Interleukin-12 (IL-12) is a heterodimeric cytokine originally defined by its ability to induce the maturation of cytolytic lymphocytes and by its capacity to effectively synergize with IL-2 in the induction of cytolytic activity. IL-12 activates NK cells, facilitates the generation of antigen-specific cytolytic T lymphocytes (CTL), maintains tumor infiltrating lymphocytes (TIL) in culture, and induces the secretion of high levels of IFN-tau (required early in the immune response) from both T cells and NK cells in vitro and in vivo. While IL-02 may promote similar functions, IL-12 is effective at 10-1000 fold lower molar concentrations than IL-2. Since IL-12 is (apparently) secreted exclusively by professional antigen presenting cells (APC); monocytes and B cells) and IL-12 receptors are expressed selectively on activated T lymphocytes and NK cells, it appears that IL-12 is well-suited to drive the expansion and maturation of the two most prevalent anti-tumor effector cells localized within tumor lesions (T cells and NK cells). Based on our previous and current observations using in vivo murine models, we believe that IL-12 is an ideal candidate for use in the immunotherapy of cancer. In particular, we believe that IL-12 therapy will be optimally effective when administered through gene transfer in a paracrine (local) manner, either alone or in conjunction with additional costimulator molecule gene therapy. As a basis for future IL-12 clinical trials we propose the following specific aims: l. ESTABLISH EFFICIENT POLYCISTRONIC GENE TRANSFER SYSTEMS FOR PRODUCING TRANSFECTANTS SECRETING BIOLOGICALLY ACTIVE IL-12 AND EXPRESSING T-12 COSTIMULATORY MOLECULES (i.e. IL-2, B7.1, B7.2). Tumor cells, fibroblasts, and antigen presenting cells (dendritic cells or the RMA-S cell line) will be transduced with novel retroviral vectors., 2. EXAMINE THE ABILITY OF PARACRINE SECRETION OF IL-12, ALONE OR IN CONJUNCTION WITH COEXPRESSED COSTIMULATORY MOLECULES, TO ENHANCE ANTI- TUMOR IMMUNE RESPONSES IN VIVO. Engineered tumors or engineered fibroblasts admixed with tumor will be evaluated for their capacity to elicit anti-tumor immunity in vivo., 3. EXAMINE THE ADJUVANTICITY OF IL-12 ENGINEERED ANTIGEN PRESENTING CELLS IN PROMOTING ANTI-TUMOR CELLULAR IMMUNE RESPONSES. Engineered antigen presenting cells (dendritic cells or RMA-S) will be evaluated for their capacity to elicit tumor peptide- specific CTL in vitro and in vivo., 4. DEVELOP CLINICAL TRIALS UTILIZING IL-12 GENE THERAPY. We will develop vectors encoding IL-12 or costimulator molecules for clinical trials in the Human Gene Therapy Applications Laboratory (Core D) and prepare a clinical protocol based on the results obtained in Specific Aims 1-3.
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