Cultured dendritic cells can prime potent cytotoxic t lymphocyte responses to chronic myelogenous leukemia

Dendritic cells are potent antigen presenting cells, which are integral to the initiation of T cell immunity. The ability to culture these cells in vitro has lead to exciting developments in the area of immunotherapy. We undertook the following study to determine if we could induce CTL responses to CML tumor cells, and then examined ways to improve levels of cytotoxicity. Chronic myelogenous leukemia is one of the first hématologie diseases with a defined cytogenetic marker. Initially we generated DC from PBMC, that were collected for autologous transplantation, by culturing adherent cells in the presence of IL-4 and OM-CSF. After 7 days the resultant dendritic cells were harvested and utilized to prime T cells. In brief, PBMC were stimulated in the presence of dendritic cells pulsed with tumor cell lysate from a CML cell line (SK562). After 7 days the cells were restimulated with another round of dendritic cells and lysate. After a further 7 days the resultant primed cells were harvested, counted and utilized in an LDH based cytotoxicity assay. We found we could generate cytotoxicity (12-75%) at a high (10:1) effector (primed cells) to target (SK562 cells) cell ratio. We were able to improve the level and consistency of cytotoxicity (60-63%) at the same E:T ratios by utilizing either whole protein or RNA from the SK562 cell line as a source of antigen. We were also able to generate appreciable cytotoxicity at lower E:T ratios (25% at 1:1 ), and had improved specificity when the CTL were tested against malignant and non-malignant controls. To further enhance the level of cytotoxicity we tried to increase the immunostimulatory ability of dendritic cells by maturing them. Subsequent to pulsing the cells with tumor lysate they were matured over night in the presence of monocyte conditioned media. At at 1:1 ratio we increased cytotoxicity levels from undetectable to 30-50%. Together this data suggests that we can generate potent cytotoxic responses to CML cells, and that by manipulating the antigen or dendritic cells we can induce responses at very low E:T ratios. These studies are ongoing.