In vitro differentiation of T-cells capable of mediating the regression of established syngeneic tumors in mice.

Previous studies have shown that successful adoptive immunotherapy of a newly induced, weakly immunogenic murine sarcoma, MCA 105, can be achieved either with fresh noncultured immune spleen cells or with immune cells after in vitro stimulation and expansion. In this study, we utilized in vivo and in vitro depletions with monoclonal antibodies (mAb) of T-cell subpopulations expressing the L3T4 or Lyt-2 antigens to investigate the phenotype of the T-cells that mediate in vivo tumor regression. The efficiency of depletion was assessed by flow microfluorometric analysis and by the ability of specifically treated spleen cell populations to generate allogeneic cytotoxic T-lymphocytes. The therapeutic efficacy of adoptively transferred fresh noncultured MCA 105 immune cells was abrogated by in vivo administration of either L3T4 or Lyt-2 mAb to mice bearing 3-day established pulmonary metastases. In vitro treatment of fresh noncultured MCA 105 immune cells with either L3T4 or Lyt-2 mAb and complement also abrogated their antitumor efficacy confirming the initial findings. However, mixing L3T4 and Lyt-2 mAb and complement-treated MCA 105 immune cells reconstituted the antitumor efficacy indicating that cellular cooperation between these two lymphoid subpopulations was essential for the regression of established tumors. Unlike fresh noncultured immune cells, the antitumor efficacy of in vitro sensitized and expanded immune cells was abrogated by in vivo treatment with Lyt-2 but not with L3T4 mAb indicating Lyt-2+ cells alone played a major role in mediating the regression of tumors. These findings provide evidence for an in vitro-induced differentiation of therapeutic T-lymphocytes. Our results thus suggest that the antitumor activities expressed by the two types of cells may represent T-cells at different stages of immunological differentiation.