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WO2012050374A2 - Immunothérapie contre les tumeurs solides - Google Patents

Immunothérapie contre les tumeurs solides Download PDF

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Publication number
WO2012050374A2
WO2012050374A2 PCT/KR2011/007621 KR2011007621W WO2012050374A2 WO 2012050374 A2 WO2012050374 A2 WO 2012050374A2 KR 2011007621 W KR2011007621 W KR 2011007621W WO 2012050374 A2 WO2012050374 A2 WO 2012050374A2
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cells
chimeric antigen
receptor
antigen receptor
chimeric
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Hyun Jin Jung
Jong Sung Choi
Jaeseung Lim
Mi-Jung Yun
Won-Ho Choi
Dong Young Kim
Ji Mi Ahn
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INNOCELL Inc
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INNOCELL Inc
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    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
    • C07K14/705—Receptors; Cell surface antigens; Cell surface determinants
    • C07K14/70503—Immunoglobulin superfamily
    • C07K14/70521—CD28, CD152
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K40/00—Cellular immunotherapy
    • A61K40/10—Cellular immunotherapy characterised by the cell type used
    • A61K40/11—T-cells, e.g. tumour infiltrating lymphocytes [TIL] or regulatory T [Treg] cells; Lymphokine-activated killer [LAK] cells
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K40/00—Cellular immunotherapy
    • A61K40/30—Cellular immunotherapy characterised by the recombinant expression of specific molecules in the cells of the immune system
    • A61K40/31—Chimeric antigen receptors [CAR]
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K40/00—Cellular immunotherapy
    • A61K40/40—Cellular immunotherapy characterised by antigens that are targeted or presented by cells of the immune system
    • A61K40/41—Vertebrate antigens
    • A61K40/42—Cancer antigens
    • A61K40/4202—Receptors, cell surface antigens or cell surface determinants
    • A61K40/4203—Receptors for growth factors
    • A61K40/4205—Her-2/neu/ErbB2, Her-3/ErbB3 or Her 4/ ErbB4
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
    • C07K14/705—Receptors; Cell surface antigens; Cell surface determinants
    • C07K14/70503—Immunoglobulin superfamily
    • C07K14/7051—T-cell receptor (TcR)-CD3 complex
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
    • C07K14/705—Receptors; Cell surface antigens; Cell surface determinants
    • C07K14/70503—Immunoglobulin superfamily
    • C07K14/70517—CD8
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
    • C07K14/705—Receptors; Cell surface antigens; Cell surface determinants
    • C07K14/70596—Molecules with a "CD"-designation not provided for elsewhere
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K2239/00—Indexing codes associated with cellular immunotherapy of group A61K40/00
    • A61K2239/31—Indexing codes associated with cellular immunotherapy of group A61K40/00 characterized by the route of administration
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K2319/00—Fusion polypeptide
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K2319/00—Fusion polypeptide
    • C07K2319/01—Fusion polypeptide containing a localisation/targetting motif
    • C07K2319/03—Fusion polypeptide containing a localisation/targetting motif containing a transmembrane segment

Definitions

  • T cells expressing exogenously introduced chimeric antigen receptor (CAR) are provided herein.
  • CAR chimeric antigen receptor
  • immunotherapy may be used in conjunction with conventional therapies to benefit advanced cancer patients by booting anti-tumor immune response, eradicating residual micrometastatic, and preventing relapse.
  • some have reported limitations of active immunotherapy using dendritic cell vaccination presumably due to the difficulties associated with effectively expanding tumor specific T cells.
  • adoptive T cell use tumor specific T cells manufactured ex vivo. Expanded T lymphocytes recognize antigen as peptide bound to MHC molecules on the surface of tumor cells, proliferate, activate and kill tumor cells by cytotoxic activity. Nonetheless, several obstacles remain with regard to induction of effective cellular immunity due to tumor escape mechanism, including antigen loss and MHC downregulation.
  • T cell based immunotherapy therefore is constrained not only by the MHC restriction of antigen presentation, but also by the requirement that an antigen presenting cell provide necessary costimulatory signals to the T cells.
  • T cells expressing exogenous chimeric antigen receptors (CARs) have been developed to redirect T cell antigen specificity to tumor associated antigens (TAAs) expressed on the tumor cell surface.
  • CARs Chimeric Antigen Receptors
  • Artificial genetically modified receptors typically consist of an extracellular domain, which is derived from immunoglobulin variable region (single chain fragment variable; scFv) or receptor ligand such as IL-13Ra2; hinge region derived, for example, from CD8a or immunoglobulin sequence; and cytoplasmic domain, which is derived from the CD3 ⁇ chain or the Fc receptor ⁇ (FcRy) chain.
  • scFv single chain fragment variable
  • IL-13Ra2 single chain fragment variable
  • hinge region derived, for example, from CD8a or immunoglobulin sequence
  • cytoplasmic domain which is derived from the CD3 ⁇ chain or the Fc receptor ⁇ (FcRy) chain.
  • First generation CARs which have 8 ⁇ 3 ⁇ structures, have been reported to be effective in redirecting cytotoxicity of cytotoxic T lymphocytes (CTLs) and inducing T cell activation.
  • CTLs cytotoxic T lymphocytes
  • Turatti et al. J. Gene Med., 7: 158-170 (2005); Gade et al, Cancer Res. , 65(19): 9080-9088 (2005); Cheadle et al, British Journal of Haematology, 142: 65-68 (2008); and Beecham et al, Journal of Immunotherapy, 23(3): 332-343 (2000).
  • rapid T cell death following expansion and insufficient in vivo persistence were observed in T cells expressing the first generation CARs.
  • next generation CARs are designed to contain costimulatory molecules such as CD28 and 4- IBB in their signaling domain.
  • costimulatory molecules such as CD28 and 4- IBB in their signaling domain.
  • CARs chimeric antigen receptors
  • the CARs provided herein are useful for the treatment and prevention of solid tumors.
  • methods of making a cell expressing an exogenous CAR comprising introducing a gene encoding the CAR into the cell. Methods of introducing the CARs to the cells are also provided in detail herein elsewhere.
  • Also provided are methods of treating, managing, and/or preventing relapse of cancer comprising administering cells that express exogenous CARs.
  • the CAR contains a tumor antigen provided herein elsewhere.
  • compositions, single unit dosage forms, dosing regimens and kits which comprise a CAR, a gene that encodes a CAR, or a cell with an expressed CAR on the surface.
  • FIG. 1 illustrates the results of FACS analysis for purified ErbB-2 (Her- 2/neu) antibodies 6D and 7G in SK-BR3 and MDA-468 cell lines.
  • FIG. 2 is a schematic illustration of pCI-6D vector encoding ErbB2 andibody 6D based chimeric antigen receptor (CAR) against ErbB2.
  • CAR chimeric antigen receptor
  • FIG. 3 illustrates EGFP expression on activated T cells following delivery of 6D-CAR mRNA by elecroporation.
  • FIG. 4 illustrates Her-2/neu expression on activated T cells following delivery of 6D-CAR mRNA by electoporation.
