Systems and methods for protein production

Abstract

The invention relates to systems and methods for producing proteins of interest. The invention employs genetically-engineered animal or plant cells that have modified protein folding or processing capacities. In one aspect, the invention features genetically-engineered cells comprising one or more recombinant expression cassettes which encode (1) a protein of interest and (2) a polypeptide that is functional in the unfolded protein response (UPR) pathway of the cells. Co-expression of the polypeptide significantly increases the yield of the protein of interest in the genetically-engineered cells. In one example, the genetically-engineered cells are animal cells, and the co-expressed polypeptide is a component or modulator of an XBP1- or ATF6-mediated UPR pathway.

1 . An animal or plant cell comprising one or more recombinant expression cassettes which encode:
 a protein of interest, and   a component or non-IRE1 modulator of an unfolded protein response (UPR) pathway.   
     
     
         2 . The cell according to  claim 1 , wherein said cell is an animal cell, and said component or non-IRE1 modulator is an XBP1 or ATF6 protein. 
     
     
         3 . The cell according to  claim 2 , wherein said one or more recombinant expression cassettes include:
 at least one recombinant expression cassette encoding said protein of interest; and   at least another recombinant expression cassette encoding said XBP1 or ATF6 protein,   
       and wherein the ratio of the total number of said at least one recombinant expression cassette to the total number of said at least another recombinant expression cassette in said cell is from 0.5:1 to 10:1. 
     
     
         4 . The cell according to  claim 3 , wherein the ratio of the total number of said at least one recombinant expression cassette to the total number of said at least another recombinant expression cassette in said cell is at least 3:1. 
     
     
         5 . The cell according to  claim 3 , wherein said cell is a mammalian cell. 
     
     
         6 . A transgenic, non-human animal comprising the cell of  claim 3 . 
     
     
         7 . The cell according to  claim 2 , wherein said cell is a hybrid cell produced by fusing an animal cell and a cancer cell. 
     
     
         8 . An animal or plant cell comprising one or more recombinant expression cassettes which encode:
 a protein of interest; and   a polypeptide capable of binding to a UPR element (UPRE) or ER-stress response element (ERSE) of said cell.   
     
     
         9 . The cell according to  claim 8 , wherein said cell is an animal cell, and said polypeptide includes a transcription activation domain. 
     
     
         10 . The cell according to  claim 9 , wherein said transcription activation domain comprises a transcription activation domain of an XBP1 or ATF6 protein. 
     
     
         11 . An animal or plant cell culture transfected or transduced with one or more expression vectors encoding:
 a protein of interest; and   a component or non-IRE1 modulator of a UPR pathway.   
     
     
         12 . The cell culture according to  claim 11 , wherein said component or non-IRE1 modulator is an XBP1 or ATF6 protein, and said one or more expression vectors include:
 at least one recombinant expression cassette encoding said protein of interest; and   at least another recombinant expression cassette encoding said XBP1 or ATF6 protein,   
       and wherein the molar ratio of said at least one recombinant expression cassette over said at least another recombinant expression cassette in said cell culture is from 0.5:1 to 10:1. 
     
     
         13 . The cell culture according to  claim 12 , wherein said cell culture is a mammalian cell culture, and the molar ratio of said at least one recombinant expression cassette over said at least another recombinant expression cassette in said cell culture is at least 3:1. 
     
     
         14 . A method comprising expressing a protein of interest in an animal or plant cell, said cell including one or more recombinant expression cassettes which encode:
 said protein of interest; and   a component or non-IRE1 modulator of a UPR pathway.   
     
     
         15 . The method according to  claim 14 , wherein said cell is an animal cell, and said component or non-IRE1 modulator is an XBP1 or ATF6 protein. 
     
     
         16 . A method comprising expressing a protein of interest in an animal or plant cell, said cell including one or more recombinant expression cassettes which encode:
 said protein of interest; and   a polypeptide capable of binding to a UPRE or ERSE of said cell.   
     
     
         17 . The method according to  claim 16 , wherein said cell is an animal cell, and said polypeptide comprises a transcription activation domain of an XBP1 or ATF6 protein. 
     
     
         18 . A method comprising expressing a protein of interest in an animal or plant cell culture, said cell culture being transfected or transduced with one or more expression vectors which include:
 at least one recombinant expression cassette encoding said protein of interest; and   at least another recombinant expression cassette encoding a component or non-IRE1 modulator of a UPR pathway.   
     
     
         19 . The method according to  claim 18 , wherein said cell culture is a mammalian cell culture, and said component or non-IRE1 modulator is an XBP1 or ATF6 protein, and wherein the molar ratio of said at least one recombinant expression cassette over said at least another recombinant expression cassette in said cell culture is from 0.5:1 to 10:1. 
     
     
         20 . An expression vector encoding:
 a therapeutic or prophylactic protein of interest, and   a component or non-IRE 1 modulator of a UPR pathway.