Membrane-associated respiratory syncytial virus F protein expressed from ahuman rhinovirus type 14 vector is immunogenic

Human rhinovirus (HRV) replicons have the potential to serve as respiratory vaccine vectors for mucosal immunization in humans. However, since many va ccine immunogens of interest are glycosylated, an important concern is whet her HRV replicons are capable of expressing glycosylated proteins. The huma n respiratory syncytial virus (RSV) fusion (F) protein was chosen as a mode l glycoprotein and the HRV replicon Delta P1FVP3 was generated by inserting the F protein-coding sequence in frame and in lieu of the 5' proximal 1489 nucleotides of the capsid-coding segment in the HRV-14 genome. When transf ected into H1-HeLa cells, Delta P1FVP3 replicated and led to the expression of the F protein. Inhibition with guanidine demonstrated that F-protein ex pression was dependent on Delta P1FVP3 replication and did not result from translation of input RNA. Although most of the F protein remained as an imm ature, glycosylated precursor (F0), a readily detectable fraction of the pr otein was processed into the mature glycosylated subunit F1, an event known to occur within the Golgi apparatus. Packaged Delta P1FVP3 replicons were generated in transfected HeLa cells by coexpression of homologous HRV capsi d proteins using the vaccinia virus/T7 RNA polymerase hybrid system. Packag ed replicon RNAs were capable of infecting fresh cells, leading to accumula tion of the F protein as in RNA-transfected cells. Mice immunized with HeLa cell lysates containing F protein expressed from Delta P1FVP3 produced neu tralizing antibodies against RSV. These results indicate that an HRV-14 rep licon can express a foreign glycosylated protein, providing further support for the potential of HRV replicons as a Vaccine delivery system. (C) 2001 Academic Press.