  • CARs chimeric antigen receptors
  • TAAs tumor associated antigens
  • scFv scFv linked to signaling domain
  • the terms “treat,” “treating” and “treatment” refer to the eradication or amelioration of a disease or disorder, or of one or more symptoms associated with the disease or disorder. In certain embodiments, the terms refer to minimizing the spread or worsening of the disease or disorder resulting from the administration of one or more therapies to a subject with such a disease or disorder. In some embodiments, the terms refer to the administration of a therapy provided herein, with or without other additional therapy or agent, after the onset. of symptoms of the particular disease.
  • the terms “prevent,” “preventing” and “prevention” refer to the prevention of the onset, recurrence or spread of a disease or disorder, or of one or more symptoms thereof.
  • the terms refer to the treatment with or administration of a therapy provided herein, with or without other additional therapy or agent, prior to the onset or recurrence of symptoms, particularly to patients at risk of disease or disorders provided herein.
  • the terms encompass the inhibition or reduction of a symptom of the particular disease.
  • Patients with familial history of a disease in particular are candidates for preventive regimens in certain embodiments.
  • patients who have a history of recurring symptoms are also potential candidates for the prevention.
  • the term “prevention” may be interchangeably used with the term “prophylactic treatment.”
  • the terms “manage,” “managing” and “management” refer to preventing or slowing the progression, spread or worsening of a disease or disorder, or of one or more symptoms thereof. Often, the beneficial effects that a subject derives from a therapy do not result in a cure of the disease or disorder. In this regard, the term “managing” encompasses treating a patient who had suffered from the particular disease in an attempt to prevent or minimize the recurrence of the disease.
  • Chimeric receptors generally comprise an exogenous antibody to specific antigen on the target cell surface and an activation/stimulation domain.
  • chimeric antigen receptor refers to an extracellular antigen- binding domain that is fused to an intracellular signaling domain capable of activating T cells.
  • the vector is a plasmid vector such as, but not limited to, pIRES2-EGFP and pcDNA 3.1 -GFP.
  • the antigen-binding domain is specific to a tumor associated antigen (TAA).
  • TAAs are differentiation antigens that are expressed at low levels in normal tissues, but are preferentially up- regulated in malignancy. Examples of TAAs from which multiple MHC class I- restricted epitopes have been identified include, but are not limited to: MAGE-1, which is described, for example, in Traversari et al, J. Exp. Med. , 176: 1453-1457 (1992); NY- ESO-1, which is described, for example, in Jager et al, J. Exp. Med.
  • the TAA is Her-2/neu.
  • Her-2/neu(c-ErbB2) is oncogene encoding an 185 kDa transmembrane receptor, which is a member of the human epidermal growth factor receptor 2.
  • Her-2/neu is expressed at a low level in normal tissues, but overexpressed in breast, ovarian, lung, gastric, oral, and prostate cancers and brain tumor.
  • Clinical data using Trastuzumab (Herceptin) approved by the FDA for the treatment of metastatic breast cancer have demonstrated improved outcome, but it is associated with increased incidence of cardiac dysfunction.
  • using Her-2/neu to construct a CAR to be expressed on the surface of immune cells reduces or minimizes the cardiac side effects associated with the direct use of the antibody, and thus, provides an advantageous treatment for cancers or tumors associated with Her-2/neu overexpression.
  • the extracellular antigen-binding domain is derived from a murine antibody. In another embodiment, the extracellular antigen-binding domain is derived from a humanized monoclonal antibody. In a specific embodiment, the antigen-binding domain is a scFv originated from a humanized anti- Her-2/neu antibody (e.g. , Herceptin).
  • molecules that exert costimulatory effect are included in the CARs.
  • examples of such molecules include, but are not limited to, CD28, CD 137 (4- IBB), CD3C,, and a combination thereof.
  • the CAR contains both CD28 and CD 137 (4- IBB).
  • the order of the costimulatory molecules is crucial to the functionality of the CARs, in particular, the extent of the background toxicities against antigen negative cells generated by a specific CAR when expressed in the cells.
  • the order of the costimulatory molecules in a CAR is CD28-4-lBB-CD3 ⁇ .
  • the CARs provided herein also contain hinge and transmembrane domain (TM) regions.
  • TM transmembrane domain
  • the hinge and TM regions of CARs provided herein consist of those originated from CD8a receptors.
  • provided herein is a cell that expresses an exogenous CAR on its surface.
  • a method of making a cell that expresses an exogenous CAR comprising introducing a gene encoding the CAR into the cell.
  • the effector cells may be any cells of immune systems.
  • examples of cells include, but are not limited to, PBMC, PBL, T cells (e.g. , cytotoxic T cells), tumor infiltrating lymphocytes (TILs), lymphokine activated killer (LAK) cells, cytokine induced killer (CIK) cells and Natural Killer (NK) cells, and mixtures thereof.
  • T cells e.g. , cytotoxic T cells
  • TILs tumor infiltrating lymphocytes
  • LAK lymphokine activated killer
  • CIK cytokine induced killer
  • NK Natural Killer
  • TILs refer to lymphocytes specific to tumor cells.
  • LAK cells refer to tumor killing lymphocytes isolated from peripheral blood and are cultured in the presence of IL-2.
  • cytotoxic T cells refer those T cells that are able to recognize and kill tumor cells.
  • CIK cells refer to those cells that maintain tumor killing efficacy regardless of the extent of expression of the MHC class molecules expressed in the tumor cells.
  • the cells are T cells. In another embodiment, the cells are CIK cells. In another embodiment, the cells are NK cells. In another embodiment, the cells are LAK cells. In certain embodiments, the cells are killer cells, which are a mixture of cytotoxic T cells (CD3 + /CD8 + ) and CIK cells (CD3 + /CD56 + ).
  • the cells are unstimulated, i.e. , in resting state.
  • the cells are stimulated, /. e. , activated.
  • the cells are activated killer cells.
  • the activated killer cells are lymphocytes, which are isolated from peripheral blood and cultured.
  • the activated killer cells are a mixture of cytotoxic T cells (CD3 + /CD8 + ) and CIK cells (CD3 + /CD56 + ).
  • the cells are NK cells.
  • NK cells are a type of cytotoxic lymphocyte, which may be activated in response to various cytokines such as interferons or macrophage-derived cytokines.
  • cytokines such as interferons or macrophage-derived cytokines.
  • NK cells are CD3 " , but express CD56 on their surface.
  • NK cells do not require antigen activation, and NK cells are not MHC restricted. 5.2.2 Delivery of CAR Genes
  • Genes containing CAR construct should be delivered to cells. In this regard, various methods such as virus-based delivery and non-viral gene delivery may be employed.
  • genes containing CAR construct provided herein are delivered using virus-based methods commonly known in the art.
  • Exemplary virus- based deliveries employ viruses such as gamma retrovirus and self inactivated lenti virus.
  • viruses such as gamma retrovirus and self inactivated lenti virus.
  • Other examples include those methods described, for example, in: Gade et al, Cancer Res., 65(19): 9080-9088 (2005); Maher et al, Nature Biotechnology, 20: 70-75 (2002); Zhao et al., The Journal of Immunology, ⁇ 83: 5563-5574 (2009); Ahmed et al., Clin. Cancer Res. , 16(2): 474-485 (2010); Ahmed et al., Cancer Res. , 67(12): 5957-5964
  • non viral methods are employed to deliver genes containing CAR construct.
