Gold Optimized CAR T-cells

Abstract

Control Devices are disclosed including RNA destabilizing elements (RDE), RNA control devices, and destabilizing elements (DE) combined with Chimeric Antigen Receptors (CARs) or other transgenes in eukaryotic cells. Multicistronic vectors are also disclosed for use in engineering host eukaryotic cells with the CARs and transgenes under the control of the control devices. These control devices can be used to optimize expression of CARs in the eukaryotic cells so that, for example, effector function is optimized. CARs and transgene payloads can also be engineered into eukaryotic cells so that the transgene payload is expressed and delivered after stimulation of the CAR on the eukaryotic cell.

1 . (canceled) 
     
     
         2 . (canceled) 
     
     
         3 . (canceled) 
     
     
         4 . (canceled) 
     
     
         5 . (canceled) 
     
     
         6 . (canceled) 
     
     
         7 . (canceled) 
     
     
         8 . (canceled) 
     
     
         9 . (canceled) 
     
     
         10 . (canceled) 
     
     
         11 . (canceled) 
     
     
         12 . (canceled) 
     
     
         13 . (canceled) 
     
     
         14 . (canceled) 
     
     
         15 . (canceled) 
     
     
         16 . (canceled) 
     
     
         17 . (canceled) 
     
     
         18 . (canceled) 
     
     
         19 . (canceled) 
     
     
         20 . (canceled) 
     
     
         21 . A bicistronic construct, comprising: a first polynucleotide encoding a chimeric antigen receptor operably linked to a first control region, a second polynucleotide encoding a transgene operably linked to an RNA destabilizing element, wherein the RNA destabilizing element is an AU rich element, and operably linked to a second control region, wherein the second nucleic acid is transcribed to make a transcript encoding the transgene operably linked to the RNA destabilizing element, and wherein the first polynucleotide and the second polynucleotide are transcribed in opposite directions in the construct. 
     
     
         22 . The bicistronic construct of  claim 21 , wherein the second control region has an inducible promoter. 
     
     
         23 . The bicistronic construct of  claim 21 , wherein the transgene encodes a cytokine, a FasL, an antibody, a growth factor, a chemokine, an enzyme that cleaves a polypeptide or a polysaccharide, a granzyme, a perforin, or a checkpoint inhibitor. 
     
     
         24 . The bicistronic construct of  claim 23 , wherein the transgene encodes an IL-2, an IL-10, an IL-12, an IL-15, an IL-18, an interferon gamma, a TNFα, or a TGF-β. 
     
     
         25 . The bicistronic construct of  claim 24 , wherein the transgene encodes an IL-12. 
     
     
         26 . The bicistronic construct of  claim 21 , wherein the CAR binds to a ligand selected from the group consisting of a CD123, a CD19, a Muc1, a CD20, a CD22, a SLAMF7, a CLL-1 or a FLT-3. 
     
     
         27 . The bicistronic construct of  claim 26 , wherein the transgene encodes an IL-2, an IL-10, an IL-12, an IL-15, an IL-18, an interferon gamma, a TNFα, or a TGF-β. 
     
     
         28 . The bicistronic construct of  claim 27 , wherein the transgene encodes an IL-12. 
     
     
         29 . The bicistronic construct of  claim 21 , wherein the chimeric antigen receptor binds to a ligand and wherein the ligand is present on a cancer cell. 
     
     
         30 . The bicistronic construct of  claim 29 , wherein the cancer cell is a solid tumor cell. 
     
     
         31 . The bicistronic construct of  claim 30 , wherein the solid tumor is an ovarian cancer. 
     
     
         32 . The bicistronic construct of  claim 30 , wherein the solid tumor is a melanoma. 
     
     
         33 . The bicistronic construct of  claim 30 , wherein the transgene encodes an IL-12. 
     
     
         34 . The bicistronic construct of  claim 21 , wherein the RNA destabilizing element is from a 3′-UTR of INFg or a 3′-UTR of IL6. 
     
     
         35 . The bicistronic construct of  claim 23 , wherein the RNA destabilizing element is from a 3′-UTR of INFg or a 3′-UTR of IL6. 
     
     
         36 . The bicistronic construct of  claim 25 , wherein the RNA destabilizing element is from a 3′-UTR of INFg or a 3′-UTR of IL6. 
     
     
         37 . The bicistronic construct of  claim 21 , wherein the transgene encodes a reporter. 
     
     
         38 . The bicistronic construct of  claim 21 , wherein the transgene encodes an imaging agent. 
     
     
         39 . The bicistronic construct of  claim 21 , wherein the transgene encodes a luciferase. 
     
     
         40 . The bicistronic construct of  claim 21 , further comprising a backbone derived from a virus.