Materials and methods for engineering cells and uses thereof in immuno-oncology
AssigneeCrispr Therapeutics AG
InventorsJonathan Alexander Terrett, Demetrios Kalaitzidis, Lawrence Klein
Life of this patent
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Abstract
Materials and methods for producing genome-edited cells engineered to express a chimeric antigen receptor (CAR) construct on the cell surface, and materials and methods for genome editing to modulate the expression, function, or activity of one or more immuno-oncology related genes in a cell, and materials and methods for treating a patient using the genome-edited engineered cells.
Claims25
An engineered T cell, comprising:
- (a) a disrupted T cell receptor alpha chain constant region (TRAC) gene and
- (b) a nucleic acid comprising a nucleotide sequence that encodes a chimeric antigen receptor (CAR);
- wherein the disrupted TRAC gene comprises a deletion of the nucleotide sequence of SEQ ID NO:
- 76, and wherein the nucleic acid of (b) is inserted in the disrupted TRAC gene.
A population of cells, comprising genetically engineered human T cells, wherein the engineered human T cells comprise:
- (a) a disrupted T cell receptor alpha chain constant region (TRAC) gene comprising a deletion of the nucleotide sequence of SEQ ID NO:76;
- and
- (b) a nucleic acid comprising a nucleotide sequence that encodes a chimeric antigen receptor (CAR);
- wherein the nucleic acid of (b) is inserted in the disrupted TRAC gene.
A method for producing a genetically engineered T cell population, the method comprising
- (a) delivering to a population of T cells
- (i) an RNA-guided nuclease and a gRNA targeting a site in a TRAC gene locus (TRAC gRNA), wherein the site in the TRAC gene locus contains the nucleotide sequence of SEQ ID NO:76;
- and
- (ii) a donor template comprising a first segment that is homologous to the TRAC gene locus left of the site targeted by the TRAC gRNA, a second segment comprising a nucleotide sequence encoding a chimeric antigen receptor (CAR), and a third segment that is homologous to the TRAC gene locus right to the site targeted by the TRAC gRNA;
- wherein homologous recombination of the donor template into the TRAC gene results in the insertion of the second segment encoding the CAR and a deletion and/or mutation of the site targeted by the TRAC gRNA in the TRAC gene;
- and
- (b) producing a genetically engineered population of T cells.
Dependencies are inferred from the claim text.