USE OF RECOMBINANT POXVIRUSES TO STIMULATE ANTIMELANOMA T-CELL REACTIVITY

Background: Dendritic cells (DC) an potent professional antigen-presen ting cells that can activate naive T lymphocytes and initiate cellular immune responses, As adjuvants, BC may be useful for enhancing immuno genicity and mediating tumor regression. Endogenous expression of anti gen by DC could offer the potential advantage of allowing prolonged co nstitutive presentation of endogenously processed epitopes and exploit ation of multiple restriction elements for the presentation of the sam e antigen. Methods: DC were prepared from the peripheral blood of HLA A0201 patients with metastatic melanoma in the presence of IL-4 (1000 IU/mL) and GMCSF (1000 IU/mL). Recombinant vaccinia and fowlpox virus es encoding the hMART-1 gene were constructed and used to infect DC, T he efficiency of infection and expression of the MART-1 antigen were a ssessed by immunohistochemistry and intracellular FAGS analyses. Cytot oxic lymphocytes (CTL) were generated by the stimulation of CD8+ T cel ls, with DC expressing the recombinant gene. Reactivity of the CTL was determined at weeks 1 and 2 by the amount of IFN-gamma released. Resu lts: DC were infected with recombinant poxviruses and demonstrated spe cific melanoma antigen expression by immunohistochemistry, immunofluor escence, and intracellular FAGS analysis. The expression by DC of MART -1 MAA after viral infection was sufficient to generate CD8+ T lymphoc ytes that recognized naturally processed epitopes on tumor cells in 10 of 11 patients. Conclusions: Human DC are receptive to infection by r ecombinant poxviruses encoding MAA genes and are capable of efficientl y processing and presenting these MAA to cytotoxic T cells. The potent ial advantage of this approach is the ability to present specific anti gen independent of the identification of the epitope or the MHC restri ction element, This strategy may be useful for the identification of r elevant epitopes for a diverse number of HLA alleles and for active im munization in patients.