Engineered cells for adoptive cell therapy

Abstract

Provided are engineered cells for adoptive therapy, including NK cells and T cells. Also provided are compositions for engineering and producing the cells, compositions containing the cells, and methods for their administration to subjects. In some aspects, features of the cells and methods provide specificity and/or efficacy. In some embodiments, the cells contain genetically engineered antigen receptors that specifically bind to antigens, such as chimeric antigen receptors (CARs) and costimulatory receptors. In some embodiments, the cells include receptors targeting multiple antigens. In some embodiments, the cells include repression of one or more gene product, for example, by disruption of a gene encoding the gene product. In some embodiments, a gene encoding an antigen recognized by the engineered antigen receptor is disrupted, reducing the likelihood of targeting of the engineered cells.

1 . An engineered immune cell comprising:
 a genetically engineered antigen receptor that specifically binds to a target antigen, wherein the target antigen is selected from the group consisting of CD26, CD92, CD148, CD150, CD261, CD262, CD362, mouse double minute 2 homolog (MDM2), cytochrome P450 1B1 (CYP1B), livin, mucin 16 (MUC16), and cyclin (D1); and   a genetic disruption resulting in reduced expression of the target antigen in the engineered immune cell.   
     
     
         2 . The engineered immune cell of  claim 1 , wherein the genetic disruption comprises a disruption in a gene encoding the target antigen. 
     
     
         3 . The engineered immune cell of  claim 1 , wherein the genetically engineered antigen receptor is a chimeric antigen receptor (CAR). 
     
     
         4 . The engineered immune cell of  claim 3 , wherein the CAR comprises an extracellular antigen-recognition domain that specifically binds to the target antigen. 
     
     
         5 . The engineered immune cell of  claim 3 , wherein the CAR comprises an extracellular antigen-recognition domain that specifically binds to a peptide of the target antigen in the context of a major histocompatibility complex (MHC) molecule. 
     
     
         6 . The engineered immune cell of  claim 3 , wherein the CAR comprises an intracellular signaling domain comprising a CD3-zeta (CD3ζ) chain. 
     
     
         7 . The engineered immune cell of  claim 6 , wherein the CAR further comprises a costimulatory signaling region. 
     
     
         8 . The engineered immune cell of  claim 7 , wherein the costimulatory signaling region comprises a signaling domain of a CD28. 
     
     
         9 . The engineered immune cell of  claim 7 , wherein the costimulatory signaling region comprises a signaling domain of a 4-1BB. 
     
     
         10 . The engineered immune cell of  claim 1 , wherein the genetically engineered antigen receptor is a T cell receptor (TCR). 
     
     
         11 . The engineered immune cell of  claim 1 , further comprising another genetically engineered antigen receptor, which is a chimeric costimulatory receptor that specifically binds to another antigen and is capable of inducing a costimulatory signal to the cell. 
     
     
         12 . The engineered immune cell of  claim 6 , further comprising another genetically engineered antigen receptor, which is a chimeric costimulatory receptor that specifically binds to another antigen and is capable of inducing a costimulatory signal to the cell. 
     
     
         13 . The engineered immune cell of  claim 1 , wherein the genetically engineered antigen receptor is capable of inducing an inhibitory or immunosuppressive or repressive signal to the cell upon recognition of the target antigen. 
     
     
         14 . The engineered immune cell of  claim 13 , wherein the genetically engineered antigen receptor is a chimeric antigen receptor (CAR) and the CAR comprises an extracellular antigen-recognition domain that specifically binds to the target antigen and an intracellular signaling domain that comprises a signaling portion of an immune checkpoint molecule. 
     
     
         15 . The engineered immune cell of  claim 13 , wherein the antigen receptor is a first genetically engineered antigen receptor, the target antigen is a first target antigen, and the immune cell further comprises a second genetically engineered antigen receptor that recognizes an antigen expressed on a disease or condition to be treated and induces a stimulatory or activating signal, which stimulatory or activating signal is dampened by a signal induced by the inhibitory or immunosuppressive or repressive signal induced by the first genetically engineered antigen receptor. 
     
     
         16 . The engineered immune cell of  claim 1 , wherein the immune cell is a T cell or an NK cell. 
     
     
         17 . The engineered immune cell of  claim 6 , wherein the immune cell is a T cell or an NK cell. 
     
     
         18 . A method of treatment, comprising administering to a subject having a disease or condition the engineered immune cell of  claim 1 . 
     
     
         19 . A method of treatment, comprising administering to a subject having a disease or condition the engineered immune cell of  claim 6 . 
     
     
         20 . A method of producing a genetically engineered immune cell, comprising:
 (a) introducing into an immune cell a genetically engineered antigen receptor that specifically binds to a target antigen, wherein the target antigen is selected from the group consisting of Wilms' tumor gene 1 (WT1), CD26, CD92, CD148, CD150, CD261, CD262, CD362, mouse double minute 2 homolog (MDM2), cytochrome P450 1B1 (CYP1B), livin, mucin 16 (MUC16), and cyclin (D1); and   (b) effecting a genetic disruption resulting in reduced expression of the target antigen in the immune cell, thereby producing a genetically engineered immune cell in which expression of the target antigen is reduced,   wherein steps (a) and (b) are carried out simultaneously or sequentially in any order.   
     
