methods Mol Biol, 2011; 680: 3-28) Examples include:

[0158] As used herein, "CD137 dimerization" refers to the dimerization of two CD137 trimers. In some embodiments, the anti-CD137 agonistic antibodies described herein In some embodiments, the antibody induces or enhances CD137 dimerization. The anti-CD137 agonistic antibody described herein induces or enhances dimerization of CD137 compared to the amount of dimerization in the absence of the anti-CD137 agonistic antibody. The anti-CD137 agonistic antibodies described herein induce or enhance dimerization of CD137 compared to the amount of dimerization in the presence of a reference anti-CD137 agonistic antibody. Thus, dimerization is increased by at least 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95% or 100%.

[0159] As used herein, "EC 50 " refers to the concentration of an antibody or antigen-binding portion thereof that induces 50% of the maximal response, i.e., a response that is halfway between the maximal response and a reference, in either an in vitro or in vivo assay.

[0160] As used herein, the term "effective dosage" or "effective dose" is defined as an amount sufficient to achieve or at least partially achieve the desired effect. The term "therapeutically effective dosage" is defined as an amount sufficient to cure or at least partially halt the disease and complications in patients already suffering from the disease. The amount effective for this purpose depends on the severity of the disorder being treated and the general state of the patient's own immune system.

[0161] As used herein, the term "epitope" or "antigenic determinant" refers to a determinant or site on an antigen (e.g., CD137) to which an antigen-binding protein (e.g., an immunoglobulin, antibody, or antigen-binding fragment) specifically binds. Epitopes can be divided into "linear epitopes" and "conformational epitopes." As used herein, the term "linear epitope" refers to an epitope formed from a continuous linear sequence of linked amino acids. Linear epitopes of protein antigens are typically maintained upon exposure to chemical denaturants (e.g., acids, bases, solvents, cross-linking reagents, chaotropic agents, disulfide bond-reducing agents) or physical denaturation (e.g., heat, radiation, or mechanical shear or stress). In some embodiments, epitopes are nonlinear and are also referred to as interrupted epitopes. As used herein, the term "conformational epitope" or "nonlinear epitope" refers to an epitope formed from noncontiguous amino acids juxtaposed by tertiary folding of a polypeptide. Conformational epitopes are typically lost upon treatment with denaturing agents. Typically, an epitope contains at least 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, or 15 amino acids in a unique spatial conformation. In some embodiments, the epitope comprises less than 25, 24, 23, 22, 21, 20, 19, 18, 17, 16, 15, 14, 13, 12, 11, 10, 9, 8, 7, 6, or 5 amino acids in a unique spatial conformation. Generally, an antibody or antigen-binding fragment thereof specific for a particular target molecule preferentially recognizes and binds to a specific epitope on the target molecule within a complex mixture of proteins and / or macromolecules. In some embodiments, the epitope is the extracellular domain of human CD137. Does not contain all amino acids of the domain.

[0162] Also encompassed by the present disclosure are antibodies that bind to epitopes on CD137, including all or part of the epitopes recognized by the specific antibodies described herein (e.g., the same or overlapping regions, or regions between or spanning regions).

[0163] As used herein, the term "epitope mapping" refers to the process or method of identifying the binding site or epitope of an antibody or antigen-binding fragment thereof, or its target protein antigen. Epitope mapping methods and techniques are presented herein.

[0164] As used herein, the term "CD137" refers to a transmembrane protein. This refers to a specific member of the tumor necrosis factor receptor (TNFR) family. Alternative names and acronyms for this protein include "tumor necrosis factor receptor superfamily member 9" (TNFRSF9), 4-1BB, and "induced by lymphocyte activation" (ILA) (Alderson et al., (1994) Eur J Immunol 24(9):2219-2227; Schwarz et al., (1993) Gene 134(2):295-298). Leader domain, transmembrane domain An example of the amino acid sequence of full-length human CD137, including the cytoplasmic domain, is shown in Table 4 (SEQ ID NO: 3) and and listed below: MGNSCYNIVATLLLVLNFERTRSLQDPCSNCPAGTFCDNNRNQICSPCPPNSFSSAGGQRTCDICRQCKGVFRTRKECSSTSNAECDCTPGFHCLGAGCSMCEQDCKQGQELTKKGCKDCCFGTFNDQ KRGICRPWTNCSLDGKSVLVNGTKERDVVCGPSPADLSPGASSVTPPAPAREPGHSPQIISFFLALTSTALLFLLFFLTLRFSVVKRGRKKLLYIFKQPFMRPVQTTQEEDGCSCRFPEEEEGGCEL.

[0165] As used herein, the term "CD137L" or "CD137 ligand" refers to a member of the tumor necrosis factor (TNF) family of transmembrane proteins. Other names and acronyms for CD137L include "tumor necrosis factor superfamily member 9" (TNFSF9) and 4-1BB ligand (4-1BBL) (Alderson et al., (1994) Eur J Immunol 24(9):2219-2227). An exemplary amino acid sequence of full-length CD137L is set forth in Table 4 (SEQ ID NO: 97).

[0166] As used herein, the term "Fc-mediated effector function" or "Fc effector function" refers to the primary function of an antibody and other biological activities of the antibody besides its intended target. For example, the effector function of a therapeutic agonistic antibody is a biological activity other than activation of a target protein or pathway. Examples of antibody effector functions include C1q binding and complement-dependent Cell damage, Fc receptor binding, antibody-dependent cell-mediated cytotoxicity (ADCC), phagocytosis, cell surface downregulation of receptors (e.g., B cell receptors), lack of activation of platelets expressing Fc receptors; and B cell activation. Many effector functions are mediated by Fc binding to Fcγ receptors. It starts from.

[0167] As used herein, the term "Fc receptor" refers to a polypeptide present on the surface of an immune effector cell that is bound to the Fc region of an antibody. In some embodiments, the Fc receptor is an Fcγ receptor. There are three subclasses of Fcγ receptors: FcγRI (CD64), FcγRII (CD32), and FcγRIII (CD16). All four IgG isotypes (IgG1, IgG2, IgG3, and IgG4) bind to the Fc receptors FcγRI, FcγRIIA, and FcγRIIIA. FcγRIIB is an inhibitory receptor, so antibody binding to this receptor does not activate complement and cellular responses. FcγRI binds to monomeric IgG. On the other hand, FcγRIIA and FcγRIIA bind to multimeric IgG. The binding of antibodies to Fc receptors and / or C1q is governed by specific residues or domains within the Fc region. Binding also depends on residues within the hinge region and CH2 portion of the antibody. In some embodiments, the agonistic and / or therapeutic activity of the antibodies described herein is dependent on binding of the Fc region to an Fc receptor (e.g., FcγR). The agonistic and / or therapeutic activity of the antibodies described herein is enhanced by binding of the Fc region to an Fc receptor (e.g., FcγR).

[0168] As used herein, the term "glycosylation pattern" is defined as the pattern of sugar units covalently attached to a protein, more specifically, an immunoglobulin protein. The glycosylation pattern of a heterologous antibody can be characterized as being substantially similar to the glycosylation pattern naturally present on antibodies produced by a non-human transgenic animal species if one skilled in the art recognizes that the glycosylation pattern of the heterologous antibody is more similar to the glycosylation pattern in the non-human transgenic animal species than to the species from which the transgenic CH gene is derived.

[0169] As used herein, the term "blood cancer" includes lymphoma, leukemia, myeloma or lymphoid malignancies, and cancer of the spleen and lymph nodes. Examples of lymphoma include both B-cell lymphoma (a blood cancer of B cells) and T-cell lymphoma. B-cell lymphomas include Hodgkin's lymphoma and most non-Hodgkin's lymphomas. Non-limiting examples of B-cell lymphomas include diffuse large B-cell lymphoma, follicular lymphoma, mucosa-associated lymphoid tissue lymphoma, small cell lymphocytic lymphoma (which overlaps with chronic lymphocytic leukemia), mantle cell lymphoma (MCL), Burkitt's lymphoma (BCL), and leukemia-associated lymphoma (LEM). lymphoma, mediastinal large B-cell lymphoma, Waldenström's macroglobulinemia, nodal Non-limiting examples of T-cell lymphomas include marginal zone B-cell lymphoma, splenic marginal zone lymphoma, intravascular large B-cell lymphoma, primary effusion lymphoma, and lymphomatoid granulomatosis. Hematological malignancies include, but are not limited to, pulmonary thrombocytopenic leukemia, extranodal T-cell lymphoma, cutaneous T-cell lymphoma, anaplastic large cell lymphoma, and angioimmunoblastic T-cell lymphoma. Hematological malignancies include, but are not limited to, myelogenous leukemia, chronic lymphocytic leukemia, acute myeloid leukemia, chronic myelogenous leukemia, and acute lymphoblastic leukemia.Hematological malignancies also include, but are not limited to, multiple myeloma and smoldering multiple myeloma.Other hematological cancers and / or B-cell-related cancers or T-cell-related cancers are also included in the term hematological malignancies.

[0170] As used herein, the term "human antibody" includes antibodies having variable and constant regions (if any) derived from human germline immunoglobulin sequences. The human antibodies of the present disclosure may include amino acid residues not encoded by human germline immunoglobulin sequences (e.g., mutations introduced by random or site-specific mutagenesis in vitro or by somatic mutation in vivo) (e.g., Lonberg et al., (1994) Nature 368(6474): 856-859; Lonberg, (1994) Handbook of Experimental (See, e.g., Pharmacology 113:49-101; Lonberg & Huszar, (1995) Intern. Rev. Immunol. 13:65-93, and Harding & Lonberg, (1995) Ann. NY Acad. Sci. 764:536-546.) However, the term "human antibody" refers to an antibody in which CDR sequences derived from the germline of another mammalian species, such as a mouse, are grafted onto human framework sequences. This does not include antibodies that have been modified (i.e., humanized antibodies).

[0171] As used herein, the term "heterologous antibody" is defined in relation to the transgenic non-human organism that produces the antibody. This term refers to an antibody that has an amino acid sequence or encoding nucleic acid sequence that corresponds to a sequence present in an organism that is not part of the transgenic non-human animal and that is generally derived from a species that is not the transgenic non-human animal.

[0172] The terms "inducing an immune response" and "enhancing an immune response" are used interchangeably and refer to the stimulation (i.e., passive or active) of an immune response to a particular antigen. The term "inducing" when used in reference to inducing CDC or ADCC refers to a specific, direct Refers to stimulation of cell killing mechanisms.

[0173] As used herein, a subject "in need of prevention," "in need of treatment," or "in need of" is one who, according to the judgment of a competent medical professional (e.g., a physician, nurse, or nursing intern in the case of a human, or a veterinarian in the case of a non-human mammal), would reasonably benefit from a given treatment (e.g., treatment with a composition comprising an anti-CD137 antibody). It refers to something.

[0174] The term "in vivo" refers to a process that occurs in a living organism.

[0175] As used herein, the term "isolated antibody" is intended to refer to an antibody that is substantially free of other antibodies having different antigen specificities (e.g., an isolated antibody that specifically binds to human CD137 is substantially free of antibodies that specifically bind to antigens other than CD137). However, isolated antibodies that specifically bind to the epitope may not be compatible with other CD137 proteins from different species. However, the antibody continues to exhibit specific binding to human CD137 in the specific binding assays described herein. Furthermore, isolated antibodies are typically substantially free of other cellular material and / or chemicals. In some embodiments, a combination of "isolated" antibodies with different CD137 specificities may be used. are combined in well-defined compositions.

[0176] As used herein, the term "isolated nucleic acid molecule" refers to a nucleic acid molecule that binds to CD137. The binding antibody or antibody moiety (e.g., V H , V L The term "antibody" refers to a nucleic acid encoding a CDR3 (CDR4), and the nucleotide sequence encoding the antibody or antibody portion is an antibody or antibody fragment that binds to an antigen other than CD137. It is intended to refer to a nucleic acid molecule that does not contain other nucleotide sequences encoding body parts, which may naturally flank the nucleic acid in human genomic DNA. For example, a sequence selected from the sequences set forth in Table 3 or Table 4 may be used in the heavy chain variable region (V) of the anti-CD137 monoclonal antibodies described herein. H ) and the light chain variable region (V L ) containing Corresponding to the octide sequence.

[0177] As used herein, "isotype" refers to the class of antibody (e.g., IgM or IgG1) encoded by heavy chain constant region genes. In some embodiments, The human monoclonal antibody of the present disclosure is an IgG1 isotype. In some embodiments, the human monoclonal antibody of the present disclosure is an IgG1 isotype and comprises a mutation. In some embodiments, the human monoclonal antibody of the present disclosure is an IgG2 isotype. In some embodiments, the human monoclonal antibody of the present disclosure is an IgG3 isotype. In some embodiments, the human monoclonal antibody of the present disclosure is an IgG4 isotype. In some embodiments, the human monoclonal antibody of the present disclosure is an IgG4 isotype and comprises a mutation. In some embodiments, the mutation is a substitution of Ser228. In some embodiments, the substitution of Ser228 is S228P.

[0178] As used herein, the term "isotype switching" refers to the phenomenon in which the class or isotype of an antibody changes from one Ig class to another Ig class isotype.

[0179] As used herein, "KD" or "K D The term "K" refers to the equilibrium dissociation constant of the binding reaction between an antibody and an antigen. D The value is a numerical expression of the ratio of the antibody dissociation rate constant (kd) to the antibody association rate constant (ka). D The K value is inversely related to the binding affinity of the antibody for the antigen. D The smaller the value, the higher the affinity of the antibody for its antigen. Affinity is the strength of binding of a single molecule to its ligand and is typically expressed as the equilibrium dissociation constant (K D ), which is used to assess and rank the strength of bimolecular interactions.

[0180] As used herein, "Kd" or "k d (or "koff" or "k offThe term kd is intended to refer to the dissociation rate constant for an antibody to dissociate from an antibody / antigen complex. The kd value is a numerical representation of the fraction of the complex that dissociates or dissociates per second. Yes, sec -1 It is expressed in units of:

[0181] As used herein, "ka" or "k a " (or "kon" or "k on The term "antibody-antigen association rate constant" is intended to refer to the association rate constant of an antibody and an antigen. The ka value is a numerical expression of the number of antibody / antigen complexes formed per second in a 1M solution of antibody and antigen, and M -1 sec -1 It is expressed in units of .

[0182] As used herein, the terms "linked," "fused," or "fusion" are used interchangeably. These terms refer to the association of two or more elements or It refers to joining components or domains by any means, including chemical conjugation or recombinant means. Methods of chemical conjugation (such as the use of heterobifunctional cross-linking agents) are known in the art.

[0183] As used herein, "local administration" or "local delivery" refers to delivery that does not rely on transport of a composition or agent to a desired target tissue or target site via the vascular system. For example, a composition may be delivered by injection or implantation of the composition or agent, or by injection or implantation of a device containing the composition or agent. After local administration in the vicinity of the target tissue or target site, the composition or agent, or one or more components thereof, may diffuse to the desired target tissue or target site.

[0184] As used herein, "MHC molecule" refers to two types of MHC molecules: MHC class I and MHC class II. MHC class I molecules present antigens to specific CD8+ T cells, and MHC class II molecules present antigens to specific CD4+ T cells. Antigens delivered exogenously to APCs are primarily expressed through MHC class I molecules. In contrast, antigens delivered endogenously to APCs are processed primarily for association with MHC class I.

