Expansion of tumor-infiltrating lymphocytes (TILs) with tumor necrosis factor receptor superfamily (TNFRSF) agonists and therapeutic combination of TILs with TNFRSF agonists

Abstract

Disclosed herein are methods for expanding tumor-infiltrating lymphocytes (TILs) using tumor necrosis factor receptor superfamily (TNFRSF) agonists, such as 4-1BB agonists, CD27 agonists, glucocorticoid-inducible TNF receptor-associated agonists, OX40 agonists, HVEM agonists, or CD95 agonists, and the use of such expanded TILs in the treatment of diseases such as cancer. Additionally, disclosed herein in some embodiments are therapeutic combinations of TILs with TNFRSF agonists useful in the treatment of diseases such as cancer, including compositions, uses, and administration regimens thereof.

Description

[Technical Field]

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS

[0001] This international application claims the benefit of priority to U.S. Provisional Patent Application No. 62 / 443,556, filed January 6, 2017, U.S. Provisional Patent Application No. 62 / 460,477, filed February 17, 2017, U.S. Provisional Patent Application No. 62 / 532,807, filed July 14, 2017, and U.S. Provisional Patent Application No. 62 / 567,151, filed October 2, 2017, which are incorporated herein by reference in their entireties.

[0002] FIELD OF THE INVENTION Disclosed herein are methods for expanding tumor-infiltrating lymphocytes (TILs) using tumor necrosis factor receptor superfamily (TNFRSF) agonists, such as 4-1BB agonists, CD27 agonists, glucocorticoid-inducible TNF receptor-associated agonists, OX40 agonists, HVEM agonists, or CD95 agonists, and uses of the expanded TILs in the treatment of diseases such as cancer. Additionally, disclosed herein are therapeutic combinations of TILs with TNFRSF agonists in the treatment of diseases such as cancer, including compositions and uses thereof. [Background technology]

[0003] Background of the Invention

[0003] Treatment of large, refractory cancers using autologous adoptive transfer of tumor-infiltrating lymphocytes (TILs) represents a powerful therapeutic approach for patients with poor prognosis. Gattinoni, et al., Nat. Rev. Immunol. 2006, 6, 383-393. TILs are predominantly T cells, and IL-2-based TIL expansion followed by the "rapid expansion process" (REP) has become the preferred method for TIL expansion due to its speed and efficiency. Dudley, et al., Science 2002, 298, 850-54; Dudley, et al., J. Clin. Oncol. 2005, 23, 2346-57; Dudley, et al., J. Clin. Oncol. 2008, 26, 5233-39; Riddell, et al., Science 1992, 257, 238-41; Dudley, et al., J. Immunother. 2003, 26, 332-42. Several approaches to improve clinical response to TIL therapy in melanoma and to extend TIL therapy to other tumor types have been investigated, but success has been limited, and this area remains challenging. Goff, et al., J. Clin. Oncol. 2016, 34, 2389-97; Dudley, et al., J. Clin. Oncol. 2008, 26, 5233-39; Rosenberg, et al., Clin. Cancer Res. 2011, 17, 4550-57. Much interest has focused on specific subsets (CD8 +Previous research has focused on selecting TILs during expansion to select for specific T cells (e.g., T cells) or to target driver mutations, such as mutant ERBB2IP epitopes or driver mutations in the KRAS oncogene. Tran, et al., N. Engl. J. Med. 2016, 375, 2255-62; Tran, et al., Science 2014, 344, 641-45. However, even if such selection methods can be developed to demonstrate efficacy in large-scale clinical trials, they significantly increase the time, complexity, and cost required to implement TIL therapy, limiting its potential for widespread use in different types of cancer.

[0004] 4-1BB (also known as CD137 and TNFRSF9) was originally identified as an inducible costimulatory receptor expressed on activated T cells and is a transmembrane glycoprotein member of the TNFRSF. Watts, Annu. Rev. Immunol. 2005, 23, 23-68. 4-1BB is expressed on activated T lymphocytes and on CD4 + than CD8 + 4-1BB is a type 2 transmembrane glycoprotein that is more abundantly expressed in T cells. It is also expressed on dendritic cells, follicular dendritic cells, natural killer (NK) cells, granulocytes, vascular mural cells at sites of inflammation, tumor vasculature, and atherosclerotic endothelium. The stimulatory ligand for 4-1BB (4-1BBL) is expressed on activated antigen-presenting cells (APCs), myeloid progenitor cells, and hematopoietic stem cells. 4-1BB is an activation-induced T cell costimulatory molecule. Signaling through 4-1BB upregulates survival genes in T cells, promotes cell division, induces cytokine production, and prevents activation-induced cell death. Current understanding of 4-1BB indicates that its expression is largely activation-dependent and encompasses a wide range of immune cell subsets, including activated NK and NK T cells (NKT cells); regulatory T cells; dendritic cells (DCs), including follicular DCs; stimulated mast cells, differentiated myeloid cells, monocytes, neutrophils, eosinophils, and activated B cells. 4-1BB is a CD8 +It potently enhances T cell proliferation and effector function. Cross-linking of 4-1BB enhances T cell proliferation, IL-2 secretion, survival, and cytolytic activity. In addition, anti-4-1BB monoclonal antibodies have potent antitumor properties, which in turn result from their potent ability to activate CD8+ T cells, produce IFN-g, and induce cytolytic markers. Vinay and Kwon, Mol. Cancer Therapeutics 2012, 11, 1062-70; Lee, et al., PLoS One, 2013, 8, e69677, 1-11.

[0005]

[0005] The interaction of 4-1BB with its ligand on activated normal human B cells upon B cell receptor engagement stimulates proliferation and enhances survival. The potential impact of 4-1BB engagement in B cell lymphoma has been investigated in at least two published studies. Evaluation of several types of human primary NHL specimens showed that 4-1BB is primarily expressed on infiltrating T cells rather than on lymphoma cells. Houot, et al., Blood, 2009, 114, 3431-38. Addition of a 4-1BB agonist to in vitro cultures of B lymphoma cells, along with rituximab and NK cells, increased lymphoma killing. Kohrt, et al., Blood, 2011, 117, 2423-32. Additionally, B cell immunophenotyping was performed in cynomolgus monkeys in two studies using PF-05082566 at doses of 0.001-100 mg / kg; as described in WO 2015 / 119923, peripheral blood B cell counts were either unchanged or decreased in these studies.

[0006]

[0006] 4-1BB is undetectable on the surface of naive T cells, but its expression increases upon activation. Activation of 4-1BB recruits two pro-survival members of the TNFR-associated factor (TRAF) family, TRAF1 and TRAF2, to the 4-1BB cytoplasmic tail, resulting in downstream activation of NFkB and the mitogen-activated protein (MAP) kinase cascade, including Erk, Jnk, and p38 MAP kinase. NFkB activation leads to upregulation of Bfl-1 and Bel-XL, pro-survival members of the Bcl-2 family. The pro-apoptotic protein Bim is downregulated in a TRAF1- and Erk-dependent manner. (Sabbagh, et al., J. Immunol. 2008, 180, 8093-8101). 4-1BB agonist monoclonal antibodies (mAbs) have been reported to increase costimulatory molecule expression and significantly enhance cytolytic T lymphocyte responses, resulting in antitumor efficacy in various models. 4-1BB agonist mAbs have demonstrated efficacy in tumor models in both preventative and therapeutic settings, as monotherapy and in combination therapy, establishing durable antitumor protective T cell memory responses. Lynch, et al., Immunol Rev., 2008, 222, 277-286. 4-1BB agonists also suppress autoimmune responses in various autoimmune models. Vinay, et al., J. Mol. Med., 2006, 84, 726-36.

[0007] The OX40 receptor (OX40) (also known as TNFRSF4, CD134, ACT-4, and ACT35) binds activated CD4 + It is a member of the TNF receptor family expressed on T cells (see WO 95 / 12673). Triggering of this receptor by the OX40 ligand, also known as OX40L, gp34, or ACT-4 ligand, present on activated B cells and dendritic cells, promotes the activation of CD4 receptors during an immune response. + T cell proliferation is increased, and CD4 +The formation of memory T cells is affected. Furthermore, the OX40-OX40L system mediates the adhesion of activated T cells to endothelial cells, which in turn mediates the activation of CD4 + Guides T cells to the site of inflammation.

[0008]

[0008] OX40 + T cells have been shown to be present within tumor lesions, including tumor-infiltrating lymphocytes, and in tumor-positive draining lymph nodes. Weinberg, et al., J. Immunol., 2000, 164, 2160-2169. In several tumor models in mice, OX40 receptor engagement during in vivo tumor priming significantly delayed or prevented tumor development compared with control-treated mice. Weinberg, et al., J. Immunol., 2000, 164, 2160-2169. Therefore, administration of OX40 receptor binding agents is intended to enhance mammalian immune responses to antigens via OX40 receptor engagement (WO 1999 / 042585; Weinberg, et al., J. Immunol., 2000, 164, 2160-2169). Preclinical studies have demonstrated that treatment of tumor-bearing hosts with OX40 agonists, including both anti-OX40 monoclonal antibodies and OX40L-Fc fusion proteins, resulted in tumor regression in several preclinical models. Linch, et al., Front. Oncol. 2015, 34, 1-14.

[0009]

[0009] CD27, also known as TNFRSF7, has overlapping activities with other TNFRSF members, including CD40, 4-1BB, and OX40. CD27 plays a crucial role in T cell survival, activation, and effector function, and also plays a role in the proliferative and cytotoxic activity of NK cells. CD27 is constitutively expressed on the majority of T cells, including naive T cells. The ligand for CD27 is CD70, which is found on T cells, B cells, and dendritic cells. Oshima, et al., Int. Immunol. 1998, 10, 517-26. CD27 acts after CD28 to sustain T effector cell survival and regulates CD4 + and CD8 + CD27 activation drives T cell expansion and is more influential in secondary than primary responses. However, CD27 activation has also been associated with tumor growth through enhancing the immunosuppressive effects of regulatory T cells. Claus, et al., Cancer Res. 2012, 72, 3664-76. Other data suggest that the immunostimulatory effects of CD27 may outweigh this tumor-promoting effect. Aulwurm, et al., Int. J. Cancer 2006, 118, 1728-35. In mouse models, agonistic CD27 monoclonal antibodies demonstrated antitumor efficacy and induced tumor immunity. He, et al., J. Immunol. 2013, 191, 4174-83.

[0010] Glucocorticoid-inducible TNFR-related protein (GITR), also known as tumor necrosis factor receptor superfamily member 18 (TNFRSF18), activation-induced TNFR family receptor (AITR), and CD357, is a costimulatory checkpoint molecule. GITR is expressed on several cell types, including regulatory T cells (Tregs) and effector T cells, B cells, NK cells, and antigen-presenting cells. Nocentini and Riccardi, Eur. J. Immunol. 2005, 35, 1016-1022. GITR is activated by its conjugate, GITR ligand (GITRL). GITR plays a role in stimulating immune responses, and antigen-binding proteins against GITR are useful in treating various GITR-associated diseases or disorders in which it is desirable to increase the immune response. Ko, et al., J. Exp. Med. 2005, 202, 885-91; Shimizu, et al., Nature Immunology 2002, 3, 135-142; Cohen, et al., Cancer Res. 2006, 66, 4904-12; Azuma, Crit. Rev. Immunol. 2010, 30, 547-57. For example, T cell stimulation through GITR attenuates Treg-mediated suppression and enhances CD4 + and CD8 + GITR enhances tumor killing by T cells. + CD25 + or CD8 + CD25 + GITR is highly constitutively expressed in CD4+ cells and is upregulated upon activation of these cells. Nocentini and Riccardi, Eur. J. Immunol. 2005, 35, 1016-1022. + and CD8 +GITR is a coactivation signal for naive T cells, inducing and enhancing proliferation and effector function, particularly in situations where T cell receptor (TCR) stimulation is suboptimal. Schaer, et al., Curr. Opin. Immunol. 2012, 24, 217-224. Enhancement of immune responses elicited by antigen-binding GITR proteins, such as fusion proteins and anti-GITR antibodies (including agonist antibodies), is beneficial in various immunotherapy applications, such as the treatment of cancer, autoimmune diseases, inflammatory diseases, or infectious diseases.

[0011] Herpesvirus entry mediator (HVEM), also known as TNFRSF14 and CD270, was originally isolated as a receptor for herpes simplex virus type 1 (HSV-1). Montgomery, et al., Cell 1996, 87, 427-36. HVEM binds to the TNF family ligands LIGHT and lymphotoxin α homotrimer (Ltα3). Mauri, et al., Immunity 1998, 8, 21-30. T cell activation can occur through HVEM-LIGHT interaction, which provides T cells with a costimulatory signal independent of CD28 signaling and can be observed in the presence of suboptimal levels of CD3 antibody (OKT-3). Tamada, et al., J. Immunol. 2000, 165, 4397-404; Harrop, et al., J. Biol. Chem. 1998, 273, 27548-56; Tamada, et al., Nat. Med. 2000, 6, 283-89; Yu, et al., Nat. Immunol. 2004, 5, 141-49. HVEM contains four cysteine-rich domains (CRDs). del Rio, et al., J. Leukoc. Biol. 2010, 87, 223-35. CRD2 and CRD3 are required for HVEM trimerization with the TNFRSF ligand LIGHT, and LIGHT transmits costimulatory signals to T cells through HVEM. In contrast, CRD1 and CRD2 bind to the co-inhibitory B and T lymphocyte attenuator (BTLA) receptor and CD160 in a monomeric manner, providing inhibitory signals to T cells. Studies of HVEM-LIGHT interaction suggest that it has a CD28-independent costimulatory effect primarily on CD8+ T cells, but also affects CD4+ T cells. Liu, et al., Int. Immunol. 2003, 75, 861-70; Scheu, et al., J. Exp. Med. 2002, 195, 1613-24.

[0012] CD95, also known as Fas, APO-1, and TNFRSF6, is a 45 kDa type I transmembrane protein that, unlike 4-1BB, OX40, GITR, CD27, and HVEM, contains a death domain. Kischkel, et al., EMBO J. 1995, 14, 5579-88; Krammer, Nature 2000, 407, 789-95. Binding of inducible CD95 ligand (CD95L) to CD95 on activated T cells leads to apoptotic cell death, and therefore it is not normally associated with the same costimulatory functions as 4-1BB, OX40, GITR, CD27, and HVEM. Strauss, et al., J. Exp. Med. 2009, 206, 1379-93. However, CD95 also behaves as a dual-function receptor, providing antiapoptotic and costimulatory effects on T cells under certain conditions. (Paulsen, et al., Cell Death Differ. 2011, 18, 619-31) CD95 engagement regulates TCR-driven signaling in a dose-dependent manner; high doses of CD95 agonists or cellular CD95L silence T cells, whereas low doses of these agonists potently enhance TCR-driven T cell activation and proliferation. Summary of the Invention [Problem to be solved by the invention]

[0013]

[0013] The present invention provides the unexpected discovery that TNFRSF agonists such as 4-1BB agonists, CD27 agonists, GITR agonists, OX40 agonists, HVEM agonists, or CD95 agonists are useful for expanding and culturing TILs from tumors known to be difficult to obtain and treat with TILs, and further useful for treating patients in combination with TIL therapy. [Means for solving the problem]

[0014] Summary of the Invention In one embodiment, the present invention provides a method of treating cancer with a tumor-infiltrating lymphocyte (TIL) population, the method comprising: (a) removing a tumor from a patient; (b) obtaining a first population of TILs from the tumor; (c) obtaining a second TIL population by initial expansion of the first TIL population in a first cell culture medium, wherein the second TIL population is at least five-fold more numerous than the first TIL population, the first cell culture medium comprising IL-2 and a tumor necrosis factor receptor superfamily (TNFRSF) agonist, and the initial expansion is for a period of 21 days or less; (d) obtaining a third TIL population by rapid expansion of the second TIL population in a second cell culture medium, wherein the third TIL population is at least 50-fold more numerous than the second TIL population 7 days after initiation of the rapid expansion; the second cell culture medium comprises IL-2, OKT-3 (anti-CD3 antibody), peripheral blood mononuclear cells (PBMCs), and optionally a TNFRSF agonist, and the rapid expansion is performed for a period of 14 days or less; (e) recovering the third population of TILs; and (f) administering a therapeutically effective amount of the third population of TILs to a patient with cancer.

[0015] In one embodiment, the present invention provides a method of treating cancer with a tumor-infiltrating lymphocyte (TIL) population, the method comprising: (a) removing a tumor from a patient; (b) obtaining a first population of TILs from the tumor; (c) obtaining a second TIL population by initial expansion of the first TIL population in a first cell culture medium, wherein the second TIL population is at least five-fold more numerous than the first TIL population, the first cell culture medium comprising IL-2 and a tumor necrosis factor receptor superfamily (TNFRSF) agonist, and the initial expansion is for a period of 21 days or less; (d) obtaining a third TIL population by rapid expansion of the second TIL population in a second cell culture medium, wherein the third TIL population is at least 50-fold more numerous than the second TIL population 7 days after initiation of the rapid expansion; the second cell culture medium comprises IL-2, OKT-3 (anti-CD3 antibody), peripheral blood mononuclear cells (PBMCs), and optionally a TNFRSF agonist, and the rapid expansion is performed for a period of 14 days or less; (e) recovering the third population of TILs; and (f) administering a therapeutically effective amount of the third population of TILs to a patient having cancer, wherein the TNFRSF agonist is selected from the group consisting of a 4-1BB agonist, an OX40 agonist, a CD27 agonist, a GITR agonist, an HVEM agonist, a CD95 agonist, and combinations thereof.

[0016] In one embodiment, the present invention provides a method of treating cancer with a tumor-infiltrating lymphocyte (TIL) population, the method comprising: (a) removing a tumor from a patient; (b) obtaining a first population of TILs from the tumor; (c) obtaining a second TIL population by initial expansion of the first TIL population in a first cell culture medium, wherein the second TIL population is at least five-fold more numerous than the first TIL population, the first cell culture medium comprising IL-2 and a tumor necrosis factor receptor superfamily (TNFRSF) agonist, and the initial expansion is for a period of 21 days or less; (d) obtaining a third TIL population by rapid expansion of the second TIL population in a second cell culture medium, wherein the third TIL population is at least 50-fold more numerous than the second TIL population 7 days after initiation of the rapid expansion; the second cell culture medium comprises IL-2, OKT-3 (anti-CD3 antibody), peripheral blood mononuclear cells (PBMCs), and optionally a TNFRSF agonist, and the rapid expansion is performed for a period of 14 days or less; (e) recovering the third population of TILs; and (f) administering a therapeutically effective amount of the third population of TILs to a patient with cancer, wherein the TNFRSF agonist is a 4-1BB agonist, and the 4-1BB agonist is selected from the group consisting of urelumab, utomilumab, EU-101, and fragments, derivatives, variants, biosimilars, and combinations thereof.

[0017] In one embodiment, the present invention provides a method of treating cancer with a tumor-infiltrating lymphocyte (TIL) population, the method comprising: (a) removing a tumor from a patient; (b) obtaining a first population of TILs from the tumor; (c) obtaining a second TIL population by initial expansion of the first TIL population in a first cell culture medium, wherein the second TIL population is at least five-fold more numerous than the first TIL population, the first cell culture medium comprising IL-2 and a tumor necrosis factor receptor superfamily (TNFRSF) agonist, and the initial expansion is for a period of 21 days or less; (d) obtaining a third TIL population by rapid expansion of the second TIL population in a second cell culture medium, wherein the third TIL population is at least 50-fold more numerous than the second TIL population 7 days after initiation of the rapid expansion; the second cell culture medium comprises IL-2, OKT-3 (anti-CD3 antibody), peripheral blood mononuclear cells (PBMCs), and optionally a TNFRSF agonist, and the rapid expansion is performed for a period of 14 days or less; (e) recovering the third population of TILs; and (f) administering to the patient having cancer a therapeutically effective amount of the third population of TILs, wherein the TNFRSF agonist is a 4-1BB agonist, and the 4-1BB agonist is a 4-1BB agonist fusion protein.

[0018] In one embodiment, the present invention provides a method of treating cancer with a tumor-infiltrating lymphocyte (TIL) population, the method comprising: (a) removing a tumor from a patient; (b) obtaining a first population of TILs from the tumor; (c) obtaining a second TIL population by initial expansion of the first TIL population in a first cell culture medium, wherein the second TIL population is at least five-fold more numerous than the first TIL population, the first cell culture medium comprising IL-2 and a tumor necrosis factor receptor superfamily (TNFRSF) agonist, and the initial expansion is for a period of 21 days or less; (d) obtaining a third TIL population by rapid expansion of the second TIL population in a second cell culture medium, wherein the third TIL population is at least 50-fold more numerous than the second TIL population 7 days after initiation of the rapid expansion; the second cell culture medium comprises IL-2, OKT-3 (anti-CD3 antibody), peripheral blood mononuclear cells (PBMCs), and optionally a TNFRSF agonist, and the rapid expansion is performed for a period of 14 days or less; (e) recovering the third population of TILs; and (f) administering a therapeutically effective amount of a third population of TILs to a patient having cancer, wherein the TNFRSF agonist is a 4-1BB agonist fusion protein, and the 4-1BB agonist fusion protein comprises (i) a first soluble 4-1BB binding domain, (ii) a first peptide linker, (iii) a second soluble 4-1BB binding domain, (iv) a second peptide linker, and (v) a third soluble 4-1BB binding domain, and further comprises additional domains at the N-terminal and / or C-terminal end, and the additional domains comprise an Fc fragment domain and a hinge domain, and the fusion protein is a dimeric structure according to Structure IA or Structure IB.

[0019] In one embodiment, the present invention provides a method of treating cancer with a tumor-infiltrating lymphocyte (TIL) population, the method comprising: (a) removing a tumor from a patient; (b) obtaining a first population of TILs from the tumor; (c) obtaining a second TIL population by initial expansion of the first TIL population in a first cell culture medium, wherein the second TIL population is at least five-fold more numerous than the first TIL population, the first cell culture medium comprising IL-2 and a tumor necrosis factor receptor superfamily (TNFRSF) agonist, and the initial expansion is for a period of 21 days or less; (d) obtaining a third TIL population by rapid expansion of the second TIL population in a second cell culture medium, wherein the third TIL population is at least 50-fold more numerous than the second TIL population 7 days after initiation of the rapid expansion; the second cell culture medium comprises IL-2, OKT-3 (anti-CD3 antibody), peripheral blood mononuclear cells (PBMCs), and optionally a TNFRSF agonist, and the rapid expansion is performed for a period of 14 days or less; (e) recovering the third population of TILs; and (f) administering a therapeutically effective amount of the third population of TILs to a patient having cancer, wherein the TNFRSF agonist is an OX40 agonist, and the OX40 agonist is selected from the group consisting of tavolixizumab, GSK3174998, MEDI6469, MEDI6383, MOXR0916, PF-04518600, CreativeBiolabs MOM-18455, and fragments, derivatives, variants, biosimilars, and combinations thereof.

[0020] In one embodiment, the present invention provides a method of treating cancer with a tumor-infiltrating lymphocyte (TIL) population, the method comprising: (a) removing a tumor from a patient; (b) obtaining a first population of TILs from the tumor; (c) obtaining a second TIL population by initial expansion of the first TIL population in a first cell culture medium, wherein the second TIL population is at least five-fold more numerous than the first TIL population, the first cell culture medium comprising IL-2 and a tumor necrosis factor receptor superfamily (TNFRSF) agonist, and the initial expansion is for a period of 21 days or less; (d) obtaining a third TIL population by rapid expansion of the second TIL population in a second cell culture medium, wherein the third TIL population is at least 50-fold more numerous than the second TIL population 7 days after initiation of the rapid expansion; the second cell culture medium comprises IL-2, OKT-3 (anti-CD3 antibody), peripheral blood mononuclear cells (PBMCs), and optionally a TNFRSF agonist, and the rapid expansion is performed for a period of 14 days or less; (e) recovering the third population of TILs; and (f) administering to the patient having cancer a therapeutically effective amount of the third population of TILs, wherein the TNFRSF agonist is an OX40 agonist, and the OX40 agonist is an OX40 agonist fusion protein.

[0021] In one embodiment, the present invention provides a method of treating cancer with a tumor-infiltrating lymphocyte (TIL) population, the method comprising: (a) removing a tumor from a patient; (b) obtaining a first population of TILs from the tumor; (c) obtaining a second TIL population by initial expansion of the first TIL population in a first cell culture medium, wherein the second TIL population is at least five-fold more numerous than the first TIL population, the first cell culture medium comprising IL-2 and a tumor necrosis factor receptor superfamily (TNFRSF) agonist, and the initial expansion is for a period of 21 days or less; (d) obtaining a third TIL population by rapid expansion of the second TIL population in a second cell culture medium, wherein the third TIL population is at least 50-fold more numerous than the second TIL population 7 days after initiation of the rapid expansion; the second cell culture medium comprises IL-2, OKT-3 (anti-CD3 antibody), peripheral blood mononuclear cells (PBMCs), and optionally a TNFRSF agonist, and the rapid expansion is performed for a period of 14 days or less; (e) recovering the third population of TILs; and (f) administering a therapeutically effective amount of a third population of TILs to a patient having cancer, wherein the TNFRSF agonist is an OX40 agonist fusion protein, and the OX40 agonist fusion protein comprises (i) a first soluble OX40 binding domain, (ii) a first peptide linker, (iii) a second soluble OX40 binding domain, (iv) a second peptide linker, and (v) a third soluble OX40 binding domain, and further comprises additional domains at the N-terminal and / or C-terminal end, and the additional domains comprise an Fc fragment domain and a hinge domain, and the fusion protein is a dimeric structure according to Structure IA or Structure IB.

[0022] In one embodiment, the present invention provides a method of treating cancer with a tumor-infiltrating lymphocyte (TIL) population, the method comprising: (a) removing a tumor from a patient; (b) obtaining a first population of TILs from the tumor; (c) obtaining a second TIL population by initial expansion of the first TIL population in a first cell culture medium, wherein the second TIL population is at least five-fold more numerous than the first TIL population, the first cell culture medium comprising IL-2 and a tumor necrosis factor receptor superfamily (TNFRSF) agonist, and the initial expansion is for a period of 21 days or less; (d) obtaining a third TIL population by rapid expansion of the second TIL population in a second cell culture medium, wherein the third TIL population is at least 50-fold more numerous than the second TIL population 7 days after initiation of the rapid expansion; the second cell culture medium comprises IL-2, OKT-3 (anti-CD3 antibody), peripheral blood mononuclear cells (PBMCs), and optionally a TNFRSF agonist, and the rapid expansion is performed for a period of 14 days or less; (e) recovering the third population of TILs; and (f) administering a therapeutically effective amount of the third population of TILs to a patient having cancer, wherein the TNFRSF agonist is a CD27 agonist, and the CD27 agonist is varlilumab, or a fragment, derivative, variant, or biosimilar thereof.

[0023] In one embodiment, the present invention provides a method of treating cancer with a tumor-infiltrating lymphocyte (TIL) population, the method comprising: (a) removing a tumor from a patient; (b) obtaining a first population of TILs from the tumor; (c) obtaining a second TIL population by initial expansion of the first TIL population in a first cell culture medium, wherein the second TIL population is at least five-fold more numerous than the first TIL population, the first cell culture medium comprising IL-2 and a tumor necrosis factor receptor superfamily (TNFRSF) agonist, and the initial expansion is for a period of 21 days or less; (d) obtaining a third TIL population by rapid expansion of the second TIL population in a second cell culture medium, wherein the third TIL population is at least 50-fold more numerous than the second TIL population 7 days after initiation of the rapid expansion; the second cell culture medium comprises IL-2, OKT-3 (anti-CD3 antibody), peripheral blood mononuclear cells (PBMCs), and optionally a TNFRSF agonist, and the rapid expansion is performed for a period of 14 days or less; (e) recovering the third population of TILs; and (f) administering to the patient having cancer a therapeutically effective amount of the third population of TILs, wherein the TNFRSF agonist is a CD27 agonist, and the CD27 agonist is a CD27 agonist fusion protein.

[0024] In one embodiment, the present invention provides a method of treating cancer with a tumor-infiltrating lymphocyte (TIL) population, the method comprising: (a) removing a tumor from a patient; (b) obtaining a first population of TILs from the tumor; (c) obtaining a second TIL population by initial expansion of the first TIL population in a first cell culture medium, wherein the second TIL population is at least five-fold more numerous than the first TIL population, the first cell culture medium comprising IL-2 and a tumor necrosis factor receptor superfamily (TNFRSF) agonist, and the initial expansion is for a period of 21 days or less; (d) obtaining a third TIL population by rapid expansion of the second TIL population in a second cell culture medium, wherein the third TIL population is at least 50-fold more numerous than the second TIL population 7 days after initiation of the rapid expansion; the second cell culture medium comprises IL-2, OKT-3 (anti-CD3 antibody), peripheral blood mononuclear cells (PBMCs), and optionally a TNFRSF agonist, and the rapid expansion is performed for a period of 14 days or less; (e) recovering the third population of TILs; and (f) administering a therapeutically effective amount of a third population of TILs to a patient having cancer, wherein the TNFRSF agonist is a CD27 agonist, and the CD27 agonist fusion protein comprises (i) a first soluble CD27-binding domain, (ii) a first peptide linker, (iii) a second soluble CD27-binding domain, (iv) a second peptide linker, and (v) a third soluble CD27-binding domain, and further comprises additional domains at the N-terminal and / or C-terminal ends, and the additional domains comprise an Fc fragment domain and a hinge domain, and the fusion protein is a dimeric structure according to Structure IA or Structure IB.

[0025] In one embodiment, the present invention provides a method of treating cancer with a tumor-infiltrating lymphocyte (TIL) population, the method comprising: (a) removing a tumor from a patient; (b) obtaining a first population of TILs from the tumor; (c) obtaining a second TIL population by initial expansion of the first TIL population in a first cell culture medium, wherein the second TIL population is at least five-fold more numerous than the first TIL population, the first cell culture medium comprising IL-2 and a tumor necrosis factor receptor superfamily (TNFRSF) agonist, and the initial expansion is for a period of 21 days or less; (d) obtaining a third TIL population by rapid expansion of the second TIL population in a second cell culture medium, wherein the third TIL population is at least 50-fold more numerous than the second TIL population 7 days after initiation of the rapid expansion; the second cell culture medium comprises IL-2, OKT-3 (anti-CD3 antibody), peripheral blood mononuclear cells (PBMCs), and optionally a TNFRSF agonist, and the rapid expansion is performed for a period of 14 days or less; (e) recovering the third population of TILs; and (f) administering a therapeutically effective amount of the third population of TILs to a patient having cancer, wherein the TNFRSF agonist is a GITR agonist, and the GITR agonist is selected from the group consisting of TRX518, 6C8, 36E5, 3D6, 61G6, 6H6, 61F6, 1D8, 17F10, 35D8, 49A1, 9E5, 31H6, 2155, 698, 70 6, 827, 1649, 1718, 1D7, 33C9, 33F6, 34G4, 35B10, 41E11, 41G5, 42A11, 44C1, 45A8, 46E11, 48H12, 48H7, 49D9, 49E2, 48A9, 5H7, 7A10, 9H6, and fragments, derivatives, variants, biosimilars, and combinations thereof.

[0026] In one embodiment, the present invention provides a method of treating cancer with a tumor-infiltrating lymphocyte (TIL) population, the method comprising: (a) removing a tumor from a patient; (b) obtaining a first population of TILs from the tumor; (c) obtaining a second TIL population by initial expansion of the first TIL population in a first cell culture medium, wherein the second TIL population is at least five-fold more numerous than the first TIL population, the first cell culture medium comprising IL-2 and a tumor necrosis factor receptor superfamily (TNFRSF) agonist, and the initial expansion is for a period of 21 days or less; (d) obtaining a third TIL population by rapid expansion of the second TIL population in a second cell culture medium, wherein the third TIL population is at least 50-fold more numerous than the second TIL population 7 days after initiation of the rapid expansion; the second cell culture medium comprises IL-2, OKT-3 (anti-CD3 antibody), peripheral blood mononuclear cells (PBMCs), and optionally a TNFRSF agonist, and the rapid expansion is performed for a period of 14 days or less; (e) recovering the third population of TILs; and (f) administering to a patient having cancer a therapeutically effective amount of the third population of TILs, wherein the TNFRSF agonist is a GITR agonist, and the GITR agonist is a GITR agonist fusion protein.

[0027] In one embodiment, the present invention provides a method of treating cancer with a tumor-infiltrating lymphocyte (TIL) population, the method comprising: (a) removing a tumor from a patient; (b) obtaining a first population of TILs from the tumor; (c) obtaining a second TIL population by initial expansion of the first TIL population in a first cell culture medium, wherein the second TIL population is at least five-fold more numerous than the first TIL population, the first cell culture medium comprising IL-2 and a tumor necrosis factor receptor superfamily (TNFRSF) agonist, and the initial expansion is for a period of 21 days or less; (d) obtaining a third TIL population by rapid expansion of the second TIL population in a second cell culture medium, wherein the third TIL population is at least 50-fold more numerous than the second TIL population 7 days after initiation of the rapid expansion; the second cell culture medium comprises IL-2, OKT-3 (anti-CD3 antibody), peripheral blood mononuclear cells (PBMCs), and optionally a TNFRSF agonist, and the rapid expansion is performed for a period of 14 days or less; (e) recovering the third population of TILs; and (f) administering a therapeutically effective amount of a third population of TILs to a patient having cancer, wherein the TNFRSF agonist is a GITR agonist fusion protein, and the GITR agonist fusion protein comprises (i) a first soluble GITR binding domain, (ii) a first peptide linker, (iii) a second soluble GITR binding domain, (iv) a second peptide linker, and (v) a third soluble GITR binding domain, and further comprises additional domains at the N-terminal and / or C-terminal end, and the additional domains comprise an Fc fragment domain and a hinge domain, and the fusion protein is a dimeric structure according to Structure IA or Structure IB.

[0028] In one embodiment, the present invention provides a method of treating cancer with a tumor-infiltrating lymphocyte (TIL) population, the method comprising: (a) removing a tumor from a patient; (b) obtaining a first population of TILs from the tumor; (c) obtaining a second TIL population by initial expansion of the first TIL population in a first cell culture medium, wherein the second TIL population is at least five-fold more numerous than the first TIL population, the first cell culture medium comprising IL-2 and a tumor necrosis factor receptor superfamily (TNFRSF) agonist, and the initial expansion is for a period of 21 days or less; (d) obtaining a third TIL population by rapid expansion of the second TIL population in a second cell culture medium, wherein the third TIL population is at least 50-fold more numerous than the second TIL population 7 days after initiation of the rapid expansion; the second cell culture medium comprises IL-2, OKT-3 (anti-CD3 antibody), peripheral blood mononuclear cells (PBMCs), and optionally a TNFRSF agonist, and the rapid expansion is performed for a period of 14 days or less; (e) recovering the third population of TILs; and (f) administering to the patient having cancer a therapeutically effective amount of the third population of TILs, wherein the TNFRSF agonist is an HVEM agonist.

