Engineered Cells Expressing Multiple Immunomodulators and Uses Thereof

Abstract

This invention relates to the field of therapeutics. Most specifically invention provides methods of generating in vitro engineered immune cells conditionally expressing interleukin-12 (IL-12) and one or more immunomodulators under the control of a gene expression modulation system in the presence of activating ligand and uses for therapeutic purposes in animals.

1 . A vector for conditionally expressing proteins having the functions of one or more immunomodulators comprising a polynucleotide encoding a gene switch, wherein said polynucleotide comprises (1) at least one transcription factor sequence which is operably linked to a promoter, wherein said at least one transcription factor sequence encodes a ligand-dependent transcription factor, and (2) a polynucleotide operably linked to a different promoter which is activated by said ligand-dependent transcription factor, the polynucleotide encoding an IL-12 polypeptide at least 90% identical to IL-12, and one or and one or more immunomodulator polypeptides at least 90% identical to a protein selected from PD-LI and CTLA4. 
     
     
         2 . The vector of  claim 1 , which is a viral vector. 
     
     
         3 . The vector of  claim 2 , wherein the viral vector is a lentiviral vector. 
     
     
         4 . The vector of  claim 1 , wherein said polynucleotide encoding said immunomodulator polypeptide or said polynucleotide encoding said IL-12 polypeptide is under control of a constitutive promoter. 
     
     
         5 . The vector of  claim 1 , wherein said polynucleotide encoding said immunomodulator polypeptide and said polynucleotide encoding said IL-12 polypeptide are under control of orthogonal gene switch promoters. 
     
     
         6 . The vector of  claim 1 , wherein said gene switch is an ecdysone receptor (EcR)-based gene switch. 
     
     
         7 . The vector of  claim 1 , wherein said polynucleotide encoding a gene switch comprises a first transcription factor sequence under the control of a first promoter and a second transcription factor sequence under the control of a second promoter, wherein the proteins encoded by said first transcription factor sequence and said second transcription factor sequence interact to form a protein complex which functions as a ligand-dependent transcription factor. 
     
     
         8 . A method of producing a population of immune cells or therapy support cells (TSC) comprising modifying the immune cells with the recombinant vector according to  claim 1 . 
     
     
         9 . The method of  claim 8 , wherein said cells are human dendritic cells. 
     
     
         10 . The method of  claim 9 , wherein said dendritic cells are bone marrow dendritic cells. 
     
     
         11 . A population of immune cells or TSC expressing proteins having the function of one or more immunomodulators, which have been modified with a recombinant vector according to  claim 1 . 
     
     
         12 . The population of  claim 11 , wherein said cells are human dendritic cells. 
     
     
         13 . The population of  claim 12 , wherein said dendritic cells are bone marrow dendritic cells. 
     
     
         14 . An in vitro engineered immune cell or a TSC comprising a vector according to  claim 1 . 
     
     
         15 . The in vitro engineered immune cell of  claim 14 , wherein said immune cell is a human dendritic cell. 
     
     
         16 . The in vitro engineered immune cell of  claim 15 , wherein said dendritic cell is a bone marrow dendritic cell. 
     
     
         17 . A pharmaceutical composition comprising the population of in vitro engineered immune cells or TSC according to  claim 11 . 
     
     
         18 . A pharmaceutical composition comprising the in vitro engineered immune cell or the TSC of  claim 14 . 
     
     
         19 - 21 . (canceled) 
     
     
         22 . A method for treating a tumor in a mammal, comprising:
 (a) administering intratumorally to tumor microenvironments the population of immune cells or TSC of  claim 11 ; and   (b) administering to said mammal a therapeutically effective amount of one or more activating ligands.   
     
     
         23 . The method of  claim 22 , wherein said tumor is a benign tumor. 
     
     
         24 . The method of  claim 22 , wherein said tumor is a malignant tumor. 
     
     
         25 . The method of  claim 22 , wherein said tumor is a glioma. 
     
     
         26 . The method of  claim 22 , wherein said tumor is a glioblastoma. 
     
     
         27 . The method of  claim 22 , wherein said in vitro engineered immune cells comprise a vector comprising a polynucleotide encoding a gene switch, wherein said polynucleotide comprises (1) at least one transcription factor sequence operably linked to a promoter, wherein said at least one transcription factor sequence encodes a ligand-dependent transcription factor, and (2) a polynucleotide encoding a protein having the function of the immunomodulator linked to a promoter which is activated by said ligand-dependent transcription factor. 
     
     
         28 - 39 . (canceled)