  • the non viral method is electroporation.
  • Electroporation is a method that transduce the genes through momentary alteration of cell membranes by applying a high-voltage to the cells. Without being limited by a particular theory, electroporation are advantageous in certain embodiments since it is possible to deliver genes into the nucleus in. primary cells whose proliferations are somewhat inactive. Further without being limited by a particular, theory, electroporation provides a high efficiency and ease of manipulation. Examples of electroporetic methods include, but are not limited to, those described in: Kahlon et al, Cancer Res. , 64: 9160-9166 (2004); Yoon et al, Cancer Gene Therapy (2008): 1-9; Park et al, Molecular Therapy, 15(4): 825-833 (2007); Till et al, Blood, 1 12: 2261-2271
  • the non viral method is a delivery using nanotechnology.
  • the method comprises delivery using bio- nanomaterials.
  • the bio-nanomaterials are those based on liposomes or dendrimers.
  • the method comprises delivery using inorganic nanoparticles.
  • the inorganic nanoparticles are carbon or metals. Without being limited to a particular theory, the inorganic nanoparticles are . advantageous due to their hollow and porous structures, and because high amount of drug loading is possible due to the ease of surface functionalization.
  • the inorganic nanomaterial is silica nanotube described, for example, in Bai et al, Nanomedicine, 3(2): 163-174 (2008), which is incorporated herein in its entirety by reference.
  • the walls of the nanotubes may be coated with fluorescent materials for the gene delivery. Without being limited by a particular theory, gene delivery using the fluorescent coated nanotubes were reported to provide detectable fluorescence when entered into the cells and cause reduced cytotoxicity. (See, e.g., Gao et al, ChemBioChem, 7: 239-242 (2006), incorporated herein by reference).
  • genes to be delivered gene expression efficiency may depend on the type of genes that are being delivered.
  • DNAs may be advantageous in that they provide a prolonged-expression. For example, it was reported that genetically modified T cells with transfection of CD19-CAR DNA by electroporation maintained over 50% of transgene expression after 10 weeks. (Cooper et al, Cryotherapy, 8(2): 105-1 17 (2006));
  • mRNAs may be advantageous in some cases because transcription is bypassed and target gene can be immediately translated into protein and detected easily protein expression. Thus, mRNAs are advantageous in that gene expression may be ascertained in a short period of time.
  • the gene to be delivered is in the form of a DNA. In another embodiment, the gene to be delivered is in the form of an mRNA.
  • a method of treating, managing, and/or preventing relapse of cancer comprising administering a therapeutically or prophylactically effective amount of a cell that expresses an exogenous CAR.
  • the CAR contains a tumor antigen binding domain, provided herein elsewhere.
  • the cell is an immune cell, provided herein elsewhere.
  • a “therapeutically effective amount” is an amount sufficient to provide a therapeutic benefit in the treatment or management of a disease or disorder, or to delay or minimize one or more symptoms associated with the disease or disorder.
  • a therapeutically effective amount means an amount of therapeutic agent (e.g., immune cells), alone or in combination with other therapies, which provides a therapeutic benefit in the treatment or management of the disease or disorder.
  • therapeutically effective amount can encompass an amount that improves overall therapy, reduces or avoids symptoms or causes of disease or disorder, or enhances the therapeutic efficacy of another therapeutic agent.
  • a prophylactically effective amount is an amount sufficient to prevent a disease or disorder, or prevent its recurrence or relapse.
  • a prophylactically effective amount means an amount of therapeutic agent (e.g. , immune cells), alone or in combination with other agents, which provides a prophylactic benefit in the prevention of the disease.
  • the term “prophylactically effective amount” can encompass an amount that improves overall prophylaxis or enhances the prophylactic efficacy of another prophylactic agent.
  • cancers include, but are not limited to, cancers of the skin, such as melanoma; lymph node; breast; cervix; uterus; gastrointestinal tract; lung; ovary; prostate; colon; rectum; mouth; brain; head and neck; throat; testes; kidney; pancreas; bone; spleen; liver; bladder; larynx; nasal passages; and AIDS-related cancers.
  • the compounds are also useful for treating cancers of the blood and bone marrow, such as multiple myeloma and acute and chronic leukemias, for example, lymphoblastic, myelogenous, lymphocytic, and myelocytic leukemias.
  • the compounds provided herein can be used for treating, preventing or managing either primary or metastatic tumors.
  • Other specific cancers include, but are not limited to, advanced malignancy, amyloidosis, neuroblastoma, meningioma, hemangiopericytoma, multiple brain metastases, glioblastoma multiforms, glioblastoma, brain stem glioma, poor prognosis malignant brain tumor, malignant glioma, recurrent malignant glioma,- anaplastic astrocytoma, anaplastic oligodendroglioma, neuroendocrine tumor, rectal adenocarcinoma, Dukes C & D colorectal cancer, unresectable colorectal carcinoma, metastatic hepatocellular carcinoma, Kaposi's sarcoma, karotype acute myeloblastic leukemia, chronic lymphocytic leukemia (CLL), Hodgkin's lymphoma, non
  • cancers are associated with overexpression of Her- 2/neu.
  • Her- 2/neu examples include, but are not limited to, solid tumors such as sarcomas, carcinomas, lymphomas, blastomas, melanomas, and in some cases, leukemias.
  • provided herein are methods of treating, preventing or managing various forms of leukemias such as chronic lymphocytic leukemia, chronic myelocytic leukemia, acute lymphoblastic leukemia, acute myelogenous leukemia and acute myeloblastic leukemia, including leukemias that are relapsed, refractory or resistant.
  • leukemias such as chronic lymphocytic leukemia, chronic myelocytic leukemia, acute lymphoblastic leukemia, acute myelogenous leukemia and acute myeloblastic leukemia, including leukemias that are relapsed, refractory or resistant.
  • leukemia refers malignant neoplasms of the blood-forming tissues.
  • the leukemia includes, but is not limited to, chronic lymphocytic leukemia, chronic myelocytic leukemia, acute lymphoblastic leukemia, acute myelogenous leukemia and acute myeloblasts leukemia.
  • the leukemia can be relapsed, refractory or resistant to conventional therapy.
  • relapsed refers to a situation where patients who have had a remission of leukemia after therapy have a return of leukemia cells in the marrow and a decrease in normal blood cells.
  • refractory or resistant refers to a circumstance where patients, even after intensive treatment, have residual leukemia cells in their marrow.
  • methods of treating, preventing or managing chronic lymphocytic leukemia In another embodiment, provided herein are methods of treating, preventing or managing acute lymphoblastic leukemia.
  • NDL Non-Hodgkin's lymphoma
  • lymphomas including Non-Hodgkin's lymphoma (NHL).
  • lymphomas include lymph nodes, bone marrow, spleen, liver and gastrointestinal tract.
