Generation of Human Anti-Leukemia CTL Vaccine Utilizing Dendritic Cell Hybrids.

Abstract The therapeutic options for patients with hematological malignancies that relapse after autologous or allogeneic hematopoietic stem cell transplantation (SCT) are limited and their overall prognosis is poor. Therefore new therapeutic approaches such as cellular-based immunotherapies are being investigated. The efficacy of cytotoxic T lymphocytes (CTLs) or dendritic cell (DC) vaccines was already established in the treatment of post transplant CMV infections and in several solid tumors. This strategy however is less effective in hematological malignancies, where the tumor specific antigens are unknown and the cellular immunotherapy is based on the induction of graft-versus-leukemia (GVL) response by allogeneic, non-antigen specific, donor derived T lymphocytes (DLI). One approach to enhance immune recognition and targeting of the leukemic cells, and consequently augment their specific killing by CTLs, is through the use of dendritic cells. DCs are potent antigen presenting cells that can be manipulated to express tumor antigens by various strategies including fusion with tumor cells. Such hybrid cells expressing tumor antigens, presented in the context of MHC class I and II and co-stimulatory molecules of the DC partner, were shown to be effective in several malignancies. Our aim was to generate human CTLs capable to mount a specific and potent in-vitro anti-leukemia activity, utilizing dendritic-leukemia cell hybrids. We have generated such hybrids by co-culture of immature human DCs with allogeneic (K562) or autologous leukemia cells in the presence of polyethylene glycol (PEG). After maturation, these dendritic-leukemia cell hybrids were utilized to generate specific CTLs, using autologous or allogeneic peripheral blood lymphocytes under IL-2 deprivation. Purified CD8+ cells were then further expanded and tested for their cytotoxicity against the target cells using Alamar Blue cytotoxicity assay. Both allogeneic and autologous dendritic-leukemia cell hybrids were shown to stimulate a significant CTL responses compared to controls. Dendritic-K562 cell hybrids stimulated a CTL response of 40% lysis of the target K562 cells, while CTLs generated by co-culture with non-fused K562 cells had no detectable cytotoxic activity against their targets. Similarly, CTLs stimulated by dendritic-autologous leukemia cell hybrids showed 15% lysis of the leukemia target cells, compared to 1.5 and 1% lysis observed by CTLs generated by co-culture with autologous tumor or DCs alone. Our results show that in-vitro generation of hybrids of human dendritic and un-manipulated primary leukemia cells is feasible, and that such hybrid cells can elicit the generation of specific autologous or allogeneic anti-leukemia CTLs. Such CTLs may be superior to the currently used non-antigen-specific donor lymphocytes, and may be useful also in the autologous setting.