The long term goal of this project is to characterize the effect of tumor gangliosides on the biology of human neuroblastoma. This goal is based upon our hypothesis that specific gangliosides are shed by tumors, act as intercellular signaling molecules, and protect tumor cells from host destruction. The foundation for studies to conclusively characterize the mechanism and consequences of tumor gangliosides in the biology of human neuroblastoma has been laid by (i) significant shedding and potent immunosuppressive activity of human neuroblastoma tumor gangliosides, (ii) inhibition of murine anti-tumor immune responses both in vivo and in vitro by these molecules, (iii) identification of antigen presenting cells as a primary tumor ganglioside targets, and (iv) preliminary data demonstrating ganglioside-induced modulation of both dendritic cell nuclear translocation of the transcription factor NF-kappaB and cytokine release. To accomplish these goals, using purified neuroblastoma and other tumor gangliosides with molecular structures confirmed by mass spectrometry, we will elucidate the transfer of gangliosides to human dendritic cells in a novel model of the tumor cell microenvironment, to link quantitatively the release (shedding) and the binding of purified tumor gangliosides to immature dendritic cells, and then to establish the biological effect of binding on cell function, including cytokine production. We will identify the cellular/molecular effects of shed tumor gangliosides on dendritic cell function and its consequences, by defining ganglioside effects on (i) initial lipid raft-associated CD154/CD40 signaling, (ii) NF-kappaB nuclear translocation, and (iii) tumor antigen-induced cytokine production and helper TH1/TH2 cell generation. Finally, to determine the biological effect of tumor gangliosides on antigen presenting cell function in vivo, we will test ganglioside effects on dendritic cell maturation and function, and helper T cell balance in vivo, using the syngeneic FBL lymphoma tumor model we have developed. By the combination of a powerful approach to ganglioside isolation, and novel in vitro and in vivo model systems to study shed gangliosides, we will directly test the hypothesis that gangliosides shed by human neuroblastoma significantly downregulate the host responses to tumor cells, in vitro and in vivo. The results obtained may identify new molecular and cellular targets for intervention in the process of ganglioside modulation of host responses to tumor, and contribute towards our longer term objective of fully elucidating the biological significance of human tumor gangliosides and their impact on human tumor pathogenesis.
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