  • NHL examples include, but are not limited to, mantle cell lymphoma (MCL), lymphocytic lymphoma of intermediate differentiation, intermediate lymphocytic lymphoma (ILL), diffuse poorly differentiated lymphocytic lymphoma (PDL), diffuse large B cell lymphoma, centrocytic lymphoma, diffuse small-cleaved cell lymphoma (DSCCL), follicular lymphoma, and any type of the mantle cell lymphomas that can be seen under the microscope (nodular, diffuse, blastic and mentle zone lymphoma).
  • MCL mantle cell lymphoma
  • ILL intermediate lymphocytic lymphoma
  • PDL diffuse poorly differentiated lymphocytic lymphoma
  • DSL diffuse large B cell lymphoma
  • centrocytic lymphoma centrocytic lymphoma
  • DSCL diffuse small-cleaved cell lymphoma
  • follicular lymphoma any type of the mant
  • Cells expressing exogenous CARs provided herein may be combined with other pharmacologically active compounds ("second active agents") or other therapies for the treatment, management, and/or prevention of relapse of cancer. Certain combinations may work synergistically in the treatment, management, and/or prevention of relapse of cancer, and conditions and symptoms associated therewith. Cells provided herein can also work to alleviate adverse effects associated with certain anti-cancer agents, and vice versa.
  • second active agents pharmacologically active compounds
  • Second active agents can be large molecules (e.g. , proteins) or small molecules (e.g., synthetic inorganic, organometallic, or organic molecules).
  • large molecule active agents include, but are not limited to, hematopoietic growth factors, cytokines, and monoclonal and polyclonal antibodies.
  • the active agents are anti-CD40 monoclonal antibodies (such as, for example, SGN-40); histone deacetlyase inhibitors (such as, for example, SAHA and LAQ 824); heat-shock protein-90 inhibitors (such as, for example, 17-AAG); insulin-like growth factor- 1 receptor kinase inhibitors; vascular endothelial growth factor receptor kinase inhibitors (such as, for example, PTK787); insulin growth factor receptor inhibitors; lysophosphatidic acid acyltransrerase inhibitors; IkB kinase inhibitors; p38MAPK inhibitors; EGFR inhibitors (such as, for example, gefitinib and erlotinib HCL); HER-2 antibodies (such as, for example, trastuzumab (Herceptin ® ) and pertuzumab (OmnitargTM)); VEGFR antibodies (such as, for example, bevacizumab (AvastinTM));
  • the second active agents include, but are not limited to: semaxanib; cyclosporin; etanercept; doxycycline; bortezomib; acivicin; aclarubicin; acodazole hydrochloride; acronine; adozelesin; aldesleukin; altretamine; ambomycin; ametantrone acetate; amsacrine; anastrozole; anthramycin; asparaginase; asperlin; azacitidine; azetepa; azotomycin; batimastat; benzodepa; bicalutamide; bisantrene hydrochloride; bisnafide dimesylate; bizelesin; bleomycin sulfate; brequinar sodium; bropirimine; busulfan; cactinomycin; calusterone; caracemide; carbetimer; carboplatin; carmus
  • talisomycin tecogalan sodium; taxotere; tegafur; teloxantrone hydrochloride; temoporfin; teniposide; teroxirone; testolactone; thiamiprine; thioguanine; thiotepa; tiazofurin; tirapazamine; toremifene citrate; trestolone acetate; triciribine phosphate; trimetrexate; trimetrexate glucuronate; triptorelin; tubulozole hydrochloride; uracil mustard; uredepa; vapreotide; verteporfin; vinblastine sulfate; vincristine sulfate; vindesine; vindesine sulfate; vinepidine sulfate; vinglycinate sulfate; vinleurosine sulfate; vinorelbine tartrate; vinrosidine sulfate; vinzolidine
  • second agents include, but are not limited to: 20-epi-l,25 dihydroxyvitamin D3; 5-ethynyluracil; abiraterone; aclarubicin; acylfulvene; adecypenol; adozelesin; aldesleukin; ALL-TK antagonists; altretamine; ambamustine; amidox; amifostine; aminolevulinic acid; amrubicin; amsacrine; anagrelide; anastrozole; andrographolide; angiogenesis inhibitors; antagonist D; antagonist G; antarelix; anti-dorsalizing morphogenetic protein- 1 ; antiandrogen, prostatic carcinoma; antiestrogen; antineoplaston; antisense oligonucleotides; aphidicolin glycinate; apoptosis gene modulators; apoptosis regulators; apurinic acid; ara-CDP-DL-PTBA;
  • the active agents include, but are not limited to, 2- methoxyestradiol, telomestatin, inducers of apoptosis in mutiple myeloma cells (such as, for example, TRAIL), statins, semaxanib, cyclosporin, etanercept, doxycycline, bortezomib, oblimersen (Genasense®), remicade, docetaxel, celecoxib, melphalan, dexamethasone (Decadron®), steroids, gemcitabine, cisplatinum, temozolomide,.
  • mutiple myeloma cells such as, for example, TRAIL
  • statins such as, for example, statins, semaxanib, cyclosporin, etanercept, doxycycline, bortezomib, oblimersen (Genasense®), remicade, docetaxel,
  • the second active agents include, but are not limited to, anthracycline, platinum, alkylating agent, oblimersen (Genasense®), cisplatinum, cyclophosphamide, temodar, carboplatin, procarbazine, gliadel, tamoxifen, topotecan, methotrexate, taxotere, irinotecan, capecitabine, cisplatin, thiotepa, fludarabine, carboplatin, liposomal daunorubicin, cytarabine, doxetaxol, pacilitaxel, vinblastine, IL-2, GM-CSF, dacarbazine, vinorelbine, zoledronic acid, palmitronate, biaxin, busulphan, prednisone, bisphosphonate, arsenic trioxide, vincristine, doxorubicin (Doxil ® ),
  • the second active agents include, but are not limited to: interleukins, such as IL-2 (including recombinant IL-II ("rIL2") and canarypox IL-2), IL-10, IL-12, and IL-18; interferons, such as interferon alfa-2a, interferon alfa-2b, interferon alfa-nl, interferon alfa-n3, interferon beta-I a, and interferon gamma-I b; and G-CSF; hydroxyurea; butyrates or butyrate derivatives; nitrous oxide; hydroxy urea; ⁇ TM (NIPRISANTM; see United States Patent No.
  • interleukins such as IL-2 (including recombinant IL-II (“rIL2") and canarypox IL-2), IL-10, IL-12, and IL-18
  • interferons such as interferon alfa-2a, interferon alfa-2b, interfer
  • Gardos channel antagonists such as clotrimazole and triaryl methane derivatives
  • Deferoxamine protein C
  • transfusions of blood, or of a blood substitute such as HemospanTM or HemospanTM PS (Sangart).
  • Administration of a cell provided herein and the second active agents to a patient can occur simultaneously or sequentially by the same or different routes of administration.
  • the suitability of a particular route of administration employed for a particular active agent will depend on the active agent itself (e.g. , whether it can be administered orally without decomposing prior to entering the blood stream) and the particular cancer being treated.
  • Routes of administration for the second active agents or ingredients are known to those of ordinary skill in the art. (See, e.g., Physicians ' Desk Reference (60 th ed., 2006)).
  • the cells provided herein may be used in combination with one or more other conventional anti-cancer therapies.
