A new form of particulate single and multiple immunogen delivery system based on recombinant bluetongue virus-derived tubules.

The development of particular vector systems for the presentation of immunogenic epitopes provides a powerful approach for the delivery of antigens. These include the core-like particles formed by recombinant bluetongue virus (BTV) capsid proteins VP3 and VP7 synthesized in insect cells by recombinant baculoviruses. Previously we have reported localization of an antigenic site on the surface of tubular structures formed by the nonstructural protein NS1 of BTV, and their potential use for epitope presentation. In this study foreign sequences ranging form 44 to 116 aa in length and representing 44 aa sequence from Clostridium difficile toxin A, 48 aa of the hepatitis B virus preS2 region, and the whole of bovine leukemia virus p15 protein were inserted at the C-terminus of BTV-10 NS1. The chimeric NS1 genes were expressed using recombinant baculoviruses and the ability of the mutated NS1 proteins to form tubules was investigated. All chimeric constructs formed tubular structures which carried the foreign antigenic sequences exposed on the surface of the tubules and were highly immunogenic. When Sf cells were coinfected with three recombinant baculoviruses expressing chimeric NS1 proteins with different epitopes, their simultaneous assembly into the same tubule was demonstrated. This observation opens up the possibility of using recombinant NS1 tubules as carriers for the delivery of multiple epitopes.