[0185] As used herein, the term "monoclonal antibody" refers to an antibody that displays a single binding specificity and affinity for a particular epitope. Thus, the term "human monoclonal antibody" refers to an antibody that displays a single binding specificity and has variable and optionally constant regions derived from human germline immunoglobulin sequences. In some embodiments, human monoclonal antibodies are produced by hybridomas comprising B cells obtained from a transgenic non-human animal, e.g., a transgenic mouse, whose genome includes human heavy chain and light chain transgenes, and fused with an immortalized cell. .

[0186] As used herein, the term "multimerization" refers to an antibody comprising three or more macromolecules, such as proteins, that are typically linked through non-covalent interactions. Multimerization refers to the formation of a complex. Methods for measuring multimerization are known to those skilled in the art and are described above with respect to dimerization. In some embodiments, the anti-CD137 agonists described herein The specific antibody induces or enhances multimerization of CD137. The anti-CD137 agonistic antibodies described herein induce or enhance multimerization of CD137 compared to the amount of multimerization in the absence of the anti-CD137 agonistic antibody. Thus, the anti-CD137 agonistic antibodies described herein induce or enhance CD137 multimerization compared to the amount of multimerization in the presence of a reference anti-CD137 agonistic antibody. In some embodiments, multimerization is increased by at least 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95% or 100%.

[0187] As used herein, the term "naturally occurring" when applied to an object refers to the fact that the object can exist in nature. For example, a polypeptide or polynucleotide sequence that exists in an organism (including viruses) that can be isolated from a source in nature and has not been intentionally modified by humans in a laboratory is naturally occurring.

[0188] As used herein, the term "nonswitched isotype" refers to a The term "non-switched" refers to the class of heavy chain isotypes produced when isotype switching has not occurred. The CH gene encoding the non-switched isotype is usually the first CH gene immediately downstream of the functionally rearranged VDJ gene. Isotype switching occurs when Isotype switching is classified as classical or non-classical. Classical isotype switching occurs through recombination events involving at least one switch sequence region in the transgene. Non-classical isotype switching is, for example, the result of human σ μ and human Σ μ Alternative non-classical switches can occur through homologous recombination (δ-associated deletion) between Mechanisms such as inter-transgene and / or inter-chromosomal recombination specifically allow isotype switching to occur and be effective.

[0189] As used herein, the term "nucleic acid" refers to deoxyribonucleotides or ribonucleotides and polymers thereof in either single- or double-stranded form. Unless otherwise limited, the term encompasses nucleic acids containing known analogs of natural nucleotides that have similar binding properties as the reference nucleic acid and are metabolized in a manner similar to natural nucleotides. Unless otherwise indicated, a particular nucleic acid sequence also encompasses implicitly conservatively modified variants thereof (e.g., degenerate codon substitutions) and complementary sequences as well as the explicitly indicated sequence. Specifically, degenerate codon substitutions refer to sequences in which the third position of one or more selected (or all) codons is replaced with a mixed-base and / or deoxyinosine residue. This can be done by creating a sequence (Batzer et al., Nucleic Acid Res. 19:5081, 1991; Ohtsuka et al., Biol. Chem. 260:2605-2608, 1985; and Cassol et al., 1992; Rossolini et al., Mol. Cell. Probes 8:91-98, 1994). For arginine and leucine, modifications at the second base can also be conservative. Nucleic Acids The terms gene, cDNA, and mRNA encoded by a gene are used interchangeably.

[0190] As used herein, a polynucleotide may be composed of any polyribonucleotide or polydeoxyribonucleotide, including unmodified RNA or unmodified ribonucleotides. It may be modified DNA, or it may be modified RNA or modified DNA. Reotides are used to identify single-stranded and double-stranded DNA, DNA that is a mixture of single- and double-stranded regions, Single-stranded and double-stranded RNA, RNA that is a mixture of single- and double-stranded regions, single-stranded or DNA that may be more typically double-stranded or is a mixture of single- and double-stranded regions and hybrid molecules containing RNA. Additionally, polynucleotides can be composed of RNA, or DNA, or triple-stranded regions containing both RNA and DNA. A polynucleotide may also contain one or more modified bases or DNA or RNA backbones modified for stability or other reasons. "Modified" bases include, for example, tritylated bases and unusual bases such as inosine. Various modifications may be made to DNA and RNA. Thus, "polynucleotide" encompasses chemically, enzymatically, or metabolically modified forms.

[0191] A nucleic acid is "operably linked" when it is placed into a functional relationship with another nucleic acid sequence. For example, a promoter or enhancer is operably linked to a coding sequence if it affects the transcription of the sequence. With respect to transcriptional regulatory sequences, operably linked means that the linked DNA sequences are contiguous and join two protein-coding regions. When necessary, operably linked means contiguous and in reading frame. With respect to switch sequences, operably linked indicates that the sequences are capable of effecting switch recombination.

[0192] As used herein, the term "paratope" or "antigen-binding site" refers to an antibody or antigen-binding fragment thereof, which recognizes and binds to an epitope on an antigen, comprising a set of complementarity-determining regions (CDRs) located within the variable heavy and variable light chains.

[0193] As used herein, "parenteral administration," "administered parenterally," and other grammatically equivalent terms refer to modes of administration other than enteral and topical administration, typically by injection, including, but not limited to, intravenous, intranasal, intraocular, intramuscular, intra-arterial, intrathecal, intracapsular, intraorbital, intracardiac, intradermal, intraperitoneal, transtracheal, subcutaneous, subcuticular, intra-articular, subcapsular, subarachnoid, intrathecal, epidural, intracerebral, intracranial, intracarotid, and substernal injection and infusion.

[0194] As used herein, the term "patient" includes human and other mammalian subjects receiving either prophylactic or therapeutic treatment.

[0195] In the context of two or more nucleic acid or polypeptide sequences, the term "percent identity" refers to two or more sequences or subsequences that have a specified percentage of identical nucleotides or amino acid residues when compared and aligned for maximum correspondence, as determined using one of the sequence comparison algorithms described below (e.g., BLASTP and BLASTN, or other algorithms available to those skilled in the art), or by visual inspection. Depending on the application, the "percent identity" may extend over a region of the compared sequences, such as a functional domain, or alternatively, over the entire length of the two sequences being compared. For sequence comparison, typically, one sequence serves as a reference sequence to which a verification sequence is compared. When using a sequence comparison algorithm, test and reference sequences are input into a computer, subsequence coordinates are designated, if necessary, and sequence algorithm program parameters are designated. The sequence comparison algorithm then calculates the percent sequence identity of the test sequence relative to the reference sequence based on the designated program parameters.

[0196] Optimal sequence alignment for comparison can be found, for example, in Smith & Waterman, Adv. Appl. Math. 2:482 (1981), by the local homology algorithm of Needleman & Wunsch, J. Mol. Biol. 48:443 (1970), by the homology alignment algorithm of Pearson & Lipman, Proc. Nat'l. Acad. Sci. USA 85:2444 (1988), The search method for these computer-implemented algorithms (Wisconsin Genetics Software Package, Genetics Computer Group, 575 Science Dr., Wisconsin This may be performed by laboratory testing (GAP, BESTFIT, FASTA, and TFASTA, Madison, MN) or by visual inspection (see generally Ausubel et al., supra).

[0197] An example of an algorithm that is suitable for determining percent sequence identity and sequence similarity is the BLAST algorithm. The BLAST algorithm is described in Altschul et al., J. Mol. Biol. 215:403-410 (1990). Software for performing BLAST analyses is publicly available through the National Center for Biotechnology Information website.

[0198] As generally used herein, "pharmaceutically acceptable" refers to those compounds, substances, compositions and / or dosage forms that are, within the scope of sound medical judgment, suitable for use in contact with the tissues, organs and / or body fluids of human beings and animals without excessive toxicity, irritation, allergic response, or other problem or complication, commensurate with a reasonable benefit / risk ratio.

[0199] As used herein, "pharmaceutically acceptable carrier" refers to and includes all physiologically compatible solvents, dispersion media, coatings, antibacterial and antifungal agents, isotonic and absorption delaying agents, etc. The compositions may also contain pharmaceutically acceptable salts, such as acid addition salts or base addition salts (e.g., as described in Berge et al. (1977) J Pharm Sci 66 :1-19).

[0200] As used herein, the terms "polypeptide," "peptide," and "protein" are used interchangeably to refer to a polymer of amino acid residues. The terms apply to amino acid polymers in which one or more amino acid residues are artificial chemical mimetics of corresponding naturally occurring amino acids, and to naturally occurring amino acid polymers and non-naturally occurring amino acid polymers.

[0201] As used herein, the term "preventing," when used in reference to a condition, refers to the administration of a composition that reduces the frequency or delays the onset of a medical condition in a subject compared to a subject not administered the composition.

[0202] As used herein, the terms "purified" or "isolated" when applied to any of the proteins (antibodies or fragments) described herein refer to a polypeptide that has been separated or purified from components (e.g., proteins or other naturally occurring biological or organic molecules) that naturally accompany it in the prokaryotic organism that expresses the protein, such as other proteins, lipids, and nucleic acids. Typically, a polypeptide is purified when it constitutes at least 60% by weight (e.g., at least 65, 70, 75, 80, 85, 90, 92, 95, 97, or 99% by weight) of the total protein in a sample.

[0203] As used herein, the term "rearranged" refers to the configuration of a heavy or light chain immunoglobulin locus in which the V segment is essentially a complete V H or V L The rearranged immunoglobulin gene loci are located adjacent to the DJ or J segments in a domain-encoding structure. The rearranged locus has at least one recombined heptamer / nonamer homology element.

[0204] As used herein, the term "receptor clustering" refers to a cellular process that results in the grouping or local accumulation of a set of receptors at a specific cellular location, often inducing or amplifying a signaling response. Many protein receptors bind to their cognate ligands and cluster together, i.e., form dimers, trimers, oligomers, or multimers upon binding to their cognate ligands. For example, members of the PDGF receptor and TNF receptor superfamily form dimers and trimers, respectively, upon binding to their ligands. and form trimers. Cognate ligand-induced clustering (e.g., dimerization, multimerization) induces signaling through the receptor. Thus, an antibody or antigen-binding fragment thereof of the present disclosure can activate a receptor by binding to two or more receptors, inducing or stabilizing dimerization, trimerization, and / or multimerization with or without cognate ligand binding.

[0205] Receptor clustering and multimerization are required for TNFR signaling (Wajant (2015) Cell Death Differ 22(11):1727-1741), particularly TNFRSF activation. 4-1BB (CD137), CD40, GITR, CD27, DR3, DR5, and Fas are some of the TNFSF receptors known to require clustering to trigger downstream signaling. Experimental evidence that the 4-1BB receptor must be crosslinked to signal has been reported by Rabu et al. These authors demonstrated that the mono-trimeric form of human 4-1BBBL has an activating effect on human T cells. However, cross-linking this protein to two or more trimers resulted in strong activation of the protein (Rabu et al., (2005) J Biol Chem 280:41472-41481). Thus, in some embodiments, the anti-CD137 agonistic antibody induces multimerization of two or more trimers of CD137.

[0206] As used herein, the term "recombinant host cell" (or simply "host cell") is intended to refer to a cell into which a recombinant expression vector has been introduced. It should, of course, be understood that the term is intended to refer not only to the particular subject cell but also to the progeny of that cell. Because certain modifications may occur in subsequent generations due to either mutation or environmental influences, such progeny may not actually be identical to the parent cell, but are still included within the scope of the term "host cell" as used herein.

[0207] As used herein, the term "recombinant human antibody" refers to any human antibody that is prepared, expressed, produced, or isolated by recombinant means, e.g., (a) transgenic or transchromosomal with respect to human immunoglobulin genes; (b) an antibody isolated from a transchromosomal animal (e.g., a mouse) or a hybridoma prepared from such an animal; (b) a host cell transformed to express the antibody; (c) antibodies isolated from, for example, transfectomas; (d) antibodies isolated from a binatorial human antibody library; and prepared by any other means, including splicing the gene sequence to another DNA sequence. Recombinant human antibodies include antibodies that have been expressed, generated, or isolated. Such recombinant human antibodies contain variable and constant regions that utilize specific human germline immunoglobulin sequences, as encoded by germline genes, but also contain subsequent rearrangements and mutations that occur, for example, during antibody maturation. These antibodies can be prepared by any method known in the art (see, e.g., Lonberg (2005) Nature Biotech. 23(9):1117-1125), the variable region contains the antigen-binding domain, and The antibodies are encoded by various genes that rearrange to form antibodies specific to foreign antigens. In addition to rearrangement, the variable regions can be further modified by multiple single amino acid changes (called somatic mutations or hypermutations) to increase the affinity of the antibody for the foreign antigen. The constant regions change in further response to the antigen (i.e., isotype switching). Thus, the nucleic acid molecules that encode light and heavy chain immunoglobulin polypeptides rearranged and somatically mutated in response to an antigen may not have sequence identity to the original nucleic acid molecules, but instead are substantially identical or similar (i.e., have at least 80% identity).

[0208] As used herein, the term "reference antibody" (interchangeable with "reference mAb") The term "reference antigen binding protein" refers to the antigen binding protein on human CD137. Refers to an antibody or antigen-binding fragment thereof that binds to a specific epitope and is used to establish a relationship between itself and one or more distinct antibodies. In some embodiments, the relationship is the binding of a reference antibody and one or more distinct antibodies to the same epitope on CD137. As used herein, the term refers to an anti-CD137 antibody useful as a competitor in a test or assay such as those described herein (e.g., a competitive binding assay), in which the assay involves binding to two or more antibodies that bind to the same epitope. The variable heavy chain (V) of an exemplary reference antibody (mAb1) is useful for discovering, identifying, or developing one or more distinct antibodies that combine V H ) and variable light chain (V L The amino acid sequences of V H 1, SEQ ID NO: 4; V H 2, SEQ ID NO: 6). In some embodiments, the term refers to a comparator. and in this case, the present invention is intended to imply an anti-CD137 antibody useful in a test or assay as a In some embodiments, the assay is useful for identifying antibody characteristics (e.g., hepatotoxicity, anti-tumor efficacy). In some embodiments, the reference antibody is urelumab. In some embodiments, the reference antibody is utomilumab.

[0209] As used herein, the terms "specific binding," "selective binding," "selectively binds," and "specifically binds" refer to an antibody that binds to an epitope on a specified antigen. Typically, the antibody binds to recombinant human CD137 as the analyte and to a target antigen as the ligand. The antibody was used to detect approximately 10 -6 Less than M, e.g., about 10 -7 Under M, 10 -8 Under M, 10 -9 Less than M or 10 -10 The equilibrium dissociation constant (K D ) and have a higher affinity for binding to nonspecific antigens (e.g., BSA, casein, etc.) other than the given antigen or closely related antigens. Binds to a given antigen with at least two times higher affinity. and "antibody specific for an antigen" are used interchangeably herein with the term "antibody that specifically binds to an antigen."

[0210] As used herein, the term "switch sequence" refers to a DNA sequence that participates in switch recombination. A "switch donor" sequence, typically a μ switch region, The "switch" region is 5' (i.e., upstream) of the construct region that is deleted during switch recombination. The "acceptor" region is located between the region of the construct to be deleted and the replacement constant region (e.g., γ, ε, etc.). The final gene sequence is usually not predictable from the construct because there are no specific sites where recombination always occurs.