  • Such therapies include, but are not limited to, surgery, chemotherapy, radiation therapy, hormonal therapy, biological therapy and immunotherapy. Without being limited by a particular theory, it is believed that using the methods provided herein in combination with other anti-cancer therapies can yield synergistic effects, resulting in higher efficacy and/or a reduced adverse effects.
  • Cells provided herein and other active ingredients discussed above can be administered to a patient prior to, during, or after the conventional therapy. 5.4 Pharmaceutical Compositions and Dosage Forms
  • compositions can be used in the preparation of individual, single unit dosage forms.
  • Pharmaceutical compositions and dosage forms provided herein comprise a cell provided herein.
  • Pharmaceutical compositions and dosage forms can further comprise one or more excipients.
  • compositions and dosage forms provided herein can also comprise one or more additional active ingredients.
  • additional active ingredients are disclosed in Section 5.3, above.
  • Single unit dosage forms provided herein are suitable for oral, mucosal
  • dosage forms include, but are not limited to: tablets; caplets; capsules, such as soft elastic gelatin capsules; cachets; troches; lozenges; dispersions; suppositories; powders; aerosols (e.g., nasal sprays or inhalers); gels; liquid dosage forms suitable for oral or mucosal administration to a patient, including suspensions (e.g.
  • aqueous or nonaqueous liquid suspensions e.g., oil-in-water emulsions, or a water-in-oil liquid emulsions), solutions, and elixirs
  • liquid dosage forms suitable for parenteral administration to a patient eye drops or other ophthalmic preparations suitable for topical administration
  • sterile solids e.g., crystalline or amorphous solids that can be reconstituted to provide liquid dosage forms suitable for parenteral administration to a patient.
  • compositions, shape, and type of dosage forms will typically vary depending on their use.
  • a dosage form used in the acute treatment of a disease may contain larger amounts of one or more of the active ingredients it comprises than a dosage form used in the chronic treatment of the same disease.
  • a parenteral dosage form may contain smaller amounts of one or more of the active ingredients it comprises than an oral dosage form used to treat the same disease.
  • Suitable excipients are well known to those skilled in the art of pharmacy, and non-limiting examples of suitable excipients are provided herein. Whether a particular excipient is suitable for incorporation into a pharmaceutical composition or dosage form depends on a variety of factors well known in the art including, but not limited to, the way in which the dosage form will be administered to a patient. For example, oral dosage forms such as tablets may contain excipients not suited for use in parenteral dosage forms. The suitability of a particular excipient may also depend on the specific active ingredients in the dosage form. For example, the decomposition of some active ingredients may be accelerated by some excipients such as lactose, or when exposed to water.
  • lactose-free compositions can comprise excipients that are well known in the art and are listed, for example, in the U.S. Pharmacopeia (USP) 25-NF20 (2002).
  • lactose-free compositions comprise active ingredients, a binder/filler, and a lubricant in pharmaceutically compatible and pharmaceutically acceptable amounts.
  • lactose-free dosage forms comprise active ingredients, microcrystalline cellulose, pre-gelatinized starch, and magnesium stearate.
  • anhydrous pharmaceutical compositions and dosage forms comprising active ingredients, since water can facilitate the degradation of some compounds.
  • water e.g., 5%
  • water is widely accepted in the pharmaceutical arts as a means of simulating long-term storage in order to determine characteristics such as shelf-life or the stability of formulations over time. See, e.g. , Jens T. Carstensen, Drug Stability: Principles & Practice, 2d. Ed., Marcel Dekker, NY, NY, 1995, pp. 379-80.
  • water and heat accelerate the decomposition of some compounds.
  • the effect of water on a formulation can be of great significance since moisture and/or humidity are commonly encountered during manufacture, handling, packaging, storage, shipment, and use of formulations.
  • Anhydrous pharmaceutical compositions and dosage forms can be prepared using anhydrous or low moisture containing ingredients and low moisture or low humidity conditions.
  • Pharmaceutical compositions and dosage forms that comprise lactose and at least one active ingredient that comprises a primary or secondary amine are preferably anhydrous if substantial contact with moisture and/or humidity during manufacturing, packaging, and/or storage is expected.
  • An anhydrous pharmaceutical composition should be prepared and stored such that its anhydrous nature is maintained.
  • anhydrous compositions are, in one embodiment, packaged using materials known to prevent exposure to water such that they can be included in suitable formulary kits. Examples of suitable packaging include, but are not limited to, hermetically sealed foils, plastics, unit dose containers (e.g., vials), blister packs, and strip packs.
  • compositions and dosage forms that comprise one or more compounds that reduce the rate by which an active ingredient (e.g. , cells provided herein or optional second active agents) will decompose.
  • active ingredient e.g. , cells provided herein or optional second active agents
  • stabilizers include, but are not limited to, antioxidants such as ascorbic acid, pH buffers, or salt buffers.
  • the amounts and specific types of active ingredients in a dosage form may differ depending on factors such as, but not limited to, the specific agent used, the route by which it is to be administered to patients, the particular cancer being treated or managed, and the amount(s) of a cell or second active agent provided herein, and any additional active agents or therapies concurrently administered to the patient.
  • compositions that are suitable for oral administration can be provided as discrete dosage forms, such as, but not limited to, tablets (e.g., chewable tablets), caplets, capsules, and liquids (e.g., flavored syrups).
  • dosage forms contain predetermined amounts of active ingredients, and may be prepared by methods of pharmacy well known to those skilled in the art. (See generally, Remington 's Pharmaceutical Sciences, 18th ed., Mack Publishing, Easton PA (1990)).
  • Oral dosage forms provided herein are prepared by combining the active ingredients in an intimate admixture with at least one excipient according to conventional pharmaceutical compounding techniques.
  • Excipients can take a wide variety of forms depending on the form of preparation desired for administration.
  • excipients suitable for use in oral liquid or aerosol dosage forms include, but are not limited to, water, glycols, oils, alcohols, flavoring agents, preservatives, and coloring agents.
  • excipients suitable for use in solid oral dosage forms include, but are not limited to, starches, sugars, micro-crystalline cellulose, diluents, granulating agents, lubricants, binders, and disintegrating agents.
  • oral dosage forms are tablets or capsules, in which case solid excipients are employed.
  • tablets can be coated by standard aqueous or nonaqueous techniques.
  • Such dosage forms can be prepared by any of the methods of pharmacy.
  • pharmaceutical compositions and dosage forms are prepared by uniformly and intimately admixing the active ingredients with liquid carriers, finely divided solid carriers, or both, and then shaping the product into the desired presentation if necessary.
  • a tablet can be prepared by compression or molding.
  • Compressed tablets can be prepared by compressing in a suitable machine the active ingredients in a free-flowing form such as powder or granules, optionally mixed with an excipient.
  • Molded tablets can be made by molding in a suitable machine a mixture of the powdered compound moistened with an inert liquid diluent.
  • excipients that can be used in oral dosage forms provided herein include, but are not limited to, binders, fillers, disintegrants, and lubricants.
