Several different experimental systems have been used to study the regulation of T-cell activation and death. To understand the complex nuclear events that follow TCR ligation, a relatively ne approach has been taken in which a transcription factor is targeted directly with a dominant negative inhibitor. We have examined the effect of a dominant negative variant of c-Jun to prob the AP-1-dependent nuclear interactions that regulate IL-2 gene transcription. The mutant Jun was potent inhibitor of endogenous AP-1 and efficiently blocked IL-2 gene transcription. Interestingly, the majority of this inhibition was found to be due to interference with the important lymphocyte specific transcription factor NF-AT. Programmed cell death (PCD) has been observed in a variety of cell types in response to physiologic signals and cytotoxic agents. How these stimuli trigger PCD, and whether there is a common PCD signal transduction pathway, is not clear. To study the role of different gene products, we have developed a simple transient transfection death assay in which particular proteins can be manipulated and the effects on PCD can be quantitated. In a T-cell hybridoma, transient expression of Bcl-2 confers protection from cytotoxic agent-induced PCD. Bcl-2 is also being examined in the NCI anticancer cell line panel to search for compounds that exhibit Bcl-2 selective toxicity. A major limitation of immune therapy for cancer is the very low frequency of cells specifically reactive with a patient's tumor cells. Bispecific antibodies (BiMabs) which simultaneously bind the T-cell receptor and a tumor antigen can potentially target all normal T cells and lead to potent cytotoxicity. We have developed a completely syngeneic mouse breast cancer model to study a molecular engineered BiMab directed against the mouse TCR and a viral glycoprotein expressed on murine breast cancer cells. This BiMab has excellent in vitro cytotoxic activity and can prevent the growth of the breast cancer in vivo. In anticipation of clinical trials, we are exploring several important issues for BiMabs: the biodistribution of the BiMab, the nature of the inflammatory cell infiltrate, and the effectiveness of this therapy in treating established tumors.