[0211] As used herein, the term "subject" includes any human or non-human animal. For example, the methods and compositions of the present invention can be used to treat subjects with immune disorders. The term "non-human animal" includes all vertebrates, such as mammals and non-mammals, including non-human primates, sheep, dogs, cows, chickens, amphibians, reptiles, etc.

[0212] With respect to nucleic acids, the term "substantial homology" indicates that two nucleic acids or designated sequences, when optimally aligned and compared, are identical in at least about 80% of the nucleotides, usually at least about 90%-95% of the nucleotides, and more preferably at least about 98%-99.5% of the nucleotides, with appropriate nucleotide insertions or deletions. Reciprocal homology exists when the segments will hybridize under selective hybridization conditions, to the complement of the strand.

[0213] The percentage identity between two sequences is a function of the number of identical positions shared by the sequences (i.e., % homology = # of identical positions / total # of positions x 100), taking into account the number of gaps and the length of each gap that need to be introduced for optimal alignment of the two sequences. The comparison of sequences and determination of the percentage identity between two sequences can be performed using mathematical algorithms, such as those described in the non-limiting examples below. It may be performed using rhythm.

[0214] The percent identity between two nucleotide sequences may be determined using the GAP program in the GCG software package (available at http: / / www.gcg.com) using the NWSgapdna.CMP matrix and gap weights of 40, 50, 60, 70, or 80, and length weights of 1, 2, 3, 4, 5, or 6. The percent identity between two nucleotide or amino acid sequences may also be determined using the algorithm of E. Meyers and W. Miller (CABIOS, 4:11-17 (1989)) incorporated into the ALIGN program (version 2.0), using a PAM120 weight residue table, a gap length penalty of 12, and a gap penalty of 4. Additionally, two The percentage identity between the amino acid sequences of the nucleotides was calculated using the GCG software package (http: / / www.gcg.com (available at http: / / www.ncbi.nlm.nih.gov / pubmed / 20170233) incorporated into the GAP program of Needleman and Wunsch (J. Mol. Biol. (48):444-453 (1970)) algorithm, using either a Blossum 62 matrix or a PAM250 matrix, and gap weights of 16, 14, 12, 10, 8, 6, or 4, and length weights of 1, 2, 3, 4, 5, or 6.

[0215] The nucleic acid and protein sequences of the present disclosure may also be used as a "query sequence" to conduct a search against public databases, for example, to identify related sequences. Such searches may be performed using the NBLAST and XBLAST programs (version 2.0) of Altschul, et al. (1990) J. Mol. Biol. 215:403-10. BLAST sequences may be used to search for related sequences. Nucleotide searches were performed using the NBLAST program, score=100, word length=12, to identify the nucleotide sequences of the present invention. Nucleotide sequences homologous to the nucleic acid molecule may be obtained. BLAST protein searches may be performed using XBLAST The program was run with score=50 and word length=3 to identify amino acids homologous to the protein molecules of the present invention. To obtain gapped alignments for comparison purposes, Gapped BLAST may be used as described in Altschul et al., (1997) Nucleic Acids Res. 25(17):3389-3402. BLAST and Gapped BLAST programs are used. In some cases, the default parameters of each program (e.g., XBLAST and NBLAST) may be used. See http: / / www.ncbi.nlm.nih.gov.

[0216] The nucleic acids may be present in whole cells, in a cell lysate, or may be present in a partially purified or substantially pure form. They may be purified from other cellular components or other contaminants, such as other cellular nucleic acids or proteins, by alkaline / SDS treatment, CsCl banding, column chromatography, or other methods. Nucleic acid is "isolated" or "substantially pure" when purified by standard techniques, including chromatography, agarose gel electrophoresis, and other methods known in the art. See F. Ausubel, et al., eds. Current Protocols in Molecular Biology, Greene Publishing and Wiley Interscience, New York (1987).

[0217] Nucleic acid compositions of the present disclosure are often naturally occurring sequences (excluding modified restriction enzyme sites, etc.) from cDNA, genomes, or mixtures thereof, but may be mutated according to standard techniques to obtain gene sequences. With respect to coding sequences, these mutations may have desirable effects on the amino acid sequence. In particular, naturally occurring V sequences, D sequences, J sequences, constant sequences, switch sequences, and sequences described herein may be mutated. DNA sequences that are substantially homologous to or derived from other such sequences are contemplated (where "derived" indicates that the sequence is identical to or modified from another sequence).

[0218] As used herein, the term "tumor microenvironment" (or "cancer microenvironment," TME) refers to a The term TME (abbreviated as TME) refers to the cellular environment or milieu in which a tumor or neoplasm resides, including surrounding blood vessels and non-cancerous cells, including but not limited to immune cells, fibroblasts, bone marrow-derived inflammatory cells, and lymphocytes. Signaling molecules and the extracellular matrix also comprise the TME. Tumors and the surrounding microenvironment are closely related and constantly interact. Tumors can influence the microenvironment by releasing extracellular signals, promoting tumor angiogenesis, and inducing immune tolerance in peripheral tissues. Meanwhile, immune cells in the microenvironment can influence the proliferation and evolution of tumor cells.

[0219] The term "T cell" refers to a type of white blood cell that can be distinguished from other white blood cells by the presence of a T cell receptor on the cell surface. There are several subsets of T cells, including but not limited to: However, T helper cells (T H cells or CD4 + T cells) and T H 1. T H 2. T H 3 , T H 17, T H 9 and T FH cells and their subtypes, including cytotoxic T cells (i.e., T C cells, CD8 + T cells, cytotoxic T lymphocytes, T-killer cells, killer T cells), memory T cells, and Central memory T cells (T CM cells), effector memory T cells (T EM and T EMRA cell) and resident memory T cells (T RM cells), and their subtypes, including regulatory T cells (T reg cell or suppressor T cells) and CD4 + FOXP3 + Treg cells, CD4 + FOXP3 - T reg cells, Tr1 cells, Th3 cells and T reg 17 cells and their subtypes, natural killer T cells (NKT cells) T cells not mentioned above or mentioned include mucosal-associated invariant T cells (MAITs), as well as gamma delta T cells (γδ T cells), including Vγ9 / Vδ2 T cells. Any one or more of the cells may be the target cell type for the methods of use of the present invention.

[0220] As used herein, the term "T cell activation" or "activation of T cells" refers to the cellular process by which mature T cells, expressing antigen-specific T cell receptors on their surface, recognize their cognate antigen and respond by entering the cell cycle, secreting cytokines or lytic enzymes, and initiating or becoming competent to exert cellular effector functions. T cell activation involves the activation of a cell by at least two signals. The first signal occurs after engagement of the T cell antigen-specific receptor (TCR) with the antigen-major histocompatibility complex (MHC), and the second signal occurs after engagement of the T cell antigen-specific receptor (TCR). These signals are transmitted to the nucleus, leading to T cell clonal expansion and upregulation of activation markers on the cell surface. The effect of the antibody is to induce cytotoxicity, differentiation into effector cells, induction of cytotoxic activity or cytokine secretion, induction of apoptosis, or a combination thereof.

[0221] As used herein, the term "T cell mediated response" includes, but is not limited to, an effector Inhibitor T cells (e.g., CD8 + cells) and helper T cells (e.g., CD4 + T cells, including T cell-mediated responses include, for example, T cell cytotoxicity and proliferation.

[0222] As used herein, the term "therapeutically effective amount" or "therapeutically effective dosage" or similar terms used herein refers to a therapeutically effective amount of an agent (e.g., an anti-CD137 antibody or antigen-binding It is intended to mean the amount of a fragment of a substance.

[0223] As used herein, the terms "treat," "treating," and "treatment" refer to therapeutic or prophylactic measures as described herein. "Treatment" methods employ administering a human antibody of the present disclosure to a subject in need of such treatment, e.g., a subject in need of an enhanced immune response to a particular antigen, or a subject who may ultimately acquire immunity to such a disorder and thereby prevent, cure, delay, reduce the severity of, or ameliorate one or more symptoms of the disorder, or prolong the subject's survival beyond that expected in the absence of such treatment.

[0224] As used herein, the term "unrearranged" or "germline configuration" refers to an arrangement in which a V segment is immediately adjacent to a D or J segment. Refers to a configuration that has not undergone any changes.

[0225] As used herein, the term "vector" is intended to refer to a nucleic acid molecule capable of transporting another nucleic acid to which it has been linked. One type of vector is a "plasmid," which refers to a circular double-stranded DNA loop into which additional DNA segments can be ligated. Another type of vector is a viral vector, in which additional DNA segments can be ligated into the viral genome. Certain vectors are self-contained. Some vectors are capable of autonomous replication in a host cell into which they are introduced (e.g., bacterial vectors having a bacterial origin of replication and episomal mammalian vectors). Other vectors (e.g., non-episomal mammalian vectors) can be integrated into the genome of a host cell upon introduction into the host cell, and thereby are replicated along with the host genome. Moreover, certain vectors are capable of directing the expression of genes to which they are operably linked. Such vectors are referred to herein as "recombinant expression vectors" (or simply "expression vectors"). Generally, the practical form of expression vectors in recombinant DNA technology is often In the form of a plasmid. As used herein, "plasmid" and "vector" can be used interchangeably as the plasmid is the most commonly used form of vector. However, the invention is intended to include other forms of expression vectors that serve equivalent functions, such as viral vectors (e.g., replication defective retroviruses, adenoviruses and adeno-associated viruses).

[0226] Unless otherwise specified, all technical and scientific terms used herein have the same meaning as commonly understood by those skilled in the art to which this application belongs. Preferred methods and materials are described below, but similar or equivalent methods and materials to those described herein can also be used to carry out or test the disclosed methods and compositions. All publications, patent applications, patents, and other references mentioned herein are incorporated by reference in their entirety.

[0227] Anti-CD137 antibodies and antigen-binding fragments thereof The present disclosure provides antibodies that specifically bind to and agonize CD137. Thus, the present disclosure provides anti-CD137 agonistic antibodies useful in the treatment of cancer. In some embodiments, the anti-CD137 agonistic antibody induces cytokine production. In some embodiments, anti-CD137 agonistic antibodies increase the number of CD8+ T cells in the tumor microenvironment. In some embodiments, anti-CD137 agonistic antibodies induce protective anti-tumor immunity. The present disclosure further provides a method for the treatment of splenic or hepatic CD4+ T cell populations and / or The present invention provides an anti-CD137 agonistic antibody that does not substantially increase the CD8+ T cell population or CD8+ T cell population.

[0228] Human CD137 is a 255 amino acid transmembrane polypeptide (SEQ ID NO: 3; Accession Nos. NM_001561; NP_001552) and is a phylogenetically conserved tumor necrosis factor receptor (TNFR) receptor. CD137 (also known as 4-1BB, TNFR superfamily 9) and its ligand (CD137L) are involved in the regulation of a wide range of immune activities. crosslinks its receptor, CD137, expressed on activated T cells and costimulates T cell activity. CD137 is an activation-induced costimulatory molecule. Recent studies have shown that CD137-mediated anti-cancer effects are due to its ability to activate T cells, particularly to induce cytotoxic T lymphocyte (CTL) responses. It has been shown that the ability to induce cytokine production, especially high levels of IFNγ, is crucial for the survival of the immune system. It was revealed (Ye et al., (2014) Clin Cancer Res 20(1):44-55). CD137 ligand is a transmembrane protein on the cell surface that transmits signals into the cell in which it is expressed (a phenomenon called "reverse signaling" or "back signaling"). Expression of the CD137 ligand is found on most types of leukocytes and some non-immune cells. In monocytic cells (monocytes, macrophages, and DCs), the CD137 ligand is expressed. Signal transmission induces activation, migration, survival, and differentiation.

[0229] Thus, in some embodiments, the isolated anti-CD137 agonistic antibodies described herein or an antigen-binding fragment thereof that binds to CD137, agonizes CD137, and enables or promotes binding of CD137L. Agonistic antibodies, or antigen-binding fragments thereof, bind to and agonize CD 137. In some embodiments, the anti-CD137 antibodies provided by the present disclosure bind to and agonize CD137 to costimulate T cell activation.

[0230] In some embodiments, an isolated anti-CD137 agonistic antibody described herein, or The antigen-binding fragment of has one or more of the following properties or characteristics: a) specifically binds to human CD137; b) binds to human and cynomolgus monkey CD137; and c) Binds to human and mouse CD137.

[0231] In some embodiments, an anti-CD137 agonistic antibody described herein, or an anti-CD137 antibody thereof The anti-CD137 agonistic antibody or antigen-binding fragment thereof described herein binds to CD137 and costimulates T cell activity. In some embodiments, the anti-CD137 agonistic antibody or antigen-binding fragment thereof described herein binds to CD137 and induces or enhances T cell activation, a cytotoxic T lymphocyte (CTL) response, T cell proliferation, cytokine production, or a combination thereof. In some embodiments, the anti-CD137 agonistic antibody or antigen-binding fragment thereof described herein binds to CD137 and induces or enhances T cell activation, a cytotoxic T lymphocyte (CTL) response, T cell proliferation, cytokine production, or a combination thereof in the tumor microenvironment. In some embodiments, the anti-CD137 antibody or antigen-binding fragment thereof described herein is intrahepatically and / or intrasplenicly In some embodiments, the anti-CD137 antibodies described herein bind to CD137 and induce the production of IFNγ, but do not significantly induce or enhance T cell activation and / or T cell proliferation. In some embodiments, the antibodies provided by the present disclosure bind to CD137 and inhibit IL-2, TNF- Induce the production of IL-α, IL-13, or a combination thereof.

[0232] In some embodiments, the anti-CD137 antibodies described herein specifically bind to and agonize CD137. In some embodiments, CD137 agonism is measured by determining the concentration of cytokines produced by immune cells. Methods for analyzing cytokine production are known in the art and are utilized in the Examples. In some embodiments, increased cytokine production by immune cells indicates CD137 agonism. In some embodiments, In some embodiments, CD137 agonism is measured by analyzing T cell proliferation. In some embodiments, increased T cell proliferation indicates CD137 agonism. Agonism of CD137 is measured by measuring the level of cell signaling either through quantification of phosphorylation of associated molecules or quantification of expression of a gene reporter following an associated promoter. In some embodiments, increased cell signaling is indicative of agonism of CD137. In some embodiments, CD137 agonism is measured by measuring tumor volume. In some embodiments, a decrease in tumor volume is indicative of CD137 agonism.

[0233] In some embodiments, the anti-CD137 antibodies described herein induce, increase, or stabilize the oligomerization, multimerization, or other higher order clustering of CD137. In some embodiments, the clustering of CD137 on the cell surface is observed via fluorescence microscopy.

[0234] As used herein, an isolated monoclonal antibody or antibody thereof that binds to and agonizes CD137 is In some embodiments, the antibody or antigen-binding fragment thereof has (i) an affinity (K D ), (ii) binds to an epitope of human CD137 described herein, and / or (iii) comprises a heavy chain CDR3 comprising the amino acid sequence DXXXXLXXXXYXYYX (SEQ ID NO: 126).