  • Binders suitable for use in pharmaceutical compositions and dosage forms include, but are not limited to, corn starch, potato starch, or other starches, gelatin, natural and synthetic gums such as acacia, sodium alginate, alginic acid, other alginates, powdered tragacanth, guar gum, cellulose and its derivatives (e.g., ethyl cellulose, cellulose acetate, carboxymethyl cellulose calcium, sodium carboxymethyl cellulose), polyvinyl pyrrolidone, methyl cellulose, pre-gelatinized starch, hydroxypropyl methyl cellulose, (e.g., Nos. 2208, 2906, 2910), microcrystalline cellulose, and mixtures thereof.
  • Suitable forms of microcrystalline cellulose include, but are not limited to, the materials sold as AVICEL-PH- 101 , AVICEL-PH-103 AVICEL RC-581 , AVICEL- PH-105 (available from FMC Corporation, American Viscose Division, Avicel Sales, Marcus Hook, PA), and mixtures thereof.
  • An specific binder is a mixture of microcrystalline cellulose and sodium carboxymethyl cellulose sold as AVICEL RC-581.
  • Suitable anhydrous or low moisture excipients or additives include AVICEL-PH-103TM and Starch 1500 LM.
  • fillers suitable for use in the pharmaceutical compositions and dosage forms provided herein include, but are not limited to, talc, calcium carbonate (e.g., granules or powder), microcrystalline cellulose, powdered cellulose, dextrates, kaolin, mannitol, silicic acid, sorbitol, starch, pre-gelatinized starch, and mixtures thereof.
  • the binder or filler in pharmaceutical compositions is, in one embodiment, present in from about 50 to about 99 weight percent of the pharmaceutical composition or dosage form.
  • Disintegrants may be used in the compositions to provide tablets that disintegrate when exposed to an aqueous environment. Tablets that contain too much disintegrant may disintegrate in storage, while those that contain too little may not disintegrate at a desired rate or under the desired conditions. Thus, a sufficient amount of disintegrant that is neither too much nor too little to detrimentally alter the release of the active ingredients may be used to form solid oral dosage forms. The amount of disintegrant used varies based upon the type of formulation, and is readily discernible to those of ordinary skill in the art. In one embodiment, pharmaceutical compositions comprise from about 0.5 to about 15 weight percent of disintegrant, or from about 1 to about 5 weight percent of disintegrant.
  • Disintegrants that can be used in pharmaceutical compositions and dosage forms include, but are not limited to, agar-agar, alginic acid, calcium carbonate, microcrystalline cellulose, croscarmellose sodium, crospovidone, polacrilin potassium, sodium starch glycolate, potato or tapioca starch, other starches, pre-gelatinized starch, other starches, clays, other algins, other celluloses, gums, and mixtures thereof.
  • Lubricants that can be used in pharmaceutical compositions and dosage forms include, but are not limited to, calcium stearate, magnesium stearate, mineral oil, light mineral oil, glycerin, sorbitol, mannitol, polyethylene glycol, other glycols, stearic acid, sodium lauryl sulfate, talc, hydrogenated vegetable oil (e.g. , peanut oil, cottonseed oil, sunflower oil, sesame oil, olive oil, corn oil, and soybean oil), zinc stearate, ethyl oleate, ethyl laureate, agar, and mixtures thereof.
  • Additional lubricants include, for example, a syloid silica gel (AEROSIL200, manufactured by W.R.
  • lubricants may be used in an amount of less than about 1 weight percent of the pharmaceutical compositions or dosage forms into which they are incorporated.
  • Active ingredients provided herein can be administered by controlled release means or by delivery devices that are well known to those of ordinary skill in the art. Examples include, but are not limited to, those described in U.S. Patent Nos.: 3,845,770; 3,916,899; 3,536,809; 3,598, 123; and 4,008,719, 5,674,533, 5,059,595, 5,591 ,767, 5, 120,548, 5,073,543, 5,639,476, 5,354,556, and 5,733,566, each of which is incorporated herein by reference.
  • Such dosage forms can be used to provide slow or controlled-release of one or more active ingredients using, for example, hydropropylmethyl cellulose, other polymer matrices, gels, permeable membranes, osmotic systems, multilayer coatings, microparticles, liposomes, microspheres, or a. combination thereof to provide the desired release profile in varying proportions.
  • Suitable controlled-release formulations known to those of ordinary skill in the art, including those described herein, can be readily selected for use with the active agents provided herein.
  • provided are single unit dosage forms suitable for oral administration such as, but not limited to, tablets, capsules, gelcaps, and caplets that are adapted for controlled-release.
  • controlled-release pharmaceutical products improve drug therapy over that achieved by their non-controlled counterparts.
  • use of a controlled-release preparation in medical treatment is characterized by a minimum of drug substance being employed to cure or control the condition in a minimum amount of time.
  • Advantages of controlled-release formulations include extended activity of the drug, reduced dosage frequency, and increased patient compliance.
  • controlled-release formulations can be used to affect the time of onset of action or other characteristics, such as blood levels of the drug, and can thus affect the occurrence of side (e.g., adverse) effects.
  • the controlled-release formulations are designed to initially release an amount of drug (active ingredient) that promptly produces the desired therapeutic or prophylactic effect, and gradually and continually release of other amounts of drug to maintain this level of therapeutic or prophylactic effect over an extended period of time.
  • the drug in order to maintain a constant level of drug in the body, the drug can be released from the dosage form at a rate that will replace the amount of drug being metabolized and excreted from the body.
  • Controlled-release of an active ingredient can be stimulated by various conditions including, but not limited to, pH, temperature, enzymes, water, or other physiological conditions or compounds.
  • Parenteral dosage forms can be administered to patients by various routes including, but not limited to, subcutaneous, intravenous (including bolus injection), intramuscular, and intraarterial.
  • administration of a parenteral dosage form bypasses patients' natural defenses against contaminants, and thus, in these embodiments, parenteral dosage forms are sterile or capable of being sterilized prior to administration to a patient.
  • parenteral dosage forms include, but are not limited to, solutions ready for injection, dry products ready to be dissolved or suspended in a pharmaceutically acceptable vehicle for injection, suspensions ready for injection, and emulsions.
  • Suitable vehicles that can be used to provide parenteral dosage forms are well known to those skilled in the art. Examples include, but are not limited to: Water for Injection USP; aqueous vehicles such as, but not limited to, Sodium Chloride Injection, Ringer's Injection, Dextrose Injection, Dextrose and Sodium Chloride Injection, and Lactated Ringer's Injection; water-miscible vehicles such as, but not limited to, ethyl alcohol, polyethylene glycol, and polypropylene glycol; and nonaqueous vehicles such as, but not limited to, corn oil, cottonseed oil, peanut oil, sesame oil, ethyl oleate, isopropyl myristate, and benzyl benzoate.
  • aqueous vehicles such as, but not limited to, Sodium Chloride Injection, Ringer's Injection, Dextrose Injection, Dextrose and Sodium Chloride Injection, and Lactated Ringer's Injection
  • cyclodextrin and its derivatives can be used to increase the solubility of a compound provided herein. See, e.g. , U.S. Patent No. 5,134,127, which is incorporated herein by reference.
