Targeted delivery to tumors of cytokine genes capable of initiating anti-tumor immune responses is a promising approach for cancer therapy. However, success has been limited due to the lack of efficient methods to ensure gene-expression selectively in established tumors. We have recently shown that adoptively transferred, IL2-activated natural killer (A-NK) cells and PHA- and IL-2-stimulated CD8+ T lymphocytes (called T-LAK cells) localize into tumor with impressive selectively. Our goal is to take advantage of this skill, and demonstrate that cytokine gene-transduced, tumor-seeking A-NK and/or T-LAK cells can be used as vehicles for the delivery of anti-cancer cytokines selectively to tumors and to show that this forced cytokine expression in the tumors will lead to strong anti-tumor immune responses. The SPECIFIC AIMS of this 3-year project are to: 1) Demonstrate that A-N K cells can be transduced with adenoviral vectors to produce significant amounts of cytokines; 2) Analyze the ability of cytokine gene-transduced T-LAK and A-NK cells to localize into tumors and to produce cytokines at the tumor site, and; 3) Demonstrate that adoptive transfer of cytokine gene-transduced A-NK and T-LAK cells leads to elimination of tumors without induction of systemic toxicity. To accomplish this, T-LAK and A-NK cells will be adeno- or retro-virally transduced with one or several genes encoding for cytokines/chemokines known to have anti tumor effect via stimulation of either NK cells, DCs, and T helper cells and CTLs, namely IL-2, IL-4, IL12, IL-15, IL-18, IFNg, GM-CSF, RANTES and lymphotactin. The ability of successfully transduced T-LAK and A-NK cells to localize into tumors and to produce cytokine at the tumor site will be tested in animal tumor models. The host response, such as increased accumulation of effector cells into the tumor tissue, induced by the intratumorally produced cytokines, will be measured. The therapeutic potential of this approach will be evaluated by analyzing tumor-reduction resulting from injection of batches of A-NK and/or T-LAK cells, each capable of producing one or several anti-cancer cytokines. The mechanism behind any anti-tumor effect observed, will be elucidated. Depending on the cytokines produced by the tumor-seeking A-NK and T-LAK cells, we anticipate that we can manipulate the host immune system to elicit strong innate and adaptive anti-tumor responses without induction of toxic side effects. On this background, it should be possible to substantially improve the efficacy of tumor-targeted cytokine gene-therapy of cancer.