[0235] Affinity for CD137 In some embodiments, an isolated anti-CD137 agonistic antibody described herein or The antigen-binding fragment has an antigen binding activity of about 30 to 100 nM (e.g., about 30 nM to about 100 nM or about 40 nM to about 100 nM). Infinity (K D ) binds to human CD137. In some embodiments, the affinity of the anti-CD137 antibody for human CD137 is at least 2-fold (e.g., at least 3, 4, 5, 6, 7, 8, 9, or 10-fold) higher than the affinity of mAb10 for mouse CD137. In some embodiments, the affinity of the anti-CD137 antibody is 500, 450, 400, 350, 300, 250, 200, 250, 200, 175, 150, 125, 110, or 100 nM or less. In some embodiments, the affinity of the anti-CD137 antibody for human CD137 is at least 2-fold (e.g., at least 3, 4, 5, 6, 7, 8, 9, or 10-fold) higher than the affinity of mAb10 for mouse CD137. (if at least 3, 4, 5, 6, 7, 8, 9 or 10 times higher) , 200, 250, 200, 175, 150, 125, 110, or 100 nM or less. The affinity of an antibody is the strength of binding to a single CD137 polypeptide. In some embodiments, the affinity is the equilibrium dissociation constant (K D ) is shown by K DThe value is inversely related to the binding affinity of the antibody for the antigen. D The lower the value, the higher the affinity of the antibody for its antigen.

[0236] Methods for determining the affinity of an antibody for its antigen are known in the art. An exemplary method for determining binding affinity employs surface plasmon resonance. Surface plasmon resonance is an optical phenomenon that allows for the analysis of biospecific interactions in real time by detecting changes in protein concentration within a biosensor matrix, for example, using the BIAcore system (Pharmacia Biosensor AB, Uppsala, Sweden, and Piscataway, New Jersey). For further details, see Jonsson, U., et al. (1993) Ann. Biol. Clin. 51: 19-26; Jonsson, U., i (1991) Biotechniques 11:620-627; Johnsson, B., et al. (1995) J. Mol. Recognit. 8: 125-131; and and Johnson, B., et al. (1991) Anal. Biochem. 198:268-277.

[0237] In some embodiments, the anti-CD137 antibodies described herein have an affinity (K D In some embodiments, the anti-CD137 antibodies described herein bind to human CD137 with an affinity (K) of about 40-100 nM. D ) binds to human CD137 In some embodiments, the anti-CD137 antibodies described herein have an affinity (K D ) binds to human CD137. In embodiments, the anti-CD137 antibodies described herein have an affinity (K D )in In some embodiments, the anti-CD137 antibodies described herein bind to human CD137 with an affinity (K D ) binds to human CD137.

[0238] In some embodiments, the anti-CD137 antibodies described herein have an affinity of about 60-90 nM. (K D In some embodiments, the anti-CD137 antibodies described herein bind to human CD137 with an affinity (K) of about 50-80 nM. D ) binds to human CD137. In some embodiments, The anti-CD137 antibodies described herein have an affinity (K D In some embodiments, the anti-CD137 antibodies described herein bind to human CD137 at an affinity of about 55-75 nM. Infinity (K D In some embodiments, the anti-CD137 antibodies described herein bind to human CD137 with an affinity (K) of about 65-75 nM. D ) binds to human CD137. Therefore, the anti-CD137 antibodies described herein have an affinity (K D In some embodiments, the anti-CD137 antibodies described herein bind to human CD137 at a binding affinity of about 55-85 nM. Affinity (K D ) binds to human CD137. In some embodiments, The anti-CD137 antibodies used have an affinity (K D In some embodiments, the anti-CD137 antibodies described herein bind to human CD137 with an affinity (K) of about 45-95 nM. D )in In some embodiments, the anti-CD137 antibodies described herein bind to human CD137 with an affinity (K D ) binds to human CD137. The anti-CD137 antibodies described have an affinity (K D In some embodiments, the anti-CD137 antibodies described herein bind to human CD137 with an affinity (K) of about 75-85 nM. D ) binds to human CD137. In some embodiments, the anti-CD137 antibodies described herein , with an affinity (K D ) binds to human CD137. The anti-CD137 antibodies described herein have an affinity (K D In some embodiments, the anti-CD137 antibodies described herein bind to human CD137 with an affinity of about 80-90 nM. Tee (K D In some embodiments, the anti-CD137 antibodies described herein bind to human CD137 with an affinity (K) of about 70-80 nM. D ) binds to human CD137. In some embodiments, The anti-CD137 antibodies described herein have an affinity (K D In some embodiments, the anti-CD137 antibodies described herein bind to human CD137 at an affinity of about 50-60 nM. Infinity (K D ) binds to human CD137. In some embodiments, Anti-CD137 antibodies have an affinity (K D In some embodiments, the anti-CD137 antibodies described herein bind to human CD137 with an affinity (K) of about 30-40 nM. D In some embodiments, the anti-CD137 antibodies described herein bind to human CD137 at a binding affinity of about 30 nM, About 31nM, about 32nM, about 33nM, about 34nM, about 35nM, about 36nM, about 37nM, about 38nM, about 39nM, about 40nM, about 41nM, about 42nM, about 43nM, about 44nM, about 45nM, about 46nM, about 47nM, about 48 nM, approximately 49nM, approximately 50nM, approximately 51nM, approximately 52nM, approximately 53nM, approximately 54nM, approximately 55nM, approximately 56nM, approximately 57nM, approximately 58nM, approximately 59nM, approximately 60nM, approximately 61nM, approximately 62nM, approximately 63nM, approximately 64nM, approximately 65nM, Approximately 66nM, approximately 67nM, approximately 68nM, approximately 69nM, approximately 70nM, approximately 71nM, approximately 72nM, approximately 73nM, approximately 74nM, approximately 75nM, approximately 76nM, approximately 77nM, approximately 78nM, approximately 79nM, approximately 80nM, approximately 81nM, approximately 82nM, approximately 83 nM, approximately 84nM, approximately 85nM, approximately 86nM, approximately 87nM, approximately 88nM, approximately 89nM, approximately 90nM, approximately 91nM, approximately 92nM, approximately 93nM, approximately 94nM, approximately 95nM, approximately 96nM, approximately 97nM, approximately 98nM, approximately 99nM, approximately 100nM , about 101 nM, about 102 nM, about 103 nM, about 104 nM, about 105 nM, about 106 nM, about 107 nM, about 108 nM, about 109 nM or about 110 nM D ) binds to human CD137.

[0239] In some embodiments, the anti-CD137 antibodies described herein have a IgG antibody titer of at least 30 nM. , less than about 110 nM, at least 31 nM but less than about 109 nM, at least 32 nM but less than about 108 nM, at least 33 nM but less than about 107 nM, at least 34 nM but less than about 106 nM at least 35 nM but less than about 105 nM; at least 36 nM but less than about 104 nM; at least 37 nM but less than about 103 nM; at least 38 nM but less than about 102 nM; at least 39 nM but less than about 101 nM; at least 40 nM but less than about 100 nM; at least 41 nM at least 42 nM but less than about 98 nM, at least 43 nM but less than about 97 nM, at least 44 nM but less than about 96 nM, at least 45 nM but less than about 95 nM, at least 46 nM but less than about 94 nM, at least 47 nM but less than about 93 nM, at least 48 nM but less than about 92 nM, at least 49 nM but less than about 91 nM, at least 50 nM but less than about 90 nM, at least 51 nM but less than about 89 nM, at least 52 nM but less than about 88 nM, at least 53 nM but less than about 87 nM, An affinity (K) of at least 54 nM but less than about 86 nM, at least 55 nM but less than about 85 nM, at least 56 nM but less than about 84 nM, at least 57 nM but less than about 83 nM, at least 58 nM but less than about 82 nM, at least 59 nM but less than about 81 nM, at least 60 nM but less than about 80 nM, at least 61 nM but less than about 79 nM, at least 62 nM but less than about 78 nM, at least 63 nM but less than about 77 nM, at least 64 nM but less than about 76 nM, or at least 65 nM but less than about 75 nM. D ) binds to human CD137. In some embodiments, The anti-CD137 antibodies used have an affinity (K) of at least 40 nM but less than about 100 nM. D ) and binds to human CD137.

[0240] In some embodiments, the anti-CD137 antibodies described herein are directed to CD137 antibodies from two or more species. In some embodiments, the anti-CD137 antibodies described herein cross-react with the polypeptide. binds to cynomolgus monkey CD137 and human CD137. In some embodiments, the anti-CD137 antibodies described herein bind to mouse CD137 and human CD137. The anti-CD137 antibodies described herein are directed against human CD137, mouse CD137, and cynomolgus monkey CD137. It binds to CD137 on monkeys.

[0241] CD137 epitope binding In some embodiments, an isolated monoclonal antibody or antibody thereof that specifically binds to human CD137. The antigen-binding portion of SEQ ID NO: 3 may be one or more (e.g., 1, 2, 3, 4) amino acids 111 to 132 of SEQ ID NO: 3. , 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24 and In some embodiments, the isolated monoclonal antibody or antigen-binding portion thereof that specifically binds to human CD137 binds to an epitope on human CD137, including all 25 of SEQ ID NO:3. In some embodiments, the present disclosure provides an isolated monoclonal antibody or antigen-binding portion thereof that specifically binds to human CD137. and the antibody or antigen-binding portion comprises all of amino acids 111 to 132 of SEQ ID NO: 3 or In some embodiments, the isolated anti-CD137 agonistic antibody described herein binds to a portion of the CD137 antibody. The antibody, or antigen-binding fragment thereof, binds to an epitope of human CD137 that includes the K114 residue of SEQ ID NO: 3. In some embodiments, the isolated anti-CD137 agonistic antibody described herein or an antigen-binding fragment thereof binds to an epitope of human CD137 that includes residues E111, T113, and K114 of SEQ ID NO: 3. In some embodiments, the isolated anti-CD137 antigen described herein is The cytosinic antibody, or antigen-binding fragment thereof, is a nucleotide sequence encoding E111, T113, K114, N126 and SEQ ID NO: 3. In some embodiments, the antibodies described herein bind to an epitope on human CD137 that includes residues I132 and I133. The described isolated anti-CD137 agonistic antibodies, or antigen-binding fragments thereof, bind to epitopes of human CD137 including E111, T113, K114, N126, I132 and P135 of SEQ ID NO:3. In embodiments, an isolated anti-CD137 agonistic antibody described herein, or an antigen thereof The binding fragment may be one or more of E111, T113, K114, N126, I132 and P135 of SEQ ID NO:3. It binds to an epitope on human CD137 containing residues

[0242] In some embodiments, an isolated anti-CD137 agonistic antibody described herein or The antigen-binding fragment binds to an epitope of human CD137 comprising a sequence of one or more amino acid residues corresponding to amino acid positions 100 to 135, 101 to 135, 102 to 135, 103 to 135, 104 to 135, 105 to 135, 106 to 135, 107 to 135, 108 to 135, 109 to 135, 110 to 135, or 111 to 135 of SEQ ID NO:3.

[0243] In some embodiments, an isolated anti-CD137 agonistic antibody described herein or The antigen-binding fragment may comprise one or more amino acids corresponding to amino acid positions 111 to 135 of SEQ ID NO:3. In some embodiments, the epitope binds to an epitope on human CD137 that comprises a sequence of acid residues. The fragments are 2, 3, 4, 5, 6, 7, 8, 9, 10, and 11, which correspond to the amino acids at positions 111 to 135 of SEQ ID NO: 3. , 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24 or 25 amino acid residues.

[0244] In some embodiments, an isolated anti-CD137 agonistic antibody described herein or The antigen-binding fragment binds to an epitope of human CD137 within amino acid positions 100-135, 101-135, 102-135, 103-135, 104-135, 105-135, 106-135, 107-135, 108-135, 109-135, 110-135, or 111-135 of SEQ ID NO: 3. In some embodiments, the isolated anti-CD137 agonistic antibody described herein, or antigen-binding fragment thereof, binds to an epitope of human CD137 within amino acid positions 111-135 of SEQ ID NO: 3. In some embodiments, the epitope is 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, and 13, which correspond to amino acids 111 to 135 in column 3 , 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24 or 25 amino acid residues.

[0245] In some embodiments, an isolated anti-CD137 agonistic antibody described herein, or The antigen-binding fragment binds to an epitope of human CD137 that includes ELTK (corresponding to amino acid residues 111 to 114 of SEQ ID NO: 3). In some embodiments, amino acid residue L112 can be another amino acid residue.

[0246] In some embodiments, the epitope is a non-linear epitope. In some embodiments, the mutation at amino acid residue K114 is a non-linear epitope of an isolated anti-CD137 agonistic antibody described herein, or or its antigen-binding fragment to human CD137.

[0247] In some embodiments, an isolated anti-CD137 agonistic antibody described herein or The antigen-binding fragment may comprise one or more amino acids corresponding to amino acid positions 111 to 135 of SEQ ID NO:3. Binds to an epitope of human CD137 that contains a sequence of acid residues, wherein the epitope The antibody or antigen-binding portion thereof comprises at least amino acid K114, and the antibody or antigen-binding portion thereof binds to mouse CD137 but not to rat CD137. In some embodiments, the epitope is a non-linear epitope. In some embodiments, the antibody or antigen-binding portion thereof binds to mouse CD137 and cynomolgus monkey CD137 but not to rat CD137. In embodiments, an isolated anti-CD137 agonistic antibody or antigen-binding fragment thereof described herein Binding of the fragments to human, mouse, rat, and cynomolgus monkey CD137 was measured by surface plasmon resonance (SPMR). (SPR) is used to determine the

[0248] In some embodiments, the antibody or antigen-binding portion thereof is an antibody directed against rat CD137. At least 10, 20, 30, 40, 50, 100, 200, 500 or 1000 times higher than the affinity In some embodiments, the antibody binds to mouse, cynomolgus monkey, or human CD137 with high affinity. The antibody or antigen-binding portion thereof comprises a lysine at position 114 relative to human CD137 of SEQ ID NO: 3. to mouse, cynomolgus monkey, or human CD137 with affinity at least 10, 20, 30, 40, 50, 100, 200, 500, or 1000 times higher than the affinity of the antibody to rat CD137 Combine.

[0249] In some embodiments, an isolated anti-CD137 agonistic antibody described herein, or The antigen-binding fragment of mAb1 binds to an epitope on human CD137 and competes with mAb1 for binding to an epitope on human CD137. In some embodiments, the isolated anti-CD137 antigen described herein is Agonistic antibodies, or antigen-binding fragments thereof, bind to and agonize CD137. In some embodiments, the anti-CD137 antibodies provided by the present disclosure bind to and agonize CD137. agonizes and co-stimulates T cell activation.

[0250] The present disclosure relates to antibodies that bind to epitopes on CD137, including all or part of the epitopes recognized by one or more specific reference antibodies (e.g., mAb1) described herein. In some embodiments, the anti-CD137 antibody binds to an epitope on human CD137 and competes with a reference antibody (e.g., mAb1) for binding to an epitope on human CD137, wherein the antibody or antigen-binding fragment thereof is at least 1x10 -6 The following equilibrium Dissociation constant K D In some embodiments, the anti-CD137 antibody binds to human CD137 at an endogenous site on CD137. In some embodiments, the antibody binds to an epitope, where one or more mutations in the epitope inhibit, reduce, or prevent binding to both the antibody and a reference antibody (e.g., mAb1). In some embodiments, the reference antibody is the mAb1 antibody described herein. In some embodiments, the reference antibody is any one of the antibodies presented in Table 3 or Table 4.

