Vaccinia virus vectors

Recombinant vaccines Vaccines that can deliver and penetrate intestinal wall via M cells Tetanus toxin, Salmonella strains, vaccinia virus vector... 

Ward GA, Stover CK, Moss B, Fuerst TR (1995), Stringent chemical and thermal regulation of recombinant gene expression by vaccinia virus vectors in mammalian cells, Proc. Natl Acad. Sci. U S A 92 6773-6777. 

Moss, B. (1992). Vaccinia virus vectors. In Vaccines New Approaches to Immunological Problems. R.W.Ellis, ed. (Boston Butterworth-Heinemann), pp. 345 362. 

Leffers, H. (1994). Expression of recombinant proteins using vaccinia virus vectors. In Cell Biology A Laboratory Handbook, Vol. 3 (Celis, J.E., ed.), Academic Press, San Diego, pp. 155-163. 

Edwards, R.H., Selby, M.J., Mobley, W.C., Weinrich, S.L., Hruby, D.E. and Rutter, W.J. (1988b) Processing and secretion of nerve growth factor expression in mammalian cells with a vaccinia virus vector. Mol. Cell. Biol. 8 2456-2464. 

Figure 4.4. Schematic illustration of directional topoisomerase cloning of PCR products into the pUNI vector. The PCR product to be cloned has the sequence 5 -CACC appended at the 5 end to direct the orientation of cloning. The Vaccinia virus topoisomerase I enzyme forms a covalent adduct with the cloning vector to create a cloning competent plasmid construct. The loxP site is 5 to the insertion site. The vector and PCR product are designed to fuse the ORF in-frame with loxP.

A number of factors render vaccinia virus a particularly attractive vector system. These include ... 

Introduction of a gene of interest into the host cell line by viral infection is a convenient method since a large number of cells can be infected simultaneously. Systems employing Semliki Forest Virus, Vaccinia Virus, and Retoviral vectors are used. However, drawbacks include the requirement for special precautions when engineering and preparing the viral... 

Early animal experiments have underlined the potential of vaccinia-based vector vaccines. Vaccinia virus-housing genes from HIV have clearly been found to elicit both humoral and cell-mediated immune responses in monkeys. Similar responses in other animals have been reported when surface polypeptides from a variety of additional pathogens have been expressed in recombinant vaccinia systems (Table 10.16). Human clinical trials are now in progress. 

Hota-Mitchell, S., Clarke, M.W., Podesta, R.B. and Dekaban, C.A. (1 999) Recombinant vaccinia viruses and gene gun vectors expressing the large subunit of Schistosoma mansoni calpain used in a murine immunization-challenge model. Vaccine 1 7, 1 338-1354. 

Carroll MW, Kovacs GR (2003), Virus-based vectors for gene expression in mammalian cells Vaccinia virus, In Makrides SC (Ed.), Gene Transfer and Expression in Mammalian Cells, Elsevier Science BV, Amsterdam, pp. 125-136. 

Mackett M, Smith GL, Moss B (1982), Vaccinia virus a selectable eukaryotic cloning and expression vector, Proc. Natl Acad. Sci. USA 79 7415-7419. 

This is not to say that cardiotoxicity is not seen with biopharmaceuticals. Cardiomyopathy is now a well-recognized complication of trastuzumab and and has been reported with bevacizumab treatment, in particular in combination with other cytotoxic cancer therapies [20]. Myocarditis and pericarditis are a well-documented complications of vaccinia immunization [21], and could also complicate use of a pox-virus vector for other therapeutics. In 1995 Genetics Institute suspended phase 2 cancer trials of Interleukin-12 for serious tox-icities including cardiac arrhythmia. However, such toxicities are best detected by incorporation of biomarkers for myocardial damage such as troponin-T into preclinical and early clinical studies, and continual ECG monitoring for arrhythmia in preclinical and early clinical studies, not by in vitro explorations of electrophysiology. 

Investigators are currently assessing the potential safety and efficacy of a mixed modality treatment regimen (i.e., combining a DNA vaccine with a viral vector). In this example preclinical toxicity and biodistribution studies were conducted in mice using a DNA vaccine in combination with a modified vaccinia virus Ankara based HIV vaccine to support a phase 1 trial in healthy HIV-1-uninfected volunteers [77,78],... 

In this example the DNA vaccine (pTHr-HIVA) is based on a novel direct gene transfer vector pTH [77], and the second component, MVA-HIVA, is based on a modified vaccinia virus Ankara. Both of these components contain most of the HIV-1 clade A gag protein coupled to conserved HIV-1 clade A CTL epitopes arranged in a polypeptide string that served as the immunogen [77]. 

Two other viral vectors are extensively used. Vaccinia virus, a large DNA-containing virus, replicates in the cytoplasm of mammalian cells, where it shuts dovm host-cell protein synthesis. Baculovirus infects insect cells, which can be conveniently cultured. Insect larvae infected with this virus can serve as efficient protein factories. Vectors based on these large-genome viruses have been engineered to express DNA inserts efficiently. 

The key function of the vector is to introduce the cDNA under the control of a strong promoter and, if a stable cell line is to be developed, also to introduce resistance to a compound that is otherwise toxic to the host cell. This facilitates selection (only vector-bearing cells grown in special media) of a minority of cells that have incorporated the vector. There are many adequate expression vectors available from a number of commercial suppliers. Several classes of vector systems appear appropriate for transporter expression as integrating, episomal, retroviral, Vaccinia virus-, and Adenovirus-based systems have all been used successfully in studies referenced in this chapter. 

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