title: Combined prokaryotic-eukaryotic delivery and expression of therapeutic factors through a primed autocatalytic positive-feedback loop.
abstract: Progress in bacterial therapy for cancer and infectious diseases is hampered by the absence of safe and efficient vectors. Sustained delivery and high gene expression levels are critical for the therapeutic efficacy. Here we developed a Salmonella typhimrium strain to maintain and safely deliver a plasmid vector to target tissues. This vector is designed to allow dual transcription of therapeutic factors, such as cytotoxic proteins, short hairpin RNAs or combinations, in the nucleus or cytoplasm of eukaryotic cells, with this expression sustained by an autocatalytic positive-feedback loop. Mechanisms to prime the system and maintain the plasmid in the bacterium are also provided. Synergistic effects of attenuated Salmonella and our inter-kingdom system allow the precise expression of Diphtheria toxin A chain (DTA) gene in tumor microenvironment and eradicate large established tumors in immunocompetent animals. In the experiments reported here, 26% of mice (n=5/19) with aggressive tumors were cured and the others all survived until the end of the experiment. We also demonstrated that ST4 packaged with shRNA-encoding plasmids has sustained knockdown effects in nude mice bearing human MDA-MB-231 xenografts. Three weeks after injection of 5×10(6) ST4/pIKT-shPlk, PLK1 transcript levels in tumors were 62.5±18.6% lower than the vector control group (P=0.015). The presence of PLK1 5' RACE-PCR cleavage products confirmed a sustained RNAi-mediated mechanism of action. This innovative technology provides an effective and versatile vehicle for efficient inter-kingdom gene delivery that can be applied to cancer therapy and other purposes.
authors.affiliation: School of Biomedical Sciences, The University of Hong Kong, Pokfulam, 999077, Hong Kong; Shenzhen Institute of Research and Innovation, The University of Hong Kong, Pokfulam, 999077, Hong Kong.
authors.forename: Lei
authors.initials: L
authors.lastname: Shi
authors.affiliation: School of Biomedical Sciences, The University of Hong Kong, Pokfulam, 999077, Hong Kong; Shenzhen Institute of Research and Innovation, The University of Hong Kong, Pokfulam, 999077, Hong Kong; Key Laboratory of Optoelectronic Devices and Systems of Ministry of Education and Guangdong Province, College of Optoelectronic Engineering, Shenzhen University, Shenzhen 518060, PR China.
authors.forename: Bin
authors.initials: B
authors.lastname: Yu
authors.affiliation: Department of Obstetrics and Gynaecology, The University of Hong Kong, Pokfulam, 999077, Hong Kong; Advanced Institute of Translational Medicine, Tongji University School of Medicine, Shanghai 200092, PR China.
authors.forename: Chun-Hui
authors.initials: CH
authors.lastname: Cai
authors.affiliation: Faculty of Biology, South University of Science and Technology of China, Shenzhen 518055, PR China.
authors.forename: Wei
authors.initials: W
authors.lastname: Huang
authors.affiliation: Department of Microbiology, The University of Hong Kong, Pokfulam, 999077, Hong Kong.
authors.forename: Bo-Jian
authors.initials: BJ
authors.lastname: Zheng
authors.affiliation: School of Public Health, The University of Hong Kong, Pokfulam, 999077, Hong Kong.
authors.forename: David Keith
authors.initials: DK
authors.lastname: Smith
authors.affiliation: School of Biomedical Sciences, The University of Hong Kong, Pokfulam, 999077, Hong Kong; Shenzhen Institute of Research and Innovation, The University of Hong Kong, Pokfulam, 999077, Hong Kong; The Centre for Synthetic Biology Engineering Research, Shenzhen Institutes of Advanced Technology, Shenzhen 518055, PR China. Electronic address: jdhuang@hku.hk.
authors.forename: Jian-Dong
authors.initials: JD
authors.lastname: Huang
mesh_terms: D000818:Animals; D018797:Cell Cycle Proteins / Q000235:genetics; D045744:Cell Line, Tumor; D004167:Diphtheria Toxin / Q000235:genetics; D025461:Feedback, Physiological; D005260:Female; D015316:Genetic Therapy / Q000379:methods*; D005822:Genetic Vectors; D006801:Humans; D051379:Mice; D008807:Mice, Inbred BALB C; D008819:Mice, Nude; D009369:Neoplasms / Q000628:therapy; D010446:Peptide Fragments / Q000235:genetics; D010957:Plasmids; D017346:Protein Serine-Threonine Kinases / Q000235:genetics; D011518:Proto-Oncogene Proteins / Q000235:genetics; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000235:genetics; D012486:Salmonella typhimurium / Q000235:genetics*; D000097869:Polo-Like Kinase 1
chemical_list: D018797:Cell Cycle Proteins; D004167:Diphtheria Toxin; D010446:Peptide Fragments; D017346:Protein Serine-Threonine Kinases; D011518:Proto-Oncogene Proteins; D034741:RNA, Small Interfering; D000097869:Polo-Like Kinase 1; C029795:diphtheria toxin fragment A
keywords: Bacterial therapy; Inter-kingdom delivery and dual expression (IKDE); Metastasis; Positive-feedback loop; Tumor-targeting Salmonella