[0251] Accordingly, the anti-CD137 antibodies provided by the present disclosure may be evaluated via x-ray crystallography of a crystal structure comprising the antibody, or a fragment or portion thereof, bound to CD137. In embodiments, the epitopes bound by the antibodies provided by the present disclosure are those present or located within 4 angstroms (Å) of the paratope residues of an antibody, e.g., mAb1. It is identified by determining the residues on the CD137 antigen.

[0252] In some embodiments, the epitope bound by an anti-CD137 antibody described herein is At least three amino acid residues. In some embodiments, the epitope bound by an anti-CD137 antibody described herein is at least four amino acid residues. In some embodiments, The epitopes bound by the anti-CD137 antibodies described herein are at least 5 amino acid residues. In some embodiments, the epitopes bound by the anti-CD137 antibodies described herein are at least 5 amino acid residues. In some embodiments, the epitope bound by an anti-CD137 antibody described herein is at least 7 amino acid residues. In some embodiments, the epitope bound by the anti-CD137 antibodies described herein is at least 8 amino acid residues. In some embodiments, the pitope is at least 9 amino acid residues. The epitope bound by the anti-CD137 antibody is at least 10 amino acid residues. In embodiments, the epitope bound by the anti-CD137 antibodies described herein is at least In some embodiments, the nucleotide sequence of ... The combined epitope is at least 3 amino acid residues. The epitope bound by the anti-CD137 antibody described herein is at least 13 amino acid residues. In some embodiments, the epitope bound by an anti-CD137 antibody described herein is , at least 14 amino acid residues. In some embodiments, the epitope bound by an anti-CD137 antibody described herein is at least 15 amino acid residues.

[0253] In some embodiments, the epitope bound by an anti-CD137 antibody described herein is fewer than 25 amino acid residues. In some embodiments, the epitope bound by an anti-CD137 antibody described herein is fewer than 24 amino acid residues. In some embodiments, the epitope bound by an anti-CD137 antibody described herein is fewer than 23 amino acid residues. In some embodiments, the epitope bound by the anti-CD137 antibodies described herein is In some embodiments, the amino acid residues bound to the anti-CD137 antibodies described herein are less than 100% amino acid residues. The epitope bound by an anti-CD137 antibody described herein is fewer than 21 amino acid residues. In some embodiments, the epitope bound by an anti-CD137 antibody described herein is fewer than 20 amino acid residues. In one embodiment, the epitope bound by the anti-CD137 antibodies described herein is 19 amino acids. In some embodiments, the anti-CD137 antibodies described herein bind to In some embodiments, the epitope bound by an anti-CD137 antibody described herein is fewer than 17 amino acid residues. In embodiments, the epitope bound by the anti-CD137 antibodies described herein is a 16 amino acid sequence. In some embodiments, the hydroxyl group conjugated to the anti-CD137 antibodies described herein has fewer than 100 acid residues. In some embodiments, the epitope bound by an anti-CD137 antibody described herein is fewer than 14 amino acid residues. In embodiments, the epitope bound by the anti-CD137 antibodies described herein is a 13 amino acid In some embodiments, the nucleotide sequence of ... In some embodiments, the epitope bound by an anti-CD137 antibody described herein is fewer than 11 amino acid residues. In this form, the epitope bound by the anti-CD137 antibodies described herein is a 10 amino acid residue In some embodiments, the hydroxyl group attached to the anti-CD137 antibody described herein is less than 1. The epitope is fewer than 9 amino acid residues. In some embodiments, the epitope bound by an anti-CD137 antibody described herein is fewer than 8 amino acid residues. In some embodiments, the epitope bound by an anti-CD137 antibody described herein is fewer than 7 amino acid residues. In some embodiments, the epitope bound by an anti-CD137 antibody described herein is fewer than 9 amino acid residues. In some embodiments, the epitope bound by an anti-CD137 antibody described herein is fewer than 5 amino acid residues.

[0254] In some embodiments, the anti-CD137 antibodies described herein are , 19, 18, 17, 16, 15, 14, 13, 12, 11, 10, 9, 8, 7, 6 or fewer than 5 amino acids It binds to an epitope comprising amino acid residue K114 of SEQ ID NO:3.

[0255] Variable region In some embodiments, provided herein is an isolated monoclonal antibody or antigen-binding fragment thereof comprising the heavy and light chain variable sequences set forth in Tables 3 and 4.

[0256] In some embodiments, the anti-CD137 antibody described herein is selected from the group consisting of: The heavy and light chain CDRs include: (a) heavy chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 48, 56 and 68, respectively, and light chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 69, 78 and 89, respectively; (b) heavy chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 48, 56 and 68, respectively; and light chain CDR1, CDR2 and CDR3 set forth in SEQ ID NOs: 70, 79 and 90, respectively; (c) heavy chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 48, 56 and 68, respectively; and light chain CDR1, CDR2 and CDR3 set forth in SEQ ID NOs: 71, 80 and 91, respectively; (d) heavy chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 48, 56 and 68, respectively; and light chain CDR1, CDR2 and CDR3 set forth in SEQ ID NOs: 72, 81 and 92, respectively; (e) heavy chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 48, 56 and 68, respectively; and light chain CDR1, CDR2 and CDR3 set forth in SEQ ID NOs: 73, 82 and 91, respectively; (f) heavy chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 48, 56 and 68, respectively; and light chain CDR1, CDR2 and CDR3 set forth in SEQ ID NOs: 74, 83 and 93, respectively; (g) heavy chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 48, 56 and 68, respectively; and light chain CDR1, CDR2 and CDR3 set forth in SEQ ID NOs: 75, 84 and 91, respectively; (h) heavy chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 48, 56 and 68, respectively; and light chain CDR1, CDR2 and CDR3 set forth in SEQ ID NOs: 74, 85 and 94, respectively; (i) heavy chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 48, 56 and 68, respectively; and light chain CDR1, CDR2 and CDR3 set forth in SEQ ID NOs: 76, 86 and 95, respectively; (j) heavy chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 48, 56 and 68, respectively; and light chain CDR1, CDR2 and CDR3 set forth in SEQ ID NOs: 77, 87 and 93, respectively; (k) heavy chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 48, 56 and 68, respectively; and light chain CDR1, CDR2 and CDR3 set forth in SEQ ID NOs: 69, 88 and 90, respectively; (l) heavy chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 49, 57 and 68, respectively; and light chain CDR1, CDR2 and CDR3 set forth in SEQ ID NOs: 69, 78 and 89, respectively; (m) heavy chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 49, 58 and 68, respectively; and light chain CDR1, CDR2 and CDR3 set forth in SEQ ID NOs: 69, 78 and 89, respectively; (n) heavy chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 49, 59 and 68, respectively; and light chain CDR1, CDR2 and CDR3 set forth in SEQ ID NOs: 69, 78 and 89, respectively; (o) heavy chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 49, 60 and 68, respectively; and light chain CDR1, CDR2 and CDR3 set forth in SEQ ID NOs: 69, 78 and 89, respectively; (p) heavy chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 50, 61 and 68, respectively; and light chain CDR1, CDR2 and CDR3 set forth in SEQ ID NOs: 69, 78 and 89, respectively; (q) heavy chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 50, 58 and 68, respectively; and light chain CDR1, CDR2 and CDR3 set forth in SEQ ID NOs: 69, 78 and 89, respectively; (r) heavy chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 51, 62 and 68, respectively, and light chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 69, 78 and 89, respectively; (s) heavy chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 52, 63 and 68, respectively; and light chain CDR1, CDR2 and CDR3 set forth in SEQ ID NOs: 69, 78 and 89, respectively; (t) heavy chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 50, 64 and 68, respectively; and light chain CDR1, CDR2 and CDR3 set forth in SEQ ID NOs: 69, 78 and 89, respectively; (u) heavy chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 50, 65 and 68, respectively; and light chain CDR1, CDR2 and CDR3 set forth in SEQ ID NOs: 69, 78 and 89, respectively; (v) heavy chain CDR1, CDR2, and CDR3 sequences set forth in SEQ ID NOs: 51, 108, and 68, respectively, and light chain CDR1, CDR2, and CDR3 sequences set forth in SEQ ID NOs: 69, 78, and 89, respectively; (w) heavy chain CDR1, CDR2, and CDR3 sequences set forth in SEQ ID NOs: 107, 56, and 68, respectively, and light chain CDR1, CDR2, and CDR3 sequences set forth in SEQ ID NOs: 69, 78, and 89, respectively; and (x) heavy chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 48, 56 and 68, respectively; and light chain CDR1, CDR2 and CDR3 set forth in SEQ ID NOs: 109, 110 and 92, respectively.

[0257] In some embodiments, the anti-CD137 antibodies described herein comprise a heavy chain variable region and and a light chain variable region, wherein the heavy chain variable region comprises an amino acid sequence selected from the group consisting of SEQ ID NOs: 4, 8, 10, 12, 14, 16, 18, 20, 22, 24, 26, 101, and 103, and the light chain variable region comprises an amino acid sequence selected from the group consisting of SEQ ID NOs: 6, 28, 30, 32, 34, 36, 38, 40, 42, 44, 46, and 105.

[0258] In some embodiments, the anti-CD137 antibodies described herein comprise heavy and light chain CDRs, wherein the heavy chain CDR3 comprises the amino acid sequence set forth in SEQ ID NO:68.

[0259] In some embodiments, the anti-CD137 antibody described herein is selected from the group consisting of: The heavy and light chain variable regions comprise the amino acid sequences: (a) SEQ ID NOs: 4 and 6, respectively; (b) SEQ ID NOs: 4 and 28, respectively; (c) SEQ ID NOs: 4 and 30, respectively; (d) SEQ ID NOs: 4 and 32, respectively; (e) SEQ ID NOs: 4 and 34, respectively; (f) SEQ ID NOs: 4 and 36, respectively; (g) SEQ ID NOs: 4 and 38, respectively; (h) SEQ ID NOs: 4 and 40, respectively; (i) SEQ ID NOs: 4 and 42, respectively; (j) SEQ ID NOs: 4 and 44, respectively; (k) SEQ ID NOs: 4 and 46, respectively; (l) SEQ ID NOs: 8 and 6, respectively; (m) SEQ ID NOs: 10 and 6, respectively; (n) SEQ ID NOs: 12 and 6, respectively; (o) SEQ ID NOs: 14 and 6, respectively; (p) SEQ ID NOs: 16 and 6, respectively; (q) SEQ ID NOs: 18 and 6, respectively; (r) SEQ ID NOs: 20 and 6, respectively; (s) SEQ ID NOs: 22 and 6, respectively; (t) SEQ ID NOs: 24 and 6, respectively; (u) SEQ ID NOs: 26 and 6, respectively; (v) SEQ ID NOs: 101 and 6, respectively; (w) SEQ ID NOs: 103 and 6, respectively; and (x) SEQ ID NOs: 4 and 105, respectively.

[0260] In some embodiments, the anti-CD137 antibodies described herein comprise a heavy chain variable region and a heavy chain variable region comprising an amino acid sequence at least 90% identical to an amino acid sequence selected from the group consisting of SEQ ID NOs: 4, 8, 10, 12, 14, 16, 18, 20, 22, 24, 26, 101, and 103, and a light chain variable region comprising an amino acid sequence at least 90% identical to an amino acid sequence selected from the group consisting of SEQ ID NOs: 6, 28, 30, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 6 , 34, 36, 38, 40, 42, 44, 46 and 105 , which contain amino acid sequences that are at least 90% identical.

[0261] In some embodiments, the anti-CD137 antibody described herein is selected from the group consisting of: The variable regions of the heavy and light chains comprise amino acid sequences that are at least 90% identical to the amino acid sequence of: (a) SEQ ID NOs: 4 and 6, respectively; (b) SEQ ID NOs: 4 and 28, respectively; (c) SEQ ID NOs: 4 and 30, respectively; (d) SEQ ID NOs: 4 and 32, respectively; (e) SEQ ID NOs: 4 and 34, respectively; (f) SEQ ID NOs: 4 and 36, respectively; (g) SEQ ID NOs: 4 and 38, respectively; (h) SEQ ID NOs: 4 and 40, respectively; (i) SEQ ID NOs: 4 and 42, respectively; (j) SEQ ID NOs: 4 and 44, respectively; (k) SEQ ID NOs: 4 and 46, respectively; (l) SEQ ID NOs: 8 and 6, respectively; (m) SEQ ID NOs: 10 and 6, respectively; (n) SEQ ID NOs: 12 and 6, respectively; (o) SEQ ID NOs: 14 and 6, respectively; (p) SEQ ID NOs: 16 and 6, respectively; (q) SEQ ID NOs: 18 and 6, respectively; (r) SEQ ID NOs: 20 and 6, respectively; (s) SEQ ID NOs: 22 and 6, respectively; (t) SEQ ID NOs: 24 and 6, respectively; (u) SEQ ID NOs: 26 and 6, respectively; (v) SEQ ID NOs: 101 and 6, respectively; (w) SEQ ID NOs: 103 and 6, respectively; and (x) SEQ ID NOs: 4 and 105, respectively.

[0262] In some embodiments, there is provided herein a method for specifically binding to human CD137, comprising heavy and light chain variable regions encoded by a nucleotide sequence selected from the group consisting of: Antibodies are provided that: (a) SEQ ID NOs: 5 and 7, respectively; (b) SEQ ID NOs: 5 and 29, respectively; (c) SEQ ID NOs: 5 and 31, respectively; (d) SEQ ID NOs: 5 and 33, respectively; (e) SEQ ID NOs: 5 and 35, respectively; (f) SEQ ID NOs: 5 and 37, respectively; (g) SEQ ID NOs: 5 and 39, respectively; (h) SEQ ID NOs: 5 and 41, respectively; (i) SEQ ID NOs: 5 and 43, respectively; (j) SEQ ID NOs: 5 and 45, respectively; (k) SEQ ID NOs: 5 and 47, respectively; (l) SEQ ID NOs: 9 and 7, respectively; (m) SEQ ID NOs: 11 and 7, respectively; (n) SEQ ID NOs: 13 and 7, respectively; (o) SEQ ID NOs: 15 and 7, respectively; (p) SEQ ID NOs: 17 and 7, respectively; (q) SEQ ID NOs: 19 and 7, respectively; (r) SEQ ID NOs: 21 and 7, respectively; (s) SEQ ID NOs: 23 and 7, respectively; (t) SEQ ID NOs: 25 and 7, respectively; (u) SEQ ID NOs: 27 and 7, respectively; (v) SEQ ID NOs: 102 and 7, respectively; (w) SEQ ID NOs: 104 and 7, respectively; and (x) SEQ ID NOs: 5 and 106, respectively.

