SUMMARYEvidence of an immune mediated graft-versus-leukaemia effect has led to the belief that T and NK cell based adoptive immunotherapy can constitute effective treatment for relapsed leukaemias. However, work on solid tumours has shown this strategy may be hampered, by an immune escape mechanism in which tumour secreted immunosuppressive factors compromise T and NK cell function. Indeed, acute myeloid leukaemia (AML) cells secrete immunosuppressive factors that block the synthesis of Th1 type cytokines in T cells. We demonstrate here that this immunosuppression, mediated by both HL60 AML cell line and primary AML blasts, inhibits T and NK cell proliferation but not cytolytic activity. Supernatants from HL60 cell line and primary AML blasts inhibited T cell proliferation to mitogenic and alloantigen stimulation but had no effect on cytolytic function. Similarly, the proliferation of NK cells to IL-2 and IL-15 stimulation was inhibited whilst their cytolytic function, shown by lysis of AML blasts, K562 and Daudi cells remained unaffected. The failure of T and NK cells to proliferate was not due to effector cell apoptosis. Indeed, removal of lymphocytes from the immunosuppressive environment partially restored their capacity to respond to mitogenic stimulation. T cells exposed to immunosuppressive supernatants did not increase expression of mitotic inhibitory proteins that arrest cell division, thereby ruling this out as a mechanism of operation for this immunosuppression. T cell expansion requires antigen stimulation, usually provided in the form of AML blasts, therefore our data suggest that NK cells may be more practical for the immunotherapy of AML.

SUMMARYWhole tumour cells are a logical basis for generating immunity against the cancers they comprise or represent. A number of human trials have been initiated using cytokine-transfected whole tumour cells of autologous (patient-derived) or allogeneic [major histocompatibility complex (MHC)-disparate] origin as vaccines. Although precedent exists for the ef®cacy of autologous-transfected cell vaccines in animal models, little preclinical evidence con®rms that these ®ndings will extrapolate to allogeneic-transfected cell vaccines. In order to address this issue a murine melanoma cell line (K1735) was transfected to secrete interleukin (IL)-2, IL-4, IL-7 or granulocyte±macrophage colony-stimulating factor (GM-CSF); cytokines currently in use in trials. The ef®cacy of these cells as irradiated vaccines was tested head-to-head in syngeneic (C3H) mice and in MHC-disparate (C57BL/6) mice, the former being subsequently challenged with K1735 cells and the latter with naturally cross-reactive B16-F10 melanoma cells. Whilst the GM-CSF-secreting vaccine was the most effective at generating protection in C3H mice, little enhancement in protection above the wild-type vaccine was seen with any of the transfections for the allogeneic vaccines, even though the wild-type vaccine was more effective than the autologous B16-F10 vaccine. Anti-tumour cytotoxic T-lymphocyte (CTL) activity was detected in both models but did not correlate well with protection, whilst in vitro anti-tumour interferon-c (IFN-c) secretion tended to be higher following the GM-CSF-secreting vaccine. Cytokine transfection of vaccines generally increased anti-tumour CTL activity and IFN-c secretion (T helper type 1 response). Further studies in other model systems are required to con®rm this apparent lack of bene®t of cytokine transduction over wild-type allogeneic vaccines, and to determine which in vitro assays will correlate best with protection in vivo.

SUMMARYThe existence of an immune based graft-versus -leukaemia (GvL) effect highlighted the prospect of managing relapsed leukaemias with T cell-based adoptive immunotherapy. Thus, various strategies have been explored for the in vitro expansion of acute myeloid leukaemia (AML)-specific T cells. In a popular approach, AML blasts have been genetically modified to express co-stimulatory molecules essential for effective T cell priming. One such tactic has been the modification of AML cells to express the B7/ CD80 co-stimulatory molecule that binds to CD28 on T cells initiating events that culminate in enhanced cytokine production, proliferation and development of effector functions by T cells. The success of these strategies has been limited by difficulties in attaining sufficient transduction efficiencies and associated high levels of CD80 expression. We demonstrate that these problems can be circumvented by using anti-CD28 monoclonal antibody. Furthermore, we show that the synergistic relationship between CD80/CD28 pathway and interleukin 12 cytokine (IL-12), documented in the generation of cytotoxic T lymphocytes (CTL) for solid tumours, also applies to AML. CD28/IL-12 synergy facilitated the proliferation of allogeneic T cells in response to stimulation with primary AML blasts. The synergy also favoured generation of a Th1-type immune response, evidenced by gamma interferon (IFNg ) secretion and facilitated naive and memory T cell proliferation. Unlike some methods of in vitro T cell expansion, use of CD28/IL-12 synergy left T cells in the physiologically appropriate CD45RA -/CCR7 -subsets known to be associated with immediate cytotoxic functions.