[0029] In one embodiment, the present invention provides a method of treating cancer with a tumor-infiltrating lymphocyte (TIL) population, the method comprising: (a) removing a tumor from a patient; (b) obtaining a first population of TILs from the tumor; (c) obtaining a second TIL population by initial expansion of the first TIL population in a first cell culture medium, wherein the second TIL population is at least five-fold more numerous than the first TIL population, the first cell culture medium comprising IL-2 and a tumor necrosis factor receptor superfamily (TNFRSF) agonist, and the initial expansion is for a period of 21 days or less; (d) obtaining a third TIL population by rapid expansion of the second TIL population in a second cell culture medium, wherein the third TIL population is at least 50-fold more numerous than the second TIL population 7 days after initiation of the rapid expansion; the second cell culture medium comprises IL-2, OKT-3 (anti-CD3 antibody), peripheral blood mononuclear cells (PBMCs), and optionally a TNFRSF agonist, and the rapid expansion is performed for a period of 14 days or less; (e) recovering the third population of TILs; and (f) administering to a patient having cancer a therapeutically effective amount of a third population of TILs, wherein the TNFRSF agonist is an HVEM agonist, and the HVEM agonist is an HVEM agonist fusion protein.

[0030] In one embodiment, the present invention provides a method of treating cancer with a tumor-infiltrating lymphocyte (TIL) population, the method comprising: (a) removing a tumor from a patient; (b) obtaining a first population of TILs from the tumor; (c) obtaining a second TIL population by initial expansion of the first TIL population in a first cell culture medium, wherein the second TIL population is at least five-fold more numerous than the first TIL population, the first cell culture medium comprising IL-2 and a tumor necrosis factor receptor superfamily (TNFRSF) agonist, and the initial expansion is for a period of 21 days or less; (d) obtaining a third TIL population by rapid expansion of the second TIL population in a second cell culture medium, wherein the third TIL population is at least 50-fold more numerous than the second TIL population 7 days after initiation of the rapid expansion; the second cell culture medium comprises IL-2, OKT-3 (anti-CD3 antibody), peripheral blood mononuclear cells (PBMCs), and optionally a TNFRSF agonist, and the rapid expansion is performed for a period of 14 days or less; (e) recovering the third population of TILs; and (f) administering a therapeutically effective amount of a third population of TILs to a patient having cancer, wherein the TNFRSF agonist is an HVEM agonist fusion protein, and the HVEM agonist fusion protein comprises (i) a first soluble HVEM-binding domain, (ii) a first peptide linker, (iii) a second soluble HVEM-binding domain, (iv) a second peptide linker, and (v) a third soluble HVEM-binding domain, and further comprises additional domains at the N-terminal and / or C-terminal ends, and the additional domains comprise an Fc fragment domain and a hinge domain, and the fusion protein is a dimeric structure according to Structure IA or Structure IB.

[0031] In one embodiment, the present invention provides a method of treating cancer with a tumor-infiltrating lymphocyte (TIL) population, the method comprising: (a) removing a tumor from a patient; (b) obtaining a first population of TILs from the tumor; (c) obtaining a second TIL population by initial expansion of the first TIL population in a first cell culture medium, wherein the second TIL population is at least five-fold more numerous than the first TIL population, the first cell culture medium comprising IL-2 and a tumor necrosis factor receptor superfamily (TNFRSF) agonist, and the initial expansion is for a period of 21 days or less; (d) obtaining a third TIL population by rapid expansion of the second TIL population in a second cell culture medium, wherein the third TIL population is at least 50-fold more numerous than the second TIL population 7 days after initiation of the rapid expansion; the second cell culture medium comprises IL-2, OKT-3 (anti-CD3 antibody), peripheral blood mononuclear cells (PBMCs), and optionally a TNFRSF agonist, and the rapid expansion is performed for a period of 14 days or less; (e) recovering the third population of TILs; and (f) administering a therapeutically effective amount of a third population of TILs to a patient having cancer, and further comprising treating the patient with a TNFRSF agonist starting the day after administration of the third population of TILs to the patient, wherein the TNFRSF agonist is administered intravenously at a dose of 0.1 mg / kg to 50 mg / kg every four weeks for up to eight cycles.

[0032] In one embodiment, the present invention provides a method of treating cancer with a tumor-infiltrating lymphocyte (TIL) population, the method comprising: (a) removing a tumor from a patient; (b) obtaining a first population of TILs from the tumor; (c) obtaining a second TIL population by initial expansion of the first TIL population in a first cell culture medium, wherein the second TIL population is at least five-fold more numerous than the first TIL population, the first cell culture medium comprising IL-2 and a tumor necrosis factor receptor superfamily (TNFRSF) agonist, and the initial expansion is for a period of 21 days or less; (d) obtaining a third TIL population by rapid expansion of the second TIL population in a second cell culture medium, wherein the third TIL population is at least 50-fold more numerous than the second TIL population 7 days after initiation of the rapid expansion; the second cell culture medium comprises IL-2, OKT-3 (anti-CD3 antibody), peripheral blood mononuclear cells (PBMCs), and optionally a TNFRSF agonist, and the rapid expansion is performed for a period of 14 days or less; (e) recovering the third population of TILs; and (f) administering a therapeutically effective amount of the third population of TILs to a patient having cancer, further comprising treating the patient with a TNFRSF agonist prior to resecting the tumor from the patient, wherein the TNFRSF agonist is administered intravenously at a dose of 0.1 mg / kg to 50 mg / kg every four weeks for up to eight cycles.

[0033] In one embodiment, the present invention provides a method of treating cancer with a tumor-infiltrating lymphocyte (TIL) population, the method comprising: (a) removing a tumor from a patient; (b) obtaining a first population of TILs from the tumor; (c) obtaining a second TIL population by initial expansion of the first TIL population in a first cell culture medium, wherein the second TIL population is at least five-fold more numerous than the first TIL population, the first cell culture medium comprising IL-2 and a tumor necrosis factor receptor superfamily (TNFRSF) agonist, and the initial expansion is for a period of 21 days or less; (d) obtaining a third TIL population by rapid expansion of the second TIL population in a second cell culture medium, wherein the third TIL population is at least 50-fold more numerous than the second TIL population 7 days after initiation of the rapid expansion; the second cell culture medium comprises IL-2, OKT-3 (anti-CD3 antibody), peripheral blood mononuclear cells (PBMCs), and optionally a TNFRSF agonist, and the rapid expansion is performed for a period of 14 days or less; (e) recovering the third population of TILs; and (f) administering a therapeutically effective amount of the third population of TILs to a patient having cancer, wherein the TNFRSF agonist is selected from the group consisting of urelumab, utomilumab, EU-101, taborixizumab, Creative Biolabs MOM-18455, and fragments, derivatives, variants, biosimilars, and combinations thereof.

[0034] In one embodiment, the present invention provides a method of treating cancer with a tumor-infiltrating lymphocyte (TIL) population, the method comprising: (a) removing a tumor from a patient; (b) obtaining a first population of TILs from the tumor; (c) obtaining a second TIL population by initial expansion of the first TIL population in a first cell culture medium, wherein the second TIL population is at least five-fold more numerous than the first TIL population, the first cell culture medium comprising IL-2 and a tumor necrosis factor receptor superfamily (TNFRSF) agonist, and the initial expansion is for a period of 21 days or less; (d) obtaining a third TIL population by rapid expansion of the second TIL population in a second cell culture medium, wherein the third TIL population is at least 50-fold more numerous than the second TIL population 7 days after initiation of the rapid expansion; the second cell culture medium comprises IL-2, OKT-3 (anti-CD3 antibody), peripheral blood mononuclear cells (PBMCs), and optionally a TNFRSF agonist, and the rapid expansion is performed for a period of 14 days or less; (e) recovering the third population of TILs; and (f) administering to the patient having cancer a therapeutically effective amount of the third population of TILs, wherein the first cell culture medium comprises a second TNFRSF agonist.

[0035] In one embodiment, the present invention provides a method of treating cancer with a tumor-infiltrating lymphocyte (TIL) population, the method comprising: (a) removing a tumor from a patient; (b) obtaining a first population of TILs from the tumor; (c) obtaining a second TIL population by initial expansion of the first TIL population in a first cell culture medium, wherein the second TIL population is at least five-fold more numerous than the first TIL population, the first cell culture medium comprising IL-2 and a tumor necrosis factor receptor superfamily (TNFRSF) agonist, and the initial expansion is for a period of 21 days or less; (d) obtaining a third TIL population by rapid expansion of the second TIL population in a second cell culture medium, wherein the third TIL population is at least 50-fold more numerous than the second TIL population 7 days after initiation of the rapid expansion; the second cell culture medium comprises IL-2, OKT-3 (anti-CD3 antibody), peripheral blood mononuclear cells (PBMCs), and optionally a TNFRSF agonist, and the rapid expansion is performed for a period of 14 days or less; (e) recovering the third population of TILs; and (f) administering a therapeutically effective amount of the third TIL population to a patient having cancer, wherein the TNFRSF agonist is added to the first cell culture medium at intervals selected from the group consisting of daily, every 2 days, every 3 days, every 4 days, every 5 days, every 6 days, every 7 days, and every 2 weeks during the initial expansion culture.

[0036] In one embodiment, the present invention provides a method of treating cancer with a tumor-infiltrating lymphocyte (TIL) population, the method comprising: (a) removing a tumor from a patient; (b) obtaining a first population of TILs from the tumor; (c) obtaining a second TIL population by initial expansion of the first TIL population in a first cell culture medium, wherein the second TIL population is at least five-fold more numerous than the first TIL population, the first cell culture medium comprising IL-2 and a tumor necrosis factor receptor superfamily (TNFRSF) agonist, and the initial expansion is for a period of 21 days or less; (d) obtaining a third TIL population by rapid expansion of the second TIL population in a second cell culture medium, wherein the third TIL population is at least 50-fold more numerous than the second TIL population 7 days after initiation of the rapid expansion; the second cell culture medium comprises IL-2, OKT-3 (anti-CD3 antibody), peripheral blood mononuclear cells (PBMCs), and optionally a TNFRSF agonist, and the rapid expansion is performed for a period of 14 days or less; (e) recovering the third population of TILs; and (f) administering a therapeutically effective amount of the third population of TILs to a patient having cancer, wherein the TNFRSF agonist is added to the second cell culture medium during the rapid expansion culture at intervals selected from the group consisting of every day, every two days, every three days, every four days, every five days, every six days, every seven days, and every two weeks.

[0037] In one embodiment, the present invention provides a method of treating cancer with a tumor-infiltrating lymphocyte (TIL) population, the method comprising: (a) removing a tumor from a patient; (b) obtaining a first population of TILs from the tumor; (c) obtaining a second TIL population by initial expansion of the first TIL population in a first cell culture medium, wherein the second TIL population is at least five-fold more numerous than the first TIL population, the first cell culture medium comprising IL-2 and a tumor necrosis factor receptor superfamily (TNFRSF) agonist, and the initial expansion is for a period of 21 days or less; (d) obtaining a third TIL population by rapid expansion of the second TIL population in a second cell culture medium, wherein the third TIL population is at least 50-fold more numerous than the second TIL population 7 days after initiation of the rapid expansion; the second cell culture medium comprises IL-2, OKT-3 (anti-CD3 antibody), peripheral blood mononuclear cells (PBMCs), and optionally a TNFRSF agonist, and the rapid expansion is performed for a period of 14 days or less; (e) recovering the third population of TILs; and (f) administering a therapeutically effective amount of the third TIL population to a patient having cancer, wherein the TNFRSF agonist is added at a concentration sufficient to achieve a concentration of 0.1 μg / mL to 100 μg / mL in the cell culture medium.

[0038] In one embodiment, the present invention provides a method of treating cancer with a tumor-infiltrating lymphocyte (TIL) population, the method comprising: (a) removing a tumor from a patient; (b) obtaining a first population of TILs from the tumor; (c) obtaining a second TIL population by initial expansion of the first TIL population in a first cell culture medium, wherein the second TIL population is at least five-fold more numerous than the first TIL population, the first cell culture medium comprising IL-2 and a tumor necrosis factor receptor superfamily (TNFRSF) agonist, and the initial expansion is for a period of 21 days or less; (d) obtaining a third TIL population by rapid expansion of the second TIL population in a second cell culture medium, wherein the third TIL population is at least 50-fold more numerous than the second TIL population 7 days after initiation of the rapid expansion; the second cell culture medium comprises IL-2, OKT-3 (anti-CD3 antibody), peripheral blood mononuclear cells (PBMCs), and optionally a TNFRSF agonist, and the rapid expansion is performed for a period of 14 days or less; (e) recovering the third population of TILs; and (f) administering a therapeutically effective amount of the third TIL population to a patient having cancer, wherein the TNFRSF agonist is added at a concentration sufficient to achieve a concentration of 20 μg / mL to 40 μg / mL in the cell culture medium.

[0039] In certain embodiments, the present invention provides a method of treating cancer with a tumor-infiltrating lymphocyte (TIL) population, the method comprising: (a) removing a tumor from a patient; (b) obtaining a first population of TILs from the tumor; (c) obtaining a second TIL population by initial expansion of the first TIL population in a first cell culture medium, wherein the second TIL population is at least five-fold more numerous than the first TIL population, the first cell culture medium comprising IL-2 and a tumor necrosis factor receptor superfamily (TNFRSF) agonist, and the initial expansion is for a period of 21 days or less; (d) obtaining a third TIL population by rapid expansion of the second TIL population in a second cell culture medium, wherein the third TIL population is at least 50-fold more numerous than the second TIL population 7 days after initiation of the rapid expansion; the second cell culture medium comprises IL-2, OKT-3 (anti-CD3 antibody), peripheral blood mononuclear cells (PBMCs), and optionally a TNFRSF agonist, and the rapid expansion is performed for a period of 14 days or less; (e) recovering the third population of TILs; and (f) administering to a patient having cancer a therapeutically effective amount of the third population of TILs, wherein IL-2 is present in the first cell culture medium at an initial concentration of about 10 to about 6000 IU / mL.

[0040] In one embodiment, the present invention provides a method of treating cancer with a tumor-infiltrating lymphocyte (TIL) population, the method comprising: (a) removing a tumor from a patient; (b) obtaining a first population of TILs from the tumor; (c) obtaining a second TIL population by initial expansion of the first TIL population in a first cell culture medium, wherein the second TIL population is at least five-fold more numerous than the first TIL population, the first cell culture medium comprising IL-2 and a tumor necrosis factor receptor superfamily (TNFRSF) agonist, and the initial expansion is for a period of 21 days or less; (d) obtaining a third TIL population by rapid expansion of the second TIL population in a second cell culture medium, wherein the third TIL population is at least 50-fold more numerous than the second TIL population 7 days after initiation of the rapid expansion; the second cell culture medium comprises IL-2, OKT-3 (anti-CD3 antibody), peripheral blood mononuclear cells (PBMCs), and optionally a TNFRSF agonist, and the rapid expansion is performed for a period of 14 days or less; (e) recovering the third population of TILs; and (f) administering to a patient having cancer a therapeutically effective amount of the third population of TILs, wherein IL-2 is present in the first cell culture medium at an initial concentration of about 3000 IU / mL.

[0041] In certain embodiments, the present invention provides a method of treating cancer with a tumor-infiltrating lymphocyte (TIL) population, the method comprising: (a) removing a tumor from a patient; (b) obtaining a first population of TILs from the tumor; (c) obtaining a second TIL population by initial expansion of the first TIL population in a first cell culture medium, wherein the second TIL population is at least five-fold more numerous than the first TIL population, the first cell culture medium comprising IL-2 and a tumor necrosis factor receptor superfamily (TNFRSF) agonist, and the initial expansion is for a period of 21 days or less; (d) obtaining a third TIL population by rapid expansion of the second TIL population in a second cell culture medium, wherein the third TIL population is at least 50-fold more numerous than the second TIL population 7 days after initiation of the rapid expansion; the second cell culture medium comprises IL-2, OKT-3 (anti-CD3 antibody), peripheral blood mononuclear cells (PBMCs), and optionally a TNFRSF agonist, and the rapid expansion is performed for a period of 14 days or less; (e) recovering the third population of TILs; and (f) administering to a patient having cancer a therapeutically effective amount of a third population of TILs, wherein IL-2 is present in the first cell culture medium at an initial concentration of about 800 to about 1100 IU / mL.

[0042] In one embodiment, the present invention provides a method of treating cancer with a tumor-infiltrating lymphocyte (TIL) population, the method comprising: (a) removing a tumor from a patient; (b) obtaining a first population of TILs from the tumor; (c) obtaining a second TIL population by initial expansion of the first TIL population in a first cell culture medium, wherein the second TIL population is at least five-fold more numerous than the first TIL population, the first cell culture medium comprising IL-2 and a tumor necrosis factor receptor superfamily (TNFRSF) agonist, and the initial expansion is for a period of 21 days or less; (d) obtaining a third TIL population by rapid expansion of the second TIL population in a second cell culture medium, wherein the third TIL population is at least 50-fold more numerous than the second TIL population 7 days after initiation of the rapid expansion; the second cell culture medium comprises IL-2, OKT-3 (anti-CD3 antibody), peripheral blood mononuclear cells (PBMCs), and optionally a TNFRSF agonist, and the rapid expansion is performed for a period of 14 days or less; (e) recovering the third population of TILs; and (f) administering to a patient having cancer a therapeutically effective amount of the third population of TILs, wherein IL-2 is present in the first cell culture medium at an initial concentration of about 1000 IU / mL.

[0043] In one embodiment, the present invention provides a method of treating cancer with a tumor-infiltrating lymphocyte (TIL) population, the method comprising: (a) removing a tumor from a patient; (b) obtaining a first population of TILs from the tumor; (c) obtaining a second TIL population by initial expansion of the first TIL population in a first cell culture medium, wherein the second TIL population is at least five-fold more numerous than the first TIL population, the first cell culture medium comprising IL-2 and a tumor necrosis factor receptor superfamily (TNFRSF) agonist, and the initial expansion is for a period of 21 days or less; (d) obtaining a third TIL population by rapid expansion of the second TIL population in a second cell culture medium, wherein the third TIL population is at least 50-fold more numerous than the second TIL population 7 days after initiation of the rapid expansion; the second cell culture medium comprises IL-2, OKT-3 (anti-CD3 antibody), peripheral blood mononuclear cells (PBMCs), and optionally a TNFRSF agonist, and the rapid expansion is performed for a period of 14 days or less; (e) recovering the third population of TILs; and (f) administering to a patient having cancer a therapeutically effective amount of the third population of TILs, wherein IL-2 is present in the second cell culture medium at an initial concentration of about 10 to about 6000 IU / mL.

[0044] In one embodiment, the present invention provides a method of treating cancer with a tumor-infiltrating lymphocyte (TIL) population, the method comprising: (a) removing a tumor from a patient; (b) obtaining a first population of TILs from the tumor; (c) obtaining a second TIL population by initial expansion of the first TIL population in a first cell culture medium, wherein the second TIL population is at least five-fold more numerous than the first TIL population, the first cell culture medium comprising IL-2 and a tumor necrosis factor receptor superfamily (TNFRSF) agonist, and the initial expansion is for a period of 21 days or less; (d) obtaining a third TIL population by rapid expansion of the second TIL population in a second cell culture medium, wherein the third TIL population is at least 50-fold more numerous than the second TIL population 7 days after initiation of the rapid expansion; the second cell culture medium comprises IL-2, OKT-3 (anti-CD3 antibody), peripheral blood mononuclear cells (PBMCs), and optionally a TNFRSF agonist, and the rapid expansion is performed for a period of 14 days or less; (e) recovering the third population of TILs; and (f) administering to a patient having cancer a therapeutically effective amount of the third population of TILs, wherein IL-2 is present in the second cell culture medium at an initial concentration of about 3000 IU / mL.

[0045] In one embodiment, the present invention provides a method of treating cancer with a tumor-infiltrating lymphocyte (TIL) population, the method comprising: (a) removing a tumor from a patient; (b) obtaining a first population of TILs from the tumor; (c) obtaining a second TIL population by initial expansion of the first TIL population in a first cell culture medium, wherein the second TIL population is at least five-fold more numerous than the first TIL population, the first cell culture medium comprising IL-2 and a tumor necrosis factor receptor superfamily (TNFRSF) agonist, and the initial expansion is for a period of 21 days or less; (d) obtaining a third TIL population by rapid expansion of the second TIL population in a second cell culture medium, wherein the third TIL population is at least 50-fold more numerous than the second TIL population 7 days after initiation of the rapid expansion; the second cell culture medium comprises IL-2, OKT-3 (anti-CD3 antibody), peripheral blood mononuclear cells (PBMCs), and optionally a TNFRSF agonist, and the rapid expansion is performed for a period of 14 days or less; (e) recovering the third population of TILs; and (f) administering to a patient having cancer a therapeutically effective amount of the third population of TILs, wherein IL-2 is present in the second cell culture medium at an initial concentration of about 800 to about 1100 IU / mL.

[0046] In certain embodiments, the present invention provides a method of treating cancer with a tumor-infiltrating lymphocyte (TIL) population, the method comprising: (a) removing a tumor from a patient; (b) obtaining a first population of TILs from the tumor; (c) obtaining a second TIL population by initial expansion of the first TIL population in a first cell culture medium, wherein the second TIL population is at least five-fold more numerous than the first TIL population, the first cell culture medium comprising IL-2 and a tumor necrosis factor receptor superfamily (TNFRSF) agonist, and the initial expansion is for a period of 21 days or less; (d) obtaining a third TIL population by rapid expansion of the second TIL population in a second cell culture medium, wherein the third TIL population is at least 50-fold more numerous than the second TIL population 7 days after initiation of the rapid expansion; the second cell culture medium comprises IL-2, OKT-3 (anti-CD3 antibody), peripheral blood mononuclear cells (PBMCs), and optionally a TNFRSF agonist, and the rapid expansion is performed for a period of 14 days or less; (e) recovering the third population of TILs; and (f) administering to a patient having cancer a therapeutically effective amount of the third population of TILs, wherein IL-2 is present in the second cell culture medium at an initial concentration of about 1000 IU / mL.

[0047] In one embodiment, the present invention provides a method of treating cancer with a tumor-infiltrating lymphocyte (TIL) population, the method comprising: (a) removing a tumor from a patient; (b) obtaining a first population of TILs from the tumor; (c) obtaining a second TIL population by initial expansion of the first TIL population in a first cell culture medium, wherein the second TIL population is at least five-fold more numerous than the first TIL population, the first cell culture medium comprising IL-2 and a tumor necrosis factor receptor superfamily (TNFRSF) agonist, and the initial expansion is for a period of 21 days or less; (d) obtaining a third TIL population by rapid expansion of the second TIL population in a second cell culture medium, wherein the third TIL population is at least 50-fold more numerous than the second TIL population 7 days after initiation of the rapid expansion; the second cell culture medium comprises IL-2, OKT-3 (anti-CD3 antibody), peripheral blood mononuclear cells (PBMCs), and optionally a TNFRSF agonist, and the rapid expansion is performed for a period of 14 days or less; (e) recovering the third population of TILs; and (f) administering to a patient having cancer a therapeutically effective amount of a third population of TILs, wherein IL-15 is present in the first cell culture medium.

[0048] In one embodiment, the present invention provides a method of treating cancer with a tumor-infiltrating lymphocyte (TIL) population, the method comprising: (a) removing a tumor from a patient; (b) obtaining a first population of TILs from the tumor; (c) obtaining a second TIL population by initial expansion of the first TIL population in a first cell culture medium, wherein the second TIL population is at least five-fold more numerous than the first TIL population, the first cell culture medium comprising IL-2 and a tumor necrosis factor receptor superfamily (TNFRSF) agonist, and the initial expansion is for a period of 21 days or less; (d) obtaining a third TIL population by rapid expansion of the second TIL population in a second cell culture medium, wherein the third TIL population is at least 50-fold more numerous than the second TIL population 7 days after initiation of the rapid expansion; the second cell culture medium comprises IL-2, OKT-3 (anti-CD3 antibody), peripheral blood mononuclear cells (PBMCs), and optionally a TNFRSF agonist, and the rapid expansion is performed for a period of 14 days or less; (e) recovering the third population of TILs; and (f) administering to a patient having cancer a therapeutically effective amount of a third population of TILs, wherein IL-15 is present in the first cell culture medium at an initial concentration of about 5 ng / mL to about 20 ng / mL.

[0049] In one embodiment, the present invention provides a method of treating cancer with a tumor-infiltrating lymphocyte (TIL) population, the method comprising: (a) removing a tumor from a patient; (b) obtaining a first population of TILs from the tumor; (c) obtaining a second TIL population by initial expansion of the first TIL population in a first cell culture medium, wherein the second TIL population is at least five-fold more numerous than the first TIL population, the first cell culture medium comprising IL-2 and a tumor necrosis factor receptor superfamily (TNFRSF) agonist, and the initial expansion is for a period of 21 days or less; (d) obtaining a third TIL population by rapid expansion of the second TIL population in a second cell culture medium, wherein the third TIL population is at least 50-fold more numerous than the second TIL population 7 days after initiation of the rapid expansion; the second cell culture medium comprises IL-2, OKT-3 (anti-CD3 antibody), peripheral blood mononuclear cells (PBMCs), and optionally a TNFRSF agonist, and the rapid expansion is performed for a period of 14 days or less; (e) recovering the third population of TILs; and (f) administering to a patient having cancer a therapeutically effective amount of a third population of TILs, wherein IL-15 is present in the second cell culture medium.

[0050] In one embodiment, the present invention provides a method of treating cancer with a tumor-infiltrating lymphocyte (TIL) population, the method comprising: (a) removing a tumor from a patient; (b) obtaining a first population of TILs from the tumor; (c) obtaining a second TIL population by initial expansion of the first TIL population in a first cell culture medium, wherein the second TIL population is at least five-fold more numerous than the first TIL population, the first cell culture medium comprising IL-2 and a tumor necrosis factor receptor superfamily (TNFRSF) agonist, and the initial expansion is for a period of 21 days or less; (d) obtaining a third TIL population by rapid expansion of the second TIL population in a second cell culture medium, wherein the third TIL population is at least 50-fold more numerous than the second TIL population 7 days after initiation of the rapid expansion; the second cell culture medium comprises IL-2, OKT-3 (anti-CD3 antibody), peripheral blood mononuclear cells (PBMCs), and optionally a TNFRSF agonist, and the rapid expansion is performed for a period of 14 days or less; (e) recovering the third population of TILs; and (f) administering to a patient having cancer a therapeutically effective amount of a third population of TILs, wherein IL-15 is present in the second cell culture medium at an initial concentration of about 5 ng / mL to about 20 ng / mL.

[0051] In certain embodiments, the present invention provides a method of treating cancer with a tumor-infiltrating lymphocyte (TIL) population, the method comprising: (a) removing a tumor from a patient; (b) obtaining a first population of TILs from the tumor; (c) obtaining a second TIL population by initial expansion of the first TIL population in a first cell culture medium, wherein the second TIL population is at least five-fold more numerous than the first TIL population, the first cell culture medium comprising IL-2 and a tumor necrosis factor receptor superfamily (TNFRSF) agonist, and the initial expansion is for a period of 21 days or less; (d) obtaining a third TIL population by rapid expansion of the second TIL population in a second cell culture medium, wherein the third TIL population is at least 50-fold more numerous than the second TIL population 7 days after initiation of the rapid expansion; the second cell culture medium comprises IL-2, OKT-3 (anti-CD3 antibody), peripheral blood mononuclear cells (PBMCs), and optionally a TNFRSF agonist, and the rapid expansion is performed for a period of 14 days or less; (e) recovering the third population of TILs; and (f) administering to a patient having cancer a therapeutically effective amount of a third population of TILs, wherein IL-21 is present in the first cell culture medium.

[0052] In certain embodiments, the present invention provides a method of treating cancer with a tumor-infiltrating lymphocyte (TIL) population, the method comprising: (a) removing a tumor from a patient; (b) obtaining a first population of TILs from the tumor; (c) obtaining a second TIL population by initial expansion of the first TIL population in a first cell culture medium, wherein the second TIL population is at least five-fold more numerous than the first TIL population, the first cell culture medium comprising IL-2 and a tumor necrosis factor receptor superfamily (TNFRSF) agonist, and the initial expansion is for a period of 21 days or less; (d) obtaining a third TIL population by rapid expansion of the second TIL population in a second cell culture medium, wherein the third TIL population is at least 50-fold more numerous than the second TIL population 7 days after initiation of the rapid expansion; the second cell culture medium comprises IL-2, OKT-3 (anti-CD3 antibody), peripheral blood mononuclear cells (PBMCs), and optionally a TNFRSF agonist, and the rapid expansion is performed for a period of 14 days or less; (e) recovering the third population of TILs; and (f) administering to a patient having cancer a therapeutically effective amount of a third population of TILs, wherein IL-21 is present in the first cell culture medium at an initial concentration of about 5 ng / mL to about 20 ng / mL.

[0053] In certain embodiments, the present invention provides a method of treating cancer with a tumor-infiltrating lymphocyte (TIL) population, the method comprising: (a) removing a tumor from a patient; (b) obtaining a first population of TILs from the tumor; (c) obtaining a second TIL population by initial expansion of the first TIL population in a first cell culture medium, wherein the second TIL population is at least five-fold more numerous than the first TIL population, the first cell culture medium comprising IL-2 and a tumor necrosis factor receptor superfamily (TNFRSF) agonist, and the initial expansion is for a period of 21 days or less; (d) obtaining a third TIL population by rapid expansion of the second TIL population in a second cell culture medium, wherein the third TIL population is at least 50-fold more numerous than the second TIL population 7 days after initiation of the rapid expansion; the second cell culture medium comprises IL-2, OKT-3 (anti-CD3 antibody), peripheral blood mononuclear cells (PBMCs), and optionally a TNFRSF agonist, and the rapid expansion is performed for a period of 14 days or less; (e) recovering the third population of TILs; and (f) administering to a patient having cancer a therapeutically effective amount of a third population of TILs, wherein IL-21 is present in the second cell culture medium.

[0054] In certain embodiments, the present invention provides a method of treating cancer with a tumor-infiltrating lymphocyte (TIL) population, the method comprising: (a) removing a tumor from a patient; (b) obtaining a first population of TILs from the tumor; (c) obtaining a second TIL population by initial expansion of the first TIL population in a first cell culture medium, wherein the second TIL population is at least five-fold more numerous than the first TIL population, the first cell culture medium comprising IL-2 and a tumor necrosis factor receptor superfamily (TNFRSF) agonist, and the initial expansion is for a period of 21 days or less; (d) obtaining a third TIL population by rapid expansion of the second TIL population in a second cell culture medium, wherein the third TIL population is at least 50-fold more numerous than the second TIL population 7 days after initiation of the rapid expansion; the second cell culture medium comprises IL-2, OKT-3 (anti-CD3 antibody), peripheral blood mononuclear cells (PBMCs), and optionally a TNFRSF agonist, and the rapid expansion is performed for a period of 14 days or less; (e) recovering the third population of TILs; and (f) administering to a patient having cancer a therapeutically effective amount of a third population of TILs, wherein IL-21 is present in the second cell culture medium at an initial concentration of about 5 ng / mL to about 20 ng / mL.

[0055] In certain embodiments, the present invention provides a method of treating cancer with a tumor-infiltrating lymphocyte (TIL) population, the method comprising: (a) removing a tumor from a patient; (b) obtaining a first population of TILs from the tumor; (c) obtaining a second TIL population by initial expansion of the first TIL population in a first cell culture medium, wherein the second TIL population is at least five-fold more numerous than the first TIL population, the first cell culture medium comprising IL-2 and a tumor necrosis factor receptor superfamily (TNFRSF) agonist, and the initial expansion is for a period of 21 days or less; (d) obtaining a third TIL population by rapid expansion of the second TIL population in a second cell culture medium, wherein the third TIL population is at least 50-fold more numerous than the second TIL population 7 days after initiation of the rapid expansion; the second cell culture medium comprises IL-2, OKT-3 (anti-CD3 antibody), peripheral blood mononuclear cells (PBMCs), and optionally a TNFRSF agonist, and the rapid expansion is performed for a period of 14 days or less; (e) recovering the third population of TILs; and (f) administering to a patient having cancer a therapeutically effective amount of the third TIL population, wherein the OKT-3 antibody is present in the second cell culture medium at an initial concentration of about 10 ng / mL to about 60 ng / mL.

[0056] In certain embodiments, the present invention provides a method of treating cancer with a tumor-infiltrating lymphocyte (TIL) population, the method comprising: (a) removing a tumor from a patient; (b) obtaining a first population of TILs from the tumor; (c) obtaining a second TIL population by initial expansion of the first TIL population in a first cell culture medium, wherein the second TIL population is at least five-fold more numerous than the first TIL population, the first cell culture medium comprising IL-2 and a tumor necrosis factor receptor superfamily (TNFRSF) agonist, and the initial expansion is for a period of 21 days or less; (d) obtaining a third TIL population by rapid expansion of the second TIL population in a second cell culture medium, wherein the third TIL population is at least 50-fold more numerous than the second TIL population 7 days after initiation of the rapid expansion; the second cell culture medium comprises IL-2, OKT-3 (anti-CD3 antibody), peripheral blood mononuclear cells (PBMCs), and optionally a TNFRSF agonist, and the rapid expansion is performed for a period of 14 days or less; (e) recovering the third population of TILs; and (f) administering to a patient having cancer a therapeutically effective amount of the third population of TILs, wherein the OKT-3 antibody is present in the second cell culture medium at an initial concentration of about 30 ng / mL.

[0057] In certain embodiments, the present invention provides a method of treating cancer with a tumor-infiltrating lymphocyte (TIL) population, the method comprising: (a) removing a tumor from a patient; (b) obtaining a first population of TILs from the tumor; (c) obtaining a second TIL population by initial expansion of the first TIL population in a first cell culture medium, wherein the second TIL population is at least five-fold more numerous than the first TIL population, the first cell culture medium comprising IL-2 and a tumor necrosis factor receptor superfamily (TNFRSF) agonist, and the initial expansion is for a period of 21 days or less; (d) obtaining a third TIL population by rapid expansion of the second TIL population in a second cell culture medium, wherein the third TIL population is at least 50-fold more numerous than the second TIL population 7 days after initiation of the rapid expansion; the second cell culture medium comprises IL-2, OKT-3 (anti-CD3 antibody), peripheral blood mononuclear cells (PBMCs), and optionally a TNFRSF agonist, and the rapid expansion is performed for a period of 14 days or less; (e) recovering the third population of TILs; and (f) administering a therapeutically effective amount of the third population of TILs to a patient with cancer, wherein the initial expansion is performed using a gas-permeable container.

[0058] In one embodiment, the present invention provides a method of treating cancer with a tumor-infiltrating lymphocyte (TIL) population, the method comprising: (a) removing a tumor from a patient; (b) obtaining a first population of TILs from the tumor; (c) obtaining a second TIL population by initial expansion of the first TIL population in a first cell culture medium, wherein the second TIL population is at least five-fold more numerous than the first TIL population, the first cell culture medium comprising IL-2 and a tumor necrosis factor receptor superfamily (TNFRSF) agonist, and the initial expansion is for a period of 21 days or less; (d) obtaining a third TIL population by rapid expansion of the second TIL population in a second cell culture medium, wherein the third TIL population is at least 50-fold more numerous than the second TIL population 7 days after initiation of the rapid expansion; the second cell culture medium comprises IL-2, OKT-3 (anti-CD3 antibody), peripheral blood mononuclear cells (PBMCs), and optionally a TNFRSF agonist, and the rapid expansion is performed for a period of 14 days or less; (e) recovering the third population of TILs; and (f) administering a therapeutically effective amount of the third population of TILs to a patient with cancer, wherein the rapid expansion is performed using a gas-permeable container.

