Integrated src kinase and costimulatory activity enhances signal transduction through single-chain chimeric receptors in T lymphocytes

Adoptive immunotherapy using receptor-modified T lymphocytes has shown prom ise in preclinical studies for the treatment of infectious and malignant di seases. These modified T cells express chimeric receptors that link ligand recognition and signal transduction domains in a single gene product. Typic ally, a single chain Fv fragment is genetically attached to the cytoplasmic domain of the T-cell receptor (TCR) zeta chain. Modulating the signaling c haracteristics of chimeric receptors will be important for their applicatio n to human immunotherapy. It was hypothesized that linking coreceptor and c ostimulatory signaling motifs together with the zeta signaling domain will enhance receptor function. The present study compares signaling characteris tics of 9 single-chain receptors consisting of the H-2K(b) extracellular an d transmembrane domains and various combinations of T cell signal transduct ion domains. Signal transduction regions studied include the TCR zeta chain , the CD4 coreceptor, the lck protein tyrosine kinase, and the CD28 costimu latory receptor. Biochemical characteristics of the receptors, analyzed usi ng calcium flux, receptor, and ZAP-70 phosphorylation, and lck association may be predicted from the known functions of receptor constituents. The com bination of zeta to gether with coreceptor and costimulatory function in a single receptor maximizes chimeric receptor sensitivity and potency. Combin ing zeta with either the costimulatory or coreceptor function independently also enhances receptor function, though to a lesser extent. It is therefor e possible to link TCR, coreceptor, and costimulatory activities in a singl e functional entity using modular domains. Such receptors demonstrate disti nct signaling properties and should prove useful in the development of chim eric receptors for therapeutic purposes. (C) 2001 by The American Society o f Hematology.