Endogenous and adoptively transferred A-NK and T-LAK cells continuously accumulate within murine metastases up to 48 h after inoculation

In murine models, therapeutic efficacy of adoptive immunotherapy (AIT) of cancer with lymphokine activated killer (LAK) cells is seen only when applied together with substantial doses of interleukin-2 (IL-2), probably because this cytokine is imperative for both motility and viability of the LAK cells. We wanted to investigate whether IL-2 in addition mediates an immunostimulatory activation and expansion of endogenous effector cells contributing to tumor regression. Using an immunoperoxidase technique, we have been able to longitudinally analyze the accumulation of tumor infiltrating lymphocytes expressing the pan-T cell/activated lymphocyte phenotype (Thy1.2), the natural killer (NK) cell phenotype (AsGM,) as well as the cytotoxic T (CD8) cell phenotype within experimental established B16 pulmonary melanoma metastases in C57BL/6 mice during the first 48 h after high dose IL-2 monotherapy. Whereas a substantial and selective infiltration of AsGM1+ lymphocytes in tumor tissue was seen (262 and 937 cells per sq.mm malignant tissue at 0 and 48 h, respectively), only a minor increase in accumulation of CD8+ cells was seen (106 and 171 cells per sq.mm tumor tissue at 0 and 48 h, respectively). The addition of adoptive transfer with lymphokine-activated adherent NK (A-NK) cells to the high-dose IL-2 treatment resulted in more than a 1.5 fold increase in infiltrating AsGM1+ cells compared to IL-2 therapy alone (1520 compared to 937 AsGM1+ cells per sq.mm malignant tissue). No substantial accumulation of CD8+ cells was observed in this setting either. In contrast, the treatment with high dose IL-2 together with adoptive transfer of mitogen-stimulated, lymphokine-activated T killer (T-LAK) cells increased the infiltration of CD8+ cells 10-fold compared to IL-2 monotherapy (2078 compared to 171 CD8+ cells per sq.mm malignant tissue, respectively). Interestingly, infiltration of both endogenous and exogenous cells continued over time, since the effector-to-tumor cell ratio in metastatic tissue dramatically increased from 1:8 and 1:6 at 16 h to 1:3 and 1:2 at 48 h after adoptive transfer of A-NK and T-LAK cells, respectively. These data underline the longevity of LAK cells in vivo and highlight the importance of IL-2 treatment in recruiting endogenous immune cells to tumor areas.