  • Topical and mucosal dosage forms include, but are not limited to, sprays, aerosols, solutions, emulsions, suspensions, eye drops or other ophthalmic preparations, or other forms known to one of skill in the art. See, e.g. , Remington 's Pharmaceutical Sciences, 16th and 18th eds., Mack Publishing, Easton PA (1980 & 1990); and Introduction to Pharmaceutical Dosage Forms, 4th ed., Lea & Febiger, Philadelphia (1985). Dosage forms suitable for treating mucosal tissues within the oral cavity can be formulated as mouthwashes or as oral gels.
  • excipients include, but are not limited to, water, acetone, ethanol, ethylene glycol, propylene glycol, butane- 1 , 3 -diol, isopropyl myristate, isopropyl palmitate, mineral oil, and mixtures thereof to form solutions, emulsions or gels, which are nontoxic and pharmaceutically acceptable.
  • Moisturizers or humectants can also be added to pharmaceutical compositions and dosage forms. Examples of additional ingredients are well known in the art. See, e.g. , Remington 's Pharmaceutical Sciences, 16th and 18th eds., Mack Publishing, Easton PA (1980 & 1990).
  • the pH of a pharmaceutical composition or dosage form may also be adjusted to improve delivery of one or more active ingredients.
  • the polarity of a solvent carrier, its ionic strength, or tonicity can be adjusted to improve delivery.
  • Compounds such as stearates can also be added to pharmaceutical compositions or dosage forms to alter the hydrophilicity or lipophilicity of one or more active ingredients so as to improve delivery.
  • stearates can serve as a lipid vehicle for the formulation, as an emulsifying agent or surfactant, or as a delivery-enhancing or penetration-enhancing agent.
  • salts, solvates, prodrugs, clathrates, or stereoisomers of the active ingredients can be used to further adjust the properties of the resulting composition. 6.
  • ErbB-2 specific human antibodies 6D and 7G were generated using methods conventionally known in the art. In order to test whether the antibodies obtained recognize Her-2/neu, the binding of these antibodies to Her-2/neu was assessed in SK-BR3 (Her-2/neu + ) and MDA-MB-468 (Her-2/neu " ) cell lines using FACS analysis.
  • Herceptin which is a Her-2/neu specific antibody, was also included as positive control.
  • antibodies 6D and 7G did not show any significant binding activities in MDA-MB-468 cells which do not express Her-2/neu, while binding acitivities were observed for these antibodies in SK-BR3 cells which express Her-2/neu.
  • Her-2 chimeric antigen receptor genes (SEQ. ID. NOs 1 and 2) were constructed based on the antibodies 6D and 7G described above. Specifically, the segment of interest was constructed by placing the single chain region of antibodies 6D and 7G on one end and signaling domain ⁇ 3 ⁇ on the other end. In between these two regions, genes for CD8 based hinge and transmembrane domain were inserted. In addition, genes for costimulatory molecules such as CD28 and 4- IBB were inserted to prolong the cell persist duration once the genes are introduced into the cells. The genes were constructed to provide the order of the costimulatory molecules to be CD28-4- 1BB-CD3 .
  • Her-2/neu CAR plasmid DNA (6D) was transfected at different levels into cultured activated T cells using electroporation. Expression of EGFP was monitored following the transfection. As showin in FIG. 3, expression of EGFP was indeed observed following the transfection. In addition, it was observed that the level of EGFP expression is proportional to the level of mRNA transfected.
  • Her-2/neu CAR was also monitored following the transfection. As shown in FIG. 4, expression of Her-2/neu CAR was indeed observed following the transfection. Similar to EGFP, it was observed that the level of Her-2/neu CAR expression is proportional to the level of mRNA transfected.
  • Her-2/neu CAR plasmid DNA (6D or 7G) is transfected into cultured activated killer cells using electroporation. Two days after transfection, efficiency and viability of the transfected cells are assessed using FACS analysis. In addition, the expression of CARs on the surface of the transfected cells is confirmed using recombinant Her-2-Fc fusion protein and goat-anti-Fc PE secondary antibodies.
  • the conditions for transfection using electroporation are optimized by varying concentrations of CAR encoding plasmid and activated killer cells.
  • the optimal conditions can be determined based on the expression of EGFP and assessment of PI using FACS analysis.
  • activated killer cells are transfected with mRNA encoding Her-2/neu CAR using procedures substantially similar to those described above.
  • the desired characteristics of the transfected cells are achieved by comparing the transfection using Her-2/neu CAR mRNA with that using Her-2/neu CAR plasmid DNA. For example, characteristics such as prolonged expression of the transfected genes and length of time required to achieve certain levels of expression are compared between the two transfection methods, and selection between plasmid and mRNA is made depending on what is desired of the resulting transfected cells.
  • Her-2/neu CAR plasmid or mRNA is transfected into activated killer cells using nanotubes.
  • Silica nanotube is efficient for gene delivery because inner surface of the nanotubes is coated with positive charges.
  • the plasmid DNA or mRNA and silica nanotube are mixed by rocking for 24 hours at 4°C.
  • the DNA or mRNA complex is added to activated T cells and incubated for 48 hours. Gene transduction efficiency is evaluated by FACS analysis.
  • the conditions for transfection using nanotubes are optimized by varying concentrations of CAR encoding plasmid and activated killer cells.
  • the optimal conditions can be determined based on the expression of EGFP and assessment of PI using FACS analysis.
  • activated killer cells are transfected with mRNA encoding Her-2/neu CAR using procedures substantially similar to those described above.
  • the desired characteristics of the transfected cells are achieved by comparing the transfection using Her-2/neu CAR mRNA with that using Her-2/neu CAR plasmid DNA. For example, characteristics such as prolonged expression of the transfected genes and length of time required to achieve certain levels of expression are compared between the two transfection methods, and selection between plasmid and mRNA is made depending on what is desired of the resulting transfected cells.
  • CAR transfected cells The antitumor activities of CAR transfected cells are assessed in vitro by determining the cytotoxicity of the transfected cells against SK-OV3 (ErbB-2 positive) and MDA-MB 468 (ErbB-2 negative).
  • Her-2/neu transfected activated killer cells or non-transduced activated killer cells are coincubated with SK-OV3 or MDA-MB 468 cells for 4 hours.
  • cytotoxicity of Her- 2/neu transfected activated killer cells and non-transduced activated killer cells is determined using any conventional methods known in the art, e.g., cytotoxicity using Calcein-AM and PI.
  • the antitumor activities of CAR transfected cells are assessed in vivo using mouse ovarian xenograft model.
  • Ovarian cancer cells SK-OV3 are transplanted i.p. into the nude mice of 6-8 weeks of age.
  • Her-2/neu transfected activated killer cells or non-transduced activated killer cells are intravenously injected once a week for 4 weeks.
  • the size of the tumor and body weight of the animals are determined and recorded at indicated time points. Tumors are excised from some of the animals 2 weeks after the final administration, and tumor weights and body weights of the animals are determined.
  • the tumor weights and body weights determined from animals that received Her-2/neu CAR transfected cells or non transduced cells are compared with those from animals that received no treatments to assess tumor growth suppression and antitumor efficacy.
  • activated killer cells that homed in tumors are imaged using immunohistochemistry.
  • mice from which the tumors are not excised are used to assess the rate of survival.