[0263] In some embodiments, provided herein is an antibody that specifically binds to human CD137, comprising heavy and light chain variable regions encoded by a nucleotide sequence having at least 90% identity to a nucleotide sequence selected from the group consisting of: (a) SEQ ID NOs: 5 and 7, respectively; (b) SEQ ID NOs: 5 and 29, respectively; (c) SEQ ID NOs: 5 and 31, respectively; (d) SEQ ID NOs: 5 and 33, respectively; (e) SEQ ID NOs: 5 and 35, respectively; (f) SEQ ID NOs: 5 and 37, respectively; (g) SEQ ID NOs: 5 and 39, respectively; (h) SEQ ID NOs: 5 and 41, respectively; (i) SEQ ID NOs: 5 and 43, respectively; (j) SEQ ID NOs: 5 and 45, respectively; (k) SEQ ID NOs: 5 and 47, respectively; (l) SEQ ID NOs: 9 and 7, respectively; (m) SEQ ID NOs: 11 and 7, respectively; (n) SEQ ID NOs: 13 and 7, respectively; (o) SEQ ID NOs: 15 and 7, respectively; (p) SEQ ID NOs: 17 and 7, respectively; (q) SEQ ID NOs: 19 and 7, respectively; (r) SEQ ID NOs: 21 and 7, respectively; (s) SEQ ID NOs: 23 and 7, respectively; (t) SEQ ID NOs: 25 and 7, respectively; (u) SEQ ID NOs: 27 and 7, respectively; (v) SEQ ID NOs: 102 and 7, respectively; (w) SEQ ID NOs: 104 and 7, respectively; and (x) SEQ ID NOs: 5 and 106, respectively.

[0264] In some embodiments, the anti-CD137 antibodies described herein comprise a heavy chain variable region and and a light chain variable region, wherein the heavy chain variable region is encoded by a nucleotide sequence selected from the group consisting of SEQ ID NOs: 5, 9, 11, 13, 15, 17, 19, 21, 23, 25, 27, 102, and 104, and the light chain variable region is encoded by a nucleotide sequence selected from the group consisting of SEQ ID NOs: 7, 29, 31, 33, 35, 37, 39, 41, 43, 45, 47, and and 106.

[0265] In some embodiments, the anti-CD137 antibodies described herein comprise a heavy chain variable region and a light chain variable region, wherein the heavy chain variable region is encoded by a nucleotide sequence having at least 90% identity to a nucleotide sequence selected from the group consisting of SEQ ID NOs: 5, 9, 11, 13, 15, 17, 19, 21, 23, 25, 27, 102, and 104, and the light chain variable region is encoded by a nucleotide sequence having at least 90% identity to a nucleotide sequence selected from the group consisting of SEQ ID NOs: 7, 29, 31, 33, 35, 37, 39, 41, 43, 45, 47, and 106.

[0266] In some embodiments, provided herein are anti-CD137 antibodies that specifically bind to human CD137 and comprise a heavy chain CDR3 having the amino acid sequence DXXXXLXXXXYXYYX (SEQ ID NO: 126), where X is any amino acid. In some embodiments, X is any amino acid except alanine. In some embodiments, mutation of residues D95, L100, Y100E, Y100G, and / or Y100H of SEQ ID NO: 126 results in loss of binding to human CD137.

[0267] In some embodiments, provided herein are anti-CD137 antibodies that specifically bind to human CD137 and comprise a heavy chain CDR3 having the amino acid sequence DXPFXLDXXYYYYYX (SEQ ID NO: 127), where X is any amino acid. In some embodiments, the heavy chain CDR3 is selected from the group consisting of F98, D100A, Y100D, and F98 of SEQ ID NO: 126. Mutation of the Y100F and / or Y100H residues to alanine inhibits binding to human CD137. In some embodiments, mutation of residues F98, D100A, Y100D and / or Y100F and / or Y100H of SEQ ID NO: 126 to any residue other than alanine results in increased binding to human CD137.

[0268] In some embodiments, the amino acid sequence DX1X2X3X4LX5X6X7X8YX9YYX described herein specifically binds to human CD137. 10 (SEQ ID NO: 128) wherein X1 is any amino acid, wherein X2 is a nonpolar amino acid, wherein X3 is a nonpolar amino acid, wherein X4 is any amino acid, wherein X5 is a polar amino acid, wherein X6 is any amino acid, wherein X7 is any amino acid, wherein X8 is a polar amino acid, wherein X9 is a polar amino acid, and wherein X 10 is any amino acid In some embodiments, X2 is proline, X3 is phenylalanine or tryptophan, X5 is aspartic acid or glutamic acid, X8 is tyrosine, and X9 is tyrosine.

[0269] The weight of an antibody or antigen-binding portion thereof in binding to a particular target (e.g., CD137) The role of amino acid residues within the heavy chain CDR3 can be determined by methods known to those skilled in the art. In some embodiments, an initial analysis using alanine scanning is completed to determine residues important for antigen binding. As described herein, alanine scanning is a technique used to determine the contribution of specific wild-type residues to the stability or function (e.g., binding affinity) of a given protein or polypeptide. This technique involves substituting an alanine residue for the wild-type residue in a polypeptide, then evaluating the stability or function (e.g., binding affinity) of the alanine-substituted derivative or mutant polypeptide and comparing it with the wild-type polypeptide. In some embodiments, residues identified as not important are further evaluated for modulating antibody binding to the antigen (e.g., increasing or decreasing binding). A non-limiting example of such an analysis is deep mutational scanning. This method allows for the evaluation of a large number of mutations. In some embodiments, each amino acid residue within the heavy chain CDR3 is mutated to each amino acid residue (except alanine) and binding is evaluated. Other methods for analyzing the effects of amino acid residue mutations are known in the art. In some embodiments, these methods are utilized to assess the role of all residues in the heavy and light chain CDRs in binding to human CD137.

[0270] Exemplary CD137-Binding Antibodies In some embodiments, the anti-CD137 antibodies described herein have an affinity (K D In some embodiments, the anti-CD137 antibodies described herein bind to human CD137 with an affinity of about 40 to 100 nM (e.g., about 40 nM to about 100 nM). (K D ) binds to human CD137. In some embodiments, the anti-CD137 antibodies described herein bind to an epitope on human CD137 described above (including, for example, K114). In some embodiments, the anti-CD137 antibodies described herein comprise a heavy chain CDR3 comprising the amino acid sequence DXXXXLXXXXYXYYX (SEQ ID NO: 126). In some embodiments, the anti-CD137 antibodies described herein have an affinity (K D ) binds to human CD137 and binds to the epitope on human CD137 described above (including, for example, K114). In the present specification, the anti-CD137 antibody has a concentration of about 30 to 100 nM (for example, about 30 nM to about 100 nM ) affinity (K D ) and comprises a heavy chain CDR3 comprising the amino acid sequence DXXXXLXXXXYXYYX (SEQ ID NO: 126). In some embodiments, the anti-CD137 antibodies described herein bind to an epitope on human CD137 as described above (e.g., comprising K114) and comprise a heavy chain CDR3 comprising the amino acid sequence DXXXXLXXXXYXYYX (SEQ ID NO: 126). The anti-CD137 antibodies described herein have an affinity of about 30 to 100 nM (e.g., about 30 nM to about 100 nM). (K D ), binds to human CD137 at the CDR3 position described above (including, for example, K114), and comprises a heavy chain CDR3 comprising the amino acid sequence DXXXXLXXXXYXYYX (SEQ ID NO: 126).

[0271] In some embodiments, the anti-CD137 antibody (i) an affinity (K) of about 30 to 100 nM (e.g., about 30 nM to about 100 nM); D ) binds to human CD137; and (ii) amino acid sequence DX1X2X3X4LX5X6X7X8YX9YYX 10wherein X1 is any amino acid, wherein X2 is a nonpolar amino acid, wherein X3 is a nonpolar amino acid, wherein X4 is any amino acid, wherein X5 is a polar amino acid, wherein X6 is any amino acid, wherein X7 is any amino acid, wherein X8 is a polar amino acid, wherein X9 is a polar amino acid, and wherein X 10 is any amino acid.

[0272] In some embodiments, the anti-CD137 antibody (i) an affinity (K) of about 30 to 100 nM (e.g., about 30 nM to about 100 nM); D ) binds to human CD137; and (ii) containing one or more of residues E111, T113, K114, N126, I132, and P135 of SEQ ID NO: 3 Binds to an epitope on human CD137.

[0273] In some embodiments, the anti-CD137 antibody (i) an affinity (K) of about 30 to 100 nM (e.g., about 30 nM to about 100 nM); D ) binds to human CD137; (ii) containing one or more of residues E111, T113, K114, N126, I132, and P135 of SEQ ID NO: 3 Binds to an epitope on human CD137. (iii) a heavy chain CDR3 comprising the amino acid sequence DXXXXLXXXXYXYYX, where X is any amino acid is; or (iv) A combination thereof.

[0274] In some embodiments, the anti-CD137 antibody (i) an affinity (K) of about 30 to 100 nM (e.g., about 30 nM to about 100 nM); D ) binds to human CD137; (ii) containing one or more of residues E111, T113, K114, N126, I132, and P135 of SEQ ID NO: 3 Binds to an epitope on human CD137. (iii) Amino acid sequence DX1X2X3X4LX5X6X7X8YX9YYX 10 wherein X1 is any wherein X2 is a nonpolar amino acid, wherein X3 is a nonpolar amino acid, wherein X4 is any amino acid, wherein X5 is a polar amino acid, wherein X6 is any amino acid, wherein X7 is any amino acid, wherein X8 is a polar amino acid, wherein X9 is a polar amino acid, and wherein X 10 is any amino acid; or (iv) A combination thereof.

[0275] In some embodiments, the anti-CD137 antibody (i) an affinity (K) of about 30 to 100 nM (e.g., about 30 nM to about 100 nM); D ) binds to human CD137; (ii) containing one or more of residues E111, T113, K114, N126, I132, and P135 of SEQ ID NO: 3 specifically binds to an epitope on human CD137; and (iii) a heavy chain CDR3 comprising the amino acid sequence DXXXXLXXXXYXYYX, where X is any amino acid is.

[0276] In some embodiments, the anti-CD137 antibody (i) an affinity (K) of about 30 to 100 nM (e.g., about 30 nM to about 100 nM); D ) binds to human CD137; (ii) containing one or more of residues E111, T113, K114, N126, I132, and P135 of SEQ ID NO: 3 binds to an epitope on human CD137; and (iii) Amino acid sequence DX1X2X3X4LX5X6X7X8YX9YYX 10 wherein X1 is any wherein X2 is a nonpolar amino acid, wherein X3 is a nonpolar amino acid, wherein X4 is any amino acid, wherein X5 is a polar amino acid, wherein X6 is any amino acid, wherein X7 is any amino acid, wherein X8 is a polar amino acid, wherein X9 is a polar amino acid, and wherein X 10 is any amino acid.

[0277] In some embodiments, the anti-CD137 antibody (i) an affinity (K) of about 30 to 100 nM (e.g., about 30 nM to about 100 nM); D ) binds to human CD137; and (ii) one or more amino acid residues corresponding to amino acids 111 to 135 of SEQ ID NO: 3 It binds to an epitope containing the sequence.

[0278] In some embodiments, the anti-CD137 antibody (i) an affinity (K) of about 30 to 100 nM (e.g., about 30 nM to about 100 nM); D ) binds to human CD137; (ii) one or more amino acid residues corresponding to amino acids 111 to 135 of SEQ ID NO: 3 It binds to an epitope containing the sequence. (iii) a heavy chain CDR3 comprising the amino acid sequence DXXXXLXXXXYXYYX, where X is any amino acid is; or (iv) A combination thereof.

[0279] In some embodiments, the anti-CD137 antibody (i) an affinity (K) of about 30 to 100 nM (e.g., about 30 nM to about 100 nM); D ) binds to human CD137; (ii) one or more amino acid residues corresponding to amino acids 111 to 135 of SEQ ID NO: 3 It binds to an epitope containing the sequence. (iii) Amino acid sequence DX1X2X3X4LX5X6X7X8YX9YYX 10 wherein X1 is any wherein X2 is a nonpolar amino acid, wherein X3 is a nonpolar amino acid, wherein X4 is any amino acid, wherein X5 is a polar amino acid, wherein X6 is any amino acid, wherein X7 is any amino acid, wherein X8 is a polar amino acid, wherein X9 is a polar amino acid, and wherein X 10 is any amino acid; or (iv) A combination thereof.

[0280] In some embodiments, the anti-CD137 antibody (i) an affinity (K) of about 30 to 100 nM (e.g., about 30 nM to about 100 nM); D ) binds to human CD137; (ii) one or more amino acid residues corresponding to amino acids 111 to 135 of SEQ ID NO: 3 binds to an epitope comprising the sequence; and (iii) a heavy chain CDR3 comprising the amino acid sequence DXXXXLXXXXYXYYX, where X is any amino acid is.

[0281] In some embodiments, the anti-CD137 antibody (i) an affinity (K) of about 30 to 100 nM (e.g., about 30 nM to about 100 nM); D ) binds to human CD137; (ii) one or more amino acid residues corresponding to amino acids 111 to 135 of SEQ ID NO: 3 binds to an epitope comprising the sequence; and (iii) Amino acid sequence DX1X2X3X4LX5X6X7X8YX9YYX 10 wherein X1 is any wherein X2 is a nonpolar amino acid, wherein X3 is a nonpolar amino acid, wherein X4 is any amino acid, wherein X5 is a polar amino acid, wherein X6 is any amino acid, wherein X7 is any amino acid, wherein X8 is a polar amino acid, wherein X9 is a polar amino acid, and wherein X 10 is any amino acid.

[0282] In some embodiments, the anti-CD137 antibody (i) binds to human CD137 with an affinity of about 30 to 100 nM (e.g., about 30 nM to about 100 nM); and (ii) an epitope containing ELTK (corresponding to amino acid residues 111 to 114 of SEQ ID NO: 3); Combine.

[0283] In some embodiments, the anti-CD137 antibody (i) an affinity (K) of about 30 to 100 nM (e.g., about 30 nM to about 100 nM); D ) binds to human CD137; (ii) an epitope containing ELTK (corresponding to amino acid residues 111 to 114 of SEQ ID NO: 3); Combine; (iii) a heavy chain CDR3 comprising the amino acid sequence DXXXXLXXXXYXYYX, where X is any amino acid is; or (iv) A combination thereof.

[0284] In some embodiments, the anti-CD137 antibody (i) an affinity (K) of about 30 to 100 nM (e.g., about 30 nM to about 100 nM); D ) binds to human CD137; (ii) an epitope containing ELTK (corresponding to amino acid residues 111 to 114 of SEQ ID NO: 3); Combine; (iii) Amino acid sequence DX1X2X3X4LX5X6X7X8YX9YYX 10wherein X1 is any wherein X2 is a nonpolar amino acid, wherein X3 is a nonpolar amino acid, wherein X4 is any amino acid, wherein X5 is a polar amino acid, wherein X6 is any amino acid, wherein X7 is any amino acid, wherein X8 is a polar amino acid, wherein X9 is a polar amino acid, and wherein X 10 is any amino acid; or (iv) A combination thereof.

[0285] In some embodiments, the anti-CD137 antibody (i) an affinity (K) of about 30 to 100 nM (e.g., about 30 nM to about 100 nM); D ) binds to human CD137; (ii) binds to an epitope containing ELTK (corresponding to amino acid residues 111 to 114 of SEQ ID NO: 3); and (iii) a heavy chain CDR3 comprising the amino acid sequence DXXXXLXXXXYXYYX, where X is any amino acid is.

