Studies Of Vaccines Against H5N1 Influenza Virus And Gene-based Drugs Against Hepatitis B Virus Delivered By Attenuated Salmonella

Attenuated strains of invasive enteric bacteria, such as Salmonella, represent promising gene delivery agents for nucleic acid-based vaccines as they can be administrated orally. In this study, we constructed a novel attenuated strain of Salmonella SL368 for the delivery and expression of the hemagglutinin (HA) and neuraminidase (NA) of a highly pathogenic H5N1 influenza virus, named Salmonella-based influenza vaccine Sal-HA-NA. We showed that the constructed Salmonella strain exhibited efficient gene transfer activity for HA and NA expression and little cytotoxicity and pathogenicity in mice. Using BALB/c mice as the model, we evaluated the immune responses and protection induced by Sal-HA-NA. Our study showed that Sal-HA-NA induced significant production of anti-HA serum IgG and mucosal IgA, and of anti-HA interferon-y producing T cells in orally vaccinated mice. Furthermore, mice orally vaccinated with the Salmonella vaccine expressing viral HA and NA proteins were completely protected from lethal challenge of highly pathogenic H5N1 as well as H1N1 influenza viruses while none of the animals treated with the Salmonella carrying the empty expression vector with no viral antigen expression was protected. These results suggest that Sal-HA-NA elicits strong antigen-specific humoral and cellular immune responses and provides effective immune protection against multiple strains of influenza viruses. Furthermore, our study demonstrates the feasibility of developing novel attenuated Salmonella strains as new oral vaccine vectors against influenza viruses.Liver-specific microRNA-122 has been shown to bind to a highly conserved HBV (Hepatitis B virus) pregenomic RNA sequence via base-pairing interactions and inhibit HBV gene expression and replication. However, the transient silencing effects of miR-122 and cytotoxic response it provokes in mammalian cells restricts its applications to the therapeutic potential. In this study, we introduced a pSuper basic vector that directs intracellular synthesis of siRNA-like transcripts for constructing pSB-miR-122 to express miR-122 sustainedly and prolong its specific suppression of HBV gene expression. Meanwhile, a novel attenuated Salmonella strain performed efficient delivery of constructed pSB-miR-122 plasmids into livers, leading to substantial expression of miR-122 with little adverse effects in mice. Moreover, suppressions of viral gene expression occurred in the HBV-infected mice that were treated with Salmonella carrying miR-122 expressing vector. These results indicate that Salmonella-mediated gene target therapy with miR-122 expressing vector could inhibit HBV gene expression in vivo.