Methods for engineering allogeneic and immunosuppressive resistant t cell for immunotherapy

A method for treating a cancer patient using immunotherapy comprising:

• providing primary human T cells from a single donor;
• co-electroporating into said primary human T-cells:
• (a) RNAs encoding two half TALE-nucleases that cleave a first gene, and
• (b) RNAs encoding two half TALE-nucleases that cleave a second gene,
• to generate a population of transfected human T cells comprising at least 15.5% doubly-inactivated human T-cells having both first and second genes inactivated;
• modifying the population of doubly inactivated human T-cells to express a chimeric antigen receptor;
• and
• infusing the population of doubly inactivated human T-cells into said cancer patient for immunotherapy treatment.

A method for treating a cancer patient using immunotherapy comprising:

• providing primary human T cells from a single donor;
• modifying the primary human T cells to express a chimeric antigen receptor;
• co-electroporating into said modified primary human T-cells:
• (a) RNAs encoding two half TALE-nucleases that cleave a first gene, and
• (b) RNAs encoding two half TALE-nucleases that cleave a second gene,
• to generate a population of transfected human T cells comprising at least 15.5% doubly-inactivated human T-cells having both first and second genes inactivated;
• and
• infusing the population of doubly inactivated human T-cells into said cancer patient for immunotherapy treatment.

A method for treating a cancer patient using immunotherapy comprising:

• providing a population of transfected human T cells from a single donor comprising at least 15.5% doubly-inactivated human T-cells having both first and second genes inactivated;
• wherein said cells are further modified to express a chimeric antigen receptor;
• and
• infusing the population of doubly inactivated human T-cells into said cancer patient for immunotherapy treatment.