Characterizing the Effects of a Fusion Binding Protein [E7/FLT3-Ligand] on the Induction and Maturation of CD11C+ Dendritic Cells

Current tumor immunotherapy research investigates fusion protein vaccines for bolstering tumor prevention. In this research, a fusion binding protein [E7/ Flt3-Ligand] and its effect on CD11c+ dendritic cell (DC) maturation were studied. This protein was constructed with E7, the epitope inducer of Human Papillomavirus oncoprotein, and Flt3-Ligand (Flt3-L), a hematopoietic catalyst, which upon binding induces DC maturation, heightening immune response. This research aimed to determine whether a) E7/Flt3-L binds to CD11c+ DCs and induces maturation under 18 and 72 hour incubations at 37°C using cell culture and flow cytometry to calculate surface and internal fluorescent antibody levels and b) to determine the specificity of E7/Flt3-L by using cell culture, conducting the competition assays on ice under 20 and 60 minute incubations of the DCs with Flt3-L and E7/Flt3-L, and measuring with flow cytometry. The 18-hour incubation displayed higher levels of fluorescence, indicating binding, and lower levels of maturation markers CD80+/CD83+, indicating lower levels of maturation. In contrast, the 72-hour incubation resulted in higher levels of maturation markers CD80+/CD83+, indicating internalization, and lower levels of E7/Flt3-L on the DCs, indicating E7/Flt3-L internalization and processing. The competitive assays indicated that E7/Flt3-L was specific for the Flt3 receptors. In conclusion, the studies displayed that E7/Flt3-L binds to CD11c+ DCs, binds specifically to Flt3 receptors on the CD11c+ DCs and induces CD11c+ DC maturation. Future experiments could study E7/Flt3-L endocytic rates and antigen presentation to activate T-cells