[0059] In certain embodiments, the present invention provides a method of treating cancer with a tumor-infiltrating lymphocyte (TIL) population, the method comprising: (a) removing a tumor from a patient; (b) obtaining a first population of TILs from the tumor; (c) obtaining a second TIL population by initial expansion of the first TIL population in a first cell culture medium, wherein the second TIL population is at least five-fold more numerous than the first TIL population, the first cell culture medium comprising IL-2 and a tumor necrosis factor receptor superfamily (TNFRSF) agonist, and the initial expansion is for a period of 21 days or less; (d) obtaining a third TIL population by rapid expansion of the second TIL population in a second cell culture medium, wherein the third TIL population is at least 50-fold more numerous than the second TIL population 7 days after initiation of the rapid expansion; the second cell culture medium comprises IL-2, OKT-3 (anti-CD3 antibody), peripheral blood mononuclear cells (PBMCs), and optionally a TNFRSF agonist, and the rapid expansion is performed for a period of 14 days or less; (e) recovering the third population of TILs; and (f) administering a therapeutically effective amount of a third population of TILs to a patient having cancer, further comprising treating the patient with a non-myeloablative lymphodepletion regimen prior to administering the third population of TILs to the patient.

[0060] In certain embodiments, the present invention provides a method of treating cancer with a tumor-infiltrating lymphocyte (TIL) population, the method comprising: (a) removing a tumor from a patient; (b) obtaining a first population of TILs from the tumor; (c) obtaining a second TIL population by initial expansion of the first TIL population in a first cell culture medium, wherein the second TIL population is at least five-fold more numerous than the first TIL population, the first cell culture medium comprising IL-2 and a tumor necrosis factor receptor superfamily (TNFRSF) agonist, and the initial expansion is for a period of 21 days or less; (d) obtaining a third TIL population by rapid expansion of the second TIL population in a second cell culture medium, wherein the third TIL population is at least 50-fold more numerous than the second TIL population 7 days after initiation of the rapid expansion; the second cell culture medium comprises IL-2, OKT-3 (anti-CD3 antibody), peripheral blood mononuclear cells (PBMCs), and optionally a TNFRSF agonist, and the rapid expansion is performed for a period of 14 days or less; (e) recovering the third population of TILs; and (f) administering a therapeutically effective amount of a third population of TILs to a patient having cancer, further comprising treating the patient with a non-myeloablative lymphodepletion regimen prior to administering the third population of TILs to the patient, wherein the non-myeloablative lymphodepletion regimen is 60 mg / m 2 / day for 2 days followed by cyclophosphamide at a dose of 25 mg / m 2 The treatment includes administering fludarabine at a dose of 100 mg / day for 5 days.

[0061] In certain embodiments, the present invention provides a method of treating cancer with a tumor-infiltrating lymphocyte (TIL) population, the method comprising: (a) removing a tumor from a patient; (b) obtaining a first population of TILs from the tumor; (c) obtaining a second TIL population by initial expansion of the first TIL population in a first cell culture medium, wherein the second TIL population is at least five-fold more numerous than the first TIL population, the first cell culture medium comprising IL-2 and a tumor necrosis factor receptor superfamily (TNFRSF) agonist, and the initial expansion is for a period of 21 days or less; (d) obtaining a third TIL population by rapid expansion of the second TIL population in a second cell culture medium, wherein the third TIL population is at least 50-fold more numerous than the second TIL population 7 days after initiation of the rapid expansion; the second cell culture medium comprises IL-2, OKT-3 (anti-CD3 antibody), peripheral blood mononuclear cells (PBMCs), and optionally a TNFRSF agonist, and the rapid expansion is performed for a period of 14 days or less; (e) recovering the third population of TILs; and (f) administering a therapeutically effective amount of a third population of TILs to a patient having cancer, further comprising treating the patient with a tapering IL-2 regimen starting the day after administering the third population of TILs to the patient, wherein the tapering IL-2 regimen includes administering 18,000,000 IU / m on day 1; 2 , 9,000,000 IU / m on day 2 2, and 4,500,000 IU / m on days 3 and 4 2 and aldesleukin, administered intravenously at a dose of

[0062] In certain embodiments, the present invention provides a method of treating cancer with a tumor-infiltrating lymphocyte (TIL) population, the method comprising: (a) removing a tumor from a patient; (b) obtaining a first population of TILs from the tumor; (c) obtaining a second TIL population by initial expansion of the first TIL population in a first cell culture medium, wherein the second TIL population is at least five-fold more numerous than the first TIL population, the first cell culture medium comprising IL-2 and a tumor necrosis factor receptor superfamily (TNFRSF) agonist, and the initial expansion is for a period of 21 days or less; (d) obtaining a third TIL population by rapid expansion of the second TIL population in a second cell culture medium, wherein the third TIL population is at least 50-fold more numerous than the second TIL population 7 days after initiation of the rapid expansion; the second cell culture medium comprises IL-2, OKT-3 (anti-CD3 antibody), peripheral blood mononuclear cells (PBMCs), and optionally a TNFRSF agonist, and the rapid expansion is performed for a period of 14 days or less; (e) recovering the third population of TILs; and (f) administering a therapeutically effective amount of the third population of TILs to a patient having cancer, further comprising treating the patient with pegylated IL-2 at a dose of 0.10 mg / day to 50 mg / day after administering the third population of TILs to the patient.

[0063] In certain embodiments, the present invention provides a method of treating cancer with a tumor-infiltrating lymphocyte (TIL) population, the method comprising: (a) removing a tumor from a patient; (b) obtaining a first population of TILs from the tumor; (c) obtaining a second TIL population by initial expansion of the first TIL population in a first cell culture medium, wherein the second TIL population is at least five-fold more numerous than the first TIL population, the first cell culture medium comprising IL-2 and a tumor necrosis factor receptor superfamily (TNFRSF) agonist, and the initial expansion is for a period of 21 days or less; (d) obtaining a third TIL population by rapid expansion of the second TIL population in a second cell culture medium, wherein the third TIL population is at least 50-fold more numerous than the second TIL population 7 days after initiation of the rapid expansion; the second cell culture medium comprises IL-2, OKT-3 (anti-CD3 antibody), peripheral blood mononuclear cells (PBMCs), and optionally a TNFRSF agonist, and the rapid expansion is performed for a period of 14 days or less; (e) recovering the third population of TILs; and (f) administering a therapeutically effective amount of a third population of TILs to a patient having cancer, and further comprising treating the patient with a high-dose IL-2 regimen starting the day after administering the third population of TILs to the patient.

[0064] In certain embodiments, the present invention provides a method of treating cancer with a tumor-infiltrating lymphocyte (TIL) population, the method comprising: (a) removing a tumor from a patient; (b) obtaining a first population of TILs from the tumor; (c) obtaining a second TIL population by initial expansion of the first TIL population in a first cell culture medium, wherein the second TIL population is at least five-fold more numerous than the first TIL population, the first cell culture medium comprising IL-2 and a tumor necrosis factor receptor superfamily (TNFRSF) agonist, and the initial expansion is for a period of 21 days or less; (d) obtaining a third TIL population by rapid expansion of the second TIL population in a second cell culture medium, wherein the third TIL population is at least 50-fold more numerous than the second TIL population 7 days after initiation of the rapid expansion; the second cell culture medium comprises IL-2, OKT-3 (anti-CD3 antibody), peripheral blood mononuclear cells (PBMCs), and optionally a TNFRSF agonist, and the rapid expansion is performed for a period of 14 days or less; (e) recovering the third population of TILs; and (f) administering a therapeutically effective amount of a third population of TILs to a patient having cancer, further comprising treating the patient with a high-dose IL-2 regimen starting the day after administration of the third population of TILs to the patient, wherein the high-dose IL-2 regimen comprises 600,000 or 720,000 IU / kg of aldesleukin, or a biosimilar or variant thereof, administered as a 15-minute bolus intravenous infusion every 8 hours to a tolerated dose.

[0065] In certain embodiments, the present invention provides a method of treating cancer with a tumor-infiltrating lymphocyte (TIL) population, the method comprising: (a) removing a tumor from a patient; (b) obtaining a first population of TILs from the tumor; (c) obtaining a second TIL population by initial expansion of the first TIL population in a first cell culture medium, wherein the second TIL population is at least five-fold more numerous than the first TIL population, the first cell culture medium comprising IL-2 and a tumor necrosis factor receptor superfamily (TNFRSF) agonist, and the initial expansion is for a period of 21 days or less; (d) obtaining a third TIL population by rapid expansion of the second TIL population in a second cell culture medium, wherein the third TIL population is at least 50-fold more numerous than the second TIL population 7 days after initiation of the rapid expansion; the second cell culture medium comprises IL-2, OKT-3 (anti-CD3 antibody), peripheral blood mononuclear cells (PBMCs), and optionally a TNFRSF agonist, and the rapid expansion is performed for a period of 14 days or less; (e) recovering the third population of TILs; and (f) administering a therapeutically effective amount of the third population of TILs to a patient having cancer, wherein the cancer is selected from the group consisting of melanoma, ovarian cancer, cervical cancer, lung cancer, bladder cancer, breast cancer, head and neck cancer, renal cell carcinoma, acute myeloid leukemia, colorectal cancer, cholangiocarcinoma, and sarcoma.

[0066] In certain embodiments, the present invention provides a method of treating cancer with a tumor-infiltrating lymphocyte (TIL) population, the method comprising: (a) removing a tumor from a patient; (b) obtaining a first population of TILs from the tumor; (c) obtaining a second TIL population by initial expansion of the first TIL population in a first cell culture medium, wherein the second TIL population is at least five-fold more numerous than the first TIL population, the first cell culture medium comprising IL-2 and a tumor necrosis factor receptor superfamily (TNFRSF) agonist, and the initial expansion is for a period of 21 days or less; (d) obtaining a third TIL population by rapid expansion of the second TIL population in a second cell culture medium, wherein the third TIL population is at least 50-fold more numerous than the second TIL population 7 days after initiation of the rapid expansion; the second cell culture medium comprises IL-2, OKT-3 (anti-CD3 antibody), peripheral blood mononuclear cells (PBMCs), and optionally a TNFRSF agonist, and the rapid expansion is performed for a period of 14 days or less; (e) recovering the third population of TILs; and (f) administering a therapeutically effective amount of the third population of TILs to a patient having cancer, wherein the cancer is selected from the group consisting of non-small cell lung cancer (NSCLC), triple-negative breast cancer, double-refractory melanoma, and uveal (ocular) melanoma.

[0067] In certain embodiments, the present invention provides a method of treating cancer with a tumor-infiltrating lymphocyte (TIL) population, the method comprising: (a) removing a tumor from a patient; (b) obtaining a first population of TILs from the tumor; (c) obtaining a second TIL population by initial expansion of the first TIL population in a first cell culture medium, wherein the second TIL population is at least five-fold more numerous than the first TIL population, the first cell culture medium comprising IL-2 and a tumor necrosis factor receptor superfamily (TNFRSF) agonist, and the initial expansion is for a period of 21 days or less; (d) obtaining a third TIL population by rapid expansion of the second TIL population in a second cell culture medium, wherein the third TIL population is at least 50-fold more numerous than the second TIL population 7 days after initiation of the rapid expansion; the second cell culture medium comprises IL-2, OKT-3 (anti-CD3 antibody), peripheral blood mononuclear cells (PBMCs), and optionally a TNFRSF agonist, and the rapid expansion is performed for a period of 14 days or less; (e) recovering the third population of TILs; and (f) administering a therapeutically effective amount of the third population of TILs to a patient having cancer, and further comprising treating the patient with a PD-1 inhibitor or a PD-L1 inhibitor prior to resecting the tumor from the patient.

[0068] In certain embodiments, the present invention provides a method of treating cancer with a tumor-infiltrating lymphocyte (TIL) population, the method comprising: (a) removing a tumor from a patient; (b) obtaining a first population of TILs from the tumor; (c) obtaining a second TIL population by initial expansion of the first TIL population in a first cell culture medium, wherein the second TIL population is at least five-fold more numerous than the first TIL population, the first cell culture medium comprising IL-2 and a tumor necrosis factor receptor superfamily (TNFRSF) agonist, and the initial expansion is for a period of 21 days or less; (d) obtaining a third TIL population by rapid expansion of the second TIL population in a second cell culture medium, wherein the third TIL population is at least 50-fold more numerous than the second TIL population 7 days after initiation of the rapid expansion; the second cell culture medium comprises IL-2, OKT-3 (anti-CD3 antibody), peripheral blood mononuclear cells (PBMCs), and optionally a TNFRSF agonist, and the rapid expansion is performed for a period of 14 days or less; (e) recovering the third population of TILs; and (f) administering a therapeutically effective amount of the third population of TILs to a patient having cancer, further comprising treating the patient with a PD-1 inhibitor or a PD-L1 inhibitor prior to resecting the tumor from the patient, wherein the PD-1 inhibitor or PD-L1 inhibitor is selected from the group consisting of nivolumab, pembrolizumab, durvalumab, atezolizumab, avelumab, and fragments, derivatives, variants, biosimilars, and combinations thereof.

[0069] In certain embodiments, the present invention provides a method of treating cancer with a tumor-infiltrating lymphocyte (TIL) population, the method comprising: (a) removing a tumor from a patient; (b) obtaining a first population of TILs from the tumor; (c) obtaining a second TIL population by initial expansion of the first TIL population in a first cell culture medium, wherein the second TIL population is at least five-fold more numerous than the first TIL population, the first cell culture medium comprising IL-2 and a tumor necrosis factor receptor superfamily (TNFRSF) agonist, and the initial expansion is for a period of 21 days or less; (d) obtaining a third TIL population by rapid expansion of the second TIL population in a second cell culture medium, wherein the third TIL population is at least 50-fold more numerous than the second TIL population 7 days after initiation of the rapid expansion; the second cell culture medium comprises IL-2, OKT-3 (anti-CD3 antibody), peripheral blood mononuclear cells (PBMCs), and optionally a TNFRSF agonist, and the rapid expansion is performed for a period of 14 days or less; (e) recovering the third population of TILs; and (f) administering a therapeutically effective amount of the third population of TILs to a patient having cancer, and further comprising treating the patient with a PD-1 inhibitor or a PD-L1 inhibitor after resecting the tumor from the patient.

[0070] In certain embodiments, the present invention provides a method of treating cancer with a tumor-infiltrating lymphocyte (TIL) population, the method comprising: (a) removing a tumor from a patient; (b) obtaining a first population of TILs from the tumor; (c) obtaining a second TIL population by initial expansion of the first TIL population in a first cell culture medium, wherein the second TIL population is at least five-fold more numerous than the first TIL population, the first cell culture medium comprising IL-2 and a tumor necrosis factor receptor superfamily (TNFRSF) agonist, and the initial expansion is for a period of 21 days or less; (d) obtaining a third TIL population by rapid expansion of the second TIL population in a second cell culture medium, wherein the third TIL population is at least 50-fold more numerous than the second TIL population 7 days after initiation of the rapid expansion; the second cell culture medium comprises IL-2, OKT-3 (anti-CD3 antibody), peripheral blood mononuclear cells (PBMCs), and optionally a TNFRSF agonist, and the rapid expansion is performed for a period of 14 days or less; (e) recovering the third population of TILs; and (f) administering a therapeutically effective amount of the third population of TILs to a patient having cancer, further comprising treating the patient with a PD-1 inhibitor or a PD-L1 inhibitor after resecting the tumor from the patient, wherein the PD-1 inhibitor or PD-L1 inhibitor is selected from the group consisting of nivolumab, pembrolizumab, durvalumab, atezolizumab, avelumab, and fragments, derivatives, variants, biosimilars, and combinations thereof.

[0071] In certain embodiments, the present invention provides a method of treating cancer with a tumor-infiltrating lymphocyte (TIL) population, the method comprising: (a) removing a tumor from a patient; (b) obtaining a first population of TILs from the tumor; (c) obtaining a second TIL population by initial expansion of the first TIL population in a first cell culture medium, wherein the second TIL population is at least five-fold more numerous than the first TIL population, the first cell culture medium comprising IL-2 and a tumor necrosis factor receptor superfamily (TNFRSF) agonist, and the initial expansion is for a period of 21 days or less; (d) obtaining a third TIL population by rapid expansion of the second TIL population in a second cell culture medium, wherein the third TIL population is at least 50-fold more numerous than the second TIL population 7 days after initiation of the rapid expansion; the second cell culture medium comprises IL-2, OKT-3 (anti-CD3 antibody), peripheral blood mononuclear cells (PBMCs), and optionally a TNFRSF agonist, and the rapid expansion is performed for a period of 14 days or less; (e) recovering the third population of TILs; and (f) administering a therapeutically effective amount of a third population of TILs to a patient having cancer, further comprising treating the patient with a PD-1 inhibitor or a PD-L1 inhibitor after administering the third population of TILs to the patient.

[0072] In certain embodiments, the present invention provides a method of treating cancer with a tumor-infiltrating lymphocyte (TIL) population, the method comprising: (a) removing a tumor from a patient; (b) obtaining a first population of TILs from the tumor; (c) obtaining a second TIL population by initial expansion of the first TIL population in a first cell culture medium, wherein the second TIL population is at least five-fold more numerous than the first TIL population, the first cell culture medium comprising IL-2 and a tumor necrosis factor receptor superfamily (TNFRSF) agonist, and the initial expansion is for a period of 21 days or less; (d) obtaining a third TIL population by rapid expansion of the second TIL population in a second cell culture medium, wherein the third TIL population is at least 50-fold more numerous than the second TIL population 7 days after initiation of the rapid expansion; the second cell culture medium comprises IL-2, OKT-3 (anti-CD3 antibody), peripheral blood mononuclear cells (PBMCs), and optionally a TNFRSF agonist, and the rapid expansion is performed for a period of 14 days or less; (e) recovering the third population of TILs; and (f) administering a therapeutically effective amount of the third population of TILs to a patient having cancer, further comprising treating the patient with a PD-1 inhibitor or a PD-L1 inhibitor after administering the third population of TILs to the patient, wherein the PD-1 inhibitor or PD-L1 inhibitor is selected from the group consisting of nivolumab, pembrolizumab, durvalumab, atezolizumab, avelumab, and fragments, derivatives, variants, biosimilars, and combinations thereof.

[0073] In one embodiment, the present invention provides a process for preparing a tumor infiltrating lymphocyte (TIL) population, the process comprising: (b) obtaining a first population of TILs; (c) obtaining a second TIL population by initial expansion of the first TIL population in a first cell culture medium, wherein the second TIL population is at least five-fold more numerous than the first TIL population, the first cell culture medium comprising IL-2 and a tumor necrosis factor receptor superfamily (TNFRSF) agonist, and the initial expansion is for a period of 21 days or less; (d) obtaining a third TIL population by rapid expansion of the second TIL population in a second cell culture medium, wherein the third TIL population is at least 50-fold more numerous than the second TIL population 7 days after initiation of the rapid expansion; the second cell culture medium comprises IL-2, OKT-3 (anti-CD3 antibody), peripheral blood mononuclear cells (PBMCs), and optionally a TNFRSF agonist, and the rapid expansion is performed for a period of 14 days or less; and (e) recovering a third population of TILs.

[0074] In one embodiment, the present invention provides (b) obtaining a first population of TILs; (c) obtaining a second TIL population by initial expansion of the first TIL population in a first cell culture medium, wherein the second TIL population is at least five-fold more numerous than the first TIL population, the first cell culture medium comprising IL-2 and a tumor necrosis factor receptor superfamily (TNFRSF) agonist, and the initial expansion is for a period of 21 days or less; (d) obtaining a third TIL population by rapid expansion of the second TIL population in a second cell culture medium, wherein the third TIL population is at least 50-fold more numerous than the second TIL population 7 days after initiation of the rapid expansion; the second cell culture medium comprises IL-2, OKT-3 (anti-CD3 antibody), peripheral blood mononuclear cells (PBMCs), and optionally a TNFRSF agonist, and the rapid expansion is performed for a period of 14 days or less; and (e) Recovering the third population of TILs The present invention provides a tumor-infiltrating lymphocyte (TIL) population obtainable from a process comprising:

[0075] In certain embodiments, the present invention provides a TIL population for use in the treatment of cancer. In certain embodiments, the present invention provides a pharmaceutical composition comprising a tumor-infiltrating lymphocyte (TIL) population for use in the treatment of cancer, wherein the tumor-infiltrating lymphocyte (TIL) population is (b) obtaining a first population of TILs; (c) obtaining a second TIL population by initial expansion of the first TIL population in a first cell culture medium, wherein the second TIL population is at least five-fold more numerous than the first TIL population, the first cell culture medium comprising IL-2 and a tumor necrosis factor receptor superfamily (TNFRSF) agonist, and the initial expansion is for a period of 21 days or less; (d) obtaining a third TIL population by rapid expansion of the second TIL population in a second cell culture medium, wherein the third TIL population is at least 50-fold more numerous than the second TIL population 7 days after initiation of the rapid expansion; the second cell culture medium comprises IL-2, OKT-3 (anti-CD3 antibody), peripheral blood mononuclear cells (PBMCs), and optionally a TNFRSF agonist, and the rapid expansion is performed for a period of 14 days or less; and (e) Recovering the third population of TILs It is obtainable by a process comprising:

[0076] In some embodiments, the first TIL population is obtained from a tumor. In some embodiments, the tumor is first resected from a patient. In some embodiments, the first TIL population is obtained from the tumor resected from the patient. In some embodiments, the TIL population is for administration in a therapeutically effective amount to a patient having cancer.

[0077] In one embodiment, the present invention provides a method for expanding a population of tumor-infiltrating lymphocytes (TILs), the method comprising: (a) removing a tumor from a patient; (b) obtaining a first population of TILs from the tumor; (c) obtaining a second TIL population by initial expansion of the first TIL population in a first cell culture medium, wherein the second TIL population is at least 5-fold more numerous than the first TIL population, the first cell culture medium comprises IL-2, and the initial expansion is for a period of 11 days or less; (d) obtaining a third TIL population by rapid expansion of the second TIL population in a second cell culture medium, wherein the third TIL population is at least 50-fold more numerous than the second TIL population 7 days after initiation of the rapid expansion; the second cell culture medium comprises IL-2, OKT-3 (anti-CD3) antibody, peripheral blood mononuclear cells (PBMCs), and a TNFRSF agonist, and the rapid expansion is performed for a period of 11 days or less; (e) recovering the third population of TILs; and (f) optionally, cryopreserving the third population of TILs in a medium containing dimethyl sulfoxide.

[0078] In certain embodiments, the present invention provides a method of treating cancer with a tumor-infiltrating lymphocyte (TIL) population, the method comprising: (a) removing a tumor from a patient; (b) obtaining a first population of TILs from the tumor; (c) obtaining a second TIL population by initial expansion of the first TIL population in a first cell culture medium, wherein the second TIL population is at least 5-fold more numerous than the first TIL population, the first cell culture medium comprises IL-2, and the initial expansion is for a period of 11 days or less; (d) obtaining a third TIL population by rapid expansion of the second TIL population in a second cell culture medium, wherein the third TIL population is at least 50-fold more numerous than the second TIL population 7 days after initiation of the rapid expansion; the second cell culture medium comprises IL-2, OKT-3 (anti-CD3) antibody, peripheral blood mononuclear cells (PBMCs), and a TNFRSF agonist, and the rapid expansion is performed for a period of 11 days or less; (e) recovering the third population of TILs; and (f) administering to the patient a therapeutically effective amount of the third population of TILs.

[0079] In certain embodiments, the present invention provides a method of treating cancer with a tumor-infiltrating lymphocyte (TIL) population, the method comprising: (a) removing a tumor from a patient; (b) obtaining a first population of TILs from the tumor; (c) obtaining a second TIL population by initial expansion of the first TIL population in a first cell culture medium, wherein the second TIL population is at least 5-fold more numerous than the first TIL population, the first cell culture medium comprises IL-2, and the initial expansion is for a period of 11 days or less; (d) obtaining a third TIL population by rapid expansion of the second TIL population in a second cell culture medium, wherein the third TIL population is at least 50-fold more numerous than the second TIL population 7 days after initiation of the rapid expansion; the second cell culture medium comprises IL-2, OKT-3 (anti-CD3) antibody, peripheral blood mononuclear cells (PBMCs), and a TNFRSF agonist, and the rapid expansion is performed for a period of 11 days or less; (e) recovering the third population of TILs; and (f) administering to the patient a therapeutically effective amount of a third population of TILs; wherein the TNFRSF agonist is selected from the group consisting of a 4-1BB agonist, an OX40 agonist, and a combination thereof.

[0080] In certain embodiments, the present invention provides a method of treating cancer with a tumor-infiltrating lymphocyte (TIL) population, the method comprising: (a) removing a tumor from a patient; (b) obtaining a first population of TILs from the tumor; (c) obtaining a second TIL population by initial expansion of the first TIL population in a first cell culture medium, wherein the second TIL population is at least 5-fold more numerous than the first TIL population, the first cell culture medium comprises IL-2, and the initial expansion is for a period of 11 days or less; (d) obtaining a third TIL population by rapid expansion of the second TIL population in a second cell culture medium, wherein the third TIL population is at least 50-fold more numerous than the second TIL population 7 days after initiation of the rapid expansion; the second cell culture medium comprises IL-2, OKT-3 (anti-CD3) antibody, peripheral blood mononuclear cells (PBMCs), and a TNFRSF agonist, and the rapid expansion is performed for a period of 11 days or less; (e) recovering the third population of TILs; and (f) administering to the patient a therapeutically effective amount of a third population of TILs; wherein the TNFRSF agonist is selected from the group consisting of a 4-1BB agonist, an OX40 agonist, and a combination thereof; and wherein the TNFRSF agonist is a 4-1BB agonist, and the 4-1BB agonist is selected from the group consisting of urelumab, utomilumab, EU-101, fusion proteins, and fragments, derivatives, variants, biosimilars, and combinations thereof.

[0081] In certain embodiments, the present invention provides a method of treating cancer with a tumor-infiltrating lymphocyte (TIL) population, the method comprising: (a) removing a tumor from a patient; (b) obtaining a first population of TILs from the tumor; (c) obtaining a second TIL population by initial expansion of the first TIL population in a first cell culture medium, wherein the second TIL population is at least 5-fold more numerous than the first TIL population, the first cell culture medium comprises IL-2, and the initial expansion is for a period of 11 days or less; (d) obtaining a third TIL population by rapid expansion of the second TIL population in a second cell culture medium, wherein the third TIL population is at least 50-fold more numerous than the second TIL population 7 days after initiation of the rapid expansion; the second cell culture medium comprises IL-2, OKT-3 (anti-CD3) antibody, peripheral blood mononuclear cells (PBMCs), and a TNFRSF agonist, and the rapid expansion is performed for a period of 11 days or less; (e) recovering the third population of TILs; and (f) administering to the patient a therapeutically effective amount of a third population of TILs; wherein the TNFRSF agonist is selected from the group consisting of a 4-1BB agonist, an OX40 agonist, and a combination thereof; and wherein the TNFRSF agonist is an OX40 agonist, and the OX40 agonist is selected from the group consisting of taborixizumab, GSK3174998, MEDI6469, MEDI6383, MOXR0916, PF-04518600, Creative Biolabs MOM-18455, and fragments, derivatives, variants, biosimilars, and combinations thereof; wherein the OX4 agonist is present at a concentration of 1 μg / mL to 30 μg / mL at the start of step (d).

[0082] In certain embodiments, the invention provides the method of any of the preceding embodiments, wherein the TNFRSF agonist is present at a concentration of 5 μg / mL to 20 μg / mL at the start of step (d).

[0083] In certain embodiments, the invention provides a method of any of the preceding embodiments, wherein the TNFRSF agonist is present at a concentration of about 10 μg / mL at the start of step (d).

[0084] In certain embodiments, the invention provides a method of any of the preceding embodiments, wherein the TNFRSF agonist is maintained at a concentration of between 1 μg / mL and 30 μg / mL throughout step (d).

[0085] In certain embodiments, the invention provides a method of any of the preceding embodiments, wherein the TNFRSF agonist is maintained at a concentration of 5 μg / mL to 20 μg / mL throughout step (d).

[0086] In certain embodiments, the invention provides a method of any of the preceding embodiments, wherein the TNFRSF agonist is maintained at a concentration of about 10 μg / mL throughout step (d).

[0087] In certain embodiments, the invention provides a method of any of the preceding embodiments, wherein the third TIL population is a population of TILs that is a reference CD8 + TIL vs CD4 + Increased CD8 compared to TIL ratio + TIL vs CD4 +The TILs exhibit a TIL ratio. In certain embodiments, the increased ratio is selected from the group consisting of at least 1% higher than the reference ratio, at least 2% higher than the reference ratio, at least 5% higher than the reference ratio, at least 10% higher than the reference ratio, at least 15% higher than the reference ratio, at least 20% higher than the reference ratio, at least 25% higher than the reference ratio, at least 30% higher than the reference ratio, at least 35% higher than the reference ratio, at least 40% higher than the reference ratio, at least 45% higher than the reference ratio, and at least 50% higher than the reference ratio. In certain embodiments, the increased ratio is 5% to 80% higher than the reference ratio. In certain embodiments, the increased ratio is 10% to 70% higher than the reference ratio. In certain embodiments, the increased ratio is 15% to 60% higher than the reference ratio. In any of the foregoing embodiments, the reference ratio is obtained from a third TIL population that is a responder to a TNFRSF agonist.

[0088] In certain embodiments, the invention provides a method of any of the preceding embodiments, wherein the cancer is selected from the group consisting of melanoma, uveal (ocular) melanoma, ovarian cancer, cervical cancer, lung cancer, bladder cancer, breast cancer, head and neck cancer (head and neck squamous cell carcinoma), renal cell carcinoma, colorectal cancer, pancreatic cancer, glioblastoma, cholangiocarcinoma, and sarcoma. In certain embodiments, the invention provides a method of any of the preceding embodiments, wherein the cancer is selected from the group consisting of cutaneous melanoma, uveal (ocular) melanoma, platinum-resistant ovarian cancer, pancreatic ductal adenocarcinoma, osteosarcoma, triple-negative breast cancer, and non-small cell lung cancer.

[0089]

[0089] In some embodiments, any of the above embodiments may be combined with any of the following embodiments.

[0090] In certain embodiments, the process is an in vitro or ex vivo process.

[0091] In certain embodiments, the TNFRSF agonist is selected from the group consisting of a 4-1BB agonist, an OX40 agonist, a CD27 agonist, a GITR agonist, an HVEM agonist, a CD95 agonist, and combinations thereof.

[0092] In certain embodiments, the TNFRSF agonist is a 4-1BB agonist.

[0093]

[0093] In one embodiment, the TNFRSF agonist is a 4-1BB agonist, and the 4-1BB agonist is selected from the group consisting of urelumab, utomilumab, EU-101, and fragments, derivatives, variants, biosimilars, and combinations thereof.

[0094] In certain embodiments, the TNFRSF agonist is a 4-1BB agonist, and the 4-1BB agonist is a 4-1BB agonist fusion protein.

[0095]

[0095] In one embodiment, the TNFRSF agonist is a 4-1BB agonist fusion protein, and the 4-1BB agonist fusion protein comprises (i) a first soluble 4-1BB binding domain, (ii) a first peptide linker, (iii) a second soluble 4-1BB binding domain, (iv) a second peptide linker, and (v) a third soluble 4-1BB binding domain, and further comprises additional domains at the N-terminal and / or C-terminal ends, and the additional domains comprise an Fc fragment domain and a hinge domain, and the fusion protein is a dimeric structure according to structure IA or structure IB.

[0096] In certain embodiments, the TNFRSF agonist is an OX40 agonist.

[0097]

[0097] In one embodiment, the TNFRSF agonist is an OX40 agonist, and the OX40 agonist is selected from the group consisting of tabolixizumab, GSK3174998, MEDI6469, MEDI6383, MOXR0916, PF-04518600, Creative Biolabs MOM-18455, and fragments, derivatives, variants, biosimilars, and combinations thereof.

[0098] In certain embodiments, the TNFRSF agonist is an OX40 agonist, and the OX40 agonist is an OX40 agonist fusion protein.

[0099]

[0099] In one embodiment, the TNFRSF agonist is an OX40 agonist fusion protein, and the OX40 agonist fusion protein comprises (i) a first soluble OX40 binding domain, (ii) a first peptide linker, (iii) a second soluble OX40 binding domain, (iv) a second peptide linker, and (v) a third soluble OX40 binding domain, and further comprises additional domains at the N-terminal and / or C-terminal ends, and the additional domains comprise an Fc fragment domain and a hinge domain, and the fusion protein is a dimeric structure according to structure IA or structure IB.

[0100] In certain embodiments, the TNFRSF agonist is a CD27 agonist.

[0101] In one embodiment, the TNFRSF agonist is a CD27 agonist, and the CD27 agonist is varlilumab, or a fragment, derivative, variant, or biosimilar thereof.

[0102] In certain embodiments, the TNFRSF agonist is a CD27 agonist, and the CD27 agonist is a CD27 agonist fusion protein.

[0103]

[0103] In one embodiment, the TNFRSF agonist is a CD27 agonist, and the CD27 agonist fusion protein comprises (i) a first soluble CD27 binding domain, (ii) a first peptide linker, (iii) a second soluble CD27 binding domain, (iv) a second peptide linker, and (v) a third soluble CD27 binding domain, and further comprises additional domains at the N-terminal and / or C-terminal ends, and the additional domains comprise an Fc fragment domain and a hinge domain, and the fusion protein is a dimeric structure according to structure IA or structure IB.

[0104] In certain embodiments, the TNFRSF agonist is a GITR agonist.

[0105]

[0105] In one embodiment, the TNFRSF agonist is a GITR agonist, and the GITR agonist is selected from the group consisting of TRX518, 6C8, 36E5, 3D6, 61G6, 6H6, 61F6, 1D8, 17F10, 35D8, 49A1, 9E5, 31H6, 2155, 698, 706, 827, 1649, 1718, 1D7, 33C9, 33F6, 34G4, 35B10, 41E11, 41G5, 42A11, 44C1, 45A8, 46E11, 48H12, 48H7, 49D9, 49E2, 48A9, 5H7, 7A10, 9H6, and fragments, derivatives, variants, biosimilars, and combinations thereof.

[0106] In certain embodiments, the TNFRSF agonist is a GITR agonist, and the GITR agonist is a GITR agonist fusion protein.

[0107]

[0107] In one embodiment, the TNFRSF agonist is a GITR agonist fusion protein, and the GITR agonist fusion protein comprises (i) a first soluble GITR binding domain, (ii) a first peptide linker, (iii) a second soluble GITR binding domain, (iv) a second peptide linker, and (v) a third soluble GITR binding domain, and further comprises additional domains at the N-terminal and / or C-terminal end, and the additional domains comprise an Fc fragment domain and a hinge domain, and the fusion protein is a dimeric structure according to structure IA or structure IB.

[0108] In certain embodiments, the TNFRSF agonist is an HVEM agonist.

[0109] In certain embodiments, the TNFRSF agonist is an HVEM agonist, and the HVEM agonist is an HVEM agonist fusion protein.

[0110]

[0110] In one embodiment, the TNFRSF agonist is an HVEM agonist fusion protein, and the HVEM agonist fusion protein comprises (i) a first soluble HVEM binding domain, (ii) a first peptide linker, (iii) a second soluble HVEM binding domain, (iv) a second peptide linker, and (v) a third soluble HVEM binding domain, and further comprises additional domains at the N-terminal and / or C-terminal ends, and the additional domains comprise an Fc fragment domain and a hinge domain, and the fusion protein is a dimeric structure according to structure IA or structure IB.

[0111]

[0111] In one embodiment, the TNFRSF agonist is selected from the group consisting of urelumab, utomilumab, EU-101, taborixizumab, Creative Biolabs MOM-18455, and fragments, derivatives, variants, biosimilars, and combinations thereof.

[0112] In certain embodiments, the first cell culture medium comprises a second TNFRSF agonist.

