Author Description

This dissertation, "Development of an Inter-kingdom Delivery and Expression System for Therapeutic Molecules Against Cancer" by Lei, Shi,, was obtained from The University of Hong Kong (Pokfulam, Hong Kong) and is being sold pursuant to Creative Commons: Attribution 3.0 Hong Kong License. The content of this dissertation has not been altered in any way. We have altered the formatting in order to facilitate the ease of printing and reading of the dissertation. All rights not granted by the above license are retained by the author. Abstract: Background: Several bacterial species such as Clostridium, E.coli and Salmonella can colonize within solid tumors and potentially be used as anticancer agents. The naturally occurring antitumor activity of Salmonella Typhimurium SL7207 strain can be further enhanced by genetic manipulations, making it a promising system for targeted delivery and expression of therapeutic molecules in cancer therapy. Methodology/Principal Findings: The engineered strain, termed ST1, was generated in the SL7207 background using the λ Red-mediated ''long homology arm'' recombineering technology. Firstly, T7 RNA polymerase (T7 RNAP) gene was integrated into the bacterial genome to enable the transcription of small RNA or exogenous gene. Secondly, an essential gene asd was replaced by a pore-forming listeriolysin O gene hlyA, which encodes a bacterial factor required for a maximal transfer of therapeutic factors in vivo. Subsequently, the asd gene with anaerobic control was cloned back at the htrA gene locus. The double mutation of asd and htrA made bacteria to grow only under anaerobic conditions while being disabled for growth in normal tissues. The final step was to relocate a small essential gene infA (coding for translation initiation factor 1) from chromosome to plasmid for stable plasmid maintenance. After genetic manipulations, a series of functional assays have been performed to test the safety and efficiency. In tumor-bearing