A RETROVIRAL VECTOR THAT DIRECTS SIMULTANEOUS EXPRESSION OF ALPHA-T-CELL AND BETA-T-CELL RECEPTOR GENES

The transfer of alpha/beta T cell receptor (TCR) genes into T lymphocy tes or their precursors could provide a means to increase frequency of tumor- or pathogen-specific cytotoxic T lymphocytes. To begin to addr ess this possibility, we have used class I MHC-restricted alpha/beta T CR cDNAs to develop a retroviral TCR expression vector. alpha- and bet a-chain cDNAs were inserted into a derivative of the LN series of retr oviral vectors, with the retroviral LTR directing expression of TCR-be ta and an internal cytomegalovirus promoter/enhancer driving TCR-alpha expression. The variable region fragments can be replaced using uniqu e restriction sites that have been introduced into the proximal consta nt regions. We have used this vector system to transfer two different pairs of alpha/beta TCR genes into an alpha- and beta-chain-deficient T cell hybridoma, TCR- hybridoma cells were transduced by coculture wi th pools off virus-producing cells, and fluorescence-activated cell so rting was used to enrich for cells expressing the transduced TCR, Tran sduction with either alpha/beta TCR restores stable, long-lived expres sion of the alpha/beta TCR complex, TCR-mediated signal transduction i s also reconstituted, as demonstrated by the ability of transduced cel ls to secrete IL-2 following stimulation with V beta-specific antibodi es. Our results suggest that alpha/beta T cell receptor gene transfer could provide a basis for new approaches to immunotherapy, and that fu rther studies examining the in vivo fate of transduced TCR are possibl e.