[0113]

[0113] In one embodiment, the TNFRSF agonist is added to the first cell culture medium during the initial expansion culture at intervals selected from the group consisting of every day, every 2 days, every 3 days, every 4 days, every 5 days, every 6 days, every 7 days, and every 2 weeks.

[0114]

[0114] In one embodiment, the TNFRSF agonist is added to the second cell culture medium during rapid expansion culture at intervals selected from the group consisting of every day, every 2 days, every 3 days, every 4 days, every 5 days, every 6 days, every 7 days, and every 2 weeks.

[0115]

[0115] In certain embodiments, the TNFRSF agonist is added at a concentration sufficient to achieve a concentration of 0.1 μg / mL to 100 μg / mL in the cell culture medium.

[0116]

[0116] In certain embodiments, the TNFRSF agonist is added at a concentration sufficient to achieve a concentration of 20 μg / mL to 40 μg / mL in the cell culture medium.

[0117]

[0117] Further details of TNFRSF agonists are provided herein.

[0118]

[0118] In certain embodiments, IL-2 is present in the first cell culture medium at an initial concentration of about 10 to about 6000 IU / mL.

[0119] In one embodiment, IL-2 is present in the first cell culture medium at an initial concentration of about 3000 IU / mL.

[0120] In certain embodiments, IL-2 is present in the first cell culture medium at an initial concentration of about 800 to about 1100 IU / mL.

[0121] In one embodiment, IL-2 is present in the first cell culture medium at an initial concentration of about 1000 IU / mL.

[0122] In certain embodiments, IL-2 is present in the second cell culture medium at an initial concentration of about 10 to about 6000 IU / mL.

[0123] In one embodiment, IL-2 is present in the second cell culture medium at an initial concentration of about 3000 IU / mL.

[0124] In certain embodiments, IL-2 is present in the second cell culture medium at an initial concentration of about 800 to about 1100 IU / mL.

[0125] In one embodiment, IL-2 is present in the second cell culture medium at an initial concentration of about 1000 IU / mL.

[0126] In certain embodiments, IL-15 is present in the first cell culture medium.

[0127]

[0127] In certain embodiments, IL-15 is present in the first cell culture medium at an initial concentration of about 5 ng / mL to about 20 ng / mL.

[0128] In certain embodiments, IL-15 is present in the second cell culture medium.

[0129]

[0129] In certain embodiments, IL-15 is present in the second cell culture medium at an initial concentration of about 5 ng / mL to about 20 ng / mL.

[0130] In certain embodiments, IL-21 is present in the first cell culture medium.

[0131]

[0131] In certain embodiments, IL-21 is present in the first cell culture medium at an initial concentration of about 5 ng / mL to about 20 ng / mL.

[0132] In certain embodiments, IL-21 is present in the second cell culture medium.

[0133]

[0133] In certain embodiments, IL-21 is present in the second cell culture medium at an initial concentration of about 5 ng / mL to about 20 ng / mL.

[0134]

[0134] In one embodiment, the OKT-3 antibody is present in the second cell culture medium at an initial concentration of about 10 ng / mL to about 60 ng / mL.

[0135]

[0135] In one embodiment, the OKT-3 antibody is present in the second cell culture medium at an initial concentration of about 30 ng / mL.

[0136]

[0136] In one embodiment, the initial expansion culture is carried out using a gas-permeable container.

[0137]

[0137] In one embodiment, rapid expansion culture is performed using a gas-permeable vessel.

[0138]

[0138] In one embodiment, the present invention provides a tumor-infiltrating lymphocyte (TIL) population for use in the treatment of cancer, wherein the tumor-infiltrating lymphocyte (TIL) population is obtainable by the process of the present invention as described herein.

[0139]

[0139] In one embodiment, the present invention provides a pharmaceutical composition comprising a tumor-infiltrating lymphocyte (TIL) population for use in a method for treating cancer, wherein the tumor-infiltrating lymphocyte (TIL) population is obtainable by the process of the present invention as described herein.

[0140] In certain embodiments, the TIL population and / or pharmaceutical composition is for use in the treatment of cancer in combination with TNFRSF.

[0141] In one embodiment, the present invention provides a combination of a TIL population obtainable by the process of the present invention as described herein with a TNFRSF for use in the treatment of cancer.

[0142]

[0142] In one embodiment, the TIL population and / or pharmaceutical composition is for use in the treatment of cancer in combination with a TNFRSF agonist, wherein the TNFRSF agonist is for administration to the patient the day after administration of the third TIL population, and wherein the TNFRSF agonist is administered intravenously at a dose of 0.1 mg / kg to 50 mg / kg every four weeks for up to eight cycles.

[0143]

[0143] In one embodiment, the TIL population and / or pharmaceutical composition is for use in the treatment of cancer in combination with a TNFRSF agonist, wherein the TNFRSF agonist is for administration prior to a step of resecting a tumor from a patient, and wherein the TNFRSF agonist is for intravenous administration at a dose of 0.1 mg / kg to 50 mg / kg every four weeks for up to eight cycles.

[0144] In certain embodiments, the TIL population and / or pharmaceutical composition is for use in the treatment of cancer in combination with a non-myeloablative lymphodepletion regimen.

[0145]

[0145] In one embodiment, the TIL population and / or pharmaceutical composition is for use in the treatment of cancer in combination with a non-myeloablative lymphodepletion regimen prior to administration of the third TIL population and / or pharmaceutical composition comprising the third TIL population to the patient.

[0146] In certain embodiments, the TIL population and / or pharmaceutical composition is for use in the treatment of cancer in combination with a non-myeloablative lymphodepletion regimen prior to administration of the third TIL population and / or pharmaceutical composition comprising the third TIL population to the patient, wherein the non-myeloablative lymphodepletion regimen comprises a dose of 60 mg / m 2 / day for 2 days followed by cyclophosphamide at a dose of 25 mg / m 2

[0033] The method includes administering fludarabine at a dose of 100 mg / day for 5 days. Further details of non-myeloablative lymphodepletion regimens are provided herein, for example, under the heading "Non-myeloablative Lymphodepletion by Chemotherapy."

[0147] In certain embodiments, the TIL population and / or pharmaceutical composition is for use in the treatment of cancer in combination with an IL-2 regimen.

[0148] In one embodiment, the IL-2 regimen is a tapering IL-2 regimen.

[0149] In certain embodiments, the TIL population and / or pharmaceutical composition is for use in the treatment of cancer in combination with a tapering IL-2 regimen starting the day after administration of the third TIL population and / or pharmaceutical composition comprising the third TIL population to the patient, wherein the tapering IL-2 regimen includes administration of 18,000,000 IU / m on day 1 2 , 9,000,000 IU / m on day 2 2 , and 4,500,000 IU / m on days 3 and 4 2 and aldesleukin, administered intravenously at a dose of

[0150] In certain embodiments, the TIL population and / or pharmaceutical composition is for use in the treatment of cancer in combination with pegylated IL-2.

[0151]

[0151] In one embodiment, the TIL population and / or pharmaceutical composition is for use in a method for treating cancer in combination with pegylated IL-2 administered at a dose of 0.10 mg / day to 50 mg / day after administration of a third TIL population and / or a pharmaceutical composition comprising the third TIL population to a patient.

[0152] In certain embodiments, the TIL population and / or pharmaceutical composition is for use in a method of treating cancer in combination with a high-dose IL-2 regimen.

[0153]

[0153] In one embodiment, the TIL population and / or pharmaceutical composition is for use in a method for treating cancer in combination with a high-dose IL-2 regimen starting the day after administration of the third TIL population and / or pharmaceutical composition comprising the third TIL population to the patient.

[0154]

[0154] In one embodiment, the TIL population and / or pharmaceutical composition is for use in the treatment of cancer in combination with a high-dose IL-2 regimen starting the day after administration of the third TIL population and / or pharmaceutical composition comprising the third TIL population to the patient, wherein the high-dose IL-2 regimen comprises 600,000 or 720,000 IU / kg aldesleukin, or a biosimilar or variant thereof, administered as a 15-minute bolus intravenous infusion every 8 hours to a tolerated dose.

[0155]

[0155] In one embodiment, the TIL population and / or pharmaceutical composition is for use in the treatment of cancer, wherein the cancer is selected from the group consisting of melanoma, ovarian cancer, cervical cancer, lung cancer, bladder cancer, breast cancer, head and neck cancer, renal cell carcinoma, acute myeloid leukemia, colorectal cancer, cholangiocarcinoma, and sarcoma.

[0156]

[0156] In one embodiment, the TIL population and / or pharmaceutical composition is for use in the treatment of cancer, wherein the cancer is selected from the group consisting of non-small cell lung cancer (NSCLC), triple-negative breast cancer, double-resistant melanoma, and uveal (ocular) melanoma.

[0157]

[0157] In certain embodiments, the TIL population and / or pharmaceutical composition is for use in the treatment of cancer in combination with a PD-1 inhibitor or a PD-L1 inhibitor.

[0158]

[0158] In one embodiment, the TIL population and / or pharmaceutical composition is for use in the treatment of cancer in combination with a PD-1 inhibitor or a PD-L1 inhibitor, wherein the PD-1 inhibitor or PD-L1 inhibitor is selected from the group consisting of nivolumab, pembrolizumab, durvalumab, atezolizumab, avelumab, and fragments, derivatives, variants, biosimilars, and combinations thereof.

[0159]

[0159] In one embodiment, the TIL population and / or pharmaceutical composition is for use in the treatment of cancer in combination with a PD-1 inhibitor or a PD-L1 inhibitor, wherein the PD-1 inhibitor or PD-L1 inhibitor is for administration prior to resection of a tumor from a patient.

[0160]

[0160] In one embodiment, the TIL population and / or pharmaceutical composition is for use in the treatment of cancer in combination with a PD-1 inhibitor or a PD-L1 inhibitor prior to resection of the tumor from the patient, wherein the PD-1 inhibitor or PD-L1 inhibitor is selected from the group consisting of nivolumab, pembrolizumab, durvalumab, atezolizumab, avelumab, and fragments, derivatives, variants, biosimilars, and combinations thereof.

[0161]

[0161] In certain embodiments, the TIL population and / or pharmaceutical composition is for use in a method for treating cancer in combination with a PD-1 inhibitor or a PD-L1 inhibitor.

[0162]

[0162] In one embodiment, the TIL population and / or pharmaceutical composition is for use in the treatment of cancer in combination with a PD-1 inhibitor or a PD-L1 inhibitor, wherein the PD-1 inhibitor or PD-L1 inhibitor is selected from the group consisting of nivolumab, pembrolizumab, durvalumab, atezolizumab, avelumab, and fragments, derivatives, variants, biosimilars, and combinations thereof.

[0163]

[0163] In one embodiment, the TIL population and / or pharmaceutical composition is for use in a method for treating cancer in combination with a PD-1 inhibitor or a PD-L1 inhibitor after resection of a tumor from a patient.

[0164]

[0164] In one embodiment, the TIL population and / or pharmaceutical composition is for use in the treatment of cancer in combination with a PD-1 inhibitor or a PD-L1 inhibitor following resection of a tumor from a patient, wherein the PD-1 inhibitor or PD-L1 inhibitor is selected from the group consisting of nivolumab, pembrolizumab, durvalumab, atezolizumab, avelumab, and fragments, derivatives, variants, biosimilars, and combinations thereof.

[0165]

[0165] In one embodiment, the TIL population and / or pharmaceutical composition is for use in the treatment of cancer in combination with a PD-1 inhibitor or a PD-L1 inhibitor, wherein the PD-1 or PD-L1 inhibitor is for administration after administration of a third TIL population and / or a pharmaceutical composition comprising the third TIL population to the patient.

[0166] In certain embodiments, the TIL population and / or pharmaceutical composition is for use in the treatment of cancer in combination with a PD-1 or PD-L1 inhibitor for administration after administration of the third TIL population to the patient, wherein the PD-1 or PD-L1 inhibitor is selected from the group consisting of nivolumab, pembrolizumab, durvalumab, atezolizumab, avelumab, and fragments, derivatives, variants, biosimilars, and combinations thereof. Further details of PD-1 and PD-L1 inhibitors are described herein, for example, under the heading "Combinations with PD-1 and PD-L1 Inhibitors." In some embodiments, the TIL population and / or pharmaceutical composition comprising the TIL population further comprises one or more features as described herein, for example, under the headings "Pharmaceutical Compositions, Dosages, and Administration Regimen of TILs" and "Pharmaceutical Compositions, Dosages, and Administration Regimen of TNFRSF Agonists."

[0167] BRIEF DESCRIPTION OF THE DRAWINGS

[0167] The foregoing summary, as well as the following detailed description of the invention, will be better understood when read in conjunction with the appended drawings. [Brief explanation of the drawings]

[0168] [Figure 1]

[0168] Figure 1 shows the TIL expansion and treatment process. The TNFRSF agonist of the present disclosure may be used in both the pre-REP stage (upper half of the figure) or the REP stage (lower half of the figure), and may be added when IL-2 is added to each cell culture. Step 1 refers to adding four tumor fragments to ten G-Rex 10 flasks. Step 2 obtains approximately 40 x 106 or more TILs. Step 3 involves splitting into 36 G-Rex 100 flasks for REP. Step 4 harvests the TILs by centrifugation. After a total processing time of approximately 43 days, step 5 yields a fresh TIL preparation, at which point the TILs can be infused into patients. [Figure 2]

[0169]

[0023] A treatment protocol for use with TILs expanded with a TNFRSF agonist of the present disclosure is shown. Surgery (and tumor resection) is performed first, and lymphodepleting chemotherapy refers to non-myeloablative lymphodepletion by chemotherapy as described elsewhere herein. A TNFRSF agonist of the present disclosure can also be used during therapy as described herein after administration of the TILs. [Figure 3]

[0170] 1 shows the results of an assay to determine whether the 4-1BB-Fc hybridoma 4B5 activates 4-1BB signaling in NF-kB-expressing Jurkat cells in a dose-dependent manner using a green fluorescent protein (GFP) reporter. "Secondary" refers to activation of the secondary antibody. [Figure 4]

[0171] 1 shows the results of an assay using a GFP reporter to determine whether the 4-1BB-Fc hybridoma 1C4 activates 4-1BB signaling in NF-kB-expressing Jurkat cells in a dose-dependent manner. "Secondary" refers to secondary antibody activation. [Figure 5]

[0172] 1 shows the results of an assay using a GFP reporter to determine whether the 4-1BB-Fc hybridoma 9B4 activates 4-1BB signaling in NF-kB-expressing Jurkat cells in a dose-dependent manner. "Secondary" refers to secondary antibody activation. [Figure 6]

[0173] 1 shows the results of an assay using a GFP reporter to determine whether the 4-1BB-Fc hybridoma 1D7 activates 4-1BB signaling in NF-kB-expressing Jurkat cells in a dose-dependent manner. "Secondary" refers to secondary antibody activation. [Figure 7]

[0174] 1 shows the results of an assay using a GFP reporter to determine whether the 4-1BB-Fc hybridoma 1D10 activates 4-1BB signaling in NF-kB-expressing Jurkat cells in a dose-dependent manner. "Secondary" refers to secondary antibody activation. [Figure 8]

[0175] 1 shows the results of an assay using a GFP reporter to determine whether the 4-1BB-Fc hybridoma 3C2 activates 4-1BB signaling in NF-kB-expressing Jurkat cells in a dose-dependent manner. "Secondary" refers to secondary antibody activation. [Figure 9]

[0176] 1 shows the results of an assay using a GFP reporter to determine whether the 4-1BB-Fc hybridoma 10D12 activates 4-1BB signaling in NF-kB-expressing Jurkat cells in a dose-dependent manner. "Secondary" refers to secondary antibody activation. [Figure 10]

[0177] 1 shows the results of an assay using a GFP reporter to determine whether 4-1BB-Fc hybridoma 8D2 activates 4-1BB signaling in NF-kB-expressing Jurkat cells in a dose-dependent manner. "Secondary" refers to secondary antibody activation. [Figure 11]

[0178] 1 shows the results of an assay using a GFP reporter to determine whether the 4-1BB-Fc hybridoma 4G6 activates 4-1BB signaling in NF-kB-expressing Jurkat cells in a dose-dependent manner. "Secondary" refers to secondary antibody activation. [Figure 12]

[0179] 1 shows the results of an assay using a GFP reporter to determine whether the 4-1BB-Fc hybridoma 8E3 activates 4-1BB signaling in NF-kB-expressing Jurkat cells in a dose-dependent manner. "Secondary" refers to secondary antibody activation. [Figure 13]

[0180] An exemplary TIL expansion and manufacturing protocol (Process 2A) is shown. [Figure 14]

[0181] 2A illustrates exemplary method steps performed in Process 2A. [Figure 15]

[0182] 1 shows an exemplary TIL expansion protocol. [Figure 16]

[0183] Figure 1 shows the binding affinity of 4-1BB agonist antibodies from Creative Biolabs (CB) and BPS Biosciences (BPS) as assessed by the percentage of 4-1BB+ cells by flow cytometry. The CB 4-1BB agonist exhibited the highest binding affinity. [Figure 17]

[0184] Figure 1 shows the binding affinity of 4-1BB agonist antibodies from Creative Biolabs (CB) and BPS Biosciences (BPS) as assessed by mean fluorescence intensity (MFI). The CB 4-1BB agonist exhibited the highest binding affinity. [Figure 18]

[0185] 1 shows the results of evaluating the activation of the NF-κB pathway by anti-4-1BB agonist antibodies. [Figure 19]

[0186] 1 shows the binding affinity of Creative Biolabs' OX40 agonist antibodies as assessed by the percentage of OX40+ cells by flow cytometry. [Figure 20]

[0187] 1 shows the binding affinity of Creative Biolabs' OX40 agonist antibodies as assessed by mean fluorescence intensity (MFI). [Figure 21]

[0188] Figure 1 shows comparable binding affinity between Creative Biolabs anti-OX40 agonist antibody (at the five concentrations shown) and a commercially available anti-OX40 (clone Ber-ACT35) agonist. The first letter of each tumor designation indicates histology: C = cervix; H = head and neck (head and neck squamous cell carcinoma); L = lung; and M = melanoma. [Figure 22]

[0189] Figure 1 shows the results of evaluating the effect of anti-OX40 agonist antibodies on NF-κB pathway activation. OX40 reporter cells were treated with either anti-OX40 alone or an isotype control at concentrations of 1, 2, 4, 8, and 16 μg / mL for 24 hours, with or without PBMC feeder cells. Cells were lysed using One-Step Luciferase Reagent, and luciferase activity was measured using a luminometer. [Figure 23]

[0190] 1 shows the experimental design for 4-1BB and OX40 agonist experiments during pre-REP. [Figure 24]

[0191] The tumor histology used in the experimental design is shown in Figure 23. [Figure 25]

[0192] 1 shows the data analysis strategy used to evaluate the effect of 4-1BB and anti-OX40 agonists used during pre-REP on TIL performance and characteristics. [Figure 26]

[0193] Total cell count results from cell expansion cultures using CB4-1BB agonists (N=3) are shown. NT=not tested (control). p-value was >0.99. [Figure 27]

[0194] Total cell count results from cell expansion cultures using CB OX40 agonists (N=5) are shown. NT=not tested (control). p-value was 0.06. [Figure 28]

[0195] Total cell number results of cell expansion cultures using CB 4-1BB agonist and OX-40 agonist (N=2) are shown. NT=not tested (control). [Figure 29]

[0196] Total CD8+ cell count results from cell expansion cultures using CB4-1BB agonist (N=3) are shown. p-value was 0.5. [Figure 30]

[0197] Total CD8+ cell count results from cell expansion cultures using CB OX40 agonist (N=5) are shown. p-value was 0.03. [Figure 31]

[0198] Total CD8+ cell count results of cell expansion cultures using CB 4-1BB agonist and OX-40 agonist (N=2) are shown. NT=not tested (control). [Figure 32]

[0199] The total CD8+ / CD4+ cell number ratio results of cell expansion cultures using CB4-1BB agonist (N=3) are shown. The p value was 0.2. [Figure 33]

[0200] The total CD8+ / CD4+ cell number ratio results of cell expansion cultures using CB OX40 agonist (N=5) are shown. The p-value was 0.12. [Figure 34]

[0201] Figure 1 shows the total CD8+ / CD4+ cell number ratio results of cell expansion cultures using CB 4-1BB agonist and OX-40 agonist (N=2). NT=not tested (control). [Figure 35]

[0202] 1 shows an experimental scheme of REP proliferation of pre-REP TILs expanded in the presence of 4-1BB or OX40 agonist. [Figure 36]

[0203] Fold expansion of TILs expanded with REP from pre-REP TILs expanded in the presence of CB4-1BB agonist compared to TILs not treated with pre-REP (NT) is shown. [Figure 37]

[0204] Fold expansion of TILs expanded with REP from pre-REP TILs expanded in the presence of CB OX40 agonist compared to TILs not treated with pre-REP (NT) is shown. [Figure 38]

[0205] Fold expansion of TILs expanded with REPs from pre-REP TILs expanded in the presence of CB 4-1BB and CB OX40 agonists compared to TILs not treated with pre-REPs (NT) is shown. [Figure 39]

[0206] 1 shows the histology of 21 TIL lines used for evaluation of CB OX40 agonists at the REP stage. [Figure 40]

[0207] 1 shows the experimental scheme for evaluation of CB OX40 agonists in the REP stage. [Figure 41]

[0208] The presence of OX40 agonistic antibody shows preferential expansion of CD8+ TILs in REP (shown as a percentage of CD3+CD4+ cells). [Figure 42]

[0209] The presence of OX40 agonistic antibody leads to preferential expansion of CD8+ TILs in REP (shown as a percentage of CD3+CD8+ cells). [Figure 43]

[0210] Non-responder TIL lines show no downregulation of OX40 in the CD4+ subset after anti-OX40 treatment. [Figure 44]

[0211] Experimental details for CB OX40 agonist dose titration in non-responder and responder TIL lines are shown. [Figure 45]

[0212] 1 shows the results of CB OX40 agonist dose titration in responder TIL lines. [Figure 46]

[0213] 1 shows the results of CB OX40 agonist dose titration in non-responder TIL lines. [Figure 47]

[0214] The equivalent TCRvb repertoire profile for responder L4005 is shown. [Figure 48]

[0215] The equivalent TCRvb repertoire profile for responder H3005 is shown. [Figure 49]

[0216] The equivalent TCRvb repertoire profile for responder M1022 is shown. DETAILED DESCRIPTION OF THE INVENTION

[0169] Brief description of the sequence listing

[0217] SEQ ID NO: 1 is the amino acid sequence of the heavy chain of muromonab.

[0170]

[0218] SEQ ID NO: 2 is the amino acid sequence of the light chain of muromonab.

[0171]

[0219] SEQ ID NO: 3 is the amino acid sequence of recombinant human IL-2 protein.

[0172]

[0220] SEQ ID NO: 4 is the amino acid sequence of aldesleukin.

[0173]

[0221] SEQ ID NO: 5 is the amino acid sequence of recombinant human IL-4 protein.

[0174]

[0222] SEQ ID NO: 6 is the amino acid sequence of recombinant human IL-7 protein.

[0175]

[0223] SEQ ID NO: 7 is the amino acid sequence of recombinant human IL-15 protein.

[0176]

[0224] SEQ ID NO: 8 is the amino acid sequence of recombinant human IL-21 protein.

[0177]

[0225] SEQ ID NO: 9 is the amino acid sequence of human 4-1BB.

[0178]

[0226] SEQ ID NO: 10 is the amino acid sequence of mouse 4-1BB.

[0179]

[0227] SEQ ID NO: 11 is the heavy chain of the 4-1BB agonist monoclonal antibody utomilumab (PF-05082566).

[0180]

[0228] SEQ ID NO: 12 is the light chain of the 4-1BB agonist monoclonal antibody utomilumab (PF-05082566).

[0181]

[0229] SEQ ID NO: 13 is the heavy chain variable region (V) of the 4-1BB agonist monoclonal antibody utomilumab (PF-05082566). H )

[0182]

[0230] SEQ ID NO: 14 is the light chain variable region (V) of the 4-1BB agonist monoclonal antibody utomilumab (PF-05082566). L )

[0183]

[0231] SEQ ID NO: 15 is the heavy chain CDR1 of the 4-1BB agonist monoclonal antibody utomilumab (PF-05082566).

[0184]

[0232] SEQ ID NO: 16 is the heavy chain CDR2 of the 4-1BB agonist monoclonal antibody utomilumab (PF-05082566).

[0185]

[0233] SEQ ID NO: 17 is the heavy chain CDR3 of the 4-1BB agonist monoclonal antibody utomilumab (PF-05082566).

[0186]

[0234] SEQ ID NO: 18 is the light chain CDR1 of the 4-1BB agonist monoclonal antibody utomilumab (PF-05082566).

[0187]

[0235] SEQ ID NO: 19 is the light chain CDR2 of the 4-1BB agonist monoclonal antibody utomilumab (PF-05082566).

[0188]

[0236] SEQ ID NO: 20 is the light chain CDR3 of the 4-1BB agonist monoclonal antibody utomilumab (PF-05082566).

[0189]

[0237] SEQ ID NO: 21 is the heavy chain of the 4-1BB agonist monoclonal antibody urelumab (BMS-663513).

[0190]

[0238] SEQ ID NO: 22 is the light chain of the 4-1BB agonist monoclonal antibody urelumab (BMS-663513).

[0191]

[0239] SEQ ID NO: 23 is the heavy chain variable region (V) of the 4-1BB agonist monoclonal antibody urelumab (BMS-663513). H )

[0192]

[0240] SEQ ID NO: 24 is the light chain variable region (V) of the 4-1BB agonist monoclonal antibody urelumab (BMS-663513). L )

[0193]

[0241] SEQ ID NO: 25 is the heavy chain CDR1 of the 4-1BB agonist monoclonal antibody urelumab (BMS-663513).

[0194]

[0242] SEQ ID NO: 26 is the heavy chain CDR2 of the 4-1BB agonist monoclonal antibody urelumab (BMS-663513).

[0195]

[0243] SEQ ID NO: 27 is the heavy chain CDR3 of the 4-1BB agonist monoclonal antibody urelumab (BMS-663513).

[0196]

[0244] SEQ ID NO: 28 is the light chain CDR1 of the 4-1BB agonist monoclonal antibody urelumab (BMS-663513).

[0197]

[0245] SEQ ID NO: 29 is the light chain CDR2 of the 4-1BB agonist monoclonal antibody urelumab (BMS-663513).

[0198]

[0246] SEQ ID NO: 30 is the light chain CDR3 of the 4-1BB agonist monoclonal antibody urelumab (BMS-663513).

[0199]

[0247] SEQ ID NO: 31 is the Fc domain of the TNFRSF agonist fusion protein.

[0200]

[0248] SEQ ID NO: 32 is the linker of the TNFRSF agonist fusion protein.

[0201]

[0249] SEQ ID NO: 33 is the linker of the TNFRSF agonist fusion protein.

[0202]

[0250] SEQ ID NO: 34 is the linker of the TNFRSF agonist fusion protein.

[0203]

[0251] SEQ ID NO: 35 is the linker of the TNFRSF agonist fusion protein.

[0204]

[0252] SEQ ID NO: 36 is the linker of the TNFRSF agonist fusion protein.

[0205]

[0253] SEQ ID NO: 37 is the linker of the TNFRSF agonist fusion protein.

[0206]

[0254] SEQ ID NO: 38 is the linker of the TNFRSF agonist fusion protein.

[0207]

[0255] SEQ ID NO: 39 is the linker of the TNFRSF agonist fusion protein.

[0208]

[0256] SEQ ID NO: 40 is the linker of the TNFRSF agonist fusion protein.

[0209]

[0257] SEQ ID NO: 41 is the linker of the TNFRSF agonist fusion protein.

[0210]

[0258] SEQ ID NO: 42 is the Fc domain of the TNFRSF agonist fusion protein.

[0211]

[0259] SEQ ID NO: 43 is the linker of the TNFRSF agonist fusion protein.

[0212]

[0260] SEQ ID NO: 44 is the linker of the TNFRSF agonist fusion protein.

[0213]

[0261] SEQ ID NO: 45 is the linker of the TNFRSF agonist fusion protein.

[0214]

[0262] SEQ ID NO: 46 is the 4-1BB ligand (4-1BBL) amino acid sequence.

[0215]

[0263] SEQ ID NO:47 is the soluble portion of the 4-1BBL polypeptide.

[0216]

[0264] SEQ ID NO: 48 is the heavy chain variable region (V) of the 4-1BB agonist antibody 4B4-1-1 version 1 H )

[0217]

[0265] SEQ ID NO: 49 is the light chain variable region (V) of the 4-1BB agonist antibody 4B4-1-1 version 1 L )

[0218]

[0266] SEQ ID NO: 50 is the heavy chain variable region (V) of the 4-1BB agonist antibody 4B4-1-1 version 2 H )

[0219]

[0267] SEQ ID NO: 51 is the light chain variable region (V) of the 4-1BB agonist antibody 4B4-1-1 version 2 L )

[0220]

[0268] SEQ ID NO: 52 is the heavy chain variable region (V H )

[0221]

[0269] SEQ ID NO: 53 is the light chain variable region (V L )

[0222]

[0270] SEQ ID NO: 54 is the amino acid sequence of human OX40.

[0223]

[0271] SEQ ID NO: 55 is the amino acid sequence of mouse OX40.

[0224]

[0272] SEQ ID NO: 56 is the heavy chain of the OX40 agonist monoclonal antibody tabolixizumab (MEDI-0562).

[0225]

[0273] SEQ ID NO: 57 is the light chain of the OX40 agonist monoclonal antibody tabolixizumab (MEDI-0562).

[0226]

[0274] SEQ ID NO: 58 is the heavy chain variable region (V) of the OX40 agonist monoclonal antibody tabolixizumab (MEDI-0562). H )

[0227]

[0275] SEQ ID NO: 59 is the light chain variable region (V) of the OX40 agonist monoclonal antibody tabolixizumab (MEDI-0562). L )

[0228]

[0276] SEQ ID NO: 60 is the heavy chain CDR1 of the OX40 agonist monoclonal antibody tabolixizumab (MEDI-0562).

[0229]

[0277] SEQ ID NO: 61 is the heavy chain CDR2 of the OX40 agonist monoclonal antibody tabolixizumab (MEDI-0562).

[0230]

[0278] SEQ ID NO: 62 is the heavy chain CDR3 of the OX40 agonist monoclonal antibody tabolixizumab (MEDI-0562).

[0231]

[0279] SEQ ID NO: 63 is the light chain CDR1 of the OX40 agonist monoclonal antibody tabolixizumab (MEDI-0562).

[0232]

[0280] SEQ ID NO: 64 is the light chain CDR2 of the OX40 agonist monoclonal antibody tabolixizumab (MEDI-0562).

[0233]

[0281] SEQ ID NO: 65 is the light chain CDR3 of the OX40 agonist monoclonal antibody tabolixizumab (MEDI-0562).

[0234]

[0282] SEQ ID NO: 66 is the heavy chain of the OX40 agonist monoclonal antibody 11D4.

[0235]

[0283] SEQ ID NO: 67 is the light chain of the OX40 agonist monoclonal antibody 11D4.

[0236]

[0284] SEQ ID NO: 68 is the heavy chain variable region (V) of the OX40 agonist monoclonal antibody 11D4 H )

[0237]

[0285] SEQ ID NO: 69 is the light chain variable region (V) of the OX40 agonist monoclonal antibody 11D4. L )

[0238]

[0286] SEQ ID NO: 70 is the heavy chain CDR1 of the OX40 agonist monoclonal antibody 11D4.

[0239]

[0287] SEQ ID NO: 71 is the heavy chain CDR2 of the OX40 agonist monoclonal antibody 11D4.

[0240]

[0288] SEQ ID NO: 72 is the heavy chain CDR3 of the OX40 agonist monoclonal antibody 11D4.

[0241]

[0289] SEQ ID NO: 73 is the light chain CDR1 of the OX40 agonist monoclonal antibody 11D4.

[0242]

[0290] SEQ ID NO: 74 is the light chain CDR2 of the OX40 agonist monoclonal antibody 11D4.

[0243]

[0291] SEQ ID NO: 75 is the light chain CDR3 of the OX40 agonist monoclonal antibody 11D4.

[0244]

[0292] SEQ ID NO: 76 is the heavy chain of the OX40 agonist monoclonal antibody 18D8.

[0245]

[0293] SEQ ID NO: 77 is the light chain of the OX40 agonist monoclonal antibody 18D8.

[0246]

[0294] SEQ ID NO: 78 is the heavy chain variable region (V) of the OX40 agonist monoclonal antibody 18D8 H )

[0247]

[0295] SEQ ID NO: 79 is the light chain variable region (V) of the OX40 agonist monoclonal antibody 18D8 L )

[0248]

[0296] SEQ ID NO: 80 is the heavy chain CDR1 of the OX40 agonist monoclonal antibody 18D8.

[0249]

[0297] SEQ ID NO: 81 is the heavy chain CDR2 of the OX40 agonist monoclonal antibody 18D8.

[0250]

[0298] SEQ ID NO: 82 is the heavy chain CDR3 of the OX40 agonist monoclonal antibody 18D8.

[0251]

[0299] SEQ ID NO: 83 is the light chain CDR1 of the OX40 agonist monoclonal antibody 18D8.

[0252]

[0300] SEQ ID NO: 84 is the light chain CDR2 of the OX40 agonist monoclonal antibody 18D8.

[0253]

[0301] SEQ ID NO: 85 is the light chain CDR3 of the OX40 agonist monoclonal antibody 18D8.

[0254]

[0302] SEQ ID NO: 86 is the heavy chain variable region (V) of the OX40 agonist monoclonal antibody Hu119-122 H )

[0255]

[0303] SEQ ID NO: 87 is the light chain variable region (V) of the OX40 agonist monoclonal antibody Hu119-122 L )

[0256]

[0304] SEQ ID NO: 88 is the heavy chain CDR1 of the OX40 agonist monoclonal antibody Hu119-122.

[0257]

[0305] SEQ ID NO: 89 is the heavy chain CDR2 of the OX40 agonist monoclonal antibody Hu119-122.

[0258]

[0306] SEQ ID NO: 90 is the heavy chain CDR3 of the OX40 agonist monoclonal antibody Hu119-122.

[0259]

[0307] SEQ ID NO: 91 is the light chain CDR1 of the OX40 agonist monoclonal antibody Hu119-122.

[0260]

[0308] SEQ ID NO: 92 is the light chain CDR2 of the OX40 agonist monoclonal antibody Hu119-122.

[0261]

[0309] SEQ ID NO: 93 is the light chain CDR3 of the OX40 agonist monoclonal antibody Hu119-122.

[0262]

[0310] SEQ ID NO: 94 is the heavy chain variable region (V) of the OX40 agonist monoclonal antibody Hu106-222 H )

[0263]

[0311] SEQ ID NO: 95 is the light chain variable region (V) of the OX40 agonist monoclonal antibody Hu106-222.L )

[0264]

[0312] SEQ ID NO: 96 is the heavy chain CDR1 of the OX40 agonist monoclonal antibody Hu106-222.

[0265]

[0313] SEQ ID NO: 97 is the heavy chain CDR2 of the OX40 agonist monoclonal antibody Hu106-222.

[0266]

[0314] SEQ ID NO: 98 is the heavy chain CDR3 of the OX40 agonist monoclonal antibody Hu106-222.

[0267]

[0315] SEQ ID NO: 99 is the light chain CDR1 of the OX40 agonist monoclonal antibody Hu106-222.