     
         21 . The method of  claim 20 , wherein the effecting the genetic disruption comprises disrupting a gene encoding the target antigen. 
     
     
         22 . The method of  claim 21 , wherein:
 the disrupting comprises introducing into the immune cell a DNA binding protein or DNA-binding nucleic acid that specifically binds to or hybridizes to the gene; or   the disrupting comprises introducing: (a) a fusion protein comprising a DNA-targeting protein and a nuclease or (b) an RNA-guided nuclease.   
     
     
         23 . The method of  claim 22 , wherein effecting the genetic disruption comprises introducing a zinc finger nuclease (ZFN), a TAL-effector nuclease (TALEN), or and a CRISPR-Cas9 combination that specifically binds to, recognizes, or hybridizes to the gene. 
     
     
         24 . The method of  claim 20 , further comprising:
 (c) introducing into the immune cell another genetically engineered antigen receptor, which is a chimeric costimulatory receptor that specifically binds to another antigen and is capable of inducing a costimulatory signal to the cell, wherein steps (a), (b) and (c) are carried out simultaneously or sequentially in any order.   
     
     
         25 . The method of  claim 20 , wherein the genetically engineered antigen receptor is a chimeric antigen receptor (CAR). 
     
     
         26 . The method of  claim 20 , wherein the genetically engineered antigen receptor is a T cell receptor (TCR). 
     
     
         27 . The method of  claim 20 , wherein the immune cell is a T cell. 
     
     
         28 . A cell produced by the method of  claim 20 . 
     
     
         29 . An engineered immune cell, comprising:
 (a) a first genetically engineered antigen receptor, which specifically binds to a first antigen and is capable of inducing an activating signal to the cell; and   (b) a second genetically engineered antigen receptor, which is a chimeric costimulatory receptor that specifically binds to a second antigen and is capable of inducing a costimulatory signal to the cell,   wherein the first and second antigens are distinct and at least one of the first or the second antigen is selected from the group consisting of Wilms' tumor gene 1 (WT1), CD26, CD92, CD148, CD150, CD261, CD262, CD362, mouse double minute 2 homolog (MDM2), cytochrome P450 1B1 (CYP1B), livin, mucin 16 (MUC16), and cyclin (D1).   
     
     
         30 . The engineered immune cell of  claim 29 , wherein:
 the first genetically engineered antigen receptor comprises an ITAM-containing sequence or comprises an intracellular signaling domain of a CD3-zeta (CD3ζ) chain; and/or   the first genetically engineered antigen receptor does not comprise a signaling domain from a T cell costimulatory molecule.   
     
     
         31 . The engineered immune cell of  claim 29 , wherein the costimulatory receptor comprises an intracellular signaling domain of a T cell costimulatory molecule. 
     
     
         32 . The engineered immune cell of  claim 29 , further comprising a genetic disruption in a gene encoding the first antigen, and/or in a gene encoding the second antigen, said genetic disruption resulting in reduced expression of the first and/or second antigen in the engineered immune cell. 
     
     
         33 . The engineered immune cell of  claim 29 , wherein the first genetically engineered antigen receptor is a chimeric antigen receptor (CAR). 
     
     
         34 . The engineered immune cell of  claim 29 , wherein the first genetically engineered antigen receptor is a T cell receptor (TCR). 
     
     
         35 . The engineered immune cell of  claim 29 , wherein the immune cell is a T cell. 
     
     
         36 . A method of treatment, comprising administering to a subject having a disease or condition the engineered immune cell of  claim 29 . 
     
     
         37 . A method of producing an engineered immune cell, the method comprising:
 (a) introducing into an immune cell a first genetically engineered antigen receptor that specifically binds to a first antigen; and   (b) introducing into the immune cell a second genetically engineered antigen receptor which is a chimeric costimulatory receptor that specifically binds to a second antigen,   thereby producing the engineered immune cell, wherein:
 said first and second antigens are distinct at least the first or the second antigen is selected from the group consisting of Wilms' tumor gene 1 (WT1), CD26, CD92, CD148, CD150, CD261, CD262, CD362, mouse double minute 2 homolog (MDM2), cytochrome P450 1B1 (CYP1B), livin, mucin 16 (MUC16), and cyclin (D1); and 
 (a) and (b) are carried out simultaneously or sequentially in any order. 
   
     
     
         38 . The method of  claim 37 , further comprising (c) disrupting a gene encoding the first antigen and/or disrupting a gene encoding the second antigen, thereby resulting in reduced expression of the first and/or second antigen in the immune cell. 
     
     
         39 . The method of  claim 38 , wherein:
 the disrupting comprises introducing into the immune cell a DNA binding protein or DNA-binding nucleic acid that specifically binds to or hybridizes to the gene; or   the disrupting comprises introducing: (a) a fusion protein comprising a DNA-targeting protein and a nuclease or (b) an RNA-guided nuclease.   
     
     
         40 . The method of  claim 38 , wherein the disrupting comprises introducing a zinc finger nuclease (ZFN), a TAL-effector nuclease (TALEN), or and a CRISPR-Cas9 combination that specifically binds to, recognizes, or hybridizes to the gene. 
     
     
         41 . A cell produced by the method of  claim 37 .