Chimeric anti-TAG72 receptors with immunoglobulin constant Fc domains and gamma or zeta signalling chains.

We recently described the generation and expression of a chimeric T cell receptor with specificity for the tumor antigen TAG72 consisting of the single chain antibody (scFv) B72.3-scFv and the gamma chain of the FcepsilonRI receptor. The corresponding chimeric receptor containing the zeta chain of the TCR as signalling unit is not functionally expressed reflecting that the requirements for functional expression of chimeric receptors containing the gamma signalling chain are apparently different compared to those containing the CD3zeta signalling chain of the TCR. We describe a novel set of chimeric anti-TAG72 receptors including in their extracellular moiety the constant immunoglobulin CH2/3 domains that allow stable expression of chimeric gamma as well as zeta receptors. We designed anti-TAG72 receptors that consist of a scFv fragment derived from an anti-TAG72 second generation antibody (CC49) and of the CH2/3 domains of the human IgG and intracellularily either of the zeta or gamma signalling chain. The recombinant CC49-CH2/3-zeta and CC49-CH2/3-gamma DNA, respectively, was transfected into MD45 T cells and expressed under control of the RSV LTR. Both receptors were found on the cell membrane of transfected cells as demonstrated by flow cytometry analysis using an anti-human IgG Fc antibody directed to the CH2/3 immunoglobulin domains of the chimeric receptor. Specific cross-linking of the chimeric zeta as well as the gamma receptor by antigen or anti-human Ig antibodies resulted in specific activation of transfected cells. Our results demonstrate that both the gamma chain and the zeta chain++ containing receptor are stably expressed and convert T cells to specificity for the TAG72 antigen. This receptor design will facilitate efficient generation of genetically modified peripheral T cells and may provide valuable tools for the cellular immunotherapy of TAG72+ tumors.