[0268]

[0316] SEQ ID NO: 100 is the light chain CDR2 of the OX40 agonist monoclonal antibody Hu106-222.

[0269]

[0317] SEQ ID NO: 101 is the light chain CDR3 of the OX40 agonist monoclonal antibody Hu106-222.

[0270]

[0318] SEQ ID NO: 102 is the OX40 ligand (OX40L) amino acid sequence.

[0271]

[0319] SEQ ID NO: 103 is the soluble portion of the OX40L polypeptide.

[0272]

[0320] SEQ ID NO: 104 is an alternative soluble portion of the OX40L polypeptide.

[0273]

[0321] SEQ ID NO: 105 is the heavy chain variable region (V H )

[0274]

[0322] SEQ ID NO: 106 is the light chain variable region (V) of OX40 agonist monoclonal antibody 008 L )

[0275]

[0323] SEQ ID NO: 107 is the heavy chain variable region (V) of OX40 agonist monoclonal antibody 011 H )

[0276]

[0324] SEQ ID NO: 108 is the light chain variable region (V) of OX40 agonist monoclonal antibody 011 L )

[0277]

[0325] SEQ ID NO: 109 is the heavy chain variable region (V) of OX40 agonist monoclonal antibody 021 H )

[0278]

[0326] SEQ ID NO: 110 is the light chain variable region (V) of the OX40 agonist monoclonal antibody 021. L )

[0279]

[0327] SEQ ID NO: 111 is the heavy chain variable region (V) of the OX40 agonist monoclonal antibody 023 H )

[0280]

[0328] SEQ ID NO: 112 is the light chain variable region (V) of the OX40 agonist monoclonal antibody 023 L )

[0281]

[0329] SEQ ID NO: 113 is the heavy chain variable region (V H )

[0282]

[0330] SEQ ID NO: 114 is the light chain variable region (V L )

[0283]

[0331] SEQ ID NO: 115 is the heavy chain variable region (V H )

[0284]

[0332] SEQ ID NO: 116 is the light chain variable region (V) of the OX40 agonist monoclonal antibody L)

[0285]

[0333] SEQ ID NO: 117 is the heavy chain variable region (V H )

[0286]

[0334] SEQ ID NO: 118 is the heavy chain variable region (V) of a humanized OX40 agonist monoclonal antibody H )

[0287]

[0335] SEQ ID NO: 119 is the light chain variable region (V) of a humanized OX40 agonist monoclonal antibody L )

[0288]

[0336] SEQ ID NO: 120 is the light chain variable region (V) of a humanized OX40 agonist monoclonal antibody L )

[0289]

[0337] SEQ ID NO: 121 is the heavy chain variable region (V H )

[0290]

[0338] SEQ ID NO: 122 is the heavy chain variable region (V H )

[0291]

[0339] SEQ ID NO: 123 is the light chain variable region (V) of a humanized OX40 agonist monoclonal antibody L )

[0292]

[0340] SEQ ID NO: 124 is the light chain variable region (V) of a humanized OX40 agonist monoclonal antibody L )

[0293]

[0341] SEQ ID NO: 125 is the heavy chain variable region (V H )

[0294]

[0342] SEQ ID NO: 126 is the light chain variable region (V) of the OX40 agonist monoclonal antibodyL )

[0295]

[0343] SEQ ID NO: 127 is the amino acid sequence of human CD27.

[0296]

[0344] SEQ ID NO: 128 is the amino acid sequence of macaque CD27.

[0297]

[0345] SEQ ID NO: 129 is the heavy chain of the CD27 agonist monoclonal antibody varlilumab (CDX-1127).

[0298]

[0346] SEQ ID NO: 130 is the light chain of the CD27 agonist monoclonal antibody varlilumab (CDX-1127).

[0299]

[0347] SEQ ID NO: 131 is the heavy chain variable region (V) of the CD27 agonist monoclonal antibody varlilumab (CDX-1127). H )

[0300]

[0348] SEQ ID NO: 132 is the light chain variable region (V) of the CD27 agonist monoclonal antibody varlilumab (CDX-1127). L )

[0301]

[0349] SEQ ID NO: 133 is the heavy chain CDR1 of the CD27 agonist monoclonal antibody varlilumab (CDX-1127).

[0302]

[0350] SEQ ID NO: 134 is the heavy chain CDR2 of the CD27 agonist monoclonal antibody varlilumab (CDX-1127).

[0303]

[0351] SEQ ID NO: 135 is the heavy chain CDR3 of the CD27 agonist monoclonal antibody varlilumab (CDX-1127).

[0304]

[0352] SEQ ID NO: 136 is the light chain CDR1 of the CD27 agonist monoclonal antibody varlilumab (CDX-1127).

[0305]

[0353] SEQ ID NO: 137 is the light chain CDR2 of the CD27 agonist monoclonal antibody varlilumab (CDX-1127).

[0306]

[0354] SEQ ID NO: 138 is the light chain CDR3 of the CD27 agonist monoclonal antibody varlilumab (CDX-1127).

[0307]

[0355] SEQ ID NO: 139 is the CD27 ligand (CD70) amino acid sequence.

[0308]

[0356] SEQ ID NO: 140 is the soluble portion of the CD70 polypeptide.

[0309]

[0357] SEQ ID NO: 141 is an alternative soluble portion of the CD70 polypeptide.

[0310]

[0358] SEQ ID NO: 142 is the amino acid sequence of human GITR (human tumor necrosis factor receptor superfamily member 18 (TNFRSF18) protein).

[0311]

[0359] SEQ ID NO: 143 is the amino acid sequence of mouse GITR (mouse tumor necrosis factor receptor superfamily member 18 (TNFRSF18) protein).

[0312]

[0360] SEQ ID NO: 144 is the amino acid sequence of the heavy chain variant HuN6C8 (glycosylated) of the 6C8 humanized GITR agonist monoclonal antibody having N (asparagine) in CDR2, corresponding to SEQ ID NO: 60 of U.S. Patent No. 7,812,135.

[0313]

[0361] SEQ ID NO: 145 is the amino acid sequence of the heavy chain variant HuN6C8 (non-glycosylated) of the 6C8 humanized GITR agonist monoclonal antibody having N (asparagine) in CDR2, corresponding to SEQ ID NO: 61 of U.S. Patent No. 7,812,135.

[0314]

[0362] SEQ ID NO: 146 is the amino acid sequence of the heavy chain variant HuQ6C8 (glycosylated) of the 6C8 humanized GITR agonist monoclonal antibody having Q (glutamine) in CDR2, corresponding to SEQ ID NO: 62 of U.S. Patent No. 7,812,135.

[0315]

[0363] SEQ ID NO: 147 is the amino acid sequence of the heavy chain variant HuQ6C8 (non-glycosylated) of the 6C8 humanized GITR agonist monoclonal antibody having Q (glutamine) in CDR2, corresponding to SEQ ID NO: 63 of U.S. Patent No. 7,812,135.

[0316]

[0364] SEQ ID NO: 148 is the amino acid sequence of the light chain of the 6C8 humanized GITR agonist monoclonal antibody, which corresponds to SEQ ID NO: 58 of US Pat. No. 7,812,135.

[0317]

[0365] SEQ ID NO: 149 is the amino acid sequence of a leader sequence that may optionally be included in a GITR agonist monoclonal antibody along with the amino acid sequence of SEQ ID NO: 144, SEQ ID NO: 145, SEQ ID NO: 146, or SEQ ID NO: 147.

[0318]

[0366] SEQ ID NO: 150 is the amino acid sequence of a leader sequence that may optionally be included in a GITR agonist monoclonal antibody along with the amino acid sequence of SEQ ID NO: 148.

[0319]

[0367] SEQ ID NO: 151 is the amino acid sequence of the heavy chain variable region of the 6C8 humanized GITR agonist monoclonal antibody, corresponding to SEQ ID NO: 1 of US Pat. No. 7,812,135.

[0320]

[0368] SEQ ID NO: 152 is the amino acid sequence of the heavy chain variable region of the 6C8 humanized GITR agonist monoclonal antibody, which corresponds to SEQ ID NO: 66 of US Pat. No. 7,812,135.

[0321]

[0369] SEQ ID NO: 153 is the amino acid sequence of the light chain variable region of the 6C8 humanized GITR agonist monoclonal antibody, corresponding to SEQ ID NO: 2 of US Pat. No. 7,812,135.

[0322]

[0370] SEQ ID NO: 154 is the amino acid sequence of the heavy chain CDR1 region of the 6C8 humanized GITR agonist monoclonal antibody, which corresponds to SEQ ID NO: 3 of US Pat. No. 7,812,135.

[0323]

[0371] SEQ ID NO: 155 is the amino acid sequence of the heavy chain CDR2 region of the 6C8 humanized GITR agonist monoclonal antibody, corresponding to SEQ ID NO: 4 of US Pat. No. 7,812,135.

[0324]

[0372] SEQ ID NO: 156 is the amino acid sequence of the heavy chain CDR2 region of the 6C8 humanized GITR agonist monoclonal antibody, corresponding to SEQ ID NO: 19 of US Pat. No. 7,812,135.

[0325]

[0373] SEQ ID NO: 157 is the amino acid sequence of the heavy chain CDR3 region of the 6C8 humanized GITR agonist monoclonal antibody, corresponding to SEQ ID NO: 5 of US Pat. No. 7,812,135.

[0326]

[0374] SEQ ID NO: 158 is the amino acid sequence of the heavy chain CDR1 region of the 6C8 humanized GITR agonist monoclonal antibody, corresponding to SEQ ID NO: 6 of US Pat. No. 7,812,135.

[0327]

[0375] SEQ ID NO: 159 is the amino acid sequence of the heavy chain CDR2 region of the 6C8 humanized GITR agonist monoclonal antibody, corresponding to SEQ ID NO: 7 of US Pat. No. 7,812,135.

[0328]

[0376] SEQ ID NO: 160 is the amino acid sequence of the heavy chain CDR3 region of the 6C8 humanized GITR agonist monoclonal antibody, corresponding to SEQ ID NO: 8 of US Pat. No. 7,812,135.

[0329]

[0377] SEQ ID NO: 161 is the amino acid sequence of the 6C8 chimeric GITR agonist monoclonal antibody heavy chain variant HuN6C8 (glycosylated) with N (asparagine) in CDR2, corresponding to SEQ ID NO: 23 of US Pat. No. 7,812,135.

[0330]

[0378] SEQ ID NO: 162 is the amino acid sequence of the heavy chain variant HuQ6C8 (non-glycosylated) of the 6C8 chimeric GITR agonist monoclonal antibody having Q (glutamine) in CDR2, corresponding to SEQ ID NO: 24 of US Pat. No. 7,812,135.

[0331]

[0379] SEQ ID NO: 163 is the amino acid sequence of the light chain of the 6C8 chimeric GITR agonist monoclonal antibody, corresponding to SEQ ID NO: 22 of US Pat. No. 7,812,135.

[0332]

[0380] SEQ ID NO: 164 is the amino acid sequence of the GITR agonist 36E5 heavy chain variable region from U.S. Patent No. 8,709,424.

[0333]

[0381] SEQ ID NO: 165 is the amino acid sequence of the GITR agonist 36E5 light chain variable region from U.S. Patent No. 8,709,424.

[0334]

[0382] SEQ ID NO: 166 is the amino acid sequence of the GITR agonist 3D6 heavy chain variable region from US Pat. No. 8,709,424.

[0335]

[0383] SEQ ID NO: 167 is the amino acid sequence of the GITR agonist 3D6 light chain variable region from U.S. Patent No. 8,709,424.

[0336]

[0384] SEQ ID NO: 168 is the amino acid sequence of the GITR agonist 61G6 heavy chain variable region from U.S. Patent No. 8,709,424.

[0337]

[0385] SEQ ID NO: 169 is the amino acid sequence of the GITR agonist 61G6 light chain variable region from U.S. Patent No. 8,709,424.

[0338]

[0386] SEQ ID NO: 170 is the amino acid sequence of the GITR agonist 6H6 heavy chain variable region from U.S. Patent No. 8,709,424.

[0339]

[0387] SEQ ID NO: 171 is the amino acid sequence of the GITR agonist 6H6 light chain variable region from U.S. Patent No. 8,709,424.

[0340]

[0388] SEQ ID NO: 172 is the amino acid sequence of the GITR agonist 61F6 heavy chain variable region from U.S. Patent No. 8,709,424.

[0341]

[0389] SEQ ID NO: 173 is the amino acid sequence of the GITR agonist 61F6 light chain variable region from U.S. Patent No. 8,709,424.

[0342]

[0390] SEQ ID NO: 174 is the amino acid sequence of the GITR agonist 1D8 heavy chain variable region from US Pat. No. 8,709,424.

[0343]

[0391] SEQ ID NO: 175 is the amino acid sequence of the GITR agonist 1D8 light chain variable region from US Pat. No. 8,709,424.

[0344]

[0392] SEQ ID NO: 176 is the amino acid sequence of the GITR agonist 17F10 heavy chain variable region from U.S. Patent No. 8,709,424.

[0345]

[0393] SEQ ID NO: 177 is the amino acid sequence of the GITR agonist 17F10 light chain variable region from U.S. Patent No. 8,709,424.

[0346]

[0394] SEQ ID NO: 178 is the amino acid sequence of the GITR agonist 35D8 heavy chain variable region from U.S. Patent No. 8,709,424.

[0347]

[0395] SEQ ID NO: 179 is the amino acid sequence of the GITR agonist 35D8 light chain variable region from U.S. Patent No. 8,709,424.

[0348]

[0396] SEQ ID NO: 180 is the amino acid sequence of the GITR agonist 49A1 heavy chain variable region from U.S. Patent No. 8,709,424.

[0349]

[0397] SEQ ID NO: 181 is the amino acid sequence of the GITR agonist 49A1 light chain variable region from U.S. Patent No. 8,709,424.

[0350]

[0398] SEQ ID NO: 182 is the amino acid sequence of the GITR agonist 9E5 heavy chain variable region from U.S. Patent No. 8,709,424.

[0351]

[0399] SEQ ID NO: 183 is the amino acid sequence of the GITR agonist 9E5 light chain variable region from U.S. Patent No. 8,709,424.

[0352]

[0400] SEQ ID NO: 184 is the amino acid sequence of the GITR agonist 31H6 heavy chain variable region from U.S. Patent No. 8,709,424.

[0353]

[0401] SEQ ID NO: 185 is the amino acid sequence of the GITR agonist 31H6 light chain variable region from U.S. Patent No. 8,709,424.

[0354]

[0402] SEQ ID NO: 186 is the amino acid sequence of the humanized GITR agonist 36E5 heavy chain variable region from U.S. Patent No. 8,709,424.

[0355]

[0403] SEQ ID NO: 187 is the amino acid sequence of the humanized GITR agonist 36E5 light chain variable region from U.S. Patent No. 8,709,424.

[0356]

[0404] SEQ ID NO: 188 is the amino acid sequence of the humanized GITR agonist 3D6 heavy chain variable region from US Pat. No. 8,709,424.

[0357]

[0405] SEQ ID NO: 189 is the amino acid sequence of the humanized GITR agonist 3D6 light chain variable region from U.S. Patent No. 8,709,424.

[0358]

[0406] SEQ ID NO: 190 is the amino acid sequence of the humanized GITR agonist 61G6 heavy chain variable region from U.S. Patent No. 8,709,424.

[0359]

[0407] SEQ ID NO: 191 is the amino acid sequence of the humanized GITR agonist 61G6 light chain variable region from U.S. Patent No. 8,709,424.

[0360]

[0408] SEQ ID NO: 192 is the amino acid sequence of the humanized GITR agonist 6H6 heavy chain variable region from U.S. Patent No. 8,709,424.

[0361]

[0409] SEQ ID NO: 193 is the amino acid sequence of the humanized GITR agonist 6H6 light chain variable region from U.S. Patent No. 8,709,424.

[0362]

[0410] SEQ ID NO: 194 is the amino acid sequence of the humanized GITR agonist 61F6 heavy chain variable region from U.S. Patent No. 8,709,424.

[0363]

[0411] SEQ ID NO: 195 is the amino acid sequence of the humanized GITR agonist 61F6 light chain variable region from U.S. Patent No. 8,709,424.

[0364]

[0412] SEQ ID NO: 196 is the amino acid sequence of the humanized GITR agonist 1D8 heavy chain variable region from US Pat. No. 8,709,424.

[0365]

[0413] SEQ ID NO: 197 is the amino acid sequence of the humanized GITR agonist 1D8 light chain variable region from US Pat. No. 8,709,424.

[0366]

[0414] SEQ ID NO: 198 is the amino acid sequence of the humanized GITR agonist 17F10 heavy chain variable region from U.S. Patent No. 8,709,424.

[0367]

[0415] SEQ ID NO: 199 is the amino acid sequence of the humanized GITR agonist 17F10 light chain variable region from U.S. Patent No. 8,709,424.

[0368]

[0416] SEQ ID NO: 200 is the amino acid sequence of the humanized GITR agonist 35D8 heavy chain variable region from US Pat. No. 8,709,424.

[0369]

[0417] SEQ ID NO: 201 is the amino acid sequence of the humanized GITR agonist 35D8 light chain variable region from US Pat. No. 8,709,424.

[0370]

[0418] SEQ ID NO: 202 is the amino acid sequence of the humanized GITR agonist 49A1 heavy chain variable region from US Pat. No. 8,709,424.

[0371]

[0419] SEQ ID NO: 203 is the amino acid sequence of the humanized GITR agonist 49A1 light chain variable region from U.S. Patent No. 8,709,424.

[0372]

[0420] SEQ ID NO:204 is the amino acid sequence of the humanized GITR agonist 9E5 heavy chain variable region from US Pat. No. 8,709,424.

[0373]

[0421] SEQ ID NO: 205 is the amino acid sequence of the humanized GITR agonist 9E5 light chain variable region from U.S. Patent No. 8,709,424.

[0374]

[0422] SEQ ID NO: 206 is the amino acid sequence of the humanized GITR agonist 31H6 heavy chain variable region from U.S. Patent No. 8,709,424.

[0375]

[0423] SEQ ID NO: 207 is the amino acid sequence of the humanized GITR agonist 31H6 light chain variable region from U.S. Patent No. 8,709,424.

[0376]

[0424] SEQ ID NO: 208 is the amino acid sequence of the GITR agonist 2155 variable heavy chain from US Patent Application Publication No. 2013 / 0108641 A1.

[0377]

[0425] SEQ ID NO: 209 is the amino acid sequence of the GITR agonist 2155 variable light chain from US Patent Application Publication No. 2013 / 0108641 A1.

[0378]

[0426] SEQ ID NO: 210 is the amino acid sequence of the GITR agonist 2155 humanized (HC1) heavy chain from U.S. Patent Application Publication No. 2013 / 0108641 A1.

[0379]

[0427] SEQ ID NO: 211 is the amino acid sequence of the GITR agonist 2155 humanized (HC2) heavy chain from U.S. Patent Application Publication No. 2013 / 0108641 A1.

[0380]

[0428] SEQ ID NO: 212 is the amino acid sequence of the GITR agonist 2155 humanized (HC3a) heavy chain from U.S. Patent Application Publication No. 2013 / 0108641 A1.

[0381]

[0429] SEQ ID NO: 213 is the amino acid sequence of the humanized (HC3b) GITR agonist heavy chain from U.S. Patent Application Publication No. 2013 / 0108641 A1.

[0382]

[0430] SEQ ID NO: 214 is the amino acid sequence of the humanized (HC4) GITR agonist heavy chain from U.S. Patent Application Publication No. 2013 / 0108641 A1.

[0383]

[0431] SEQ ID NO: 215 is the amino acid sequence of the 2155 humanized (LC1) GITR agonist light chain from U.S. Patent Application Publication No. 2013 / 0108641 A1.

[0384]

[0432] SEQ ID NO: 216 is the amino acid sequence of the 2155 humanized (LC2a) GITR agonist light chain from U.S. Patent Application Publication No. 2013 / 0108641 A1.

[0385]

[0433] SEQ ID NO: 217 is the amino acid sequence of the 2155 humanized (LC2b) GITR agonist light chain from U.S. Patent Application Publication No. 2013 / 0108641 A1.

[0386]

[0434] SEQ ID NO: 218 is the amino acid sequence of the 2155 humanized (LC3) GITR agonist light chain from U.S. Patent Application Publication No. 2013 / 0108641 A1.

[0387]

[0435] SEQ ID NO: 219 is the amino acid sequence of the GITR agonist 698 variable heavy chain from US Patent Application Publication No. 2013 / 0108641 A1.

[0388]

[0436] SEQ ID NO: 220 is the amino acid sequence of the GITR agonist 698 variable light chain from U.S. Patent Application Publication No. 2013 / 0108641 A1.

[0389]

[0437] SEQ ID NO: 221 is the amino acid sequence of the GITR agonist 706 variable heavy chain from US Patent Application Publication No. 2013 / 0108641 A1.

[0390]

[0438] SEQ ID NO: 222 is the amino acid sequence of the GITR agonist 706 variable light chain from U.S. Patent Application Publication No. 2013 / 0108641 A1.

[0391]

[0439] SEQ ID NO: 223 is the amino acid sequence of the GITR agonist 827 variable heavy chain from US Patent Application Publication No. 2013 / 0108641 A1.

[0392]

[0440] SEQ ID NO: 224 is the amino acid sequence of the GITR agonist 827 variable light chain from U.S. Patent Application Publication No. 2013 / 0108641 A1.

[0393]

[0441] SEQ ID NO: 225 is the amino acid sequence of the GITR agonist 1718 variable heavy chain from US Patent Application Publication No. 2013 / 0108641 A1.

[0394]

[0442] SEQ ID NO: 226 is the amino acid sequence of the GITR agonist 1718 variable light chain from U.S. Patent Application Publication No. 2013 / 0108641 A1.

[0395]

[0443] SEQ ID NO: 227 is the amino acid sequence of the GITR agonist 2155 heavy chain CDR3 from U.S. Patent Application Publication No. 2013 / 0108641 A1.

[0396]

[0444] SEQ ID NO: 228 is the amino acid sequence of GITR agonist 2155 heavy chain CDR2 from U.S. Patent Application Publication No. 2013 / 0108641 A1.

[0397]

[0445] SEQ ID NO: 229 is the amino acid sequence of GITR agonist 2155 heavy chain CDR1 from U.S. Patent Application Publication No. 2013 / 0108641 A1.

[0398]

[0446] SEQ ID NO: 230 is the amino acid sequence of the GITR agonist 2155 light chain CDR3 from U.S. Patent Application Publication No. 2013 / 0108641 A1.

[0399]

[0447] SEQ ID NO: 231 is the amino acid sequence of GITR agonist 2155 light chain CDR2 from U.S. Patent Application Publication No. 2013 / 0108641 A1.

[0400]

[0448] SEQ ID NO: 232 is the amino acid sequence of GITR agonist 2155 light chain CDR1 from U.S. Patent Application Publication No. 2013 / 0108641 A1.

[0401]

[0449] SEQ ID NO: 233 is the amino acid sequence of the GITR agonist 698 and 706 heavy chain CDR3 from U.S. Patent Application Publication No. 2013 / 0108641 A1.

[0402]

[0450] SEQ ID NO: 234 is the amino acid sequence of GITR agonist 698 and 706 heavy chain CDR2 from U.S. Patent Application Publication No. 2013 / 0108641 A1.

[0403]

[0451] SEQ ID NO: 235 is the amino acid sequence of GITR agonist 698 and 706 heavy chain CDR1 from U.S. Patent Application Publication No. 2013 / 0108641 A1.

[0404]

[0452] SEQ ID NO: 236 is the amino acid sequence of the GITR agonist 698 light chain CDR3 from U.S. Patent Application Publication No. 2013 / 0108641 A1.

[0405]

[0453] SEQ ID NO: 237 is the amino acid sequence of the light chain CDR2 of GITR agonists 698, 706, 827, and 1649 from U.S. Patent Application Publication No. 2013 / 0108641 A1.

[0406]

[0454] SEQ ID NO: 238 is the amino acid sequence of the light chain CDR1 of GITR agonists 698, 706, 827, and 1649 from U.S. Patent Application Publication No. 2013 / 0108641 A1.

[0407]

[0455] SEQ ID NO: 239 is the amino acid sequence of the light chain CDR3 of GITR agonists 706, 827, and 1649 from U.S. Patent Application Publication No. 2013 / 0108641 A1.

[0408]

[0456] SEQ ID NO: 240 is the amino acid sequence of the GITR agonist 827 and 1649 heavy chain CDR3 from U.S. Patent Application Publication No. 2013 / 0108641 A1.

[0409]

[0457] SEQ ID NO: 241 is the amino acid sequence of GITR agonist 827 heavy chain CDR2 from U.S. Patent Application Publication No. 2013 / 0108641 A1.

[0410]

[0458] SEQ ID NO: 242 is the amino acid sequence of GITR agonist 1649 heavy chain CDR2 from U.S. Patent Application Publication No. 2013 / 0108641 A1.

[0411]

[0459] SEQ ID NO: 243 is the amino acid sequence of the GITR agonist 1718 heavy chain CDR3 from U.S. Patent Application Publication No. 2013 / 0108641 A1.

[0412]

[0460] SEQ ID NO: 244 is the amino acid sequence of GITR agonist 1718 heavy chain CDR2 from U.S. Patent Application Publication No. 2013 / 0108641 A1.

[0413]

[0461] SEQ ID NO: 245 is the amino acid sequence of GITR agonist 1718 heavy chain CDR1 from U.S. Patent Application Publication No. 2013 / 0108641 A1.

[0414]

[0462] SEQ ID NO: 246 is the amino acid sequence of the GITR agonist 1718 light chain CDR3 from U.S. Patent Application Publication No. 2013 / 0108641 A1.

[0415]

[0463] SEQ ID NO: 247 is the amino acid sequence of GITR agonist 1718 light chain CDR2 from U.S. Patent Application Publication No. 2013 / 0108641 A1.

[0416]

[0464] SEQ ID NO: 248 is the amino acid sequence of GITR agonist 1718 light chain CDR1 from U.S. Patent Application Publication No. 2013 / 0108641 A1.

[0417]

[0465] SEQ ID NO: 249 is the amino acid sequence of the GITR agonist 827 and 1649 heavy chain CDR1 from U.S. Patent Application Publication No. 2013 / 0108641 A1.

[0418]

[0466] SEQ ID NO: 250 is the amino acid sequence of the GITR agonist 1D7 heavy chain from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0419]

[0467] SEQ ID NO: 251 is the amino acid sequence of the GITR agonist 1D7 light chain from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0420]

[0468] SEQ ID NO: 252 is the amino acid sequence of the GITR agonist 1D7 variable heavy chain from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0421]

[0469] SEQ ID NO: 253 is the amino acid sequence of the GITR agonist 1D7 variable light chain from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0422]

[0470] SEQ ID NO: 254 is the amino acid sequence of GITR agonist 1D7 heavy chain CDR1 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0423]

[0471] SEQ ID NO: 255 is the amino acid sequence of GITR agonist 1D7 heavy chain CDR2 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0424]

[0472] SEQ ID NO: 256 is the amino acid sequence of the GITR agonist 1D7 heavy chain CDR3 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0425]

[0473] SEQ ID NO: 257 is the amino acid sequence of GITR agonist 1D7 light chain CDR1 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0426]

[0474] SEQ ID NO: 258 is the amino acid sequence of the GITR agonist 1D7 light chain CDR2 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0427]

[0475] SEQ ID NO: 259 is the amino acid sequence of the GITR agonist 1D7 light chain CDR3 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0428]

[0476] SEQ ID NO: 260 is the amino acid sequence of the GITR agonist 33C9 heavy chain from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0429]

[0477] SEQ ID NO: 261 is the amino acid sequence of the GITR agonist 33C9 light chain from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0430]

[0478] SEQ ID NO: 262 is the amino acid sequence of the GITR agonist 33C9 variable heavy chain from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0431]

[0479] SEQ ID NO: 263 is the amino acid sequence of the GITR agonist 33C9 variable light chain from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0432]

[0480] SEQ ID NO: 264 is the amino acid sequence of the GITR agonist 33C9 heavy chain CDR1 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0433]

[0481] SEQ ID NO: 265 is the amino acid sequence of the GITR agonist 33C9 heavy chain CDR2 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0434]

[0482] SEQ ID NO: 266 is the amino acid sequence of the GITR agonist 33C9 heavy chain CDR3 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0435]

[0483] SEQ ID NO: 267 is the amino acid sequence of the GITR agonist 33C9 light chain CDR1 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0436]

[0484] SEQ ID NO: 268 is the amino acid sequence of the GITR agonist 33C9 light chain CDR2 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0437]

[0485] SEQ ID NO: 269 is the amino acid sequence of the GITR agonist 33C9 light chain CDR3 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0438]

[0486] SEQ ID NO: 270 is the amino acid sequence of the GITR agonist 33F6 heavy chain from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0439]

[0487] SEQ ID NO: 271 is the amino acid sequence of the GITR agonist 33F6 light chain from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0440]

[0488] SEQ ID NO: 272 is the amino acid sequence of the GITR agonist 33F6 variable heavy chain from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0441]

[0489] SEQ ID NO: 273 is the amino acid sequence of the GITR agonist 33F6 variable light chain from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0442]

[0490] SEQ ID NO: 274 is the amino acid sequence of the GITR agonist 33F6 heavy chain CDR1 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0443]

[0491] SEQ ID NO: 275 is the amino acid sequence of the GITR agonist 33F6 heavy chain CDR2 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0444]

[0492] SEQ ID NO: 276 is the amino acid sequence of the GITR agonist 33F6 heavy chain CDR3 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0445]

[0493] SEQ ID NO: 277 is the amino acid sequence of the GITR agonist 33F6 light chain CDR1 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0446]

[0494] SEQ ID NO: 278 is the amino acid sequence of the GITR agonist 33F6 light chain CDR2 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0447]

[0495] SEQ ID NO: 279 is the amino acid sequence of the GITR agonist 33F6 light chain CDR3 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0448]

[0496] SEQ ID NO: 280 is the amino acid sequence of the GITR agonist 34G4 heavy chain from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0449]

[0497] SEQ ID NO: 281 is the amino acid sequence of the GITR agonist 34G4 light chain from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0450]

[0498] SEQ ID NO: 282 is the amino acid sequence of the GITR agonist 34G4 variable heavy chain from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0451]

[0499] SEQ ID NO: 283 is the amino acid sequence of the GITR agonist 34G4 variable light chain from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0452]

[0500] SEQ ID NO: 284 is the amino acid sequence of the GITR agonist 34G4 heavy chain CDR1 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0453]

[0501] SEQ ID NO: 285 is the amino acid sequence of the GITR agonist 34G4 heavy chain CDR2 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0454]

[0502] SEQ ID NO: 286 is the amino acid sequence of the GITR agonist 34G4 heavy chain CDR3 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0455]

[0503] SEQ ID NO: 287 is the amino acid sequence of the GITR agonist 34G4 light chain CDR1 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0456]

[0504] SEQ ID NO: 288 is the amino acid sequence of the GITR agonist 34G4 light chain CDR2 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0457]

[0505] SEQ ID NO: 289 is the amino acid sequence of the GITR agonist 34G4 light chain CDR3 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0458]

[0506] SEQ ID NO: 290 is the amino acid sequence of the GITR agonist 35B10 heavy chain from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0459]

[0507] SEQ ID NO: 291 is the amino acid sequence of the GITR agonist 35B10 light chain from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0460]

[0508] SEQ ID NO: 292 is the amino acid sequence of the GITR agonist 35B10 variable heavy chain from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0461]

[0509] SEQ ID NO: 293 is the amino acid sequence of the GITR agonist 35B10 variable light chain from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0462]

[0510] SEQ ID NO: 294 is the amino acid sequence of the GITR agonist 35B10 heavy chain CDR1 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0463]

[0511] SEQ ID NO: 295 is the amino acid sequence of the GITR agonist 35B10 heavy chain CDR2 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0464]

[0512] SEQ ID NO: 296 is the amino acid sequence of the GITR agonist 35B10 heavy chain CDR3 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0465]

[0513] SEQ ID NO: 297 is the amino acid sequence of the GITR agonist 35B10 light chain CDR1 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0466]

[0514] SEQ ID NO: 298 is the amino acid sequence of the GITR agonist 35B10 light chain CDR2 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0467]

[0515] SEQ ID NO: 299 is the amino acid sequence of the GITR agonist 35B10 light chain CDR3 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0468]

[0516] SEQ ID NO: 300 is the amino acid sequence of the GITR agonist 41E11 heavy chain from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0469]

[0517] SEQ ID NO: 301 is the amino acid sequence of the GITR agonist 41E11 light chain from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0470]

[0518] SEQ ID NO: 302 is the amino acid sequence of the GITR agonist 41E11 variable heavy chain from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0471]

[0519] SEQ ID NO: 303 is the amino acid sequence of the GITR agonist 41E11 variable light chain from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0472]

[0520] SEQ ID NO: 304 is the amino acid sequence of GITR agonist 41E11 heavy chain CDR1 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0473]

[0521] SEQ ID NO: 305 is the amino acid sequence of GITR agonist 41E11 heavy chain CDR2 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0474]

[0522] SEQ ID NO: 306 is the amino acid sequence of the GITR agonist 41E11 heavy chain CDR3 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0475]

[0523] SEQ ID NO: 307 is the amino acid sequence of GITR agonist 41E11 light chain CDR1 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0476]

[0524] SEQ ID NO: 308 is the amino acid sequence of the GITR agonist 41E11 light chain CDR2 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0477]

[0525] SEQ ID NO: 309 is the amino acid sequence of the GITR agonist 41E11 light chain CDR3 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0478]

[0526] SEQ ID NO: 310 is the amino acid sequence of the GITR agonist 41G5 heavy chain from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0479]

[0527] SEQ ID NO: 311 is the amino acid sequence of the GITR agonist 41G5 light chain from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0480]

[0528] SEQ ID NO: 312 is the amino acid sequence of the GITR agonist 41G5 variable heavy chain from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0481]

[0529] SEQ ID NO: 313 is the amino acid sequence of the GITR agonist 41G5 variable light chain from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0482]

[0530] SEQ ID NO: 314 is the amino acid sequence of the GITR agonist 41G5 heavy chain CDR1 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0483]

[0531] SEQ ID NO: 315 is the amino acid sequence of the GITR agonist 41G5 heavy chain CDR2 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0484]

[0532] SEQ ID NO: 316 is the amino acid sequence of the GITR agonist 41G5 heavy chain CDR3 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0485]

[0533] SEQ ID NO: 317 is the amino acid sequence of the GITR agonist 41G5 light chain CDR1 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0486]

[0534] SEQ ID NO: 318 is the amino acid sequence of the GITR agonist 41G5 light chain CDR2 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0487]

[0535] SEQ ID NO: 319 is the amino acid sequence of the GITR agonist 41G5 light chain CDR3 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0488]

[0536] SEQ ID NO: 320 is the amino acid sequence of the GITR agonist 42A11 heavy chain from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0489]

[0537] SEQ ID NO: 321 is the amino acid sequence of the GITR agonist 42A11 light chain from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0490]

[0538] SEQ ID NO: 322 is the amino acid sequence of the GITR agonist 42A11 variable heavy chain from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0491]

[0539] SEQ ID NO: 323 is the amino acid sequence of the GITR agonist 42A11 variable light chain from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0492]

[0540] SEQ ID NO: 324 is the amino acid sequence of GITR agonist 42A11 heavy chain CDR1 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0493]

[0541] SEQ ID NO: 325 is the amino acid sequence of GITR agonist 42A11 heavy chain CDR2 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0494]

[0542] SEQ ID NO: 326 is the amino acid sequence of the GITR agonist 42A11 heavy chain CDR3 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0495]

