Polyoma and Papilloma Virus Vectors For Cancer Gene Therapy
The studies carried out to date using polyoma and papillomavirus based carriers demonstrate their considerable promise, although numerous aspects of the systems require further attention. Present vectors vary widely in efficacy of DNA transfer, and parame
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POLYOMA AND PAPILLOMA VIRUS VECTORS FOR CANCER GENE THERAPY Nina Krauzewicz and Beverly E. Griffin Department of Infectious Diseases (7th Floor) Imperial College School of Medicine Du Cane Road, London W12 ONN UK
1. INTRODUCTION Gene therapy is becoming increasingly accepted as a strategem for treating human disease. Over 60% of all therapies approved for clinical trials are directed towards cancer targets, the majority of which have used retroviral or adenoviral vectors to deliver genes to the target tissues (Verma and Somia, 1997; Anderson, 1998). Viral vectors are engineered to carry a gene expression cassette and only be capable of replication in a helper system. Results with these vectors have been encouraging, however, concerns for safety and adverse immunological responses in vivo may ultimately limit their use in the clinic. Lipid and colloid based non-viral vectors are chemically better defined than viral vectors, but lack the specificity required to target cancer cells (Cooper, 1996). The limitations of the currently available “vectorology” has led several groups to focus on developing alternative systems, which retain the targetting and cell uptake properties of viruses, whilst having the potential to be totally synthesised in vitro. One such approach uses viral like particles, or pseudocapsids.
2. PSEUDOVIRIONS AND PSEUDOCAPSIDS 2.1. Early Studies In the early 1970s, Aposhian and co-workers proposed that “empty” particles from the non-enveloped mouse virus, polyoma, could be co-assembled with heterologous DNA in a cell free system and used as carriers of genes into mammalian cells (Osterman et al., 1971; Qasba and Aposhian, 1971). In their experiments, capsids devoid of DNA and composed only of protein were isolated by density gradient centrifugation from infected Cancer Gene Therapy: Past Achievements and Future Challenges, edited by Habib Kluwer Academic/Plenum Publishers, New York, 2000.
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cells. These particles could be encouraged to create polyoma-like particles (PLPs), by incubating the mixture at low ionic strength with supercoiled polyoma viral DNA, and these were able to enter rat cells, resulting in viral protein expression (Slilaty and Aposhian, 1983). Subsequently, it was shown that, using the closely related simian polyomavirus, SV40, empty capsids could be dis-assembled under mild reducing conditions and reformed around foreign DNA (Colomar et al., 1993). These experiments demonstrated that: first, it is possible stably to interact DNA with viral coat protein assemblies; secondly, polyoma-like particles could be assembled from the basic subunits of the virus and DNA and, thirdly, these complexes could transfer DNA to cells in a biologically active form.
2.2. In Vitro Expression Systems A second line of research, developing in parallel with the reconstitution studies, centred around generating viral coat protein genes in in vitro expression systems to analyse coat protein functions and assembly pathways. Such studies included protein production in
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