[0286] In some embodiments, the anti-CD137 antibody (i) an affinity (K) of about 30 to 100 nM (e.g., about 30 nM to about 100 nM); D ) binds to human CD137; (ii) an epitope containing ELTK (corresponding to amino acid residues 111 to 114 of SEQ ID NO: 3); combine; and (iii) Amino acid sequence DX1X2X3X4LX5X6X7X8YX9YYX 10 wherein X1 is any wherein X2 is a nonpolar amino acid, wherein X3 is a nonpolar amino acid, wherein X4 is any amino acid, wherein X5 is a polar amino acid, wherein X6 is any amino acid, wherein X7 is any amino acid, wherein X8 is a polar amino acid, wherein X9 is a polar amino acid, and wherein X 10is any amino acid.

[0287] In some embodiments, the anti-CD137 antibody described above comprises a heavy chain and a nucleotide sequence selected from the group consisting of: and light chain CDRs: (a) heavy chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 48, 56 and 68, respectively; and light chain CDR1, CDR2 and CDR3 set forth in SEQ ID NOs: 69, 78 and 89, respectively; and (b) Heavy chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 51, 108 and 68, respectively, and light chain CDR1, CDR2 and CDR3 sequences set forth in SEQ ID NOs: 69, 78 and 89, respectively.

[0288] In some embodiments, the anti-CD137 antibody comprises a heavy chain variable region and a light chain variable region, In some cases, the heavy chain variable region comprises an amino acid sequence selected from the group consisting of SEQ ID NOs: 4 and 101, and in some cases, the light chain variable region comprises the amino acid sequence of SEQ ID NO: 6. Includes:

[0289] In some embodiments, the anti-CD137 antibody has an amino acid sequence selected from the group consisting of: The heavy and light chain variable regions include: (a) SEQ ID NOs: 4 and 6, respectively; and (b) SEQ ID NOs: 101 and 6, respectively.

[0290] In some embodiments, the anti-CD137 antibody has an amino acid sequence selected from the group consisting of: The heavy and light chain variable regions include: (a) SEQ ID NOs: 4 and 6, respectively; (b) SEQ ID NOs: 101 and 6, respectively; and (c) SEQ ID NOs: 26 and 6, respectively.

[0291] In some embodiments, the anti-CD137 antibody comprises a nucleotide sequence selected from the group consisting of: The heavy and light chain variable regions are encoded by the sequences: (a) SEQ ID NOs: 5 and 7, respectively; and (b) SEQ ID NOs: 102 and 7, respectively.

[0292] In some embodiments, the anti-CD137 antibody comprises a nucleotide sequence selected from the group consisting of: The heavy and light chain variable regions are encoded by the sequences: (a) SEQ ID NOs: 5 and 7, respectively; (b) SEQ ID NOs: 102 and 7, respectively; and (c) SEQ ID NOs: 27 and 7, respectively.

[0293] In some embodiments, the anti-CD137 antibody comprises a heavy chain variable region and a light chain variable region, wherein the heavy chain variable region comprises an amino acid sequence at least 90% identical to an amino acid sequence selected from the group consisting of SEQ ID NOs: 4 and 101, and The light chain variable region comprises an amino acid sequence that is at least 90% identical to the amino acid sequence of SEQ ID NO:6.

[0294] In some embodiments, the anti-CD137 antibody comprises a heavy chain variable region and a light chain variable region, In some cases, the heavy chain variable region comprises an amino acid sequence that is at least 90% identical to an amino acid sequence selected from the group consisting of SEQ ID NOs: 4, 26, and 101, and in some cases, wherein the light chain variable region comprises an amino acid sequence that is at least 90% identical to the amino acid sequence of SEQ ID NO:6.

[0295] In some embodiments, the anti-CD137 antibody comprises a heavy chain variable region and a light chain variable region, in which case the heavy chain variable region is encoded by a nucleotide sequence that is at least 90% identical to a nucleotide sequence selected from the group consisting of SEQ ID NOs: 5 and 102, and and wherein the light chain variable region has at least one nucleotide sequence that is different from the nucleotide sequence of SEQ ID NO:7. Both are encoded by nucleotide sequences that are 90% identical.

[0296] In some embodiments, the anti-CD137 antibody comprises a heavy chain variable region and a light chain variable region, in which case the heavy chain variable region is encoded by a nucleotide sequence that is at least 90% identical to a nucleotide sequence selected from the group consisting of SEQ ID NOs: 5, 27, and 102; and in this case, the light chain variable region has at least one nucleotide sequence different from that of SEQ ID NO:7. are encoded by nucleotide sequences that are at least 90% identical.

[0297] In some embodiments, the anti-CD137 antibody has an amino acid sequence selected from the group consisting of: (a) heavy and light chain variable regions comprising amino acid sequences at least 90% identical to: (a) SEQ ID NOs: 4 and 6, respectively; and (b) SEQ ID NOs: 101 and 6, respectively.

[0298] In some embodiments, the anti-CD137 antibody has an amino acid sequence selected from the group consisting of: The heavy and light chain variable regions comprise amino acid sequences that are at least 90% identical to: (a) SEQ ID NOs: 4 and 6, respectively; (b) SEQ ID NOs: 101 and 6, respectively; and (c) SEQ ID NOs: 26 and 6, respectively.

[0299] In some embodiments, the anti-CD137 antibody comprises a nucleotide sequence selected from the group consisting of: The heavy and light chain variable regions are encoded by nucleotide sequences that are at least 90% identical to the sequence: (a) SEQ ID NOs: 5 and 7, respectively; and (b) SEQ ID NOs: 102 and 7, respectively.

[0300] In some embodiments, the anti-CD137 antibody comprises a nucleotide sequence selected from the group consisting of: The heavy and light chain variable regions are encoded by nucleotide sequences that are at least 90% identical to the sequence: (a) SEQ ID NOs: 5 and 7, respectively; (b) SEQ ID NOs: 102 and 7, respectively; and (c) SEQ ID NOs: 27 and 7, respectively.

[0301] In some embodiments, the anti-CD137 antibodies described herein are mAb1 (i.e., In some embodiments, the functional properties of the antibodies described herein include, but are not limited to, the following: inhibition of CD137 dimerization; ... Induction or enhancement of CD137 multimerization; induction or enhancement of CD137-mediated T cell activation Induction or enhancement of CD137-mediated cytotoxic T cell responses; Induction or enhancement of CD137-mediated T cell proliferation; Induction or enhancement of CD137-mediated cytokine production; Induction or enhancement of CD137-mediated cytokine production in the liver and / or or a lack of induction or enhancement of T cell activation and / or T cell proliferation within the spleen; and a reduction or inhibition of tumor growth.

[0302] In some embodiments, the anti-CD137 antibodies described herein are mAb1 (i.e., 4 and 6, respectively), mab8 (i.e., an antibody comprising the heavy and light chain variable sequences of SEQ ID NOs: 101 and 6, respectively), or mAb10 (i.e., an antibody comprising the heavy and light chain variable sequences of SEQ ID NOs: 26 and 6, respectively). D It binds to human CD137.

[0303] In some embodiments, the anti-CD137 antibodies described herein comprise a human IgG1 heavy chain constant region or or a human IgG4 heavy chain constant region. The antibody comprises a human wild-type IgG1 heavy chain constant region or a human wild-type IgG4 heavy chain constant region. In some embodiments, the anti-CD137 antibody described herein comprises a human wild-type IgG1 heavy chain constant region set forth in SEQ ID NO: 1. In some embodiments, the anti-CD137 antibody described herein comprises a human wild-type IgG1 heavy chain constant region set forth in SEQ ID NO: 1. In some embodiments, the anti-CD137 antibodies described herein comprise a mutated IgG1 heavy chain constant region or a mutated IgG4 heavy chain constant region, in which The mutant IgG4 heavy chain constant region comprises an amino acid substitution at residue Ser228. In some embodiments, the amino acid substitution at Ser228 is S228P. In some embodiments, the mutant IgG4 heavy chain constant region comprises an amino acid substitution at residue Ser228. In some embodiments, the amino acid substitution at residue Ser228 is S228P. The anti-CD137 antibodies described herein comprise an IgG4 heavy chain constant region in which the C-terminal lysine residue has been removed. In some embodiments, the anti-CD137 antibodies described herein comprise an IgG4 heavy chain constant region in which the C-terminal lysine residue has been removed and include a S228P amino acid substitution. In the present specification, the anti-CD137 antibodies comprise the IgG4 heavy chain constant region set forth in SEQ ID NO:2.

[0304] In some embodiments, the anti-CD137 antibodies described herein comprise heavy and light chains comprising the amino acid sequences set forth in SEQ ID NOs: 129 and 133, respectively.

[0305] In some embodiments, the anti-CD137 antibodies described herein comprise heavy and light chains comprising the amino acid sequences set forth in SEQ ID NOs: 130 and 133, respectively.

[0306] In some embodiments, the anti-CD137 antibodies described herein comprise heavy and light chains comprising the amino acid sequences set forth in SEQ ID NOs: 131 and 133, respectively.

[0307] In some embodiments, the anti-CD137 antibodies described herein comprise heavy and light chains comprising the amino acid sequences set forth in SEQ ID NOs: 132 and 133, respectively.

[0308] In some embodiments, the anti-CD137 antibodies described herein have a homology ratio of at least 80%, at least 85%, at least 90%, at least 95%, or at least 100% to SEQ ID NOs: 129 and 133, respectively. Amino acids with 96%, at least 97%, at least 98%, or at least 99% identity In some embodiments, the anti-CD137 antibody comprises a heavy chain and a light chain comprising the sequence The antibody may have a sequence identity of at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or less to SEQ ID NOs: 130 and 133, respectively. In some embodiments, the anti-CD137 antibodies described herein comprise heavy and light chains that comprise amino acid sequences that share at least 99% identity with SEQ ID NOs: 131 and 133, respectively. heavy chains comprising amino acid sequences that have 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% identity with each other; In some embodiments, the anti-CD137 antibodies described herein each comprise a light chain having the sequence 132 and 133. The antibody comprises a heavy chain and a light chain containing the same amino acid sequence.

[0309] Characterization and function of CD137-binding antibodies I. Affinity In some embodiments, the anti-CD137 antibodies described herein are In some embodiments, the anti-CD137 antibodies described herein bind to human CD137 with an affinity (KD) of about 40 to 100 nM (e.g., about 40 nM to about 100 nM), as determined by antigen binding assays. In some embodiments, the anti-CD137 antibodies described herein bind to human CD137 with an affinity (KD) of about 30 to 100 nM (e.g., about 30 nM to about 100 nM), as determined by antigen binding assays. In some embodiments, the anti-CD137 antibodies described herein bind to human CD137 with an affinity (KD) of about 45 to 95 nM, 50 to 90 nM, 55 to 85 nM, 60 to 80 nM, 65 to 75 nM, 55 to 75 nM, 40 to 70 nM, 50 to 80 nM, or 60 to 90 nM, as determined by antigen binding assays.

[0310] In some embodiments, the antigen binding assay determines the binding affinity of the anti-CD137 antibody to a CD137 polypeptide. In some embodiments, the antigen binding assay is surface plasmon resonance. Thus, in some embodiments, the anti-CD137 antibodies described herein are As determined using surface plasmon resonance, the antibodies bind to human CD137 with an affinity (KD) of about 40 to 100 nM (e.g., about 40 nM to about 100 nM). The anti-CD137 antibodies described herein bind to human CD137 with an affinity (KD) of about 30 to 100 nM (e.g., about 30 nM to about 100 nM) as determined using surface plasmon resonance. In some embodiments, the anti-CD137 antibodies described herein bind to human CD137 with an affinity (KD) of about 30 to 100 nM as determined using surface plasmon resonance. Binds to human CD137 with an affinity (KD) of approximately 45-95nM, 50-90nM, 55-85nM, 60-80nM, 65-75nM, 55-75nM, 40-70nM, 50-80nM, or 60-90nM.

[0311] The term "surface plasmon resonance" refers to, for example, a BIAcore system (Pharmacia Biosensor The use of optical sensors (such as Jonsson, U., ... et al. (1993) Ann. Biol. Clin. 51:19-26; Jonsson, U., et al. (1991) Biotechniques 11:620-627; Johnsson, B., et al. (1995) J. Mol. Recognit. 8:125-131; and Johnson, B., et al. (1991) Anal. Biochem. 198:268-277. In some embodiments, the antigen binding assay is biolayer interferometry (BLI). Thus, in some embodiments, the anti-CD137 antibodies described herein bind to human CD137 with an affinity (KD) of about 40 to 100 nM (e.g., about 40 nM to about 100 nM) as determined using biolayer interferometry. In some embodiments, the anti-CD137 antibodies described herein bind to human CD137 with an affinity (KD) of about 40 to 100 nM (e.g., about 40 nM to about 100 nM) as determined using biolayer interferometry. When the antibody was used, it bound to human CD137 with an affinity (KD) of about 30 to 100 nM (e.g., about 30 nM to about 100 nM). In some embodiments, the anti-CD137 antibodies described herein are combined with biolayer intercalators. Binds to human CD137 with an affinity (KD) of approximately 45-95nM, 50-90nM, 55-85nM, 60-80nM, 65-75nM, 55-75nM, 40-70nM, 50-80nM, or 60-90nM, as determined using coherence tomography (CIP) assays do.

[0312] The term "Bio-Layer Interferometry" or "BLI" refers to the thickness of the detection surface of its optical layer. In some embodiments, the optical phenomenon allows measurement of sub-nanometer changes in the thickness of the optical layer. In some embodiments, biomolecules bind to the sensor surface, causing a change in the thickness of the optical layer. The magnitude of the change in the thickness of the optical layer is proportional to the mass or molecular weight of the bound molecule. In some embodiments, CD137 is immobilized on the sensor surface, and antibody binding is measured. In this case, binding is A change in molecular weight produces a corresponding change in the thickness of the optical layer. In some embodiments, BLI is performed in an OCTET system (ForteBio).

[0313] II. Immune cell effects In some embodiments, the anti-CD137 antibodies described herein are useful in cytokine assays. In some embodiments, cytokine assays are performed to determine whether or not the antibody induces or enhances cytokine production by immune cells. In some embodiments, cytokine assays are performed to determine whether or not the antibody induces or enhances cytokine production by immune cells secreted by immune cells contacted with the anti-CD137 antibody. determining the amount of at least one cytokine, wherein an increase in the amount of the at least one cytokine indicates induction or enhancement of cytokine production by the anti-CD137 antibody; In some embodiments, increased cytokine production is indicative of increased cytokine production in response to a control antibody (e.g., CD137 at least 1-fold, 2-fold, or 3-fold higher than antibodies that do not bind to or induce cytokine production , four or five times higher.

[0314] In some embodiments, the anti-CD137 antibodies described herein are useful in cytokine assays. wherein the cytokine assay comprises the steps of: (i) contacting immune cells with an anti-CD137 antibody; and (ii) determining the amount of at least one cytokine produced by the immune cells; In this case, the increase in the amount of the at least one cytokine indicates that the anti-CD137 antibody , suggesting that cytokine production by the immune cells was induced or enhanced.