[0543] SEQ ID NO: 327 is the amino acid sequence of the GITR agonist 42A11 light chain CDR1 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0496]

[0544] SEQ ID NO: 328 is the amino acid sequence of the GITR agonist 42A11 light chain CDR2 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0497]

[0545] SEQ ID NO: 329 is the amino acid sequence of the GITR agonist 42A11 light chain CDR3 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0498]

[0546] SEQ ID NO: 330 is the amino acid sequence of the GITR agonist 44C1 heavy chain from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0499]

[0547] SEQ ID NO: 331 is the amino acid sequence of the GITR agonist 44C1 light chain from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0500]

[0548] SEQ ID NO: 332 is the amino acid sequence of the GITR agonist 44C1 variable heavy chain from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0501]

[0549] SEQ ID NO: 333 is the amino acid sequence of the GITR agonist 44C1 variable light chain from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0502]

[0550] SEQ ID NO: 334 is the amino acid sequence of the GITR agonist 44C1 heavy chain CDR1 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0503]

[0551] SEQ ID NO: 335 is the amino acid sequence of the GITR agonist 44C1 heavy chain CDR2 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0504]

[0552] SEQ ID NO: 336 is the amino acid sequence of the GITR agonist 44C1 heavy chain CDR3 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0505]

[0553] SEQ ID NO: 337 is the amino acid sequence of the GITR agonist 44C1 light chain CDR1 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0506]

[0554] SEQ ID NO: 338 is the amino acid sequence of the GITR agonist 44C1 light chain CDR2 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0507]

[0555] SEQ ID NO: 339 is the amino acid sequence of the GITR agonist 44C1 light chain CDR3 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0508]

[0556] SEQ ID NO: 340 is the amino acid sequence of the GITR agonist 45A8 heavy chain from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0509]

[0557] SEQ ID NO: 341 is the amino acid sequence of the GITR agonist 45A8 light chain from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0510]

[0558] SEQ ID NO: 342 is the amino acid sequence of the GITR agonist 45A8 variable heavy chain from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0511]

[0559] SEQ ID NO: 343 is the amino acid sequence of the GITR agonist 45A8 variable light chain from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0512]

[0560] SEQ ID NO: 344 is the amino acid sequence of GITR agonist 45A8 heavy chain CDR1 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0513]

[0561] SEQ ID NO: 345 is the amino acid sequence of the GITR agonist 45A8 heavy chain CDR2 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0514]

[0562] SEQ ID NO: 346 is the amino acid sequence of the GITR agonist 45A8 heavy chain CDR3 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0515]

[0563] SEQ ID NO: 347 is the amino acid sequence of the GITR agonist 45A8 light chain CDR1 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0516]

[0564] SEQ ID NO: 348 is the amino acid sequence of the GITR agonist 45A8 light chain CDR2 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0517]

[0565] SEQ ID NO: 349 is the amino acid sequence of the GITR agonist 45A8 light chain CDR3 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0518]

[0566] SEQ ID NO: 350 is the amino acid sequence of the GITR agonist 46E11 heavy chain from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0519]

[0567] SEQ ID NO: 351 is the amino acid sequence of the GITR agonist 46E11 light chain from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0520]

[0568] SEQ ID NO: 352 is the amino acid sequence of the GITR agonist 46E11 variable heavy chain from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0521]

[0569] SEQ ID NO: 353 is the amino acid sequence of the GITR agonist 46E11 variable light chain from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0522]

[0570] SEQ ID NO: 354 is the amino acid sequence of GITR agonist 46E11 heavy chain CDR1 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0523]

[0571] SEQ ID NO: 355 is the amino acid sequence of the GITR agonist 46E11 heavy chain CDR2 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0524]

[0572] SEQ ID NO: 356 is the amino acid sequence of the GITR agonist 46E11 heavy chain CDR3 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0525]

[0573] SEQ ID NO: 357 is the amino acid sequence of the GITR agonist 46E11 light chain CDR1 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0526]

[0574] SEQ ID NO: 358 is the amino acid sequence of the GITR agonist 46E11 light chain CDR2 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0527]

[0575] SEQ ID NO: 359 is the amino acid sequence of the GITR agonist 46E11 light chain CDR3 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0528]

[0576] SEQ ID NO: 360 is the amino acid sequence of the GITR agonist 48H12 heavy chain from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0529]

[0577] SEQ ID NO: 361 is the amino acid sequence of the GITR agonist 48H12 light chain from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0530]

[0578] SEQ ID NO: 362 is the amino acid sequence of the GITR agonist 48H12 variable heavy chain from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0531]

[0579] SEQ ID NO: 363 is the amino acid sequence of the GITR agonist 48H12 variable light chain from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0532]

[0580] SEQ ID NO: 364 is the amino acid sequence of the GITR agonist 48H12 heavy chain CDR1 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0533]

[0581] SEQ ID NO: 365 is the amino acid sequence of the GITR agonist 48H12 heavy chain CDR2 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0534]

[0582] SEQ ID NO: 366 is the amino acid sequence of the GITR agonist 48H12 heavy chain CDR3 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0535]

[0583] SEQ ID NO: 367 is the amino acid sequence of the GITR agonist 48H12 light chain CDR1 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0536]

[0584] SEQ ID NO: 368 is the amino acid sequence of the GITR agonist 48H12 light chain CDR2 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0537]

[0585] SEQ ID NO: 369 is the amino acid sequence of the GITR agonist 48H12 light chain CDR3 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0538]

[0586] SEQ ID NO: 370 is the amino acid sequence of the GITR agonist 48H7 heavy chain from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0539]

[0587] SEQ ID NO: 371 is the amino acid sequence of the GITR agonist 48H7 light chain from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0540]

[0588] SEQ ID NO: 372 is the amino acid sequence of the GITR agonist 48H7 variable heavy chain from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0541]

[0589] SEQ ID NO: 373 is the amino acid sequence of the GITR agonist 48H7 variable light chain from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0542]

[0590] SEQ ID NO: 374 is the amino acid sequence of the GITR agonist 48H7 heavy chain CDR1 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0543]

[0591] SEQ ID NO: 375 is the amino acid sequence of the GITR agonist 48H7 heavy chain CDR2 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0544]

[0592] SEQ ID NO: 376 is the amino acid sequence of the GITR agonist 48H7 heavy chain CDR3 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0545]

[0593] SEQ ID NO: 377 is the amino acid sequence of the GITR agonist 48H7 light chain CDR1 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0546]

[0594] SEQ ID NO: 378 is the amino acid sequence of the GITR agonist 48H7 light chain CDR2 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0547]

[0595] SEQ ID NO: 379 is the amino acid sequence of the GITR agonist 48H7 light chain CDR3 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0548]

[0596] SEQ ID NO: 380 is the amino acid sequence of the GITR agonist 49D9 heavy chain from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0549]

[0597] SEQ ID NO: 381 is the amino acid sequence of the GITR agonist 49D9 light chain from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0550]

[0598] SEQ ID NO: 382 is the amino acid sequence of the GITR agonist 49D9 variable heavy chain from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0551]

[0599] SEQ ID NO: 383 is the amino acid sequence of the GITR agonist 49D9 variable light chain from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0552]

[0600] SEQ ID NO: 384 is the amino acid sequence of the GITR agonist 49D9 heavy chain CDR1 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0553]

[0601] SEQ ID NO: 385 is the amino acid sequence of the GITR agonist 49D9 heavy chain CDR2 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0554]

[0602] SEQ ID NO: 386 is the amino acid sequence of the GITR agonist 49D9 heavy chain CDR3 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0555]

[0603] SEQ ID NO: 387 is the amino acid sequence of the GITR agonist 49D9 light chain CDR1 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0556]

[0604] SEQ ID NO: 388 is the amino acid sequence of the GITR agonist 49D9 light chain CDR2 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0557]

[0605] SEQ ID NO: 389 is the amino acid sequence of the GITR agonist 49D9 light chain CDR3 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0558]

[0606] SEQ ID NO: 390 is the amino acid sequence of the GITR agonist 49E2 heavy chain from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0559]

[0607] SEQ ID NO: 391 is the amino acid sequence of the GITR agonist 49E2 light chain from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0560]

[0608] SEQ ID NO: 392 is the amino acid sequence of the GITR agonist 49E2 variable heavy chain from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0561]

[0609] SEQ ID NO: 393 is the amino acid sequence of the GITR agonist 49E2 variable light chain from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0562]

[0610] SEQ ID NO: 394 is the amino acid sequence of the GITR agonist 49E2 heavy chain CDR1 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0563]

[0611] SEQ ID NO: 395 is the amino acid sequence of the GITR agonist 49E2 heavy chain CDR2 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0564]

[0612] SEQ ID NO: 396 is the amino acid sequence of the GITR agonist 49E2 heavy chain CDR3 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0565]

[0613] SEQ ID NO: 397 is the amino acid sequence of the GITR agonist 49E2 light chain CDR1 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0566]

[0614] SEQ ID NO: 398 is the amino acid sequence of the GITR agonist 49E2 light chain CDR2 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0567]

[0615] SEQ ID NO: 399 is the amino acid sequence of the GITR agonist 49E2 light chain CDR3 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0568]

[0616] SEQ ID NO: 400 is the amino acid sequence of the GITR agonist 48A9 heavy chain from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0569]

[0617] SEQ ID NO: 401 is the amino acid sequence of the GITR agonist 48A9 light chain from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0570]

[0618] SEQ ID NO: 402 is the amino acid sequence of the GITR agonist 48A9 variable heavy chain from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0571]

[0619] SEQ ID NO: 403 is the amino acid sequence of the GITR agonist 48A9 variable light chain from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0572]

[0620] SEQ ID NO: 404 is the amino acid sequence of the GITR agonist 48A9 heavy chain CDR1 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0573]

[0621] SEQ ID NO: 405 is the amino acid sequence of the GITR agonist 48A9 heavy chain CDR2 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0574]

[0622] SEQ ID NO: 406 is the amino acid sequence of the GITR agonist 48A9 heavy chain CDR3 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0575]

[0623] SEQ ID NO: 407 is the amino acid sequence of the GITR agonist 48A9 light chain CDR1 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0576]

[0624] SEQ ID NO: 408 is the amino acid sequence of the GITR agonist 48A9 light chain CDR2 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0577]

[0625] SEQ ID NO: 409 is the amino acid sequence of the GITR agonist 48A9 light chain CDR3 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0578]

[0626] SEQ ID NO: 410 is the amino acid sequence of the GITR agonist 5H7 heavy chain from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0579]

[0627] SEQ ID NO: 411 is the amino acid sequence of the GITR agonist 5H7 light chain from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0580]

[0628] SEQ ID NO: 412 is the amino acid sequence of the GITR agonist 5H7 variable heavy chain from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0581]

[0629] SEQ ID NO: 413 is the amino acid sequence of the GITR agonist 5H7 variable light chain from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0582]

[0630] SEQ ID NO: 414 is the amino acid sequence of the GITR agonist 5H7 heavy chain CDR1 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0583]

[0631] SEQ ID NO: 415 is the amino acid sequence of the GITR agonist 5H7 heavy chain CDR2 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0584]

[0632] SEQ ID NO: 416 is the amino acid sequence of the GITR agonist 5H7 heavy chain CDR3 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0585]

[0633] SEQ ID NO: 417 is the amino acid sequence of the GITR agonist 5H7 light chain CDR1 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0586]

[0634] SEQ ID NO: 418 is the amino acid sequence of the GITR agonist 5H7 light chain CDR2 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0587]

[0635] SEQ ID NO: 419 is the amino acid sequence of the GITR agonist 5H7 light chain CDR3 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0588]

[0636] SEQ ID NO: 420 is the amino acid sequence of the GITR agonist 7A10 heavy chain from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0589]

[0637] SEQ ID NO: 421 is the amino acid sequence of the GITR agonist 7A10 light chain from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0590]

[0638] SEQ ID NO: 422 is the amino acid sequence of the GITR agonist 7A10 variable heavy chain from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0591]

[0639] SEQ ID NO: 423 is the amino acid sequence of the GITR agonist 7A10 variable light chain from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0592]

[0640] SEQ ID NO: 424 is the amino acid sequence of GITR agonist 7A10 heavy chain CDR1 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0593]

[0641] SEQ ID NO: 425 is the amino acid sequence of the GITR agonist 7A10 heavy chain CDR2 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0594]

[0642] SEQ ID NO: 426 is the amino acid sequence of the GITR agonist 7A10 heavy chain CDR3 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0595]

[0643] SEQ ID NO: 427 is the amino acid sequence of the GITR agonist 7A10 light chain CDR1 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0596]

[0644] SEQ ID NO: 428 is the amino acid sequence of the GITR agonist 7A10 light chain CDR2 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0597]

[0645] SEQ ID NO: 429 is the amino acid sequence of the GITR agonist 7A10 light chain CDR3 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0598]

[0646] SEQ ID NO: 430 is the amino acid sequence of the GITR agonist 9H6 heavy chain from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0599]

[0647] SEQ ID NO: 431 is the amino acid sequence of the GITR agonist 9H6 light chain from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0600]

[0648] SEQ ID NO: 432 is the amino acid sequence of the GITR agonist 9H6 variable heavy chain from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0601]

[0649] SEQ ID NO: 433 is the amino acid sequence of the GITR agonist 9H6 variable light chain from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0602]

[0650] SEQ ID NO: 434 is the amino acid sequence of the GITR agonist 9H6 heavy chain CDR1 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0603]

[0651] SEQ ID NO: 435 is the amino acid sequence of the GITR agonist 9H6 heavy chain CDR2 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0604]

[0652] SEQ ID NO: 436 is the amino acid sequence of the GITR agonist 9H6 heavy chain CDR3 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0605]

[0653] SEQ ID NO: 437 is the amino acid sequence of the GITR agonist 9H6 light chain CDR1 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0606]

[0654] SEQ ID NO: 438 is the amino acid sequence of the GITR agonist 9H6 light chain CDR2 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0607]

[0655] SEQ ID NO: 439 is the amino acid sequence of the GITR agonist 9H6 light chain CDR3 from U.S. Patent Application Publication No. 2015 / 0064204 A1.

[0608]

[0656] SEQ ID NO: 440 is the GITR ligand (GITRL) amino acid sequence.

[0609]

[0657] SEQ ID NO:441 is the soluble portion of the GITRL polypeptide.

[0610]

[0658] SEQ ID NO: 442 is the amino acid sequence of human HVEM (CD270).

[0611]

[0659] SEQ ID NO: 443 is the HVEM ligand (LIGHT) amino acid sequence.

[0612]

[0660] SEQ ID NO:444 is a soluble portion of the LIGHT polypeptide.

[0613]

[0661] SEQ ID NO:445 is an alternative soluble portion of the LIGHT polypeptide.

[0614]

[0662] SEQ ID NO:446 is an alternative soluble portion of the LIGHT polypeptide.

[0615]

[0663] SEQ ID NO: 447 is the amino acid sequence of human CD95 isoform 1.

[0616]

[0664] SEQ ID NO: 448 is the amino acid sequence of human CD95 isoform 2.

[0617]

[0665] SEQ ID NO: 449 is the amino acid sequence of human CD95 isoform 3.

[0618]

[0666] SEQ ID NO: 450 is the amino acid sequence of human CD95 isoform 4.

[0619]

[0667] SEQ ID NO: 451 is the heavy chain variable region (V) of the CD95 agonist monoclonal antibody E09 H )

[0620]

[0668] SEQ ID NO: 452 is the light chain variable region (V) of the CD95 agonist monoclonal antibody E09 L )

[0621]

[0669] SEQ ID NO: 453 is the heavy chain CDR1 of the CD95 agonist monoclonal antibody E09.

[0622]

[0670] SEQ ID NO: 454 is the heavy chain CDR2 of the CD95 agonist monoclonal antibody E09.

[0623]

[0671] SEQ ID NO: 455 is the heavy chain CDR3 of the CD95 agonist monoclonal antibody E09.

[0624]

[0672] SEQ ID NO: 456 is the light chain CDR1 of the CD95 agonist monoclonal antibody E09.

[0625]

[0673] SEQ ID NO: 457 is the light chain CDR2 of the CD95 agonist monoclonal antibody E09.

[0626]

[0674] SEQ ID NO: 458 is the light chain CDR3 of the CD95 agonist monoclonal antibody E09.

[0627]

[0675] SEQ ID NO: 459 is the CD95 ligand (CD95L) amino acid sequence.

[0628]

[0676] SEQ ID NO:460 is the soluble portion of the CD95L polypeptide.

[0629]

[0677] SEQ ID NO:461 is an alternative soluble portion of the CD95L polypeptide.

[0630]

[0678] SEQ ID NO:462 is an alternative soluble portion of the CD95L polypeptide.

[0631]

[0679] SEQ ID NO: 463 is the heavy chain amino acid sequence of the PD-1 inhibitor nivolumab.

[0632]

[0680] SEQ ID NO: 464 is the light chain amino acid sequence of the PD-1 inhibitor nivolumab.

[0633]

[0681] SEQ ID NO: 465 is the heavy chain variable region (V H ) amino acid sequence.

[0634]

[0682] SEQ ID NO: 466 represents the light chain variable region (V) of the PD-1 inhibitor nivolumab L ) amino acid sequence.

[0635]

[0683] SEQ ID NO: 467 is the heavy chain CDR1 amino acid sequence of the PD-1 inhibitor nivolumab.

[0636]

[0684] SEQ ID NO: 468 is the heavy chain CDR2 amino acid sequence of the PD-1 inhibitor nivolumab.

[0637]

[0685] SEQ ID NO: 469 is the heavy chain CDR3 amino acid sequence of the PD-1 inhibitor nivolumab.

[0638]

[0686] SEQ ID NO: 470 is the light chain CDR1 amino acid sequence of the PD-1 inhibitor nivolumab.

[0639]

[0687] SEQ ID NO: 471 is the light chain CDR2 amino acid sequence of the PD-1 inhibitor nivolumab.

[0640]

[0688] SEQ ID NO: 472 is the light chain CDR3 amino acid sequence of the PD-1 inhibitor nivolumab.

[0641]

[0689] SEQ ID NO: 473 is the heavy chain amino acid sequence of the PD-1 inhibitor pembrolizumab.

[0642]

[0690] SEQ ID NO: 474 is the light chain amino acid sequence of the PD-1 inhibitor pembrolizumab.

[0643]

[0691] SEQ ID NO: 475 is the heavy chain variable region (V H ) amino acid sequence.

[0644]

[0692] SEQ ID NO: 476 is the light chain variable region (V L ) amino acid sequence.

[0645]

[0693] SEQ ID NO: 477 is the heavy chain CDR1 amino acid sequence of the PD-1 inhibitor pembrolizumab.

[0646]

[0694] SEQ ID NO: 478 is the heavy chain CDR2 amino acid sequence of the PD-1 inhibitor pembrolizumab.

[0647]

[0695] SEQ ID NO: 479 is the heavy chain CDR3 amino acid sequence of the PD-1 inhibitor pembrolizumab.

[0648]

[0696] SEQ ID NO: 480 is the light chain CDR1 amino acid sequence of the PD-1 inhibitor pembrolizumab.

[0649]

[0697] SEQ ID NO: 481 is the light chain CDR2 amino acid sequence of the PD-1 inhibitor pembrolizumab.

[0650]

[0698] SEQ ID NO: 482 is the light chain CDR3 amino acid sequence of the PD-1 inhibitor pembrolizumab.

[0651]

[0699] SEQ ID NO: 483 is the heavy chain amino acid sequence of the PD-L1 inhibitor durvalumab.

[0652]

[0700] SEQ ID NO: 484 is the light chain amino acid sequence of the PD-L1 inhibitor durvalumab.

[0653]

[0701] SEQ ID NO: 485 is the heavy chain variable region (V H ) amino acid sequence.

[0654]

[0702] SEQ ID NO: 486 is the light chain variable region (V) of the PD-L1 inhibitor durvalumab L ) amino acid sequence.

[0655]

[0703] SEQ ID NO: 487 is the heavy chain CDR1 amino acid sequence of the PD-L1 inhibitor durvalumab.

[0656]

[0704] SEQ ID NO: 488 is the heavy chain CDR2 amino acid sequence of the PD-L1 inhibitor durvalumab.

[0657]

[0705] SEQ ID NO: 489 is the heavy chain CDR3 amino acid sequence of the PD-L1 inhibitor durvalumab.

[0658]

[0706] SEQ ID NO: 490 is the light chain CDR1 amino acid sequence of the PD-L1 inhibitor durvalumab.

[0659]

[0707] SEQ ID NO: 491 is the light chain CDR2 amino acid sequence of the PD-L1 inhibitor durvalumab.

[0660]

[0708] SEQ ID NO: 492 is the light chain CDR3 amino acid sequence of the PD-L1 inhibitor durvalumab.

[0661]

[0709] SEQ ID NO: 493 is the heavy chain amino acid sequence of the PD-L1 inhibitor avelumab.

[0662]

[0710] SEQ ID NO: 494 is the light chain amino acid sequence of the PD-L1 inhibitor avelumab.

[0663]

[0711] SEQ ID NO: 495 is the heavy chain variable region (V H ) amino acid sequence.

[0664]

[0712] SEQ ID NO: 496 is the light chain variable region (V L ) amino acid sequence.

[0665]

[0713] SEQ ID NO: 497 is the heavy chain CDR1 amino acid sequence of the PD-L1 inhibitor avelumab.

[0666]

[0714] SEQ ID NO: 498 is the heavy chain CDR2 amino acid sequence of the PD-L1 inhibitor avelumab.

[0667]

[0715] SEQ ID NO: 499 is the heavy chain CDR3 amino acid sequence of the PD-L1 inhibitor avelumab.

[0668]

[0716] SEQ ID NO: 500 is the light chain CDR1 amino acid sequence of the PD-L1 inhibitor avelumab.

[0669]

[0717] SEQ ID NO: 501 is the light chain CDR2 amino acid sequence of the PD-L1 inhibitor avelumab.

[0670]

[0718] SEQ ID NO: 502 is the light chain CDR3 amino acid sequence of the PD-L1 inhibitor avelumab.

[0671]

[0719] SEQ ID NO: 503 is the heavy chain amino acid sequence of the PD-L1 inhibitor atezolizumab.

[0672]

[0720] SEQ ID NO: 504 is the light chain amino acid sequence of the PD-L1 inhibitor atezolizumab.

[0673]

[0721] SEQ ID NO: 505 is the heavy chain variable region (V H ) amino acid sequence.

[0674]

[0722] SEQ ID NO: 506 is the light chain variable region (V L ) amino acid sequence.

[0675]

[0723] SEQ ID NO: 507 is the heavy chain CDR1 amino acid sequence of the PD-L1 inhibitor atezolizumab.

[0676]

[0724] SEQ ID NO: 508 is the heavy chain CDR2 amino acid sequence of the PD-L1 inhibitor atezolizumab.

[0677]

[0725] SEQ ID NO: 509 is the heavy chain CDR3 amino acid sequence of the PD-L1 inhibitor atezolizumab.

[0678]

[0726] SEQ ID NO: 510 is the light chain CDR1 amino acid sequence of the PD-L1 inhibitor atezolizumab.

[0679]

[0727] SEQ ID NO: 511 is the light chain CDR2 amino acid sequence of the PD-L1 inhibitor atezolizumab.

[0680]

[0728] SEQ ID NO: 512 is the light chain CDR3 amino acid sequence of the PD-L1 inhibitor atezolizumab.

[0681] Detailed Description of the Invention

[0729] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. All patents and publications mentioned herein are incorporated by reference in their entirety.

[0682] definition

[0730] The terms "co-administration," "co-administer," "administered in conjunction with," "administered in conjunction with," "concurrently," and "concurrently," as used herein, encompass administration of two or more active pharmaceutical ingredients (e.g., at least one TNFRSF agonist and multiple TILs) to a subject such that both active pharmaceutical ingredients and / or their metabolites are present in the subject at the same time. Co-administration includes simultaneous administration in separate compositions, administration at different times in separate compositions, or administration in a composition in which two or more active pharmaceutical ingredients are present. Simultaneous administration in separate compositions and administration in a composition in which both agents are present are preferred.

[0683]

[0731] The term "rapid expansion" refers to an increase in the number of antigen-specific TILs of at least about 3-fold (or 4-fold, 5-fold, 6-fold, 7-fold, 8-fold, or 9-fold) in a one-week period, more preferably at least about 10-fold (or 20-fold, 30-fold, 40-fold, 50-fold, 60-fold, 70-fold, 80-fold, or 90-fold) in a one-week period, or most preferably at least about 100-fold in a one-week period. Several rapid expansion protocols are described herein.

[0684]

[0732] As used herein, "tumor infiltrating lymphocytes" or "TILs" refers to a population of cells, originally obtained as white blood cells, that have left the bloodstream of a subject and migrated into a tumor. TILs include, but are not limited to, CD8 + Cytotoxic T cells (lymphocytes), Th1 and Th17 CD4 +These include T cells, natural killer cells, dendritic cells, and M1 macrophages. TILs include both primary and secondary TILs. "Primary TILs" are those obtained from a patient tissue sample as outlined herein (sometimes referred to as "freshly harvested"), and "secondary TILs" are any TIL cell population that has been expanded or grown as discussed herein, including, but not limited to, bulk TILs and expanded TILs ("REP TILs" or "post-REP TILs").

[0685]

[0733] As used herein, a "cell population" (including TILs) refers to a large number of cells that share a common trait. Generally, a population is roughly 1 x 10 6 ~1×10 10 The range is 1 x 10, with different TIL populations containing different numbers. For example, the initial growth of primary TILs in the presence of IL-2 is approximately 1 x 10 8 REP expansion cultures generally yield a bulk TIL population of 1.5 x 10 cells. 9 ~1.5×10 10 This is done to provide a population of cells for injection.

[0686]

[0734] The term "central memory T cells" refers to cells that are CD45R0+ and CCR7 (CCR7 hi ) and CD62L (CD62 hi Central memory T cells are a subset of T cells that constitutively express CD4 receptors (CD4, CD127, CD127R, and CD15R). The surface phenotype of central memory T cells also includes TCR, CD3, CD127 (IL-7R), and IL-15R. Transcription factors of central memory T cells include BCL-6, BCL-6B, MBD2, and BMI1. After TCR triggering, central memory T cells primarily secrete IL-2 and CD40L as effector molecules. Central memory T cells are predominant in the CD4 compartment of the blood and are proportionally concentrated in lymph nodes and tonsils in humans.

[0687]

[0735] The term "anti-CD3 antibody" refers to an antibody or variant thereof directed against the CD3 receptor in the T cell antigen receptor of mature T cells, including monoclonal antibodies and human, humanized, chimeric, or murine antibodies. Anti-CD3 antibodies include OKT-3, also known as muromonab. Anti-CD3 antibodies also include the UHCT1 clone, also known as T3 and CD3ε. Other anti-CD3 antibodies include, for example, otelixizumab, teplizumab, and visilizumab.

[0688]

[0736] The term "OKT-3" (also referred to herein as "OKT3") refers to a monoclonal antibody against the CD3 receptor in the T cell antigen receptor of mature T cells, or a biosimilar or variant thereof, including a human, humanized, chimeric, or murine antibody, and includes commercially available forms such as OKT-3 (30 ng / mL, MACS GMP CD3 pure, Miltenyi Biotech, Inc., San Diego, CA, USA) and muromonab or its variants, conservative amino acid substitutions, glycoforms, or biosimilars. The amino acid sequences of the heavy and light chains of muromonab are provided in Table 1 (SEQ ID NO: 1 and SEQ ID NO: 2).

[0689] [Table 1]

[0690]

[0737] The term "IL-2" (also referred to herein as "IL2") refers to the T cell growth factor known as interleukin-2 and includes all forms of IL-2, including its human and mammalian forms, conservative amino acid substitutions, glycoforms, biosimilars, and variants. IL-2 is described, for example, in Nelson, J. Immunol. 2004, 172, 3983-88 and Malek, Annu. Rev. Immunol. 2008, 26, 453-79, the disclosures of which are incorporated herein by reference. The amino acid sequence of recombinant human IL-2 suitable for use in the present invention is provided in Table 2 (SEQ ID NO: 3). For example, the term IL-2 encompasses human recombinant forms of IL-2, such as aldesleukin (PROLEUKIN, commercially available from multiple suppliers at 22 million IU per single-use vial), as well as forms of recombinant IL-2 commercially supplied by CellGenix, Inc., Portsmouth, NH, USA (CELLGRO GMP) or ProSpec-Tany TechnoGene Ltd., East Brunswick, NJ, USA (catalog number CYT-209-b), and other commercially available equivalents from other vendors. Aldesleukin (des-alanyl-1, serine-125 human IL-2) is a non-glycosylated human recombinant form of IL-2 with a molecular weight of approximately 15 kDa. The amino acid sequence of an aldesleukin suitable for use in the present invention is provided in Table 2 (SEQ ID NO: 4). The term IL-2 also encompasses pegylated forms of IL-2 as described herein, including the pegylated IL2 prodrug NKTR-214 available from Nektar Therapeutics, South San Francisco, Calif., USA. NKTR-214 and pegylated IL-2 suitable for use in the present invention are described in U.S. Patent Application Publication No. 2014 / 0328791 A1 and WO 2012 / 065086 A1, the disclosures of which are incorporated herein by reference.Other forms of conjugated IL-2 suitable for use in the present invention are described in U.S. Patent Nos. 4,766,106, 5,206,344, 5,089,261, and 4,902,502, the disclosures of which are incorporated herein by reference. Formulations of IL-2 suitable for use in the present invention are described in U.S. Patent No. 6,706,289, the disclosure of which is incorporated herein by reference.

[0691] [Table 2]

[0692]

[0738] The term "IL-4" (also referred to herein as "IL4") refers to the cytokine known as interleukin 4, which is produced by Th2 T cells, as well as by eosinophils, basophils, and mast cells. IL-4 regulates the differentiation of naive helper T cells (Th0 cells) into Th2 T cells. Steinke and Borish, Respir. Res. 2001, 2, 66-70. Upon activation by IL-4, Th2 T cells subsequently produce additional IL-4 in a positive feedback loop. IL-4 also stimulates B cell proliferation and class II MHC expression and induces class switching from B cells to IgE and IgG1 expression. Recombinant human IL-4 suitable for use in the present invention is commercially available from several suppliers, including ProSpec-Tany TechnoGene Ltd., East Brunswick, NJ, USA (catalog number CYT-211) and ThermoFisherScientific, Inc., Waltham, MA, USA (human IL-15 recombinant protein, catalog number Gibco CTP0043). The amino acid sequence of recombinant human IL-4 suitable for use in the present invention is provided in Table 2 (SEQ ID NO: 5).

[0693]

[0739] The term "IL-7" (also referred to herein as "IL7") refers to the glycosylated tissue-derived cytokine known as interleukin-7, which is available from stromal and epithelial cells as well as dendritic cells. Fry and Mackall, Blood 2002, 99, 3892-904. IL-7 can stimulate T cell development. IL-7 binds to the heterodimeric IL-7 receptor, consisting of the IL-7 receptor α and the common γ chain receptor, which provides a series of signals important for T cell development in the thymus and survival in the periphery. Recombinant human IL-7 suitable for use in the present invention is commercially available from several suppliers, including ProSpec-Tany TechnoGene Ltd., East Brunswick, NJ, USA (catalog number CYT-254) and ThermoFisherScientific, Inc., Waltham, MA, USA (human IL-7 recombinant protein, catalog number Gibco PHC0071). The amino acid sequence of a recombinant human IL-7 suitable for use in the present invention is provided in Table 2 (SEQ ID NO: 6).

[0694]

[0740] The term "IL-15" (also referred to herein as "IL15") refers to the T-cell growth factor known as interleukin-15 and includes all forms of IL-15, including its human and mammalian forms, conservative amino acid substitutions, glycoforms, biosimilars, and variants. IL-15 is described, for example, in Fehniger and Caligiuri, Blood 2001, 97, 14-32, the disclosure of which is incorporated herein by reference. IL-15 shares β and γ signaling receptor subunits with IL-2. Recombinant human IL-15 is a single, non-glycosylated polypeptide chain containing 114 amino acids (plus an N-terminal methionine) with a molecular mass of 12.8 kDa. Recombinant human IL-15 is commercially available from several suppliers, including ProSpec-Tany TechnoGene Ltd., East Brunswick, NJ, USA (catalog number CYT-230-b) and ThermoFisherScientific, Inc., Waltham, MA, USA (human IL-15 recombinant protein, catalog number 34-8159-82). The amino acid sequence of recombinant human IL-15 suitable for use in the present invention is provided in Table 2 (SEQ ID NO: 7).

[0695]

[0741] The term "IL-21" (also referred to herein as "IL21") refers to the pleiotropic cytokine protein known as interleukin-21, and includes all forms of IL-21, including its human and mammalian forms, conservative amino acid substitutions, glycoforms, biosimilars, and variants. IL-21 is described, for example, in Spolski and Leonard, Nat. Rev. Drug. Disc. 2014, 13, 379-95, the disclosure of which is incorporated herein by reference. IL-21 primarily stimulates natural killer T cells and activated human CD4 +It is produced by T cells. Recombinant human IL-21 is a single, non-glycosylated polypeptide chain containing 132 amino acids with a molecular mass of 15.4 kDa. Recombinant human IL-21 is commercially available from several suppliers, including ProSpec-Tany TechnoGene Ltd., East Brunswick, NJ, USA (catalog number CYT-408-b) and ThermoFisherScientific, Inc., Waltham, MA, USA (human IL-21 recombinant protein, catalog number 14-8219-80). The amino acid sequence of recombinant human IL-21 suitable for use in the present invention is provided in Table 2 (SEQ ID NO: 8).

[0696]

[0742] The term "in vivo" refers to events that take place inside the body of a mammalian subject.

[0697]

[0743] The term "ex vivo" refers to events that take place outside the body of a mammalian subject in an artificial environment.

[0698]

[0744] The term "in vitro" refers to events that take place in a test system. In vitro assays include cell-based assays, which may utilize living or dead cells, and can also include cell-free assays, which do not utilize intact cells.

[0699]

[0745] The term "effective amount" or "therapeutically effective amount" refers to an amount of a compound or combination of compounds as described herein sufficient to achieve the intended application, including but not limited to disease treatment. A therapeutically effective amount may vary depending on the intended application (in vitro or in vivo), or the subject and disease condition under treatment (e.g., the subject's weight, age, and sex), the severity of the disease condition, or the method of administration. The term also applies to a dose that will produce a particular response in target cells (e.g., reduced platelet adhesion and / or cell migration). The specific dose will vary depending on the particular compound selected, the administration regimen to be followed, whether the compound is administered in combination with other compounds, the timing of administration, the tissue to which it is administered, and the physical delivery system that carries the compound.

[0700]

[0746] A "therapeutic effect," as that term is used herein, encompasses a therapeutic benefit and / or a prophylactic benefit. A prophylactic benefit includes delaying or eliminating the appearance of a disease or condition, delaying or eliminating the onset of symptoms of a disease or condition, slowing, halting, or reversing the progression of a disease or condition, or any combination thereof.

[0701]

[0747] The terms "QD," "qd," or "qd" mean quaque die, once a day, or once daily. The terms "BID," "bid," or "bid" mean bis in die, twice a day, or twice daily. The terms "TID," "tid," or "tid" mean ter in die, three times a day, or three times daily. The terms "QID," "qid," or "qid" mean quater in die, four times a day, or four times daily.

