Regulated expression of adenoviral vectors-based gene therapies: therapeutic expression of toxins and immune-modulators.

Abstract

Regulatable promoter systems allow gene expression to be tightly controlled in vivo. This is highly desirable for the development of safe, efficacious adenoviral vectors that can be used to treat human diseases in the clinic. Ideally, regulatable cassettes should have minimal gene expression in the "OFF" state, and expression should quickly reach therapeutic levels in the "ON" state. In addition, the components of regulatable cassettes should be non-toxic at physiological concentrations and should not be immunogenic, especially when treating chronic illness that requires long-lasting gene expression. In this chapter, we will describe in detail protocols to develop and validate first generation (Ad) and high-capacity adenoviral (HC-Ad) vectors that express therapeutic genes under the control of the TetON regulatable system. Our laboratory has successfully used these protocols to regulate the expression of marker genes, immune stimulatory genes, and toxins for cancer gene therapeutics, i.e., glioma that is a deadly form of brain cancer. We have shown that this third generation TetON regulatable system, incorporating a doxycycline (DOX)-sensitive rtTA(2)S-M2 inducer and tTS(Kid) silencer, is non-toxic, relatively non-immunogenic, and can tightly regulate reporter transgene expression downstream of a TRE promoter from adenoviral vectors in vitro and also in vivo.

• Authors: James F Curtin, Marianela Candolfi, Mariana Puntel, Weidong Xiong, A K M Muhammad, Kurt Kroeger, Sonali Mondkar, Chunyan Liu, Niyati Bondale, Pedro R Lowenstein, Maria G Castro
• Journal: Methods in molecular biology (Clifton, N.J.) (Methods Mol Biol) Vol. 434 Pg. 239-66 ( 2008) ISSN: 1064-3745 [Print] United States
• PMID: 18470649 (Publication Type: Journal Article, Research Support, N.I.H., Extramural, Research Support, Non-U.S. Gov't)

Chemical References

• Anti-Bacterial Agents
• Bacterial Toxins
• Diphtheria Toxin
• Exotoxins
• Virulence Factors
• ADP Ribose Transferases
• Pseudomonas aeruginosa exotoxin A
• Tetracycline
• Doxycycline

Topics

• ADP Ribose Transferases (genetics, metabolism)
• Adenoviridae (genetics)
• Anti-Bacterial Agents (therapeutic use)
• Bacterial Toxins (genetics, metabolism)
• Blotting, Southern
• Cells, Cultured
• Diphtheria Toxin (genetics, metabolism)
• Doxycycline (therapeutic use)
• Exotoxins (genetics, metabolism)
• Gene Expression Regulation
• Genetic Therapy (methods)
• Genetic Vectors (therapeutic use)
• Glioma (genetics, metabolism, therapy)
• Humans
• Tetracycline (therapeutic use)
• Transgenes (physiology)
• Virulence Factors (genetics, metabolism)