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Abstract

L'invention concerne des récepteurs d'antigènes chimériques spécifiques de tumeurs associées à des antigènes et des procédés d'expression de ces récepteurs d'antigènes chimériques dans des cellules tueuses activées. Elle concerne également des procédés et des compositions pour traiter un cancer employant les récepteurs d'antigènes chimériques.
PCT/KR2011/007621 2010-10-13 2011-10-13 Immunothérapie contre les tumeurs solides Ceased WO2012050374A2 (fr)

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* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US9181527B2 (en) 2009-10-29 2015-11-10 The Trustees Of Dartmouth College T cell receptor-deficient T cell compositions
US9273283B2 (en) 2009-10-29 2016-03-01 The Trustees Of Dartmouth College Method of producing T cell receptor-deficient T cells expressing a chimeric receptor
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US10336804B2 (en) 2004-09-24 2019-07-02 Trustees Of Dartmouth College Chimeric NK receptor and methods for treating cancer
WO2019136288A1 (fr) * 2018-01-05 2019-07-11 Maxcyte, Inc. Traitement chronique du cancer par les car
US10464988B2 (en) 2015-10-23 2019-11-05 Eureka Therapeutics, Inc. Antibody/T-cell receptor chimeric constructs and uses thereof
EP3436037A4 (fr) * 2016-03-31 2019-12-04 University of Southern California Dosage de rapporteur hautement sensible à base de luciférase spécifique pour la détection d'antigènes
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US10869888B2 (en) 2018-04-17 2020-12-22 Innovative Cellular Therapeutics CO., LTD. Modified cell expansion and uses thereof
US10874693B2 (en) 2012-05-25 2020-12-29 Cellectis CD19 specific chimeric antigen receptor and uses thereof
WO2020257885A1 (fr) * 2019-06-28 2020-12-30 Commonwealth Scientific And Industrial Research Organisation Système de distribution à médiation par des nanofils et procédés les comprenant
US10918667B2 (en) 2018-11-20 2021-02-16 Innovative Cellular Therapeutics CO., LTD. Modified cell expressing therapeutic agent and uses thereof
US10967005B2 (en) 2013-03-15 2021-04-06 Celgene Corporation Modified T lymphocytes comprising a BAFF antibody-inducible caspase and methods of apoptosis
US11077144B2 (en) 2013-05-13 2021-08-03 Cellectis CD19 specific chimeric antigen receptor and uses thereof
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US11130820B2 (en) 2012-12-20 2021-09-28 Celgene Corporation Chimeric antigen receptors
US11161913B2 (en) 2018-08-30 2021-11-02 Innovative Cellular Therapeutics Holdings, Ltd. Chimeric antigen receptor cells for treating solid tumor
US11331380B2 (en) 2016-10-20 2022-05-17 Celgene Corporation Cereblon-based heterodimerizable chimeric antigen receptors
US11331344B2 (en) 2008-04-09 2022-05-17 Maxcyte Inc. Engineering and delivery of therapeutic compositions of freshly isolated cells
US11613573B2 (en) 2017-04-26 2023-03-28 Eureka Therapeutics, Inc. Chimeric antibody/T-cell receptor constructs and uses thereof
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Publication number Priority date Publication date Assignee Title
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US11014989B2 (en) 2015-01-26 2021-05-25 Cellectis Anti-CLL1 specific single-chain chimeric antigen receptors (scCARs) for cancer immunotherapy
WO2016120216A1 (fr) 2015-01-26 2016-08-04 Cellectis Systèmes de récepteurs d'antigènes chimériques dirigés par des mab pour trier/appauvrir les cellules immunitaires génétiquement modifiées
EP3277314A4 (fr) * 2015-04-03 2018-08-29 Eureka Therapeutics, Inc. Constructions ciblant des complexes peptide afp/cmh et leurs utilisations
TWI786034B (zh) * 2015-04-03 2022-12-11 美商優瑞科生物技術公司 靶向afp肽/mhc複合體之構築體及其用途
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US11976105B2 (en) 2015-10-23 2024-05-07 Eureka Therapeutics, Inc. Antibody/T-cell receptor chimeric constructs and uses thereof
US10822389B2 (en) 2015-10-23 2020-11-03 Eureka Therapeutics, Inc. Antibody/T-cell receptor chimeric constructs and uses thereof
US11421013B2 (en) 2015-10-23 2022-08-23 Eureka Therapeutics, Inc. Antibody/T-cell receptor chimeric constructs and uses thereof
US10464988B2 (en) 2015-10-23 2019-11-05 Eureka Therapeutics, Inc. Antibody/T-cell receptor chimeric constructs and uses thereof
US11197919B2 (en) 2015-11-04 2021-12-14 City Of Hope Chimeric antigen receptors targeting HER2
RU2753695C2 (ru) * 2015-11-04 2021-08-19 Сол Дж. ПРАЙСМЕН Химерные рецепторы антигена, нацеленные на her2
EP3436037A4 (fr) * 2016-03-31 2019-12-04 University of Southern California Dosage de rapporteur hautement sensible à base de luciférase spécifique pour la détection d'antigènes
US11768203B2 (en) 2016-03-31 2023-09-26 University Of Southern California Highly sensitive and specific luciferase based reporter assay for antigen detection
CN106222201A (zh) * 2016-08-27 2016-12-14 北京艺妙神州医疗科技有限公司 一种制备car‑t细胞的方法以及制得的car‑t细胞及其应用
US11331380B2 (en) 2016-10-20 2022-05-17 Celgene Corporation Cereblon-based heterodimerizable chimeric antigen receptors
US11613573B2 (en) 2017-04-26 2023-03-28 Eureka Therapeutics, Inc. Chimeric antibody/T-cell receptor constructs and uses thereof
US11965021B2 (en) 2017-04-26 2024-04-23 Eureka Therapeutics, Inc. Cells expressing chimeric activating receptors and chimeric stimulating receptors and uses thereof
WO2019002633A1 (fr) 2017-06-30 2019-01-03 Cellectis Immunothérapie cellulaire pour une administration répétitive
WO2019136288A1 (fr) * 2018-01-05 2019-07-11 Maxcyte, Inc. Traitement chronique du cancer par les car
US10869888B2 (en) 2018-04-17 2020-12-22 Innovative Cellular Therapeutics CO., LTD. Modified cell expansion and uses thereof
US12150960B2 (en) 2018-04-17 2024-11-26 Innovative Cellular Therapeutics Holdings, Ltd. Modified cell expansion and uses thereof
US12240915B2 (en) 2018-08-30 2025-03-04 Innovative Cellular Therapeutics Holdings, Ltd. Chimeric antigen receptor cells for treating solid tumor
US11161913B2 (en) 2018-08-30 2021-11-02 Innovative Cellular Therapeutics Holdings, Ltd. Chimeric antigen receptor cells for treating solid tumor
US10918667B2 (en) 2018-11-20 2021-02-16 Innovative Cellular Therapeutics CO., LTD. Modified cell expressing therapeutic agent and uses thereof
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US12076343B2 (en) 2020-02-19 2024-09-03 Innovative Cellular Therapeutics Holdings, Ltd. Engineered safety in cell therapy
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