[0702]

[0748] The term "pharmaceutically acceptable salts" refers to salts derived from various organic and inorganic counterions known in the art. Pharmaceutically acceptable acid addition salts can be formed with inorganic and organic acids. Preferred inorganic acids from which salts can be derived include, for example, hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, and phosphoric acid. Preferred organic acids from which salts can be derived include, for example, acetic acid, propionic acid, glycolic acid, pyruvic acid, oxalic acid, maleic acid, malonic acid, succinic acid, fumaric acid, tartaric acid, citric acid, benzoic acid, cinnamic acid, mandelic acid, methanesulfonic acid, ethanesulfonic acid, p-toluenesulfonic acid, and salicylic acid. Pharmaceutically acceptable base addition salts can be formed with inorganic and organic bases. Inorganic bases from which salts can be derived include, for example, sodium, potassium, lithium, ammonium, calcium, magnesium, iron, zinc, copper, manganese, and aluminum. Organic bases from which salts can be derived include, for example, primary, secondary, and tertiary amines, substituted amines including naturally occurring substituted amines, cyclic amines, and basic ion exchange resins. Specific examples include isopropylamine, trimethylamine, diethylamine, triethylamine, tripropylamine, and ethanolamine. In some embodiments, the pharmaceutically acceptable base addition salt is selected from ammonium salts, potassium salts, sodium salts, calcium salts, and magnesium salts. The term "cocrystal" refers to a molecular complex derived from any of several cocrystal formers known in the art. Unlike salts, cocrystals typically do not involve hydrogen transfer between the cocrystal and the drug, but instead involve intermolecular interactions such as hydrogen bonding, aromatic ring stacking, or dispersive forces between the cocrystal former and the drug in the crystal structure.

[0703]

[0749] The term "pharmaceutically acceptable carrier" or "pharmaceutically acceptable excipient" is intended to include any solvents, dispersion media, coatings, antibacterial and antifungal agents, isotonic and absorption delaying agents, and inactive ingredients. The use of such pharmaceutically acceptable carriers or pharmaceutically acceptable excipients for active pharmaceutical ingredients is well known in the art. Except insofar as any conventional pharmaceutically acceptable carrier or pharmaceutically acceptable excipient is incompatible with the active pharmaceutical ingredient, its use in the therapeutic compositions of the invention is contemplated. Additional active pharmaceutical ingredients, such as other drugs, can also be incorporated into the compositions, processes, and methods described.

[0704]

[0750] The term "antigen" refers to a substance that induces an immune response. In some embodiments, an antigen is a molecule that, when presented by a major histocompatibility complex (MHC) molecule, has the ability to be bound by an antibody or a T cell receptor (TCR). The term "antigen," as used herein, also encompasses T cell epitopes. An antigen additionally has the ability to be recognized by the immune system. In some embodiments, an antigen has the ability to induce a humoral or cellular immune response that leads to the activation of B and / or T lymphocytes. In some cases, this may require that the antigen contain or be linked to a Th cell epitope. An antigen may also have one or more epitopes (e.g., B- and T-epitopes). In some embodiments, an antigen preferably reacts with its corresponding antibody or TCR, typically with high specificity and selectivity, and will not react with many other antibodies or TCRs that may be induced by other antigens.

[0705]

[0751] The terms "antibody" and its plural "antibodies" refer to whole immunoglobulins and any antigen-binding fragments ("antigen-binding portions") or single chains thereof. "Antibody" also refers to a glycoprotein comprising at least two heavy (H) chains and two light (L) chains inter-connected by disulfide bonds, or antigen-binding portions thereof. Each heavy chain contains a heavy chain variable region (referred to herein as V H Each light chain comprises a light chain variable region (abbreviated herein as V) and a heavy chain constant region. The heavy chain constant region comprises three domains, CH1, CH2, and CH3. L The light chain constant region comprises one domain, C L Contains the V of the antibody H and V L The regions can be further subdivided into regions of hypervariability called complementarity determining regions (CDRs) or hypervariable regions (HVRs), and may be interspersed with more highly conserved regions called framework regions (FRs). H and V L contains three CDRs and four FRs arranged from amino-terminus to carboxy-terminus in the following order: FR1, CDR1, FR2, CDR2, FR3, CDR3, FR4. The variable regions of the heavy and light chains contain binding domains that interact with one or more antigen epitopes. The constant region of the antibody may mediate the binding of the immunoglobulin to host tissues or factors, including various cells of the immune system (e.g., effector cells) and the first component (C1q) of the classical complement system.

[0706]

[0752] The terms "monoclonal antibody," "mAb," "monoclonal antibody composition," or plural forms thereof, refer to a preparation of antibody molecules of single molecular composition. A monoclonal antibody composition exhibits a single binding specificity and affinity for a particular epitope. Monoclonal antibodies specific to a TNFRSF receptor can be prepared using knowledge and skill in the art by injecting a test subject with an appropriate antigen and then isolating hybridomas expressing antibodies with the desired sequence or functional properties. DNA encoding the monoclonal antibody is readily isolated and sequenced using conventional procedures (e.g., by using oligonucleotide probes capable of specifically binding to genes encoding the heavy and light chains of the monoclonal antibody). Hybridoma cells serve as a preferred source of such DNA. After isolation, the DNA may be placed into an expression vector, which is then transfected into host cells that do not naturally express immunoglobulin proteins, such as Escherichia coli (E. coli) cells, simian COS cells, Chinese hamster ovary (CHO) cells, or myeloma cells, to achieve monoclonal antibody synthesis in the recombinant host cells. Recombinant production of antibodies is described in further detail below.

[0707]

[0753] The term "antigen-binding portion" or "antigen-binding fragment" of an antibody (or simply "antibody portion" or "fragment"), as used herein, refers to one or more fragments of an antibody that retain the ability to specifically bind to an antigen. It has been shown that the antigen-binding function of an antibody can be performed by fragments of a full-length antibody. Examples of binding fragments that fall within the term "antigen-binding portion" of an antibody include: (i) V L , V H , C L (ii) a Fab fragment, which is a monovalent fragment consisting of two Fab fragments linked by a disulfide bridge at the hinge region; (iii) a V H and an Fd fragment consisting of the CH1 domain; (iv) a V of a single arm of an antibody. L and V H Fv fragment consisting of domains, (v) V H or V Land (vi) isolated complementarity-determining regions (CDRs). In addition, two domains of an Fv fragment, V, and VB, can be included. L and V H are encoded by separate genes, which can be joined by synthetic linkers using recombinant methods to form V L and V H These regions can be paired together to form a single protein chain that forms a monovalent molecule known as a single-chain Fv (scFv); see, e.g., Bird, et al., Science 1988, 242, 423-426; and Huston, et al., Proc. Natl. Acad. Sci. USA 1988, 85, 5879-5883). Such scFv antibodies are also intended to be encompassed within the term "antigen-binding portion" or "antigen-binding fragment" of an antibody. These antibody fragments are obtained using conventional techniques known to those skilled in the art, and the fragments are screened for utility in the same manner as intact antibodies.

[0708]

[0754] The term "human antibody," as used herein, is intended to include antibodies having variable regions in which both the framework and CDR regions are derived from human germline immunoglobulin sequences. Furthermore, if the antibody contains a constant region, the constant region also is derived from human germline immunoglobulin sequences. The human antibodies of the invention 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). The term "human antibody," as used herein, is not intended to include antibodies in which CDR sequences derived from the germline of another mammalian species, such as a mouse, have been grafted onto human framework sequences.

[0709]

[0755] The term "human monoclonal antibody" refers to antibodies displaying a single binding specificity which have variable regions in which both the framework and CDR regions are derived from human germline immunoglobulin sequences. In certain 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 comprises human heavy chain and light chain transgenes fused to an immortalized cell.

[0710]

[0756] The term "recombinant human antibody," as used herein, includes any human antibody prepared, expressed, produced, or isolated by recombinant means, such as (a) antibodies isolated from animals (such as mice) transgenic or transchromosomal for human immunoglobulin genes or hybridomas prepared therefrom (described further below); (b) antibodies isolated from host cells, e.g., transfectomas, transformed to express a human antibody; (c) antibodies isolated from a recombinant combinatorial human antibody library; and (d) antibodies prepared, expressed, produced, or isolated by any other means involving splicing human immunoglobulin gene sequences to other DNA sequences. Such recombinant human antibodies have variable regions in which the framework and CDR regions are derived from human germline immunoglobulin sequences. In certain embodiments, however, such recombinant human antibodies can be subjected to in vitro mutagenesis (or, when animals transgenic for human Ig sequences are used, in vivo somatic mutagenesis) to thereby modify the V and CDR regions of the recombinant antibody. H and V L The amino acid sequence of the region is human germline V H and V L These are sequences that are derived from and related to sequences, but may not naturally occur in the human antibody germline repertoire in vivo.

[0711]

[0757] As used herein, "isotype" refers to the antibody class (e.g., IgM or IgG1) that is encoded by heavy chain constant region genes.

[0712]

[0758] The phrases "antibody that recognizes an antigen" and "antibody that is specific for an antigen" are used interchangeably herein with the term "antibody that specifically binds to an antigen."

[0713]

[0759] The term "human antibody derivative" refers to any modified form of a human antibody, including a conjugate of the antibody with another active pharmaceutical ingredient or antibody. The terms "conjugate," "antibody-drug conjugate," "ADC," or "immunoconjugate" refer to an antibody, or fragment thereof, conjugated to another therapeutic moiety, which can be conjugated to the antibodies described herein using methods available in the art.

[0714]

[0760] The terms "humanized antibody," "humanized antibodies," and "humanization" are intended to refer to antibodies in which CDR sequences derived from the germline of another mammalian species, such as mouse, have been grafted onto human framework sequences. Additional framework region modifications may be made within the human framework sequences. Humanized forms of non-human (e.g., murine) antibodies are chimeric antibodies containing minimal sequence derived from non-human immunoglobulin. In most cases, humanized antibodies are human immunoglobulins (recipient antibodies) in which residues from 15 hypervariable regions of a non-human species (donor antibody) such as mouse, rat, rabbit, or non-human primate having the desired specificity, affinity, and capacity have been replaced with residues from the recipient's hypervariable region. In some instances, Fv framework region (FR) residues of the human immunoglobulin are replaced with corresponding non-human residues. Furthermore, humanized antibodies may comprise residues that are not found in the recipient antibody or the donor antibody. These modifications are made to further refine antibody performance. Generally, a humanized antibody will comprise substantially all of at least one, and typically two, variable domains, in which all or substantially all of the hypervariable loops correspond to those of a non-human immunoglobulin and all or substantially all of the FR regions are those of a human immunoglobulin sequence. The humanized antibody will optionally also comprise at least a portion of an immunoglobulin constant region (Fc), typically that of a human immunoglobulin. For further details, see Jones, et al., Nature 1986, 321, 522-525; Riechmann, et al., Nature 1988, 332, 323-329; and Presta, Curr. Op. Struct. Biol. 1992, 2, 593-596. The TNFRSF agonists described herein may also be modified to utilize any Fc variants known to confer improved (e.g., reduced) effector function and / or FcR binding.Examples of Fc variants include WO 1988 / 07089 A1, WO 1996 / 14339 A1, WO 1998 / 05787 A1, WO 1998 / 23289 A1, WO 1999 / 51642 A1, WO 99 / 58572 A1, and WO 2000 / 09560. A2, 2000 / 32767 A1, 2000 / 42072 A2, 2002 / 44215 A2, 2002 / 060919 A2, 2003 / 074569 A2, 2004 / 016750 A2, 2004 / 029207 A2, 2004 / 035752 A2, 2004 / 063351 A2, 2004 / 074455 A2, 2004 / 099249 A2, 2005 / 040217 A2, 2005 / 070963 A1, 2005 / 077981 A2, 2005 / 092925 A2, 2005 / 123780 A2, 2006 / 019447 A1, 2006 / 047350 A2, and 2006 / 085967 A2; and any one of the amino acid substitutions disclosed in U.S. Patent Nos. 5,648,260; 5,739,277; 5,834,250; 5,869,046; 6,096,871; 6,121,022; 6,194,551; 6,242,195; 6,277,375; 6,528,624; 6,538,124; 6,737,056; 6,821,505; 6,998,253; and 7,083,784; the disclosures of which are incorporated herein by reference.

[0715]

[0761] The term "chimeric antibody" is intended to refer to an antibody in which the variable region sequences are derived from one species and the constant region sequences are derived from another species, such as an antibody in which the variable region sequences are derived from a murine antibody and the constant region sequences are derived from a human antibody.

[0716]

[0762] A "diabody" is a small antibody fragment that has two antigen-binding sites. Such a fragment consists of a heavy chain variable domain (V H ) is the light chain variable domain (VL ) connected to the same polypeptide chain (V H -V L or V L -V H (The term "diabodies" refers to a diabody that contains a complementary domain on a different chain.) By using a linker that is too short to allow pairing between the two domains on the same chain, the domains are forced to pair with complementary domains on another chain, creating two antigen-binding sites. Diabodies are described in more detail in, for example, EP 404,097; WO 93 / 11161; and Bolliger, et al., Proc. Natl. Acad. Sci. USA 1993, 90, 6444-6448.

[0717]

[0763] The term "glycosylation" refers to a modified derivative of an antibody. An aglycoslated antibody lacks glycosylation. Glycosylation can be altered to, for example, increase the affinity of the antibody for an antigen. Such carbohydrate modifications can be achieved, for example, by altering one or more glycosylation sites within the antibody sequence. For example, one or more amino acid substitutions can be made that result in the elimination of one or more variable region framework glycosylation sites, thereby eliminating glycosylation at those sites. Aglycosylation can increase the affinity of the antibody for the antigen, as described in U.S. Pat. Nos. 5,714,350 and 6,350,861. Additionally or alternatively, antibodies can be generated with altered types of glycosylation, such as hypofucosylated antibodies with reduced amounts of fucosyl residues or antibodies with increased bisecting GlcNac structures. Such altered glycosylation patterns have been demonstrated to increase antibody potency. Such carbohydrate modifications can be achieved, for example, by expressing the antibody in a host cell with altered glycosylation machinery. Cells with altered glycosylation machinery have been described in the art and can be used as host cells to express the recombinant antibodies of the invention, thereby producing antibodies with altered glycosylation. For example, the cell lines Ms704, Ms705, and Ms709 lack the fucosyltransferase gene FUT8 (α(1,6)fucosyltransferase), and therefore antibodies expressed in the Ms704, Ms705, and Ms709 cell lines lack fucose on their carbohydrates. The Ms704, Ms705, and Ms709 FUT8- / - cell lines were generated by targeted disruption of the FUT8 gene in CHO / DG44 cells using two replacement vectors (see, e.g., U.S. Patent Application Publication No. 2004 / 0110704 or Yamane-Ohnuki, et al., Biotechnol. Bioeng., 2004, 87, 614-622).As another example, European Patent No. 1,176,195 describes a cell line in which the function of the FUT8 gene encoding fucosyltransferase has been disrupted, and antibodies expressed therein exhibit hypofucosylation due to reduced or absent α1,6-linkage-related enzyme activity. The patent also describes cell lines that have low or no enzymatic activity for the addition of fucose to N-acetylglucosamine bound to the Fc region of antibodies, such as the rat myeloma cell line YB2 / 0 (ATCC CRL 1662). WO 03 / 035835 describes a mutant CHO cell line, Lec 13 cells, that has a reduced ability to attach fucose to Asn(297)-linked carbohydrates, thereby also resulting in hypofucosylation of antibodies expressed in the host cells (see also Shields, et al., J. Biol. Chem. 2002, 277, 26733-26740). WO 99 / 54342 describes engineered cell lines that express glycoprotein-modifying glycosyltransferases (e.g., β(1,4)-N-acetylglucosaminyltransferase III (GnTIII)), such that antibodies expressed therein exhibit an increase in bisecting GlcNac structures, which results in increased ADCC activity of the antibodies (Umana, et al., Nat. Biotech. 1999, 17, (See also, e.g., J. Am. Soc. 1999, 14, 176-180.) Alternatively, the fucose residues of the antibody may be removed by cleavage with a fucosidase enzyme. For example, the fucosidase α-L-fucosidase removes fucosyl residues from antibodies as described in Tarentino, et al., Biochem. 1975, 14, 5516-5523.

[0718]

[0764] "PEGylation" refers to a modified antibody or fusion protein, or fragment thereof, that is typically reacted with polyethylene glycol (PEG), such as a reactive ester or aldehyde derivative of PEG, under conditions such that one or more PEG groups become attached to the antibody or antibody fragment. PEGylation can, for example, increase the biological (e.g., serum) half-life of the antibody. Preferably, PEGylation is carried out by an acylation reaction or an alkylation reaction with a reactive PEG molecule (or an analogous reactive water-soluble polymer). As used herein, the term "polyethylene glycol" refers to a mono(C1-C 10 PEG is intended to encompass any of the forms of PEG used to derivatize other proteins, such as alkoxy- or aryloxy-polyethylene glycol or polyethylene glycol-maleimide. The protein or antibody to be PEGylated may be a non-glycosylated protein or antibody. Pegylation methods are known in the art, e.g., as described in European Patent Nos. 0154316 and 0401384 and U.S. Patent No. 5,824,778 (the disclosures of each of which are incorporated herein by reference), and can be applied to the antibodies of the invention.

[0719]

[0765] The term "fusion protein" or "fusion polypeptide" refers to a protein that combines the properties of two or more individual proteins. Such proteins have at least two heterologous polypeptides covalently linked, either directly or via an amino acid linker. The polypeptides forming the fusion protein are typically linked C-terminally to N-terminally, but may also be linked C-terminally to C-terminally, N-terminally to N-terminally, or N-terminally to C-terminally. The polypeptides of a fusion protein may be in any order and may contain two or more of either or both of the constituent polypeptides. The term encompasses conservatively modified variants, polymorphic variants, alleles, mutants, subsequences, interspecies homologs, and immunogenic fragments of the antigens that make up the fusion protein. Fusion proteins of the present disclosure may also contain additional copies of the constituent antigens or immunogenic fragments thereof. Fusion proteins contain one or more binding domains linked together, and may be further linked to an Fc domain, such as an IgG Fc domain. Fusion proteins may also be further linked together to mimic monoclonal antibodies, providing six or more binding domains. Fusion protein can be produced by recombinant methods as known in the art.The preparation of fusion protein is known in the art, and is described in, for example, WO 1995 / 027735 A1, WO 2005 / 103077 A1, WO 2008 / 025516 A1, WO 2009 / 007120 A1, WO 2010 / 003766 A1, WO 2010 / 010051 A1, WO 2010 / 078966 A1, U.S. Patent Application Publication No. 2015 / 0125419 A1 and U.S. Patent Application Publication No. 2016 / 0272695 A1, and U.S. Patent No. 8,921,519 (the disclosure of each of which is incorporated herein by reference).

[0720]

[0766] The term "heterologous," when used with reference to a portion of a nucleic acid or protein, indicates that the nucleic acid or protein comprises two or more subsequences that are not found in the same relationship to each other in nature. For example, nucleic acids are typically produced recombinantly, having two or more sequences from unrelated genes arranged to create a novel functional nucleic acid, e.g., a promoter from one source and a coding region from another source, or coding regions from different sources. Similarly, a heterologous protein indicates that the protein comprises two or more subsequences that are not found in the same relationship to each other in nature (e.g., a fusion protein).

[0721]

[0767] The term "conservative amino acid substitution" refers to an amino acid sequence modification that does not abolish the binding of an antibody or fusion protein to an antigen. Conservative amino acid substitutions include the substitution of an amino acid from one class with an amino acid from the same class, where a class is defined by common physicochemical amino acid side chain properties and high substitution frequency in naturally occurring homologous proteins, as determined, for example, by standard Dayhoff frequency exchange matrices or BLOSUM matrices. Six general amino acid side chain classes have been classified, including class I (Cys); class II (Ser, Thr, Pro, Ala, Gly); class III (Asn, Asp, Gln, Glu); class IV (His, Arg, Lys); class V (Ile, Leu, Val, Met); and class VI (Phe, Tyr, Trp). For example, substituting Asp for another class III residue, such as Asn, Gln, or Glu, is a conservative substitution. Thus, a predicted non-essential amino acid residue in an antibody is preferably replaced with another amino acid residue from the same class. Methods for identifying conservative amino acid substitutions that do not abolish antigen binding are well known in the art (see, e.g., Brummell, et al., Biochemistry 1993, 32, 1180-1187; Kobayashi, et al., Protein Eng. 1999, 12, 879-884 (1999); and Burks, et al., Proc. Natl. Acad. Sci. USA 1997, 94, 412-417).

[0722]

[0768] The terms "sequence identity," "percent identity," and "percent sequence identity" (or their equivalents, e.g., "99% identical") in the context of two or more nucleic acids or polypeptides refer to two or more sequences or subsequences that are the same or have a specified percentage of identical nucleotides or amino acid residues when compared and aligned (introducing gaps as necessary) to maximize correspondence, not considering any conservative amino acid substitutions as part of the sequence identity. Percent identity can be measured using sequence comparison software or algorithms or by visual inspection. Various algorithms and software that can be used to achieve alignment of amino acid or nucleotide sequences are known in the art. Suitable programs for determining percent sequence identity include, for example, the BLAST suite of programs available from the U.S. government's National Center for Biotechnology Information BLAST website. Comparison between two sequences can be performed using either the BLASTN or BLASTP algorithm. BLASTN is used to compare nucleic acid sequences, while BLASTP is used to compare amino acid sequences. ALIGN, ALIGN-2 (Genentech, South San Francisco, California), or MegAlign available from DNASTAR are further publicly available software programs that can be used to align sequences. Those skilled in the art can determine appropriate parameters to maximize alignment with a particular alignment software. In certain embodiments, the default parameters of the alignment software are used.

[0723]

[0769] Certain embodiments of the present invention include variants of antibodies or fusion proteins. As used herein, the term "variant" includes, but is not limited to, antibodies or fusion proteins that contain an amino acid sequence that differs from the amino acid sequence of a reference antibody by one or more substitutions, deletions, and / or additions at specific positions within or adjacent to the amino acid sequence of the reference antibody. A variant may contain one or more conservative substitutions in its amino acid sequence compared to the amino acid sequence of the reference antibody. Conservative substitutions may involve, for example, the substitution of similarly charged or uncharged amino acids. A variant retains the ability of the reference antibody to specifically bind to the antigen.

[0724]

[0770] A nucleic acid sequence implicitly encompasses conservatively modified variants thereof (e.g., degenerate codon substitutions) and complementary sequences, as well as the explicitly designated sequence. Specifically, degenerate codon substitutions can be achieved by generating sequences in which the third position of one or more selected (or all) codons is substituted with mixed-base and / or deoxyinosine residues. Batzer, et al., Nucleic Acid Res. 1991, 19, 5081; Ohtsuka, et al., J. Biol. Chem. 1985, 260, 2605-2608; Rossolini, et al., Mol. Cell. Probes 1994, 8, 91-98. The term nucleic acid is used interchangeably with cDNA, mRNA, oligonucleotide, and polynucleotide.

[0725]

[0771] The term "biosimilar" refers to a biologic, including a monoclonal antibody or fusion protein, that is highly similar to an originator biologic approved in the United States, despite minor differences in clinically inactive components, and for which there are no clinically meaningful differences between the biologic and the originator in terms of pharmaceutical safety, purity, and potency. Furthermore, a generic biologic or "biosimilar" drug is a biologic that is similar to another biologic already approved for use by the European Medicines Agency. The term "biosimilar" is also used interchangeably by regulatory agencies in other countries and regions. A biologic or biopharmaceutical is a drug made by or derived from a biological source, such as bacteria or yeast. It can consist of relatively small molecules, such as human insulin or erythropoietin, or complex molecules, such as monoclonal antibodies. For example, if the originator monoclonal antibody is rituximab, a biosimilar monoclonal antibody approved by a medicines regulatory authority similar to rituximab would be a "biosimilar" to rituximab or a "biosimilar of" rituximab. In Europe, a generic or "biosimilar" biopharmaceutical is a biopharmaceutical that is similar to another biopharmaceutical that has already been authorized for use by the European Medicines Agency (EMA). The relevant legal basis for generic biopharmaceutical applications in Europe is Article 6 of Regulation (EC) No. 726 / 2004 and Article 10(4) of Directive 2001 / 83 / EC, as amended. Accordingly, in Europe, biosimilars may be authorized, granted authorization, or be the subject of an authorization application under Article 6 of Regulation (EC) No. 726 / 2004 and Article 10(4) of Directive 2001 / 83 / EC. The original biopharmaceutical product that has already been licensed may be referred to as the "originator product" in Europe. Some of the requirements for a product to be considered a biosimilar are outlined in the CHMP Guideline on Similar Biological Medicinal Products.In addition, product-specific guidelines, including guidelines for monoclonal antibody biosimilars, are provided by the EMA for each product and are published on its website. A biosimilar as described herein may be similar to the originator drug product in terms of quality attributes, biological activity, mechanism of action, safety profile, and / or efficacy. Additionally, a biosimilar may be used or intended for use in treating the same condition as the originator drug product. Thus, a biosimilar as described herein may be considered to have similar or highly similar quality attributes to the originator drug product. Alternatively, or in addition, a biosimilar as described herein may be considered to have similar or highly similar biological activity to the originator drug product. Alternatively, or in addition, a biosimilar as described herein may be considered to have similar or highly similar safety profile to the originator drug product. Alternatively, or in addition, a biosimilar as described herein may be considered to have similar or highly similar efficacy to the originator drug product. As described herein, biosimilars in Europe are compared to originator drug products approved by the EMA. However, in some cases, biosimilars may be compared to biopharmaceutical products approved outside the European Economic Area (non-EEA-authorized "comparator") in certain studies. Such studies include, for example, certain clinical trials and in vivo preclinical studies. As used herein, the term "biosimilar" also relates to biopharmaceutical products that have been compared to or can be compared to non-EEA-authorized comparators. Some biosimilars are proteins, such as antibodies, antibody fragments (e.g., antigen-binding portions), and fusion proteins. Protein biosimilars may have amino acid sequences with minor amino acid structural modifications (e.g., including amino acid deletions, additions, and / or substitutions) that do not significantly affect the function of the polypeptide. Biosimilars may include amino acid sequences that share 97% or more, e.g., 97%, 98%, 99%, or 100% sequence identity with the amino acid sequence of their originator drug product.A biosimilar may contain one or more post-translational modifications, such as, but not limited to, glycosylation, oxidation, deamidation, and / or truncation, that differ from those of the originator pharmaceutical formulation, provided that the differences do not result in a change in the safety and / or efficacy of the pharmaceutical formulation. A biosimilar may have the same or a different glycosylation pattern as the originator pharmaceutical formulation. In particular, but not limited to, a biosimilar may have a different glycosylation pattern if the difference addresses or is intended to address safety concerns associated with the originator pharmaceutical formulation. In addition, a biosimilar may deviate from the originator pharmaceutical formulation, for example, in terms of its strength, pharmaceutical form, formulation, excipients, and / or presentation, provided that the safety and efficacy of the pharmaceutical formulation are not compromised. In some embodiments, a biosimilar is provided as a composition further comprising one or more excipients, which may be the same or different from the excipients contained in the originator pharmaceutical formulation or originator biologic. Biosimilars may contain differences compared to the originator drug product, for example, in terms of pharmacokinetic (PK) and / or pharmacodynamic (PD) profile, but are still considered sufficiently similar to the originator drug product to be considered for approval or suitable for approval. In certain circumstances, biosimilars exhibit different binding properties compared to the originator drug product, where the different binding properties are not considered by regulatory authorities, such as the EMA, to preclude approval as a generic biologic. The term "biosimilar" is also used interchangeably by regulatory agencies in other countries and regions.

[0726]

[0772] As used herein, the term "4-1BB agonist" may refer to any antibody or protein that specifically binds to the 4-1BB (CD137) antigen. "Specifically binds" means that the binding molecule exhibits essentially no background binding to non-4-1BB molecules. The 4-1BB agonist may be any 4-1BB agonist known in the art. In particular, it is one of the 4-1BB agonists described in more detail herein. However, an isolated binding molecule that specifically binds to 4-1BB may have cross-reactivity with 4-1BB molecules from other species. 4-1BB agonist antibodies and proteins may also specifically bind to human 4-1BB (h4-1BB or hCD137), for example, on T cells.

[0727]

[0773] As used herein, the term "OX40 agonist" can refer to any antibody or protein that specifically binds to the OX40 (CD134) antigen. "Specifically binds" means that the binding molecule exhibits essentially no background binding to non-OX40 molecules. The OX40 agonist can be any OX40 agonist known in the art. In particular, it is one of the OX40 agonists described in more detail herein. However, an isolated binding molecule that specifically binds to OX40 may have cross-reactivity with OX40 molecules from other species. OX40 agonist antibodies and proteins can also specifically bind to human OX40 (hOX40 or hCD134), for example, on T cells.

[0728]

[0774] As used herein, the term "CD27 agonist" can refer to any antibody or protein that specifically binds to the CD27 antigen. "Specifically binds" means that the binding molecule exhibits essentially no background binding to non-CD27 molecules. The CD27 agonist can be any CD27 agonist known in the art. In particular, it is one of the CD27 agonists described in more detail herein. However, an isolated binding molecule that specifically binds to CD27 may have cross-reactivity with CD27 molecules from other species. CD27 agonist antibodies and proteins can also specifically bind to human CD27 (hCD27), for example, on T cells.

[0729]

[0775] As used herein, the term "GITR agonist" includes molecules comprising at least one antigen-binding site that specifically binds to GITR (CD357). "Specifically binds" means that the binding molecule exhibits essentially no background binding to non-GITR molecules. A GITR agonist can be any GITR agonist known in the art. In particular, it is one of the GITR agonists described in more detail herein. However, an isolated binding molecule that specifically binds to GITR may have cross-reactivity with GITR molecules from other species. GITR agonist antibodies and proteins can also specifically bind to human GITR (hGITR), for example, on T cells and dendritic cells.

[0730]

[0776] As used herein, the term "HVEM agonist" includes molecules comprising at least one antigen-binding site that specifically binds to HVEM (CD270). "Specifically binds" means that the binding molecule exhibits essentially no background binding to non-HVEM molecules. The HVEM agonist may be any HVEM agonist known in the art. In particular, it is one of the HVEM agonists described in more detail herein. However, an isolated binding molecule that specifically binds to HVEM may have cross-reactivity with HVEM molecules from other species. HVEM agonist antibodies and proteins may also specifically bind to human HVEM (hHVEM), for example, on T cells.

[0731]

[0777] The term "hematological malignancies" refers to cancers and tumors of hematopoietic and lymphoid tissues in mammals, including, but not limited to, blood, bone marrow, lymph nodes, and lymphatic tissues. Hematological malignancies are also referred to as "liquid tumors." Hematological malignancies include, but are not limited to, acute lymphoblastic leukemia (ALL), chronic lymphocytic lymphoma (CLL), small lymphocytic lymphoma (SLL), acute myeloid leukemia (AML), chronic myelocytic leukemia (CML), acute monocytic leukemia (AMoL), Hodgkin's lymphoma, and non-Hodgkin's lymphoma. The term "B-cell hematological malignancies" refers to hematological malignancies affecting B cells.

[0732]

[0778] The term "solid tumor" refers to an abnormal mass of tissue that usually does not contain cysts or fluid areas. Solid tumors can be benign or malignant. The term "solid tumor cancer" refers to a malignant, neoplastic, or cancerous solid tumor. Solid tumor cancers include, but are not limited to, sarcomas, carcinomas, and lymphomas, such as lung cancer, breast cancer, prostate cancer, colon cancer, rectal cancer, and bladder cancer. The histology of a solid tumor comprises interdependent tissue compartments, including parenchyma (cancer cells) and supporting stromal cells, which may provide a supportive microenvironment within which the cancer cells are dispersed.

[0733]

[0779] The term "microenvironment," as used herein, may refer to the entire microenvironment of a solid tumor or hematological tumor or to individual cell subsets within the microenvironment. As used herein, the tumor microenvironment refers to a complex mixture of "cells, soluble factors, signaling molecules, extracellular matrix, and mechanical cues that promote neoplastic transformation, support tumor growth and invasion, protect tumors from host immunity, promote therapeutic resistance, and provide a niche for overt metastasis development," as described in Swartz, et al., Cancer Res., 2012, 72, 2473. Tumors express antigens that should be recognized by T cells, but tumor clearance by the immune system is rare due to immunosuppression by the microenvironment.

[0734]

[0780] For the avoidance of doubt, it is intended that any particular feature (e.g., integer, property, value, use, disease, formula, compound, or group) described herein in conjunction with a particular aspect, embodiment, or example of the invention be understood to be applicable to any other aspect, embodiment, or example described herein, except where inconsistent therewith. Accordingly, such feature may, where appropriate, be used in conjunction with any of the definitions, claims, or embodiments defined herein. All features disclosed in this specification (including any accompanying claims, abstract, and drawings), and / or all steps of any method or process so disclosed, may be combined in any combination, except combinations in which at least some of the features and / or steps are mutually exclusive. The invention is not limited to any details of any disclosed embodiment. The invention extends to any novel one or novel combination of features disclosed in this specification (including any accompanying claims, abstract, and drawings), or to any novel one or novel combination of steps of any method or process so disclosed.

[0735]

[0781] The terms "about" and "approximately" mean within a statistically meaningful range of a value. Such a range may be within an order of magnitude of a given value or range, preferably within 50%, more preferably within 20%, more preferably still within 10%, and even more preferably within 5%. The allowable variation encompassed by the terms "about" or "approximately" depends on the specific system under study and can be readily understood by one of ordinary skill in the art. Furthermore, as used herein, the terms "about" and "approximately" mean that dimensions, sizes, formulations, parameters, shapes, and other quantities and characteristics are not, and may not be, exactly that, and may be approximate and / or larger or smaller, as appropriate, reflecting tolerances, conversion factors, rounding, measurement errors, etc., and other factors known to those of ordinary skill in the art. In general, a dimension, size, formulation, parameter, shape, or other quantity or characteristic is "about" or "approximately" whether or not expressly stated as such. It is noted that embodiments of widely different sizes, shapes, and dimensions may use the described configurations.

[0736]

[0782] The transitional phrases "comprising," "consisting essentially of," and "consisting of," when used in the accompanying claims in their original and amended form, define the claim in terms of what, if any, additional unrecited claim elements or steps are excluded from the claim. The term "comprising" is intended to be inclusive or open-ended and does not exclude any additional unrecited elements, methods, steps, or materials. The term "consisting of" excludes any element, step, or material other than that specified in the claim, and in the latter instance, also excludes impurities normally associated with the specified material. The term "consisting essentially of" limits the claim to the specified elements, steps, or materials and those that do not materially affect the basic and novel characteristics of the claimed invention. All compositions, methods, and kits described herein embodying the present invention may, in alternative embodiments, be more specifically defined by any of the transitional phrases "comprising," "consisting essentially of," and "consisting of."