[0315] In some embodiments, the anti-CD137 antibodies described herein are useful in cytokine assays. wherein the cytokine assay comprises the steps of: (i) contacting immune cells with an anti-CD137 antibody; and (ii) determining the amount of at least one cytokine produced by the immune cells; and (iii) measuring the amount of the at least one cytokine produced by the immune cell using the reference comparing the amount secreted by immune cells; In this case, the reference immune cells are contacted with a control antibody, and in this case, an increase in the amount of the at least one cytokine produced by the immune cells compared to the reference immune cells indicates induction or enhancement of human CD137-mediated cytokine production.

[0316] In some embodiments, the anti-CD137 antibodies described herein are useful in cytokine assays. Induce or enhance cytokine production by immune cells, as determined by the cytokine assay, wherein the cytokine assay comprises the steps of: (i) contacting immune cells with an anti-CD137 antibody; (ii) determining the amount of at least one cytokine produced by the immune cells; and (iii) measuring the amount of the at least one cytokine produced by the immune cell using the reference comparing the amount or level secreted by immune cells; In this case, the reference immune cells have not been contacted with an anti-CD137 antibody, and in this case an increase in the amount of the at least one cytokine produced by the immune cells compared to the reference immune cells in the presence of human CD137-mediated cytokine production by the immune cells; This suggests the induction or strengthening of

[0317] In some embodiments, the at least one cytokine is IL-2, IFNγ, TNFα, IL-13 and combinations thereof. In some embodiments, the cytokine is IL-2. In some embodiments, the cytokine is IFNγ. In some embodiments, the cytokine is TNFα. In some embodiments, the cytokine is IL-13. In some embodiments, the anti-CD137 antibody induces or enhances the production of IL-2. In embodiments, the anti-CD137 antibody induces or enhances the production of TNFα. In some embodiments, the anti-CD137 antibody induces or enhances the production of IL-13. In some aspects, the cytokine produced is IL-2. In some aspects, the cytokine produced is TNFα. In some embodiments, the cytokine produced is IL-13. In some embodiments, the cytokine produced is IFNγ. The cytokines are IL-2 and TNFα. In some embodiments, the cytokines produced are IL-2 and IL-13. In some embodiments, the cytokines produced are TNFα and IL-13. In some embodiments, the cytokines produced are TNFα and IFNγ. In some embodiments, the cytokines produced are IL-13 and IFNγ. In some embodiments, the cytokines produced are IL-2, TNFα, and IL-13. In some embodiments, the cytokines produced are IL-2, TNFα, and IFNγ. In some embodiments, the cytokines produced are IFNγ, TNFα, and IL-13.

[0318] In some embodiments, the immune cell is a T cell. In some embodiments, the reference immune cell are T cells. In some embodiments, the T cells are CD8+ T cells.

[0319] In some embodiments, the cytokine assay is a cytokine bead array assay. A cytokine bead array assay is a bead-based immunoassay that allows for the flow cytometric determination of multiple analytes of multiple cytokines in a sample. The use of microspheres of different sizes or colors is the basis of a cytokine bead array assay, with each microsphere (or "bead") coated with an antibody that specifically binds to an antigen (e.g., a cytokine). The antibody-coated beads are then introduced into the sample in combination with a detection antibody. The bead:antigen:detection antibody complex is then analyzed by flow cytometry. Commercially available cytokine bead array assays include, but are not limited to, BD™ Cytometric Bead Array Systems (BD Biosciences) and Luminex® Assays (R&D Systems). In some embodiments, the induction or enhancement of human CD137-mediated cytokine production is achieved by the induction or enhancement of cytokine production in a cell line comprising a cytoplasmic domain of the cytoplasmic domain. In some embodiments, human CD137-mediated Induction or enhancement of cytokine production is determined by Luminex® Assay.

[0320] In some embodiments, the cytokine assay is performed using a Meso Scale Discovery (MSD) assay. The MSD assay is manufactured by Meso Scale Diagnostics, Inc. (Rockville, MD). MSD assays are commercially available assays based on the detection of electrochemiluminescently labeled antibodies that specifically bind to antigens of interest (e.g., cytokines). The MSD assay contains a high-binding carbon electrode at the bottom of the microplate well to which a specific capture antibody (specific capture antibody) can be attached. The MSD assay uses an electrochemiluminescent label conjugated to a detection antibody. The solution is added to the microplate wells, and the MSD device applies electricity to the plate electrodes. This causes the electrochemiluminescent label to emit light. The intensity of the light is measured to quantify the analyte (e.g., cytokine) in the sample. In some embodiments, human CD137-mediated Induction or enhancement of cytokine production was determined by Meso Scale Discovery (MSD) assay. is determined.

[0321] In some embodiments, the anti-CD137 antibodies described herein induce or enhance T cell activation as determined by a T cell activation assay. In some embodiments, the T cell activation assay determines the amount of at least one cytokine secreted by T cells contacted with an anti-CD137 antibody described herein, wherein an increase in the amount of the at least one cytokine indicates induction or enhancement of T cell activation. In some embodiments, the anti-CD137 antibodies described herein induce or enhance T cell activation. The increase in cytokine production is due to the presence of a control antibody (e.g., a control antibody that does not bind to CD137 and does not bind to cytokine production). at least 1-fold, 2-fold, 3-fold, 4-fold or 5-fold higher than that of an antibody that does not induce

[0322] In some embodiments, the anti-CD137 antibodies described herein induce or enhance T cell activation as determined by a T cell activation assay, wherein the T cell activation assay comprises the steps of: (i) isolating T cells from a subject; (ii) contacting the T cells with an anti-CD137 antibody; and (iii) after (ii), determining the amount of at least one cytokine secreted by the T cells; determining the In this case, an increase in the level of the at least one cytokine is associated with the anti-CD137 antibody. This suggests that the body has induced or enhanced T cell activation.

[0323] In some embodiments, the anti-CD137 antibodies described herein induce or enhance T cell activation as determined by a T cell activation assay, wherein the T cell activation assay comprises the steps of: (i) isolating T cells from a subject; (ii) contacting the T cells with an anti-CD137 antibody; (iii) determining the amount of at least one cytokine secreted by the T cells; and (iv) comparing the amount of the at least one cytokine produced by the T cells with the amount or level secreted by a reference T cell; In this case, the reference T cells have not been contacted with an anti-CD137 antibody, and in this case, an increase in the amount of the at least one cytokine produced by the T cells compared to the reference T cells indicates that the anti-CD137 antibody has induced or enhanced T cell activation.

[0324] In some embodiments, the anti-CD137 antibodies described herein induce or enhance T cell activation as determined by a T cell activation assay, wherein the T cell activation assay comprises the steps of: (i) isolating T cells from a subject; (ii) contacting the T cells with an anti-CD137 antibody; (iii) determining the amount of at least one cytokine secreted by the T cells; and (iv) comparing the amount of the at least one cytokine produced by the T cells with the amount secreted by a reference T cell; In this case, the reference T cells are contacted with a control antibody, and in this case, the reference T cells are An increase in the amount of the at least one cytokine produced by the T cells compared to control T cells indicates that the anti-CD137 antibody induced or enhanced T cell activation.

[0325] In some embodiments, the T cell activation assay determines the level of at least one cytokine secreted by T cells after contact with an anti-CD137 antibody described herein. In some embodiments, the cytokine is IL-2. In some embodiments, the cytokine is IFNγ. In some embodiments, the cytokine is TNFα. In some embodiments, the cytokine is IL-13. In some embodiments, the T cell activation assay comprises at least one T cell activation assay, e.g., as described herein. In some embodiments, the cytokine produced is IL-2. In some embodiments, the cytokine produced is TNFα. In some embodiments, the cytokine produced is IL-13. In some embodiments, the cytokine produced is IFNγ. In some embodiments, the cytokine produced is IL-2. In some embodiments, the cytokine produced is TNFα. In some embodiments, the cytokine produced is IFNγ. In some embodiments, the cytokines produced are IL-2 and TNFα. The cytokines produced are IL-2 and IL-13. The cytokines are IL-2 and IFNγ. In some embodiments, the cytokines produced are TNFα and IL-13. In some embodiments, the cytokines produced are IL-2, TNFα, and IL-13. In some embodiments, the cytokines produced are IL-13 and IFNγ. In some embodiments, the cytokines produced are IL-2, TNFα, and IL-13. In some embodiments, the cytokines produced are IL-2, TNFα, and IFNγ. In some embodiments, the cytokines produced are IFNγ, TNFα, and IL-13. .

[0326] In some embodiments, the anti-CD137 antibodies described herein induce or enhance T cell activation as determined by a T cell activation assay, wherein the T cell activation assay includes detecting surface expression of at least one activation marker on T cells. In some embodiments, "increased surface expression" refers to the induction or enhancement of T cell activation. means at least 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70% increase in surface expression compared to surface expression in the presence of a control antibody or in the absence of antibody. , refers to an increase of 75%, 80%, 85%, 90%, 95% or 100%.

[0327] In some embodiments, the anti-CD137 antibodies described herein induce or enhance T cell activation as determined by an in vitro T cell activation assay, wherein The T cell activation assay comprises the following steps: (i) isolating T cells from a subject; (ii) contacting the T cells with an anti-CD137 antibody; and (iii) detecting surface expression of at least one activation marker on the T cells; In this case, an increase in surface expression of the at least one activation marker indicates that the anti-CD137 antibody has induced or enhanced T cell activation.

[0328] In some embodiments, the anti-CD137 antibodies described herein induce or enhance T cell activation as determined by a T cell activation assay, wherein the T cell activation assay comprises the steps of: (i) isolating T cells from a subject; (ii) contacting the T cells with an anti-CD137 antibody; (iii) determining the surface expression of at least one activation marker on the T cells; and (iv) comparing the surface expression of at least one activation marker on the T cells with the surface expression of the at least one activation marker on a reference T cell; In this case, the reference T cells have not been contacted with an anti-CD137 antibody, and in this case, an increase in surface expression of at least one activation marker on the T cells compared to the reference T cells indicates that the anti-CD137 antibody has induced or enhanced T cell activation.

[0329] In some embodiments, the anti-CD137 antibodies described herein induce or enhance T cell activation as determined by a T cell activation assay, wherein the T cell activation assay comprises the steps of: (i) isolating T cells from a subject; (ii) contacting the T cells with an anti-CD137 antibody; (iii) determining the surface expression of at least one activation marker on the T cells; (iv) comparing the surface expression of at least one activation marker on the T cells with the surface expression of the at least one activation marker on a reference T cell; In this case, the reference T cells are contacted with a control antibody, and in this case, the reference T cells are An increase in the surface expression of at least one activation marker on the T cells compared to the surface expression of at least one activation marker on control T cells indicates that the anti-CD137 antibody induced or enhanced T cell activation.

[0330] In some embodiments, the anti-CD137 antibodies described herein induce or enhance T cell activation as determined by an in vivo T cell activation assay, in which case the The T cell activation assay comprises the following steps: (i) administering an anti-CD137 antibody to a subject; (ii) isolating T cells from the subject; and (iii) detecting surface expression of at least one activation marker on the T cells; In this case, an increase in surface expression of the at least one activation marker indicates that the anti-CD137 antibody induced or enhanced CD137-mediated T cell activation.

[0331] In some embodiments, the anti-CD137 antibodies described herein induce or enhance T cell activation as determined by a T cell activation assay, wherein the T cell activation assay comprises the steps of: (i) administering an anti-CD137 antibody to a subject; (ii) isolating T cells from the subject; (iii) then determining the surface expression of at least one activation marker on the T cells; and (iv) comparing the surface expression of at least one activation marker on the T cells with the surface expression of the at least one activation marker on a reference T cell; In this case, the reference T cells are isolated from a subject that has not been administered the anti-CD137 antibody, and in this case, an increase in surface expression of at least one activation marker on the T cells compared to the reference T cells indicates that the anti-CD137 antibody has induced or enhanced T cell activation.

[0332] In some embodiments, the anti-CD137 antibodies described herein induce or enhance T cell activation as determined by a T cell activation assay, wherein the T cell activation assay comprises the steps of: (i) administering an anti-CD137 antibody to a subject; (ii) isolating T cells from the subject; (iii) determining the surface expression of at least one activation marker on the T cells; and (iv) comparing the surface expression of at least one activation marker on the T cells with the surface expression of the at least one activation marker on a reference T cell; In this case, the reference T cells are isolated from a subject that has been contacted with a control antibody, and In this case, the surface expression of at least one activation marker on the reference T cells is compared. In comparison, an increase in surface expression of at least one activation marker on the T cells suggests that the anti-CD137 antibody induced or enhanced T cell activation.

[0333] In some embodiments, the anti-CD137 antibodies described herein do not induce or enhance intrahepatic T cell activation as determined by an in vivo T cell activation assay, wherein the T cell activation assay comprises the following steps: (i) administering anti-CD137 to a subject; (ii) isolating T cells from the liver of the subject; (iii) detecting surface expression of at least one activation marker on the T cells; and (iv) comparing the surface expression of at least one activation marker on the T cells with the surface expression of the at least one activation marker on a reference T cell; In this case, the reference T cells are isolated from a subject who has not received an anti-CD137 antibody; optionally, the reference T cells are isolated from a subject who has received a control antibody; and In this case, the surface expression of at least one activation marker on the reference T cells is compared. no increase in surface expression of at least one activation marker on the T cells compared to This suggests that anti-CD137 antibodies induced or enhanced T cell activation.

[0334] In some embodiments, the anti-CD137 antibodies described herein do not induce or enhance intrasplenic T cell activation as determined by an in vivo T cell activation assay, wherein the T cell activation assay comprises the following steps: (i) administering anti-CD137 to a subject; (ii) isolating T cells from the subject's spleen; (iii) detecting surface expression of at least one activation marker on the T cells; and (iv) comparing the surface expression of at least one activation marker on the T cells with the surface expression of the at least one activation marker on a reference T cell; In this case, the reference T cells are isolated from a subject who has not received an anti-CD137 antibody; optionally, the reference T cells are isolated from a subject who has received a control antibody; and In this case, the surface expression of at least one activation marker on the reference T cells is compared. no increase in surface expression of at least one activation marker on the T cells compared to This suggests that anti-CD137 antibodies induced or enhanced T cell activation.

[0335] In some embodiments, "does not induce or enhance" refers to the absence of an activity (e.g., T cell activation) or lack of an increase in activity compared to the increase caused by a reference antibody. It is intended that

[0336] In some embodiments, the surface expression of a T cell activation marker is determined by measuring the surface expression in the absence of an antibody. In some embodiments, surface expression of a T cell activation marker is equivalent to the presence of a reference antibody. The reference antibody induces or enhances surface expression by at least 1-fold, 5-fold, 10-fold, 50-fold, or 100-fold compared to surface expression in the absence of antibody.

[0337] In some embodiments, the at least one activation marker is selected from CD25, CD69, and CD40L. In some embodiments, the one or more activation markers is CD25.

[0338] In some embodiments, the T cells are isolated from a subject with a tumor. In some embodiments, T cells are isolated from the tumor. In some embodiments, the control antibody is an isotype It is a target antibody.

[0339] In some embodiments, the anti-CD137 antibodies described herein are useful in immune cell infiltration assays. In some embodiments, the anti-CD137 antibodies described herein induce or enhance the infiltration of one or more immune cells into the tumor microenvironment, as determined by the immunohistochemistry. The antibody reduces the infiltration of one or more immune cells into the tumor microenvironment as determined by an immunoassay.

[0340] In s...

Claims

[Claim 1] The method described in the specification.

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