Uses of mesothelin-targeted chimeric antigen receptors

Abstract

The presently disclosed subject matter provides for methods and compositions for enhancing the immune response toward cancers and pathogens. It relates to chimeric antigen receptors (CARs) that specifically target human mesothelin, and immunoresponsive cells comprising such CARs. The presently disclosed mesothelin-targeted CARs have enhanced immune-activating properties, including anti-tumor activity.

1 .- 61 . (canceled) 
     
     
         62 . A method of increasing immune-activating cytokine production in response to a cancer cell, comprising administering an effective amount of immunoresponsive cells comprising a chimeric antigen receptor (CAR), wherein the CAR comprises an extracellular antigen-binding domain, a transmembrane domain, and an intracellular domain, wherein the extracellular antigen-binding domain specifically binds to human mesothelin comprises:
 (a) a heavy chain variable region comprising a CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 11, a CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 12, and a CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 13; and   (b) a light chain variable region comprising a CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 14, a CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 15, and a CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 16.   
     
     
         63 . The method of  claim 62 , wherein the immune-activating cytokine is selected from the group consisting of (GM-CSF), IFN-α, IFN-β, IFN-γ, TNF-α, IL-2, IL-3, IL-6, IL-11, IL-7, IL-12, IL-15, IL-21, interferon regulatory factor 7 (IRF7), and combinations thereof. 
     
     
         64 . The method of  claim 63 , wherein the immune-activating cytokine is selected from the group consisting of (GM-CSF), IFN-γ, TNF-α, and combinations thereof. 
     
     
         65 . The method of  claim 62 , further comprising administering at least one immunomodulatory agent. 
     
     
         66 . The method of  claim 65 , wherein the at least one immunomodulatory agent is selected from the group consisting of immunostimulatory agents, checkpoint immune blockade agents, radiation therapy agents, chemotherapy agents, and combinations thereof. 
     
     
         67 . The method of  claim 66 , wherein the immunostimulatory agents are selected from the group consisting of IL-12, an agonist costimulatory monoclonal antibody, and combinations thereof. 
     
     
         68 . The method of  claim 67 , wherein the immunostimulatory agent is IL-12. 
     
     
         69 . The method of  claim 66 , wherein the agonist costimulatory monoclonal antibody is selected from the group consisting of an anti-4-1BB antibody, an anti-OX40 antibody, an anti-ICOS antibody, and combinations thereof. 
     
     
         70 . The method of  claim 69 , wherein the agonist costimulatory monoclonal antibody is an anti-4-1BB antibody. 
     
     
         71 . The method of  claim 66 , wherein the checkpoint immune blockade agents are selected from the group consisting of anti-PD-L1 antibodies, anti-CTLA-4 antibodies, anti-PD-1 antibodies, anti-LAG3 antibodies, anti-B7-H3 antibodies, anti-TIM3 antibodies, and combinations thereof. 
     
     
         72 . The method of  claim 71 , wherein the checkpoint immune blockade agent is an anti-PD-L1 antibody. 
     
     
         73 . The method of  claim 62 , wherein the subject is a human. 
     
     
         74 . The method of  claim 62 , wherein the immunoresponsive cell is pleurally administered to the subject. 
     
     
         75 .- 97 . (canceled) 
     
     
         98 . The method of  claim 62 , wherein the extracellular antigen-binding domain recognizes human mesothelin with a mesothelin expression level of about 1,000 or more mesothelin binding sites/cell. 
     
     
         99 . The method of  claim 62 , wherein (a) the heavy chain variable region comprises an amino acid sequence that is at least about 95% identical to amino acids 1-119 of SEQ ID NO: 1, wherein SEQ ID NOS: 11-13 are invariable; and/or (b) the light chain variable region comprises an amino acid sequence that is at least about 95% identical to amino acids 1-10 7  of SEQ ID NO: 3, wherein SEQ ID NOS: 14-16 are invariable. 
     
     
         100 . The method of  claim 62 , wherein (a) the heavy chain variable region comprises amino acids 1-119 of SEQ ID NO: 1, and/or (b) the light chain variable region comprises amino acids 1-10 7  of SEQ ID NO: 3. 
     
     
         101 . The method of  claim 62 , wherein the extracellular antigen-binding domain comprises a single-chain variable fragment (scFv). 
     
     
         102 . The method of  claim 101 , wherein the scFv is a human scFv. 
     
     
         103 . The method of  claim 62 , wherein a linker is positioned between the heavy chain variable region and the light chain variable region. 
     
     
         104 . The method of  claim 103 , wherein the linker comprises the amino acid sequence set forth in SEQ ID NO: 17. 
     
     
         105 . The method of  claim 62 , wherein the transmembrane domain comprises a CD8 polypeptide, a CD28 polypeptide, a CD3ζ polypeptide, a CD4 polypeptide, a 4-1BB polypeptide, an OX40 polypeptide, an ICOS polypeptide, a CTLA-4 polypeptide, a PD-1 polypeptide, a LAG-3 polypeptide, a 2B4 polypeptide, a BTLA polypeptide, or a combination thereof. 
     
     
         106 . The method of  claim 105 , wherein the transmembrane domain comprises a CD28 polypeptide. 
     
     
         107 . The method of  claim 62 , wherein the intracellular domain comprises a CD3ζ polypeptide. 
     
     
         108 . The method of  claim 62 , wherein the intracellular domain further comprises at least one co-stimulatory signaling region. 
     
     
         109 . The method of  claim 108 , wherein the at least one co-stimulatory signaling region comprises a CD28 polypeptide, a 4-1BB polypeptide, an OX40 polypeptide, an ICOS polypeptide, or a combination thereof. 
     
     
         110 . The method of  claim 62 , wherein the intracellular domain comprises (i) a CD3ζ polypeptide and a co-stimulatory signaling domain comprising a CD28 polypeptide, or (ii) a CD3ζ polypeptide and a co-stimulatory signaling domain comprising a 4-1BB polypeptide. 
     
     
         111 . The method of  claim 62 , wherein the cell is selected from the group consisting of a T cell, a Natural Killer (NK) cell, and a pluripotent stem cell from which a lymphoid cell may be differentiated. 
     
     
         112 . The method of  claim 111 , wherein the cell is a T cell. 
     
     
         113 . The method of  claim 112 , wherein the T cell is selected from the group consisting of a cytotoxic T lymphocyte (CTL), a regulatory T cell, and combinations thereof. 
     
     
         114 . The method of  claim 113 , wherein the T cell is a CD4 +  T cell or a CD8 +  T cell. 
     
     
         115 . The method of  claim 62 , wherein the cancer cell is a solid tumor cancer cell. 
     
     
         116 . The method of  claim 115 , wherein the solid tumor is selected from the group consisting of mesothelioma, lung cancer, pancreatic cancer, ovarian cancer, breast cancer, colon cancer, pleural tumor, glioblastoma, esophageal cancer, gastric cancer, synovial sarcoma, thymic carcinoma, endometrial carcinoma, stomach cancer, cholangiocarcinoma, and a combination thereof. 
     
     
         117 . The method of  claim 62 , wherein the effective amount is at least about 1×10 5  of the cells.