[0737] 4-1BB (CD137) agonist

[0783] In one embodiment, the TNFRSF agonist is a 4-1BB (CD137) agonist. The 4-1BB agonist may be any 4-1BB binding molecule known in the art. The 4-1BB binding molecule may be a monoclonal antibody or fusion protein capable of binding to human or mammalian 4-1BB. The 4-1BB agonist or 4-1BB binding molecule may comprise an immunoglobulin heavy chain of any isotype (e.g., IgG, IgE, IgM, IgD, IgA, and IgY), class (e.g., IgG1, IgG2, IgG3, IgG4, IgA1, and IgA2), or subclass of immunoglobulin molecule. The 4-1BB agonist or 4-1BB binding molecule may have both a heavy chain and a light chain. As used herein, the term binding molecule also includes antibodies (including full-length antibodies), monoclonal antibodies (including full-length monoclonal antibodies), polyclonal antibodies, multispecific antibodies (e.g., bispecific antibodies), human, humanized, or chimeric antibodies that bind to 4-1BB, and antibody fragments, such as Fab fragments, F(ab') fragments, fragments produced by an Fab expression library, epitope-binding fragments, and engineered forms of antibodies, such as scFv molecules, of any of the above. In certain embodiments, the 4-1BB agonist is an antigen-binding protein that is a fully human antibody. In certain embodiments, the 4-1BB agonist is an antigen-binding protein that is a humanized antibody. In some embodiments, the 4-1BB agonist used in the methods and compositions of the present disclosure includes an anti-4-1BB antibody, a human anti-4-1BB antibody, a mouse anti-4-1BB antibody, a mammalian anti-4-1BB antibody, a monoclonal anti-4-1BB antibody, a polyclonal anti-4-1BB antibody, a chimeric anti-4-1BB antibody, an anti-4-1BB Adnectin, an anti-4-1BB domain antibody, a single-chain anti-4-1BB fragment, a heavy-chain anti-4-1BB fragment, a light-chain anti-4-1BB fragment, an anti-4-1BB fusion protein, and fragments, derivatives, conjugates, variants, or biosimilars thereof. Agonistic anti-4-1BB antibodies are known to induce potent immune responses. Lee, et al., PLOS One 2013, 8, e69677.In a preferred embodiment, the 4-1BB agonist is an agonist anti-4-1BB humanized or fully human monoclonal antibody (i.e., an antibody derived from a single cell line). In one embodiment, the 4-1BB agonist is EU-101 (Eutilex Co. Ltd.), utomilumab, or urelumab, or a fragment, derivative, conjugate, variant, or biosimilar thereof. In a preferred embodiment, the 4-1BB agonist is utomilumab or urelumab, or a fragment, derivative, conjugate, variant, or biosimilar thereof.

[0738]

[0784] In a preferred embodiment, the 4-1BB agonist or 4-1BB binding molecule may also be a fusion protein. In a preferred embodiment, a multimeric 4-1BB agonist, such as a trimeric or hexameric 4-1BB agonist (having three or six ligand-binding domains), can induce superior receptor (4-1BBL) clustering and intracellular signaling complex formation compared to agonist monoclonal antibodies, which typically have two ligand-binding domains. Trimeric (trivalent) or hexameric (or hexavalent) or higher fusion proteins comprising three TNFRSF-binding domains and IgG1-Fc, and optionally further linking two or more of these fusion proteins, are described, for example, in Gieffers, et al., Mol. Cancer Therapeutics 2013, 12, 2735-47.

[0739]

[0785] Agonistic 4-1BB antibodies and fusion proteins are known to induce potent immune responses. In preferred embodiments, the 4-1BB agonist is a monoclonal antibody or fusion protein that specifically binds to the 4-1BB antigen in a manner sufficient to reduce toxicity. In some embodiments, the 4-1BB agonist is an agonistic 4-1BB monoclonal antibody or fusion protein that neutralizes antibody-dependent cellular cytotoxicity (ADCC), e.g., NK cell cytotoxicity. In some embodiments, the 4-1BB agonist is an agonistic 4-1BB monoclonal antibody or fusion protein that neutralizes antibody-dependent cellular phagocytosis (ADCP). In some embodiments, the 4-1BB agonist is an agonistic 4-1BB monoclonal antibody or fusion protein that neutralizes complement-dependent cytotoxicity (CDC). In some embodiments, the 4-1BB agonist is an agonistic 4-1BB monoclonal antibody or fusion protein that neutralizes Fc region function.

[0740]

[0786] In some embodiments, the 4-1BB agonist is characterized by high affinity binding and agonistic activity to human 4-1BB (SEQ ID NO: 9). In certain embodiments, the 4-1BB agonist is a binding molecule that binds to human 4-1BB (SEQ ID NO: 9). In certain embodiments, the 4-1BB agonist is a binding molecule that binds to mouse 4-1BB (SEQ ID NO: 10). The amino acid sequences of the 4-1BB antigens to which the 4-1BB agonists or binding molecules bind are summarized in Table 3.

[0741] [Table 3]

[0742]

[0787] In some embodiments, the compositions, processes, and methods described have a K of about 100 pM or less with human or mouse 4-1BB. D binds to human or mouse 4-1BB with a K of approximately 90 pM or less D binds to human or mouse 4-1BB with a K of approximately 80 pM or less Dbinds to human or mouse 4-1BB with a K of approximately 70 pM or less D binds to human or mouse 4-1BB with a K of approximately 60 pM or less D binds to human or mouse 4-1BB with a K of about 50 pM or less D binds to human or mouse 4-1BB with a K of about 40 pM or less D or binds to human or mouse 4-1BB with a K of about 30 pM or less D This includes 4-1BB agonists that bind at

[0743]

[0788] In some embodiments, the compositions, processes, and methods described deliver approximately 7.5×10 5 1 / M s or faster k assoc Approximately 7.5 × 10 5 1 / M s or faster k assoc Approximately 8 × 10 binds to human or mouse 4-1BB. 5 1 / M s or faster k assoc Approximately 8.5 × 10 5 1 / M s or faster k assoc Approximately 9 × 10 binds to human or mouse 4-1BB. 5 1 / M s or faster k assoc Approximately 9.5 × 10 5 1 / M s or faster k assoc or approximately 1 x 10 to human or mouse 4-1BB. 6 1 / M s or faster k assoc This includes 4-1BB agonists that bind at

[0744]

[0789] In some embodiments, the compositions, processes, and methods described deliver approximately 2×10 -5 1 / s or slower k dissoc Approximately 2.1 × 10 binds to human or mouse 4-1BB. -5 1 / s or slower k dissoc Approximately 2.2 × 10-5 1 / s or slower k dissoc Approximately 2.3 × 10 -5 1 / s or slower k dissoc Approximately 2.4 × 10 -5 1 / s or slower k dissoc Approximately 2.5 × 10 -5 1 / s or slower k dissoc Approximately 2.6 × 10 -5 1 / s or slower k dissoc or approximately 2.7 x 10 to human or mouse 4-1BB. -5 1 / s or slower k dissoc Approximately 2.8 × 10 -5 1 / s or slower k dissoc Approximately 2.9 × 10 -5 1 / s or slower k dissoc or approximately 3 x 10 to human or mouse 4-1BB. -5 1 / s or slower k dissoc This includes 4-1BB agonists that bind at

[0745]

[0790] In some embodiments, the compositions, processes and methods described have an IC50 of about 10 nM or less for human or mouse 4-1BB. 50 Binds to human or mouse 4-1BB with an IC of approximately 9 nM or less 50 Binds to human or mouse 4-1BB with an IC of approximately 8 nM or less 50 Binds to human or mouse 4-1BB with an IC of approximately 7 nM or less 50 Binds to human or mouse 4-1BB with an IC of about 6 nM or less 50 Binds to human or mouse 4-1BB with an IC of about 5 nM or less 50 Binds to human or mouse 4-1BB with an IC of approximately 4 nM or less 50 Binds to human or mouse 4-1BB with an IC of approximately 3 nM or less 50 Binds to human or mouse 4-1BB with an IC of approximately 2 nM or less50 or binds to human or mouse 4-1BB with an IC of about 1 nM or less 50 This includes 4-1BB agonists that bind at

[0746]

[0791] In a preferred embodiment, the 4-1BB agonist is utomilumab, also known as PF-05082566 or MOR-7480, or a fragment, derivative, variant, or biosimilar thereof. Utomilumab is available from Pfizer, Inc. Utomilumab is an immunoglobulin G2-λ, anti-[human (Homo sapiens) TNFRSF9 (tumor necrosis factor receptor (TNFR) superfamily member 9, 4-1BB, T cell antigen ILA, CD137)], human (Homo sapiens) (fully human) monoclonal antibody. The amino acid sequence of utomilumab is shown in Table 4. Utomilumab contains glycosylation sites at Asn59 and Asn292; positions 22-96 (V H -V L ), 143~199(C H 1-C L ), 256~316(C H 2) and 362-420(C H 3) heavy chain intrachain disulfide bridge; positions 22' to 87' (V H -V L ) and 136'~195'(C H 1-C L), interchain heavy-heavy disulfide bridges at IgG2A isoform positions 218-218, 219-219, 222-222, and 225-225, at IgG2A / B isoform positions 218-130, 219-219, 222-222, and 225-225, and at IgG2B isoform positions 219-130(2), 222-222, and 225-225; and interchain heavy-light disulfide bridges at IgG2A isoform position 130-213'(2), at IgG2A / B isoform positions 218-213' and 130-213', and at IgG2B isoform position 218-213'(2). The preparation and properties of utomilumab and its variants and fragments are described in U.S. Patent Nos. 8,821,867; 8,337,850; and 9,468,678, and International Publication No. WO 2012 / 032433 A1, the disclosures of each of which are incorporated herein by reference. Preclinical characterization of utomilumab is described in Fisher, et al., Cancer Immunolog. & Immunother. 2012, 61, 1721-33. Current clinical trials of utomilumab in various hematological and solid tumor indications include U.S. National Institutes of Health clinicaltrials.gov identification numbers NCT02444793, NCT01307267, NCT02315066, and NCT02554812.

[0747]

[0792] In some embodiments, the 4-1BB agonist comprises a heavy chain provided by SEQ ID NO: 11 and a light chain provided by SEQ ID NO: 12. In some embodiments, the 4-1BB agonist comprises heavy and light chains having the sequences set forth in SEQ ID NO: 11 and SEQ ID NO: 12, respectively, or antigen-binding fragments, Fab fragments, single-chain variable fragments (scFv), variants, or conjugates thereof. In some embodiments, the 4-1BB agonist comprises heavy and light chains, each at least 99% identical to the sequences set forth in SEQ ID NO: 11 and SEQ ID NO: 12, respectively. In some embodiments, the 4-1BB agonist comprises heavy and light chains, each at least 98% identical to the sequences set forth in SEQ ID NO: 11 and SEQ ID NO: 12, respectively. In some embodiments, the 4-1BB agonist comprises heavy and light chains, each at least 97% identical to the sequences set forth in SEQ ID NO: 11 and SEQ ID NO: 12, respectively. In some embodiments, the 4-1BB agonist comprises heavy and light chains, each at least 96% identical to the sequences set forth in SEQ ID NO: 11 and SEQ ID NO: 12, respectively. In certain embodiments, the 4-1BB agonist comprises a heavy chain and a light chain that are each at least 95% identical to the sequences set forth in SEQ ID NO:11 and SEQ ID NO:12, respectively.

[0748]

[0793] In some embodiments, the 4-1BB agonist comprises the heavy and light chain CDRs or variable regions (VRs) of utomilumab. In some embodiments, the 4-1BB agonist comprises the heavy chain variable region (VR) of utomilumab. H ) comprises the sequence set forth in SEQ ID NO: 13, and the 4-1BB agonist light chain variable region (V L ) comprises the sequence set forth in SEQ ID NO: 14, and includes conservative amino acid substitutions thereof. In one embodiment, the 4-1BB agonist comprises a V that is at least 99% identical to the sequences set forth in SEQ ID NO: 13 and SEQ ID NO: 14, respectively. H and V L In one embodiment, the 4-1BB agonist comprises a V region that is at least 98% identical to the sequences set forth in SEQ ID NO: 13 and SEQ ID NO: 14, respectively. H and V LIn one embodiment, the 4-1BB agonist comprises a V region that is at least 97% identical to the sequences set forth in SEQ ID NO: 13 and SEQ ID NO: 14, respectively. H and V L In one embodiment, the 4-1BB agonist comprises a V region that is at least 96% identical to the sequences set forth in SEQ ID NO: 13 and SEQ ID NO: 14, respectively. H and V L In one embodiment, the 4-1BB agonist comprises a V region that is at least 95% identical to the sequences set forth in SEQ ID NO: 13 and SEQ ID NO: 14, respectively. H and V L In one embodiment, the 4-1BB agonist comprises a V region that is at least 99% identical to the sequences set forth in SEQ ID NO: 13 and SEQ ID NO: 14, respectively. H and V L The present invention also includes scFv antibodies containing the domain.

[0749]

[0794] In one embodiment, the 4-1BB agonist comprises heavy chain CDR1, CDR2 and CDR3 domains having the sequences set forth in SEQ ID NO: 15, SEQ ID NO: 16, and SEQ ID NO: 17, respectively, and conservative amino acid substitutions thereof, and light chain CDR1, CDR2 and CDR3 domains having the sequences set forth in SEQ ID NO: 18, SEQ ID NO: 19, and SEQ ID NO: 20, respectively, and conservative amino acid substitutions thereof.

[0750]

[0795] In some embodiments, the 4-1BB agonist is a 4-1BB agonist biosimilar monoclonal antibody approved by a drug regulatory authority similar to utomilumab. In some embodiments, the biosimilar monoclonal antibody comprises a 4-1BB antibody comprising an amino acid sequence having at least 97% sequence identity, e.g., 97%, 98%, 99%, or 100% sequence identity, to the amino acid sequence of the originator pharmaceutical product or originator biological product, and comprising one or more post-translational modifications compared to the originator pharmaceutical product or originator biological product (wherein the originator pharmaceutical product or originator biological product is utomilumab). In some embodiments, the one or more post-translational modifications are selected from one or more of the following: glycosylation, oxidation, deamidation, and truncation. In some embodiments, the biosimilar is a 4-1BB agonist antibody that has been approved or for which approval is pending, and the 4-1BB agonist antibody is provided in a formulation that differs from that of the originator pharmaceutical formulation or originator biologic, where the originator pharmaceutical formulation or originator biologic is utomilumab. The 4-1BB agonist antibody may be approved by a drug regulatory authority, such as the U.S. FDA and / or the European Union EMA. In some embodiments, the biosimilar is provided as a composition further comprising one or more excipients, which are the same or different from the excipients included in the originator pharmaceutical formulation or originator biologic, where the originator pharmaceutical formulation or originator biologic is utomilumab. In some embodiments, the biosimilar is provided as a composition further comprising one or more excipients, which are the same or different from the excipients included in the originator pharmaceutical formulation or originator biologic, where the originator pharmaceutical formulation or originator biologic is utomilumab.

[0751] [Table 4]

[0752]

[0796] In a preferred embodiment, the 4-1BB agonist is the monoclonal antibody urelumab, also known as BMS-663513 and 20H4.9.h4a, or a fragment, derivative, variant, or biosimilar thereof. Urelumab is available from Bristol-Myers Squibb, Inc., and CreativeBiolabs, Inc. Urelumab is an immunoglobulin G4-κ, anti-[human (Homo sapiens) TNFRSF9 (tumor necrosis factor receptor superfamily member 9, 4-1BB, T cell antigen ILA, CD137)], human (Homo sapiens) (fully human) monoclonal antibody. The amino acid sequence of urelumab is shown in Table 5. Urelumab contains an N-glycosylation site at position 298 (and 298''); positions 22-95 (V H -V L ), 148-204(C H 1-C L ), 262~322(C H 2) and 368-426(C H 3) (and at positions 22' to 95', 148' to 204', 262' to 322', and 368' to 426') heavy chain intrachain disulfide bridges; positions 23' to 88' (V H -V L ) and 136'~196'(C H 1-C L), light chain intrachain disulfide bridges at positions 23'''-88''' and 136'''-196'''; interchain heavy chain-heavy chain disulfide bridges at positions 227-227'' and 230-230''; and interchain heavy chain-light chain disulfide bridges at positions 135-216' and 135''-216'''. The preparation and properties of urelumab and variants and fragments thereof are described in U.S. Pat. Nos. 7,288,638 and 8,962,804, the disclosures of which are incorporated herein by reference. Preclinical and clinical characteristics of urelumab are described in Segal, et al., Clin. Cancer Res. 2016, available at http: / / dx.doi.org / 10.1158 / 1078-0432.CCR-16-1272. Current clinical trials of urelumab in various hematological and solid tumor indications include US National Institutes of Health clinicaltrials.gov identifiers NCT01775631, NCT02110082, NCT02253992, and NCT01471210.

[0753]

[0797] In some embodiments, the 4-1BB agonist comprises a heavy chain provided by SEQ ID NO:21 and a light chain provided by SEQ ID NO:22. In some embodiments, the 4-1BB agonist comprises heavy and light chains having the sequences set forth in SEQ ID NO:21 and SEQ ID NO:22, respectively, or antigen-binding fragments, Fab fragments, single-chain variable fragments (scFv), variants, or conjugates thereof. In some embodiments, the 4-1BB agonist comprises heavy and light chains, each at least 99% identical to the sequences set forth in SEQ ID NO:21 and SEQ ID NO:22, respectively. In some embodiments, the 4-1BB agonist comprises heavy and light chains, each at least 98% identical to the sequences set forth in SEQ ID NO:21 and SEQ ID NO:22, respectively. In some embodiments, the 4-1BB agonist comprises heavy and light chains, each at least 97% identical to the sequences set forth in SEQ ID NO:21 and SEQ ID NO:22, respectively. In some embodiments, the 4-1BB agonist comprises heavy and light chains, each at least 96% identical to the sequences set forth in SEQ ID NO:21 and SEQ ID NO:22, respectively. In certain embodiments, the 4-1BB agonist comprises a heavy chain and a light chain that are each at least 95% identical to the sequences set forth in SEQ ID NO:21 and SEQ ID NO:22, respectively.

[0754]

[0798] In some embodiments, the 4-1BB agonist comprises the heavy and light chain CDRs or variable regions (VRs) of urelumab. In some embodiments, the 4-1BB agonist comprises the heavy chain variable region (VR) of urelumab. H ) comprises the sequence set forth in SEQ ID NO: 23, and the 4-1BB agonist light chain variable region (V L ) comprises the sequence set forth in SEQ ID NO: 24, and includes conservative amino acid substitutions thereof. In one embodiment, the 4-1BB agonist comprises a V that is at least 99% identical to the sequences set forth in SEQ ID NO: 23 and SEQ ID NO: 24, respectively. H and V L In one embodiment, the 4-1BB agonist comprises a V region that is at least 98% identical to the sequences set forth in SEQ ID NO: 23 and SEQ ID NO: 24, respectively. H and V LIn one embodiment, the 4-1BB agonist comprises a V region that is at least 97% identical to the sequences set forth in SEQ ID NO:23 and SEQ ID NO:24, respectively. H and V L In one embodiment, the 4-1BB agonist comprises a V region that is at least 96% identical to the sequences set forth in SEQ ID NO: 23 and SEQ ID NO: 24, respectively. H and V L In one embodiment, the 4-1BB agonist comprises a V region that is at least 95% identical to the sequences set forth in SEQ ID NO: 23 and SEQ ID NO: 24, respectively. H and V L In one embodiment, the 4-1BB agonist comprises a V region that is at least 99% identical to the sequences set forth in SEQ ID NO:23 and SEQ ID NO:24, respectively. H and V L The present invention also includes scFv antibodies containing the domain.

[0755]

[0799] In one embodiment, the 4-1BB agonist comprises heavy chain CDR1, CDR2 and CDR3 domains having the sequences set forth in SEQ ID NO: 25, SEQ ID NO: 26, and SEQ ID NO: 27, respectively, and conservative amino acid substitutions thereof, and light chain CDR1, CDR2 and CDR3 domains having the sequences set forth in SEQ ID NO: 28, SEQ ID NO: 29, and SEQ ID NO: 30, respectively, and conservative amino acid substitutions thereof.

[0756]

[0800] In some embodiments, the 4-1BB agonist is a 4-1BB agonist biosimilar monoclonal antibody approved by a drug regulatory agency similar to urelumab. In some embodiments, the biosimilar monoclonal antibody comprises a 4-1BB antibody comprising an amino acid sequence having at least 97% sequence identity, e.g., 97%, 98%, 99%, or 100% sequence identity, to the amino acid sequence of the originator pharmaceutical product or originator biological product, and comprising one or more post-translational modifications compared to the originator pharmaceutical product or originator biological product (wherein the originator pharmaceutical product or originator biological product is urelumab). In some embodiments, the one or more post-translational modifications are selected from one or more of the following: glycosylation, oxidation, deamidation, and truncation. In some embodiments, the biosimilar is a 4-1BB agonist antibody that has been approved or for which approval is pending, and the 4-1BB agonist antibody is provided in a formulation that differs from that of the originator pharmaceutical formulation or originator biologic, where the originator pharmaceutical formulation or originator biologic is urelumab. The 4-1BB agonist antibody may be approved by a drug regulatory authority, such as the U.S. FDA and / or the European Union EMA. In some embodiments, the biosimilar is provided as a composition further comprising one or more excipients, which are the same or different from the excipients in the originator pharmaceutical formulation or originator biologic, where the originator pharmaceutical formulation or originator biologic is urelumab. In some embodiments, the biosimilar is provided as a composition further comprising one or more excipients, which are the same or different from the excipients in the originator pharmaceutical formulation or originator biologic, where the originator pharmaceutical formulation or originator biologic is urelumab.

[0757] [Table 5]

[0758]

[0801] In certain embodiments, the 4-1BB agonist is selected from the group consisting of 1D8, 3Elor, 4B4 (BioLegend 309809), H4-1BB-M127 (BD Pharmingen 552532), BBK2 (Thermo Fisher MS621PABX), 145501 (Leinco Technologies B591), the antibody produced by the cell line deposited under ATCC number HB-11248, and disclosed in U.S. Pat. No. 6,974,863, 5F4 (BioLegend 31 1503), C65-485 (BD Pharmingen No. 559446), antibodies disclosed in U.S. Patent Application Publication No. 2005 / 0095244, antibodies disclosed in U.S. Patent No. 7,288,638 (such as 20H4.9-IgG1 (BMS-663031)), antibodies disclosed in U.S. Patent No. 6,887,673 (such as 4E9 or BMS-554271), antibodies disclosed in U.S. Patent No. 7,214,493, antibodies disclosed in U.S. Patent No. 6,303,121, antibodies disclosed in U.S. Patent No. 6,569,997, antibodies disclosed in U.S. Patent No. 6,905,685 (such as 4E9 or BMS-554271), antibodies disclosed in U.S. Patent No. 6,362,325 (1D8 or BMS-469492; 3H3 or BMS-4694 No. 6,210,669 (such as 1D8, 3B8, or 3E1), an antibody disclosed in U.S. Pat. No. 6,974,863 (such as 53A2); an antibody disclosed in U.S. Pat. No. 6,210,669 (such as 1D8, 3B8, or 3E1), an antibody described in U.S. Pat. No. 5,928,893, an antibody disclosed in U.S. Pat. No. 6,303,121, an antibody disclosed in U.S. Pat. No. 6,569,997, an antibody disclosed in WO 2012 / 177788, WO 2015 / 119923, and WO 2010 / 042433, and fragments, derivatives, conjugates, variants, or biosimilars thereof (the disclosures of each of the foregoing patents or patent application publications are incorporated herein by reference).

[0759]

[0802] In certain embodiments, the 4-1BB agonist is a compound disclosed in WO 2008 / 025516 A1, WO 2009 / 007120 A1, WO 2010 / 003766 A1, WO 2010 / 010051 A1, and WO 2010 / 078966 A1; U.S. Patent Application Publication Nos. 2011 / 0027218 A1, 2015 / 0126709 A1, 2011 / 0111494 A1, 2015 / 0110734 A1, and 2015 / 0126710 A1; and 4-1BB agonist fusion proteins described in U.S. Patent Nos. 9,359,420, 9,340,599, 8,921,519, and 8,450,460, the disclosures of which are incorporated herein by reference.

[0760]

[0803] In one embodiment, the 4-1BB ...

Claims

1. 1. Use of a tumor infiltrating lymphocyte (TIL) population for the manufacture of a pharmaceutical composition comprising a therapeutically effective amount of the TIL population for treating cancer, said treatment comprising: (a) resecting a tumor from a patient; (b) obtaining a first population of TILs from the tumor; (c) obtaining a second TIL population by initial expansion of the first TIL population in a first cell culture medium, wherein the second TIL population is at least 5-fold more numerous than the first TIL population, the first cell culture medium comprises IL-2, and the initial expansion is for a period of 21 days or less; (d) obtaining a third TIL population by rapid expansion of the second TIL population in a second cell culture medium, wherein the third TIL population is at least 50-fold more numerous than the second TIL population 7 days after initiation of the rapid expansion; the second cell culture medium comprising IL-2, OKT-3, peripheral blood mononuclear cells (PBMCs), and a first tumor necrosis factor receptor superfamily (TNFRSF) agonist, the first TNFRSF agonist comprising an OX40 agonist antibody or a soluble OX40L fusion protein, and the rapid expansion is performed for a period of 14 days or less; (e) the third population of tumor-infiltrating lymphocytes (TILs) as a TIL population for the preparation of a pharmaceutical composition, wherein the reference CD8 ＋ TIL vs. CD4 ＋ Increased CD8 compared to TIL ratio ＋ TIL vs. CD4 ＋ recovering said third population of TILs exhibiting a TIL ratio; and (f) administering to said patient a therapeutically effective amount of said third population of TILs as a therapeutic population of TILs. Including, use.

2. 2. The use of claim 1, wherein the second cell culture medium further comprises a second TNFRSF agonist selected from the group consisting of a 4-1BB agonist, an OX40 agonist, a CD27 agonist, a GITR agonist, an HVEM agonist, a CD95 agonist, and combinations thereof.

3. 2. The use of claim 1, wherein the first cell culture medium further comprises the first TNFRSF agonist and a second TNFRSF agonist selected from the group consisting of a 4-1BB agonist, an OX40 agonist, a CD27 agonist, a GITR agonist, an HVEM agonist, a CD95 agonist, and combinations thereof.

4. 4. The use of claim 2 or 3, wherein the second TNFRSF agonist is a 4-1BB agonist, and the 4-1BB agonist is selected from the group consisting of urelumab, utomilumab, EU-101, and combinations thereof.

5. 5. The use of any one of claims 2 to 4, wherein the second TNFRSF agonist is an OX40 agonist, and the OX40 agonist is selected from the group consisting of taborixizumab, GSK3174998, MEDI6469, MEDI6383, MOXR0916, PF-04518600, Creative Biolabs MOM-18455, OX40 agonist fusion proteins, and combinations thereof.

6. The use of any one of claims 2 to 5, wherein the second TNFRSF agonist is a CD27 agonist, and the CD27 agonist is selected from the group consisting of varlilumab and a CD27 agonist fusion protein.

7. 7. The use of any one of claims 2 to 6, wherein the second TNFRSF agonist is a GITR agonist, and the GITR agonist is selected from the group consisting of TRX518, 6C8, 36E5, 3D6, 61G6, 6H6, 61F6, 1D8, 17F10, 35D8, 49A1, 9E5, 31H6, 2155, 698, 706, 827, 1649, 1718, 1D7, 33C9, 33F6, 34G4, 35B10, 41E11, 41G5, 42A11, 44C1, 45A8, 46E11, 48H12, 48H7, 49D9, 49E2, 48A9, 5H7, 7A10, 9H6, and combinations thereof.

8. 8. The use of any one of claims 1 to 7, wherein said treatment further comprises treating said patient with said first TNFRSF agonist starting the day after administration of said third population of TILs to said patient, wherein said first TNFRSF agonist is administered intravenously at a dose of 0.1 mg / kg to 50 mg / kg every four weeks for up to eight cycles.

9. The use according to any one of claims 1 to 8, wherein the OX40 agonist antibody is selected from the group consisting of urelumab, utomilumab, EU-101, taborixizumab, Creative Biolabs MOM-18455, and combinations thereof.

10. 10. The use of any one of claims 1, 2, or 4-9, wherein the first cell culture medium additionally comprises a third TNFRSF agonist selected from the group consisting of a 4-1BB agonist, an OX40 agonist, a CD27 agonist, a GITR agonist, an HVEM agonist, a CD95 agonist, and combinations thereof.

11. 11. The use according to any one of claims 1 to 10, wherein the first TNFRSF agonist is added to the first cell culture medium at intervals selected from the group consisting of every day, every 2 days, every 3 days, every 4 days, every 5 days, every 6 days, every 7 days, and every 2 weeks during the initial expansion culture.

12. 12. The use according to any one of claims 1 to 11, wherein the first TNFRSF agonist is added to the second cell culture medium at intervals selected from the group consisting of every day, every 2 days, every 3 days, every 4 days, every 5 days, every 6 days, every 7 days, and every 2 weeks during the rapid expansion culture.

13. The use according to any one of claims 1 to 12, wherein the first TNFRSF agonist is added at a concentration sufficient to achieve a concentration of 0.1 μg / mL to 100 μg / mL in the cell culture medium.

14. 14. The use according to any one of claims 1 to 13, wherein IL-2 is present in the first cell culture medium at an initial concentration of about 10 to about 6000 IU / mL.

15. 15. The use according to any one of claims 1 to 14, wherein IL-2 is present in the second cell culture medium at an initial concentration of about 10 to about 6000 IU / mL.

16. The use according to any one of claims 1 to 15, wherein the OKT-3 antibody is present in the second cell culture medium at an initial concentration of about 10 ng / mL to about 60 ng / mL.

17. The use according to any one of claims 1 to 16, wherein the initial expansion culture and the rapid expansion culture are carried out using a gas-permeable container.

18. The use of any one of claims 1 to 17, further comprising treating said patient with a non-myeloablative lymphodepletion regimen prior to administering said third population of TILs to said patient.

19. 19. The use according to any one of claims 1 to 18, wherein the cancer is selected from the group consisting of melanoma, cutaneous melanoma, double resistant melanoma, uveal (ocular) melanoma, ovarian cancer, platinum resistant ovarian cancer, cervical cancer, lung cancer, non-small cell lung cancer (NSCLC), bladder cancer, breast cancer, triple negative breast cancer, head and neck cancer, head and neck squamous cell carcinoma, renal cell carcinoma, acute myeloid leukemia, colorectal cancer, pancreatic ductal adenocarcinoma, glioblastoma, cholangiocarcinoma, osteosarcoma and sarcoma.

20. The use of any one of claims 1 to 19, wherein the patient is treated with a PD-1 inhibitor or a PD-L1 inhibitor after administering the third population of TILs to the patient.

21. 21. The use of claim 20, wherein the PD-1 inhibitor or PD-L1 inhibitor is selected from the group consisting of nivolumab, pembrolizumab, durvalumab, atezolizumab, avelumab, and combinations thereof.

22. The use according to any one of claims 1 to 21, wherein the initial expansion culture is carried out for a period of 11 days or less.

23. The use according to any one of claims 1 to 22, wherein the rapid expansion culture is carried out for a period of 11 days or less.

24. 1. A pharmaceutical composition comprising a tumor infiltrating lymphocyte (TIL) population for use in the treatment of cancer, said tumor infiltrating lymphocyte (TIL) population comprising: (b) obtaining a first population of TILs; (c) obtaining a second TIL population by initial expansion of the first TIL population in a first cell culture medium, wherein the second TIL population is at least 5-fold more numerous than the first TIL population, the first cell culture medium comprises IL-2, and the initial expansion is for a period of 21 days or less; (d) obtaining a third TIL population by rapid expansion of the second TIL population in a second cell culture medium, wherein the third TIL population is at least 50-fold more numerous than the second TIL population 7 days after initiation of the rapid expansion; the second cell culture medium comprises IL-2, OKT-3, peripheral blood mononuclear cells (PBMCs), and a first tumor necrosis factor receptor superfamily (TNFRSF) agonist, the first TNFRSF agonist comprising an OX40 agonist antibody or a soluble OX40L fusion protein, and the rapid expansion is performed for a period of 14 days or less; and (e) Reference CD8 in the second TIL population ＋ TIL vs. CD4 ＋ Increased CD8 compared to TIL ratio ＋ TIL vs. CD4 ＋ recovering said third population of TILs exhibiting a TIL ratio wherein said pharmaceutical composition comprises said third population of TILs.

25. 25. The pharmaceutical composition of claim 24, wherein the first cell culture medium further comprises the first TNFRSF agonist.

26. 26. The pharmaceutical composition of claim 24 or 25, wherein the second cell culture medium further comprises a second TNFRSF agonist selected from the group consisting of a 4-1BB agonist, an OX40 agonist, a CD27 agonist, a GITR agonist, an HVEM agonist, a CD95 agonist, and combinations thereof.

27. 27. The pharmaceutical composition of any one of claims 24 to 26, wherein the first TNFRSF agonist is added to the first cell culture medium at intervals selected from the group consisting of every day, every second day, every third day, every fourth day, every fifth day, every sixth day, every seventh day, and every two weeks during the initial expansion culture.

28. 28. The pharmaceutical composition of any one of claims 24-27, wherein the first TNFRSF agonist is added to the second cell culture medium at intervals selected from the group consisting of every day, every 2 days, every 3 days, every 4 days, every 5 days, every 6 days, every 7 days, and every 2 weeks during the rapid expansion culture.

29. 29. The pharmaceutical composition of any one of claims 24 to 28, wherein the first TNFRSF agonist is added at a concentration sufficient to achieve a concentration of 0.1 μg / mL to 100 μg / mL in the cell culture medium.

30. 30. The pharmaceutical composition of any one of claims 24 to 29, wherein IL-2 is present in the first cell culture medium at an initial concentration of about 10 to about 6000 IU / mL.

31. 31. The pharmaceutical composition of any one of claims 24 to 30, wherein the OKT-3 antibody is present in the second cell culture medium at an initial concentration of about 10 ng / mL to about 60 ng / mL.

32. The pharmaceutical composition according to any one of claims 24 to 31, wherein the initial expansion culture and the rapid expansion culture are carried out using a gas-permeable container.

33. A pharmaceutical composition for use in the treatment of cancer according to any one of claims 24 to 32 for use in combination with a non-myeloablative lymphodepletion regimen.

34. 33. The pharmaceutical composition for use in treating cancer according to any one of claims 24 to 32, wherein the composition is for use in combination with a myeloablative lymphodepletion regimen prior to administration of the third population of TILs to the patient.

35. The pharmaceutical composition for use in treating cancer according to any one of claims 24 to 34, which is for use in combination with a PD-1 inhibitor or a PD-L1 inhibitor.

36. 35. The pharmaceutical composition for use in treating cancer according to any one of claims 24 to 34, wherein the pharmaceutical composition is for use in combination with a PD-1 inhibitor or a PD-L1 inhibitor, wherein the PD-1 inhibitor or PD-L1 inhibitor is selected from the group consisting of nivolumab, pembrolizumab, durvalumab, atezolizumab, avelumab, and combinations thereof.

37. 37. The pharmaceutical composition for use in treating cancer of claim 36, wherein the PD-1 inhibitor or PD-L1 inhibitor is administered before or after resection of the tumor from the patient.

38. 33. The pharmaceutical composition for use in treating cancer of any one of claims 24 to 32, wherein the pharmaceutical composition is for use in combination with a PD-1 inhibitor or a PD-L1 inhibitor, wherein the PD-1 inhibitor or PD-L1 inhibitor is administered after administration of the third population of TILs to the patient.

39. 39. The pharmaceutical composition for use in the treatment of cancer according to any one of claims 24 to 38, wherein the cancer is selected from the group consisting of melanoma, cutaneous melanoma, double resistant melanoma, uveal (ocular) melanoma, ovarian cancer, platinum resistant ovarian cancer, cervical cancer, lung cancer, non-small cell lung cancer (NSCLC), bladder cancer, breast cancer, triple negative breast cancer, head and neck cancer, head and neck squamous cell carcinoma, renal cell carcinoma, acute myeloid leukemia, colorectal cancer, pancreatic ductal adenocarcinoma, glioblastoma, cholangiocarcinoma, osteosarcoma and sarcoma.

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