Immobilization of plasmid DNA in bacterial ghosts.

The development of novel delivery vehicles is crucial for the improvement of DNA vaccine efficiency. In this report, we describe a new platform technology, which is based on the immobilization of plasmid DNA in the cytoplasmic membrane of a bacterial carrier. This technology retains plasmid DNA (Self-Immobilizing Plasmid, pSIP) in the host envelope complex due to a specific protein/DNA interaction during and after protein E-mediated lysis. The resulting bacterial ghosts (empty bacterial envelopes) loaded with pDNA were analyzed in detail by real time PCR assays. We could verify that pSIP plasmids were retained in the pellets of lysed Escherichia coli cultures indicating that they are efficiently anchored in the inner membrane of bacterial ghosts. In contrast, a high percentage of control plasmids that lack essential features of the self-immobilization system were expelled in the culture broth during the lysis process. We believe that the combination of this plasmid immobilization procedure and the protein E-mediated lysis technology represents an efficient in vivo technique for the production of non-living DNA carrier vehicles. In conclusion, we present a "self-loading", non-living bacterial DNA delivery vector for vaccination endowed with intrinsic adjuvant properties of the Gram-negative bacterial cell envelope.

Bacterial ghosts--biological particles as delivery systems for antigens, nucleic acids and drugs.

Despite the exponential rate of discovery of new antigens and DNA vaccines resulting from modern molecular biology and proteomics, the lack of effective delivery technology is a major limiting factor in their application. The bacterial ghost system represents a platform technology for antigen, nucleic acid and drug delivery. Bacterial ghosts have significant advantages over other engineered biological delivery particles, owing to their intrinsic cellular and tissue tropic abilities, ease of production and the fact that they can be stored and processed without the need for refrigeration. These particles have found both veterinary and medical applications for the vaccination and treatment of tumors and various infectious diseases.